Objective To discuss the construction of a NOD-signal regulatory protein α （SIRPA） gene knock-in mouse model based on clustered regularly interspaced short palindromic repeats （CRISPR）/CRISPR-associated protein （Cas） 9 technology， and to provide the reference for the establishment of more advanced humanized mouse models. Methods Based on CRISPR/Cas9 technology， the NOD-background SIRPA gene was knocked into the BALB/c mouse fertilized eggs， and homozygous mice with stable genotypes （SIRPA-KI mice） were obtained through expansion and breeding for experiments； PCR primers were designed， and mouse genotypes were identified by nucleic acid electrophoresis. The mice were divided into C57BL/6 group， BALB/c group， and SIRPA-KI group according to their strains， and 3 mice with similar ages were randomly selected from each group for experiments. Mature bone marrow-derived macrophages were co-incubated with human CD47-Fc fusion protein， stained with Streptavidin PE/Cy7， and the mean fluorescence intensity （MFI） of PE/Cy7 was detected by flow cytometry to compare the ability of SIRPA in the mice from various groups to bind human CD47 Fc. Each mouse was intravenously injected with 2×10? carboxyfluorescein diacetate， succinimidyl ester （CFSE）-labeled human red blood cells， and the phagocytosis of human red blood cells by macrophages in various groups was detected by in vivo phagocytosis assay. One BALB/c mouse and one SIRPA-KI mouse were randomly selected to induce mature bone marrow-derived macrophages， and the phagocytosis of human red blood cells by macrophages in various groups was detected by in vitro phagocytosis assay. Results BLAB/c SIRPA^NOD/NOD homozygous mice were successfully obtained. The flow cytometry results showed that compared with C57BL/6 group， the MFI of the mice in BALB/c group was significantly increased （P<0.01）； compared with C57BL/6 group and BALB/c group， the MFI of the mice in SIRPA-KI group was significantly increased （P<0.01）. The in vivo phagocytosis assay results showed that the macrophages in C57BL/6 group exhibited the fastest overall clearance rate of human red blood cells； at 6 h of macrophage phagocytosis， compared with SIRPA-KI group， the residual percentage of the human red blood cells in C57BL/6 group was significantly decreased （P<0.01） and was closed to 0%； compared with SIRPA-KI group， the residual percentage of the human red blood cells in BALB/c group was significantly decreased （P<0.05）. The in vitro phagocytosis assay results showed that the macrophages in BALB/c group significantly phagocytosed the human red blood cells， with a high phagocytic index of 30.7%； compared with BALB/c group， the phagocytic index of the macrophages in SIRPA-KI group was significantly decreased （P<0.01）. Conclusion The study successfully constructs a mouse model with enhanced affinity for human CD47 and significantly inhibited phagocytosis of human red blood cells by knocking the NOD-background SIRPA gene into the BALB/c strain using CRISPR/Cas9 technology， providing a superior human cell xenotransplantation efficiency.

Objective To discuss the role of nuclear factor of activated T-cells 5 （NFAT5） inhibitor KRN5 in high salt-induced senescence of mouse vascular smooth muscle cells （VSMCs）， and to clarify its mechanism. Methods Thirty 8-week-old male ApoE^-/- mice were divided into normal group， senescence group and high-salt treatment senecence group， with 10 mice in each group； the mice in senescence group and high-salt treatment senecence group were used to establish natural senecence mouse models； the mouse VSMCs were isolated and cultured， and divided into normal group， senescence group， high-salt treatment senecence group and high-salt treatment senecence+KRN5 group. β-galactosidase （Sa-β-gal） staining was used to detect the senescence of aortic tissues and VSMCs in various groups； immunofluorescence method was used to detect the expressions of NFAT5 and phosphorylated histone H2A variant X （γ-H2AX） proteins in mouse aortic tissues and VSMCs in various groups； real-time fluorescence quantitative PCR （RT-qPCR） method was used to detect the mRNA expression levels of NFAT5， γ-H2AX， cyclin-dependent kinase inhibitor 2A （P16） and cyclin-dependent kinase inhibitor 1A （P21） in the cells in various groups； Western blotting method was used to detect the protein expression levels of NFAT5， γ-H2AX， P16 and P21 in VSMCs in various groups. Results The Sa-β-gal staining results showed that compared with normal group， the proportions of senescence-positive area in aortic tissues of the mice in senescence group and high-salt treatment senecence group were significantly increased （P<0.05）， and the proportion of senescence-positive cells in the VSMCs of the mice was significantly increased （P<0.05）； compared with senecence group， the proportion of senescence-positive cells in the VSMCs mice in high-salt treatment senecence group was significantly increased （P<0.05）； compared with high-salt treatment senecence group， the proportion of senescence-positive cells in the VSMCs of the mice in high-salt treatment senecence+KRN5 group was significantly decreased （P<0.01）. The immunofluorescence results showed that compared with normal group， the expression level of γ-H2AX protein in mouse VSMCs of the mice in senescence group was significantly increased （P<0.05）； compared with senescence group， the expression levels of SA-β-gal staining and NFAT5 protein in aortic tissue of the mice in high-salt treatment senecence group were significantly increased （P<0.05）； compared with normal group， the expression level of NFAT5 protein in the VSMCs of the mice in senecence group and high-salt treatment senecence group was significantly increased （P<0.05）； compared with senecence group， the expression level of NFAT5 protein in the VSMCs of the mice in high-salt treatment senecence group was significantly increased （P<0.05）. The RT-qPCR results showed that compared with normal group， the expression levels of NFAT5， γ-H2AX， P16， and P21 mRNA in the VSMCs of the mice in senescence group and high-salt treatment senecence group were significantly increased （P<0.05）； compared with senecence group， the mRNA expression levels of NFAT5， γ-H2AX， P16， and P21 mRNA in the VSMCs of the mice in high-salt treatment senecence group were significantly increased （P<0.05）； compared with senecence group， the expression levels of NFAT5， γ-H2AX， P16， and P21 mRNA in the VSMCs of the mice in high-salt treatment senecence group and high-salt treatment senecence+KRN5 group were significantly increased （P<0.05）； compared with high-salt treatment senecence group， the mRNA expression levels of NFAT5， γ-H2AX， P16， and P21 in the VSMCs of the mice in high-salt treatment senecence+KRN5 group were significantly decreased （P<0.05）. The Western blotting results showed that compared with normal group， the expression levels of NFAT5， γ-H2AX， P16， and P21 proteins in the VSMCs of the mice in senescence group and high-salt treatment senecence group were significantly increased （P<0.05）； compared with senescence group， the expression levels of NFAT5， γ-H2AX， P16， and P21 proteins in the VSMCs of the mice in high-salt treatment senecence group were significantly increased （P<0.05）； compared with senecence group， the expression levels of NFAT5， γ-H2AX， P16， and P21 proteins in the VSMCs of the mice in high-salt treatment senecence group and high-salt treatment senecence+KRN5 group were significantly increased （P<0.05）； compared with high-salt treatment senecence group， the expression levels of NFAT5， γ-H2AX， P16， and P21 proteins in the VSMCs of the mice in high-salt treatment senecence+KRN5 group were significantly decreased （P<0.05）. Conclusion NFAT5 may play a promoting role in high salt-induced senescence of the mouse VSMCs.

Objective To discuss the enhancing effect of cordycepin on ferroptosis inducer RSL3-induced ferroptosis in the hepatocellular carcinoma HepG2 cells， and to clarify its potential mechanism. Methods The HepG2 cells were divided into control group， RSL3 group， low， medium and high doses of cordycepin groups， RSL3+low， medium and high doses of cordycepin groups， RSL3+medium-dose cordycepin+ferroptosis inhibitor Ferrostatin-1 （Fer-1） group， and RSL3+medium-dose cordycepin+ferroptosis inhibitor Liproxstatin-1 （Lip-1） group. The HepG2， Huh-7 and HCCLM3 cells were treated with 0， 1， 5， 10， 15 and 20 μmol·L^-1 RSL3 for 24， 48 and 72 h， respectively. Cell counting kit-8（CCK-8） method was used to detect cell viability and determine the optimal concentration and treatment time of RSL3. The HepG2 cells were treated with 0， 50， 100， 200， 400， 600， 800， 1 000， and 1 200 μmol·L^-1 cordycepin for 24， 48 and 72 h， respectively. CCK-8 method was used to detect the survival rate of the cells and the half maximal inhibitory concentration （IC₅₀） was calculated to determine the optimal concentration and treatment time of cordycepin； the apoptosis inhibitor Z-VAD-FMK， autophagy inhibitor Chloroquine （CQ）， necroptosis inhibitor Necrostatin-1 （Nec-1）， Fer-1， Lip-1， Deferasirox and 2，2，6，6-tetramethylpiper idinoxy（TEMPO） were used to treat HepG2 cells， and the survival rate of the cells was calculated； 2'，7'-Dichlorodihydrofluorescein diacetate （DCFH-DA） fluorescence probe was used to detect reactive oxygen species （ROS） levels in the HepG2 cells in various groups； C11 BODIPY 581/591 fluorescence probe was used to detect lipid peroxidation （LPO） levels in the HepG2 cells in various groups； FeRhoNox-1 fluorescent probe was used to detect ferrous ion （Fe²⁺） levels in the HepG2 cells in various groups； kits were used to detect glutathione （GSH） and malondialdehyde （MDA） levels in the HepG2 cells； Western blotting method was used to detect the expression levels of ferroptosis-related proteins， nuclear factor erythroid 2-related factor 2 （Nrf2） and heme oxygenase-1 （HO-1） proteins in the hepG2 cells in various groups； transmission electron microscope was used to observe the ultrastructural morphology of the HepG2 cells in various groups. Results The CCK-8 results showed that when the cells were treated with 0.56 μmol·L?1 RSL3， the viabilities of the three cell types differed significantly. Compared with 0 μmol·L?1 RSL3 group， the survival rates of the cells in 6.4 and 12.8 μmol·L^-1 RSL3 groups were significantly decreased （P<0.05）. The HepG2 cells had the highest IC₅₀ value and were selected for subsequent experiments. Compared with 0 μmol·L^-1 cordycepin group， the survival rates of the HepG2 cells in 200， 400， 600， 800， 1 000， and 2 000 μmol·L^-1 cordycepin groups were significantly decreased （P<0.05 or P<0.01）. 0.5×IC₅₀ （267.9 μmol·L^-1）， 1×IC₅₀ （535.8 μmol·L^-1） and 1.5×IC₅₀ （803.7 μmol·L^-1） were selected as low， medium and high doses of cordycepin groups， respectively， with an intervention time of 24 h. Compared with control group， the survival rates of the HepG2 cells in low， medium， and high doses of cordycepin groups were significantly decreased （P<0.05）. Compared with low， medium， and high doses of cordycepin groups， the survival rates of the HepG2 cells in Z-VAD-FMK+low， medium， and high doses of cordycepin groups were significantly increased （P<0.01）， and those in Fer-1+medium and high doses of cordycepin groups and Lip-1+low， medium， and high doses of cordycepin groups were significantly increased （P<0.05）. Compared with control group， the survival rates of the HepG2 cells in RSL3 group， RSL3+low， medium， and high doses of cordycepin groups， RSL3+cordycepin+Fer-1 group and RSL3+cordycepin+Lip-1 group were significantly decreased （P<0.05 or P<0.01）. Compared with RSL3 group， the survival rates of the HepG2 cells in RSL3+low， medium， and high doses of cordycepin groups were significantly decreased （P<0.05）. The DCFH-DA results showed that compared with control group， the ROS levels in the cells in medium and high doses of cordycepin groups， RSL3 group and RSL3+low， medium， and high doses of cordycepin groups were significantly increased （P<0.05 or P<0.01）. The C11 BODIPY 581/591 results showed that compared with control group， the LPO levels in the HepG2 cells in medium and high doses of cordycepin groups， RSL3 group and RSL3+low， medium and high doses of cordycepin groups were significantly increased （P<0.05 or P<0.01）. Compared with RSL3 group， the LPO levels in the HepG2 cells in RSL3+low， medium， and high doses of cordycepin groups were significantly increased （P<0.05 or P<0.01）. The FeRhoNox-1 results showed that compared with control group， the Fe²⁺ levels in the HepG2 cells in medium and high doses of cordycepin groups， RSL3 group and RSL3+low， medium， and high doses of cordycepin groups were significantly increased （P<0.05 or P<0.01）. Compared with RSL3 group， the Fe²⁺ levels in the HepG2 cells in RSL3+low， medium， and high doses of cordycepin groups were significantly increased （P<0.05 or P<0.01）. Compared with control group， the MDA levels in the HepG2 cells in high doses of cordycepin group， RSL3 group and RSL3+low， medium， and high doses of cordycepin groups were significantly increased （P<0.05 or P<0.01）. Compared with RSL3 group， the MDA levels in the HepG2 cells in RSL3+low， medium， and high doses of cordycepin groups were significantly increased （P<0.05 or P<0.01）. Compared with control group， the GSH levels in the HepG2 cells in medium and high doses of cordycepin groups， RSL3 group and RSL3+low， medium， and high doses of cordycepin groups were significantly decreased （P<0.05 or P<0.01）. Compared with RSL3 group， the GSH levels in the HepG2 cells in RSL3+low， medium， and high doses cordycepin groups were significantly decreased （P<0.05 or P<0.01）. Compared with control group， the ultrastructure of the HepG2 cells in low and medium doses of cordycepin groups showed no significant changes， while the cells in high dose of cordycepin group exhibited reduced mitochondrial cristae， mild swelling and increased membrane density， with slightly distorted inner membrane structure. The cells in RSL3 group and RSL3+low， medium， and high doses of cordycepin groups all showed ultrastructural changes characteristic of ferroptosis. Compared with RSL3 group， the cells in RSL3+low， medium， and high doses of cordycepin groups exhibited ruptured mitochondrial membranes with increased membrane density， abnormally twisted or expanded inner membrane structures， and reduced or even disappeared mitochondrial cristae. The Western blotting results showed that compared with control group， the expression levels of FTH1 and GPX4 proteins in the HepG2 cells in medium and high doses of cordycepin groups were significantly decreased （P<0.05 or P<0.01）， while the expression levels of Nrf2 and HO-1 proteins were significantly decreased （P<0.05 or P<0.01）. Compared with control group， the expression levels of GPX4 protein in the HepG2 cells in low， medium and high doses of cordycepin groups， RSL3 group， and RSL3+cordycepin+Fer-1 group were significantly decreased （P<0.05）. Compared with RSL3 group， the expression levels of GPX4 protein in the HepG2 cells in RSL3+low， medium， and high doses of cordycepin groups were significantly decreased （P<0.05）. Conclusion Cordycepin can significantly enhance RSL3-induced ferroptosis in the hepatocellular carcinoma HepG2 cells and down-regulate the expression of Nrf2 and HO-1 proteins in the HepG2 cells.

Objective To discuss the effect of sericin on the phosphatidylinositol 3-kinase （PI3K）/protein kinase B （Akt）/nuclear factor-κB （NF-κB） signaling pathway and apoptosis in the streptozotocin （STZ）-damaged INS-1 cells， and to clarify its mechanism. Methods The INS-1 cells were cultured with complete medium containing 0， 0.1， 0.3， 1.0， 3.0， and 10.0 μmol·L^-1 Akt1 inhibitor A-674563， 10 mmol·L^-1 STZ， and 600 mg·L^-1 sericin， and divided into 0， 0.1， 0.3， 1.0， 3.0， and 10.0 μmol·L^-1 A-674563 groups， and the control group （complete medium without drugs） was set up. Cell counting kit-8 （CCK-8） method was used to detect the survival rates of the INS-1 cells， and the half-maximal inhibitory concentration （IC₅₀） value was calculated to determine the optimal inhibitory concentration of A-674563， which was further verified by Western blotting method. The INS-1 cells were divided into normal control group （complete medium）， model group （10 mmol·L^-1 STZ+complete medium）， and low， medium， and high doses of sericin groups （10 mmol·L^-1 STZ+150 mg·L^-1 sericin+complete medium， 10 mmol·L^-1 STZ+300 mg·L^-1 sericin+complete medium， and 10 mmol·L^-1 STZ+600 mg·L^-1 sericin+complete medium）. CCK-8 method was used to detect the survival rates of the INS-1 cells in various groups to determine the optimal concentration of sericin. Additionally， the INS-1 cells were divided into normal control group （complete medium）， model group （10 mmol·L^-1 STZ+complete medium）， sericin group （10 mmol·L^-1 STZ+600 mg·L^-1 sericin + complete medium）， and A-674563 group （10 mmol·L^-1 STZ+600 mg·L^-1 sericin+0.3 μmol·L^-1 A-674563+complete medium）. Flow cytometry was used to detect the apoptotic rates of the INS-1 cells in various groups； real-time fluorescence quantitative PCR （RT-qPCR ） method was used to detect the expression levels of Akt1， NF-κB， tumor necrosis factor-α （TNF-α）， and interleukin-6 （IL-6） mRNA in the INS-1 cells in various groups； Western blotting method was used to detect the expression levels of phosphorylated Akt1 （p-Akt1） and NF-κB proteins in the INS-1 cells in various groups； enzyme linked immunosorbent assay（ELISA） method was used to detect the levels of TNF-α and IL-6 in the INS-1 cells in various groups. Results The survival rates of the INS-1 cells in control group was 100.00%±0.00%； in 0， 0.1， 0.3， 1.0， 3.0， and 10.0 μmol·L^-1 A-674563+10 mmol·L^-1 STZ+600 mg·L^-1 sericin+complete medium groups， which were 82.50%±2.28%， 69.47%±1.94%， 51.51%±1.74%， 38.94%±1.57%， 24.79%±1.14%， and 19.85%±1.03%， respectively. The IC?? value of A-674563 for INS-1 cells was 0.3 μmol·L^-1， and 0.3 μmol·L^-1 A-674563 was selected for subsequent experiments. Compared with 0 μmol·L^-1 A-674563， the expression level of p-Akt1 protein in the INS-1 cells after treated with 0.3 μmol·L^-1 A-674563+10 mmol·L^-1 STZ + 600 mg·L^-1 sericin+complete medium was significantly decreased （P<0.05）. The CCK-8 results showed that compared with normal control group， the survival rate of the INS-1 cells in model group was significantly decreased （P<0.05）； compared with model group， the survival rates of the INS-1 cells in low， medium， and high doses of sericin groups were significantly increased （P<0.05）； compared with low and medium doses of sericin groups， the survival rate of the INS-1 cells in high dose of sericin group was significantly increased （P<0.05）. Thus， 600 mg·L^-1 sericin was selected for subsequent experiments. The CCK-8 results showed that compared with normal control group， the survival rate of the INS-1 cells in model group was significantly decreased （P<0.05）； compared with model group， the survival rate of the INS-1 cells in sericin group was significantly increased （P<0.05）； compared with sericin group， the survival rate of the INS-1 cells in A-674563 group was significantly decreased （P<0.05）. The flow cytometry results showed that compared with normal control group， the apoptotic rate of the INS-1 cells in model group was significantly increased （P<0.05）； compared with model group， the apoptotic rate of the INS-1 cells in sericin group was significantly decreased （P<0.05）； compared with sericin group， the apoptotic rate of the INS-1 cells in A-674563 group was significantly increased （P<0.05）. The RT-qPCR results showed that compared with normal control group， the expression level of Akt1 mRNA in the INS-1 cells in model group was significantly decreased （P<0.05）； compared with model group， the expression levels of Akt1 mRNA in the INS-1 cells in low， medium， and high doses of sericin groups were significantly increased （P<0.05）； compared with low and medium doses of sericin groups， the expression level of Akt1 mRNA in the INS-1 cells in high dose of sericin group was significantly increased （P<0.05）. Compared with normal control group， the expression levels of NF-κB， TNF-α， and IL-6 mRNA in the INS-1 cells in model group were significantly increased （P<0.05）； compared with model group， the expression levels of NF-κB， TNF-α， and IL-6 mRNA in the INS-1 cells in sericin group were significantly decreased （P<0.05）； compared with sericin group， the expression level of NF-κB mRNA in the INS-1 cells in A-674563 group was significantly increased （P<0.05）. The Western blotting results showed that compared with normal control group， the expression level of p-Akt1 protein in the INS-1 cells in model group was significantly decreased （P<0.05）； compared with model group， the expression levels of p-Akt1 protein in the INS-1 cells in low， medium， and high doses of sericin groups were significantly increased （P<0.05）； compared with low and medium doses of sericin groups， the expression level of p-Akt1 protein in the INS-1 cells in high dose of sericin group was significantly increased （P<0.05）. Compared with normal control group， the expression level of NF-κB protein in the INS-1 cells in model group was significantly increased （P<0.05）； compared with model group， the expression level of NF-κB protein in the INS-1 cells in sericin group was significantly decreased （P<0.05）； compared with sericin group， the expression level of NF-κB protein in the INS-1 cells in A-674563 group was significantly increased （P<0.05）. The ELISA results showed that compared with normal control group， the levels of TNF-α and IL-6 in the INS-1 cells in model group were significantly increased （P<0.05）； compared with model group， the levels of TNF-α and IL-6 in the INS-1 cells in sericin group were significantly decreased （P<0.05）； compared with sericin group， the levels of TNF-α and IL-6 in the INS-1 cells in A-674563 group were significantly increased （P<0.05）. Conclusion Sericin alleviates the PI3K/Akt/NF-κB signaling pathway-mediated inflammatory response and apoptosis by targeting Akt1， exerting a protective effect against STZ-induced damage in INS-1 cells.

Objective To discuss the effect of zanubrutinib （BGB-3111） combined with bortezomib （Btz） on the apoptosis of the human multiple myeloma （MM） cells， and to clarify its possible mechanism. Methods The human MM cell lines U266， PS-R， RPMI8226， KMS28-PE， KMS28-BM， and H929 were cultured in vitro. Western blotting method was used to detect the expression level of Bruton’s tyrosine kinase （BTK） protein in various cells； cell counting kit-8（CCK-8） method was used to detect the survival rates of the RPMI8226， U266， and KMS28-BM cells after treated with 0， 10， 20， 30， 40， and 50 μmol·L?1 BGB-3111. The RPMI8226， U266， and KMS28-BM cells at the logarithmic growth phase were selected and divided into control group， BGB-3111 group， Btz group， and BGB-3111+Btz group. Flow cytometry was used to detect the apoptotic rates of the cells in various groups； Western blotting method was used to detect the expression levels of myeloid cell leukemia 1 （MCL-1）， B-cell lymphoma-2（Bcl-2）， Bcl-2-interacting mediator of cell death （Bim）， phosphorylated Bim （p-Bim）， P65， phosphorylated P65 （p-P65）， tumor necrosis factor receptor-associated factor （TRAF） 2， and tumor necrosis factor alpha-induced protein 3 （A20） in different kinds of cells. The U266 cells were divided into A20 overexpression group （A20-OE group） and empty vector control group （EV group）. Each group was further divided into control group， BGB-3111 group， Btz group， and BGB-3111+Btz group. The corresponding plasmids were transfected； Western blotting method was used to detect the transfection efficiency of the cells in various groups； flow cytometry was used to detect the apoptotic rates of the cells in various groups after over-expression of A20. Results The Western blotting results showed that compared with KMS28-BM cells， the expression levels of BTK protein in the U266， RPMI8226， and H929 cells were significantly increased （P<0.05 or P<0.01）. The CCK-8 results showed that compared with 0 μmol·L?1 BGB-3111 group， the survival rates of the RPMI8226 and U266 cells in 10， 20， 30， 40， and 50 μmol·L?1 BGB-3111 groups were significantly decreased （P<0.05 or P<0.01）， and the survival rates of the KMS28-BM cells in 20， 30， 40， and 50 μmol·L?1 BGB-3111 groups were significantly decreased （P<0.05）. Compared with RPMI8226 and U266 cells， the survival rates of the KMS28-BM cells in 20， 30， and 40 μmol·L?1 BGB-3111 groups were significantly increased （P<0.05）. Therefore， 10 μmol·L?1 BGB-3111 was selected for subsequent experiments. The flow cytometry results showed that compared with control group， the apoptotic rates of the RPMI8226 and U266 cells in BGB-3111 group， Btz group， and BGB-3111+Btz group were significantly increased （P<0.05 or P<0.01）； compared with BGB-3111 group and Btz group， the apoptotic rates of the RPMI8226 and U266 cells in BGB-3111+Btz group were significantly increased （P<0.01）； compared with control group， the apoptotic rates of the KMS28-BM cells in Btz group and BGB-3111+Btz group were significantly increased （P<0.01）； compared with BGB-3111 group， the apoptotic rate of the KMS28-BM cells in BGB-3111+Btz group was significantly increased （P<0.01）； compared with EV group， the apoptotic rates of the cells in A20-OE group in Btz group and BGB-3111+Btz group were significantly increased （P<0.05）. The Western blotting results showed that compared with control group， the expression levels of Bim protein in the RPMI8226 and U266 cells in BGB-3111 group， Btz group， and BGB-3111+Btz group were significantly increased （P<0.05）， while the expression levels of MCL-1， p-Bim， and Bcl-2 proteins in the RPMI8226 and U266 cells in Btz group and BGB-3111+Btz group were significantly decreased （P<0.05）； compared with BGB-3111 group and Btz group， the expression levels of Bim protein in the RPMI8226 and U266 cells in BGB-3111+Btz group were significantly increased （P<0.05）， while the expression levels of MCL-1， p-Bim， and Bcl-2 proteins were significantly decreased （P<0.05）. Compared with control group， the expression levels of p-P65 protein in the RPMI8226 and U266 cells in Btz group and BGB-3111+Btz group were significantly increased （P<0.05）， while the expression levels of TRAF2 and A20 proteins were significantly decreased （P<0.05）； compared with BGB-3111 group and Btz group， the expression levels of p-P65 protein in the RPMI8226 and U266 cells in BGB-3111+Btz group were significantly increased （P<0.05）， while the expression levels of TRAF2 and A20 proteins were significantly decreased （P<0.05）. The flow cytometry results showed that compared with EV group， the expression level of A20 protein in A20-OE group cells was significantly increased （P<0.01）. Conclusion BGB-3111 induces apoptosis in the MM cells by inhibiting BTK activity and enhances the pro-apoptotic effect of Btz. Over-expression of A20 increases the sensitivity of the MM cells to the combined treatment. The antitumor effect may be related to the inhibition of the nuclear factor kappa B （NF-κB） signaling pathway.

Objective To discuss the effects of bone marrow-derived mesenchymal stem cells （BMSCs） transplantation on mitophagy in the vascular dementia （VaD） rats through reactive oxygen species （ROS）/nuclear factor erythroid 2-related factor 2 （Nrf2） signaling， and to clarify its mechanism. Methods Forty-five male adult SD rats were randomly divided into sham operation group， model group， unloaded group， BMSCs group， and MSCs+ML385 （Nrf2 inhibitor） group （combination group）， and there were 9 rats in each group. After intraperitoneal anesthesia， the VaD models were established in all groups except sham operation group. Morris water maze test was used to detect the learning and memory abilities of the rats in various groups； HE staining was used to observe the histopathological morphology of brain tissue of the rats in various groups； Nissl staining was used to observe the changes of Nissl bodies in hippocampus region of brain tissue of the rats in various groups； transmission electron microscope was used to observe the ultrastructure of hippocampus region of the rats in various groups； fluorescence probe method was used to detect the ROS levels in hippocampus neurons in various groups； Western blotting method was used to detect the expression levels of Nrf2， heme oxygenase-1 （HO-1）， PTEN-induced putative kinase 1 （PINK1）， parkin RBR E3 ubiquitin protein ligase （Parkin）， Beclin-1， ubiquitin-binding protein p62 （P62）， and microtubule-associated protein 1A/1B-light chain 3 （LC3-Ⅱ/LC3-Ⅰ） ratio in brain tissue of the rats in various groups. Results The Morris water maze results showed that compared with sham operation group， the escape latency of the rats in model group was significantly increased （P<0.01）， while the number of crossing time and residence time were significantly decreased （P<0.01）. Compared with model group， the escape latency of the rats in BMSCs group was significantly decreased （P<0.01）， while the number of crossing time and residence time were significantly increased （P<0.01）. Compared with BMSCs group， the escape latency of the rats in combination group was significantly increased （P<0.01）， while the number of crossing time and residence time were significantly decreased （P<0.01）. The HE staining results showed that hippocampus neurons of the rats in sham operation group were normal in quantity and morphology， with uniform staining and clear structure. Compared with sham operation group， the hippocampus tissue of the rats in model group showed sparse arrangement， disordered structure， reduced neuronal quantity， varied morphology， uneven staining， nuclear pyknosis， and partial neuronal necrosis. Compared with model group， the neuronal damage of the rats in hippocampus regio in BMSCs group was alleviated， with restored morphology and improved neuronal loss. Compared with BMSCs group， the neurons of the rats in hippocampus region in combination group showed irregular morphology， disordered structure， unclear cell boundaries， uneven staining， and nuclear pyknosis. The Nissl staining results showed that the hippocampal neurons in sham operation group were tightly arranged with intact morphology， obvious nucleoli， and abundant darkly stained Nissl bodies. Compared with sham operation group， the neurons in hippocampus region of the rats in model group showed pyknosis， vacuolization， and sparse Nissl bodies. Compared with model group， the BMSCs group showed reduced neuronal pyknosis， relatively intact morphology， and increased Nissl bodies. Compared with BMSCs group， the combination group showed neuronal pyknosis， loss of morphological integrity， and fragmented Nissl bodies. The transmission electron microscope results showed that mitochondria in sham operation group exhibited oval shape with intact double-membrane structure and cristae. Compared with sham operation group， the mitochondria in model group showed swelling， disrupted membranes， broken cristae， and numerous autophagosomes. Compared with model group， the BMSCs group showed improved mitochondrial structure and reduced autophagosomes. Compared with BMSCs group， the combination group showed swollen mitochondria， disrupted membranes， broken cristae， and visible autophagosomes. The fluorescence probe results showed that compared with sham operation group， the ROS levels in the hippocampus neurons in brain tissue of the rats in model group were significantly increased （P<0.01）； compared with model group， the ROS levels in hippocampus neurons in brain tissue of the rats in BMSCs group were significantly decreased （P<0.01）； compared with BMSCs group， the ROS levels in hippocampus neurons in brain tissue of the rats in combination group were significantly increased （P<0.01）. The Western blotting results showed that compared with sham operation group， the expression levels of Nrf2 and HO-1 proteins in brain tissue of the rats in model group were significantly decreased （P<0.01）； compared with model group， the expression levels of Nrf2 and HO-1 proteins in brain tissue of the rats in BMSCs group were significantly increased （P<0.01）； compared with BMSCs group， the expression levels of Nrf2 and HO-1 proteins in brain tissue of the rats in combination group were significantly decreased （P<0.01）； compared with sham operation group， the expression levels of Parkin， PINK1， and Beclin-1 proteins， and LC3-Ⅱ/LC3-Ⅰ ratio of the rats in model group were significantly increased （P<0.01）， while the expression level of P62 protein was significantly decreased （P<0.01）； compared with model group， the expression levels of Parkin， PINK1， and Beclin-1 proteins， as well as the LC3-Ⅱ/LC3-Ⅰ ratio， of the rats in BMSCs group were significantly decreased （P<0.01）， while the expression level of P62 protein was significantly increased （P<0.01）； compared with BMSCs group， the expression levels of Parkin， PINK1， and Beclin-1 proteins， as well as the LC3-Ⅱ/LC3-Ⅰ ratio， of the rats in combination group were significantly increased （P<0.01）， while the expression level of P62 protein was significantly decreased （P<0.01）. Conclusion BMSCs can alleviate the hippocampal neuronal pathological changes and improve cognitive function in the VaD rats， and its mechanism may be related to the regulation of ROS/Nrf2 signaling pathway to inhibit mitophagy.

Objective To discuss the improvement effect of exosomes （Exo） derived from human adipose-derived stem cells （hADSCs） and human dermal fibroblasts （HDFs） on ultraviolet-induced skin wrinkles in photoaged nude mice， and to clarify its effect. Methods The Exo were isolated from hADSCs and HDFs， respectively， and identified by Western blotting method， designated as hADSCs-Exo and HDFs-Exo. Twenty-eight nude mice were randomly divided into control group， model group ［injected with Hank’s balanced salt solution （HBSS）］， hADSCs-Exo group （injected with hADSCs-Exo）， and HDFs-Exo group （injected with HDFs-Exo）， and there were 7 mice in each group. Except for control group， the other three groups were used to establish a photoaging model on the dorsal skin. After 4 weeks， the gross morphological changes of dorsal skin of the nude mice in various groups were observed， and the wrinkle severity scores were evaluated； HE staining was used to observe the pathomorphology of skin tissue of the nude mice in various groups； ELISA was used to detect the levels of interleukin （IL）-1β， IL-6， and tumor necrosis factor α （TNF-α） in skin tissue of the nude mice in various groups. Real-time fluorescence quantitative PCR （RT-qPCR） and Western blotting method were used to detect the expression levels of collagen Ⅰ （Col Ⅰ）， matrix metalloproteinase （MMP）-1， MMP-2， MMP-3， MMP-9， and MMP-13 mRNA and protein in skin tissue of the nude mice in various groups； immunohistochemical staining was used to observe the protein expression of Col Ⅰ， tropoelastin， and fibrillin-1 in skin tissue of the nude mice in various groups. Results The particles isolated from hADSCs and HDFs exhibited typical vesicle-like structures with diameters of 50-100 nm， and highly expressed CD81， CD63， heat shock protein 70 （HSP70）， and tumor susceptibility gene 101 protein （TSG101）， indicating successful isolation of hADSCs-Exo and HDFs-Exo. The dorsal skin of the nude mice in control group was smooth without looseness or wrinkles， while severe wrinkles， rough epidermis， dryness， and pigmentation were observed in model group. Compared with model group， the dorsal skin of the nude mice in hADSCs-Exo group showed fewer deep wrinkles and mild looseness， whereas the wrinkles in HDFs-Exo group were significantly alleviated compared with control group， the wrinkle severity score in model group was significantly increased （P<0.05）， compared with model group， the wrinkle severity socres of the mice in hADSCs-Exo and HDFs-Exo groups were significantly decreased （P<0.05）. Compared with hADSCs-Exo group， the wrinkle severity score of the mice in HDFs-Exo group was decreased （P<0.05）. The HE staining results showed that clear skin tissue stratification and thin epidermis in control group， while disordered structure， loose arrangement， and thickened epidermis were observed in model group. Compared with model group， the skin lesions in hADSCs-Exo and HDFs-Exo groups were alleviated， with thinner epidermis， clearer stratification， and normalized structure in HDFs-Exo group. The ELISA results showed that compared with control group， the levels of IL-1β， IL-6， and TNF-α in skin tissue of the mice in model group were significantly increased （P<0.05）， and compared with model group， the levels of I6-1β， IL-6， and TNF-α in skin tissue of the mice in hADSCs-Exo and HDFs-Exo groups were significantly decreased （P<0.05）. Compared with hADSCs-Exo group， the levels of IL-1β， IL-6， and TNF-α in skin tissue of the mice in HDFs-Exo group were decreased （P<0.05）. The RT-qPCR and Western blotting results showed that compared with control group， the expression levels of Col Ⅰ mRNA and protein in skin tissue of the mice in model group were significantly decreased （P<0.05）， while the levels of MMP-1， MMP-2， MMP-3， MMP-9， and MMP-13 were significantly increased （P<0.05）. Compared with model group， the expression levels of Col Ⅰ in skin tissue of the mice in hADSCs-Exo and HDFs-Exo groups were significantly increased （P<0.05）， while the levels of MMP-1， MMP-2， MMP-3， MMP-9， and MMP-13 were significantly decreased （P<0.05）. Compared with hADSCs-Exo group， the expression levels of Col Ⅰ in skin tissue of the mice in HDFs-Exo group were increased （P<0.05）， while the levels of MMP-1， MMP-2， MMP-3， MMP-9， and MMP-13 were further decreased （P<0.05）. The immunohistochemical staining results showed that compared with control group the staining intensities of tropoelastin， fibrillin-1， and Col Ⅰ in skin tissue of the mice in model group were significantly weakened， and compared with model group， the staining intensities of tropoelastin， fibrillin-1， and Col Ⅰ in hADSCs-Exo and HDFs-Exo groups were enhanced. Compared with hADSCs-Exo group， the staining intensities in HDFs-Exo group were stronger. Conclusion The Exo derived from hADSCs and HDFs significantly improve ultraviolet-induced skin wrinkles in the photoaged nude mice， with HDFs-Exo exhibiting superior effects compared with hADSCs-Exo.

Objective To discuss the regulatory effect of velvet antler polypeptide on mitochondrial energy metabolism efficiency during myoblast differentiation， and to clarify its related mechanism. Methods The C2C12 myoblasts were maintained in growth medium （GM） and then induced to differentiate in differentiation medium（DM） for 1， 3， and 5 d. The differentiated C2C12 myoblasts were divided into control group and velvet antler polypeptide group. The VAP group was treated with 80 mg·L^-1 velvet antler polypeptide， while the control group was given an equal volume of phosphate buffer saline （PBS）. Cell counting kit-8 （CCK-8） method was used to detect the proliferation rate of the undifferentiated cells in various groups after treated with 0， 5， 10， 20， 40， 60， 80， 120， 160， and 200 mg·L^-1 velvet antler polypeptide； immunofluorescence staining was used to detect the fusion indexes of the cells in various groups； Western blotting method was used to detect the expression levels of myosin， myogenin， myogenin：ubiquinone oxidoreductase subunit B8 （NDUFB8）， mitochondrially encoded cytochrome c oxidase Ⅰ （MT-CO1）， and succinate dehydrogenase complex flavoprotein subunit A （SDHA） proteins in the cells in various groups； 2'，7'-dichlorodihydrofluorescein diacetate （DCFH-DA） fluorescence probe assay was used to detect the reactive oxygen species （ROS） level in the cells in various groups； JC-1 mitochondrial membrane potential assay kit was used to detect the mitochondrial membrane potential of the cells in various groups； commercial kits were used to detect the adenosine triphosphate （ATP） level， glucose uptake， and lactate level in the cells in various groups. Results The CCK-8 results showed that compared with 0 mg·L^-1 velvet antler polypeptide group， the proliferation rates of undifferentiated C2C12 myoblasts in 60， 80， 120， 160， and 200 mg·L^-1 velvet antler polypeptide groups were significantly increased （P<0.05）. The immunofluorescence results showed that compared with control group at the same differentiation stage， the fusion indices of the C2C12 myoblasts in velvet antler polypeptide group at 3 and 5 d of differentiation were significantly decreased （P<0.05）， and the number of multinucleated myotubes was significantly decreased. The Western blotting results showed that compared with control group at the same differentiation stage， the expression levels of myosin in the cells in velvet antler polypeptide group at 1， 3， and 5 d of differentiation were significantly decreased （P<0.05）； the expression levels of myogenin protein in the cells in velvet antler polypeptide group at 3 and 5 d of differentiation were significantly decreased （P<0.05 or P<0.01）. Compared with control group at the same differentiation stage， the expression levels of NDUFB8 and MT-CO1 proteins in the cells in velvet antler polypeptide group at 5 d of differentiation were significantly decreased （P<0.05 or P<0.01）； the expression levels of SDHA protein in the cells in velvet antler polypeptide group at 1， 3， and 5 d of differentiation were significantly decreased （P<0.05 or P<0.01）. The DCFH-DA results showed that compared with control group at the same differentiation stage， the ROS level in velvet antler polypeptide group at 5 d of differentiation was significantly decreased （P<0.01）. The JC-1 assay results showed that compared with control group at the same differentiation stage， the mitochondrial membrane potentials in the cells in velvet antler polypeptide group at 3 and 5 d of differentiation were significantly increased （P<0.05）. Compared with control group at the same differentiation stage， the glucose uptake in the cells in velvet antler polypeptide group at 1 d of differentiation was significantly decreased （P<0.01）， and the glucose uptake at 5 d of differentiation was significantly increased （P<0.05）； there were no significant differences in lactate levels in the cells in velvet antler polypeptide group at 1， 3， and 5 d of differentiation （P>0.05）； the ATP levels in velvet antler polypeptide group at 1 and 3 d of differentiation were significantly increased （P<0.05）. Conclusion Velvet antler polypeptide inhibits the differentiation of the C2C12 myoblasts， and its mechanism may be related to the down-regulation of mitochondrial oxidative phosphorylation complex subunit expression and the improvement of mitochondrial oxidative phosphorylation efficiency.

Objective To discuss the role of M2 macrophages in epithelial-mesenchymal transition （EMT） and cisplatin （DDP） resistance in the non-small cell lung cancer （NSCLC）， and to clarify the regulatory mechanism of nuclear factor κB （NF-κB） signaling pathway. Methods The human monocytic leukemia THP-1 cells were selected and differentiated into M0 macrophages by phorbol myristate acetate （PMA） induction， followed by M2 macrophage polarization through interleukin （IL）-4 and IL-13 stimulation. Western blotting and immunofluorescence methods were used to detect the protein expression levels of CD163， CD86， and arginase-1 （Arg-1） in M0 and M2 macrophages.The human NSCLC A549 cells were co-cultured non-contactly with M0 or M2 macrophages in Transwell chambers， and the cells were divided into A549+M0 group （A549 cells co-cultured with M0 macrophages）， A549+M2 group （A549 cells co-cultured with M2 macrophages）， and A549+M2+BAY11-7082 group （A549 cells co-cultured with M2 macrophages and treated with 10 mmol·L^-1 NF-κB inhibitor BAY11-7082）. Wound healing assay was used to detect the wound healing rate of the A549 cells in various groups； Transwell assay was used to detect the number of invasion A549 cells in various groups； cell counting kit-8 （CCK-8） assay was used to detect the inhibitory rate of proliferation and half maximal inhibitory concentration （IC₅₀） value of the A549 cells after treated with DDP in the co-culture system； Western blotting method was used to detect the expression levels of vimentin， E-cadherin， N-cadherin， transcription factor Snail， phosphorylated P65 （p-P65）， P-glycoprotein （P-gp）， and programmed death-ligand 1 （PD-L1） proteins in the A549 cells in various groups. Results The Western blotting results showed that compared with M0 group， the expression levels of CD163 and Arg-1 proteins in the macrophages in M2 group were significantly increased （P<0.05）， while the expression level of CD86 protein was significantly decreased （P<0.05）. The immunofluorescence results showed that compared with M0 group， the expression of CD163 protein in the macrophages in M2 group was enhanced and the expression of CD86 protein was weakened. The wound healing assay results showed that at 24 and 48 h of culture， compared with A549+M0 group， the wound healing rate of the A549 cells in A549+M2 group was significantly increased （P<0.05）； in the co-culture system， compared with A549+M0 group， the wound healing rate of the A549 cells in A549+M2 group was significantly increased （P<0.05）； compared with A549+M2 group， the wound healing rate of the A549 cells in A549+M2+BAY11-7082 group was significantly decreased （P<0.05）. The Transwell assay results showed that compared with A549+M0 group， the number of invasion A549 cells in A549+M2 group was significantly increased （P<0.05）； compared with A549+M2 group， the number of invasion A549 cells in A549+M2+BAY11-7082 group was significantly decreased （P<0.05）； in the co-culture system， compared with A549+M0 group， the number of invasion A549 cells in A549+M2 group was significantly increased （P<0.05）. The CCK-8 assay results showed that after treated with 2.50， 5.00， 10.00， 20.00， and 40.00 mg·L^-1 DDP， compared with A549+M0 group， the inhibitory rate of proliferation of the A549 cells in A549+M2 group was significantly decreased （P<0.05 or P<0.01）， and the IC₅₀ value was significantly increased （P<0.01）； in the co-culture system， compared with A549+M0 group， the inhibitory rate of proliferation of the A549 cells in A549+M2 group was significantly decreased （P<0.05 or P<0.01）， and the IC₅₀ value was significantly increased （P<0.01）； compared with A549+M2 group， the inhibitory rate of proliferation of the A549 cells in A549+M2+BAY11-7082 group was significantly increased （P<0.05）， and the IC₅₀ value was significantly decreased （P<0.05）. The Western blotting results showed that compared with A549+M0 group， the expression level of E-cadherin proteins in the A549 cells in A549+M2 group was significantly decreased （P<0.05）， while the expression levels of N-cadherin， vimentin， and Snail proteins were significantly increased （P<0.05）； in the co-culture system， compared with A549+M0 group， the expression levels of vimentin， Snail， N-cadherin， and p-P65 proteins in the A549 cells in A549+M2 group were significantly increased （P<0.05）， while the expression level of E-cadherin proteins was significantly decreased （P<0.05）； compared with A549+M2 group， the expression levels of vimentin， N-cadherin， and p-P65 proteins in the A549 cells in A549+M2+BAY11-7082 group were significantly decreased （P<0.05）， while the expression level of E-cadherin proteins was significantly increased （P<0.05）； compared with A549+M0 group， the expression levels of P-gp and PD-L1 proteins in the A549 cells in A549+M2 group were significantly increased （P<0.05）； in the co-culture system， compared with A549+M0 group， the expression levels of P-gp and PD-L1 proteins in the A549 cells in A549+M2 group were significantly increased （P<0.05）； compared with A549+M2 group， the expression levels of P-gp and PD-L1 proteins in the A549 cells in A549+M2+BAY11-7082 group were significantly decreased （P<0.05）. Conclusion The M2 macrophages can regulate EMT in the NSCLC cells to promote the invasion and metastasis of tumor， and modulate the expressions of P-gp and PD-L1 to enhance DDP resistance， which is associated with the NF-κB signaling pathway.

Objective To discuss the effect of circadion rhythm gene TIMELESS （TIM） silencing on immune escape of the ovarian cancer cells， and to clarify its related mechanism. Methods The CD8+T lymphocytes were isolated and identified by flow cytometry to detect the proportion of CD3+/CD8+ cell subsets. The human ovarian cancer SK-OV-3 cells were cultured in vitro and divided into interference plasmid transfected with TIM small interfering （siRNA） （si-TIM）， negative control plasmid （si-NC）， programmed death ligand 1 （PD-L1） over-expression plasmid （oe-PD-L1）， and negative control plasmid （oe-NC） groups. The cells were further divided into blank control group （BC group， non-transfection）， si-NC group （transfected with si-NC）， si-TIM group（transfected with si-TIM）， si-NC+oe-NC group （transfected with si-NC and oe-NC）， and si-TIM+oe-PD-L1 group （transfected with si-TIM and oe-PD-L1）. Real-time fluorescence quantitative PCR（RT-qPCR） and Western blotting methods were used to detect the expression levels of TIM mRNA and protein in the SK-OV-3 cells to verify TIM gene silencing. The transfected SK-OV-3 cells were co-cultured with activated CD8+T lymphocytes and divided into BC group （SK-OV-3 cells cultured alone）， BC/T group， si-NC/T group， si-TIM/T group， si-NC+oe-NC/T group， and si-TIM+oe-PD-L1/T group. CCK-8 method was used to detect the survival rates of the SK-OV-3 cells in various groups； flow cytometry was used to detect the apoptotic rates of the SK-OV-3 cells and positive expression rate of PD-L1 on surface of the cells in various groups； enzyme-linked immunosorbent assay （ELISA） method was used to detect the levels of interferon-γ （IFN-γ） and tumor necrosis factor-α （TNF-α） in the co-culture supernatant； lactate dehydrogenase （LDH） release assay was used to detect the cytotoxicity of the CD8+T lymphocytes in various groups； RT-qPCR method was used to detect the expression levels of TIM and PD-L1 mRNA in the SK-OV-3 cells in various groups； Western blotting method was used to detect the expression levels of TIM and PD-L1 proteins in the SK-OV-3 cells in various groups. Results After scparated with immune magnetic bead method， the proportion of CD8+T lymphocyte （CD3+/CD8+） subsets was （96.56%±0.59%）， indicating high purity of the extracted CD8+T lymphocytes. Compared with BC group， the expression levels of TIM mRNA and protein in the cells in si-TIM group were significantly decreased （P<0.01）， suggesting successful TIM gene silencing in the ovarian cancer SK-OV-3 cells. The CCK-8 results showed that compared with BC group， the survival rate of the SK-OV-3 cells in BC/T group was significantly decreased （P<0.01）； compared with BC/T group， the survival rate of the SK-OV-3 cells in si-TIM/T group was significantly decreased （P<0.01）. The flow cytometry results showed that compared with BC group， the apoptotic rate of the SK-OV-3 cells in BC/T group was significantly increased （P<0.01）； compared with BC/T group， the apoptotic rate of the SK-OV-3 cells in si-TIM/T group was significantly increased （P<0.01）； compared with si-TIM/T group， the apoptotic rate of the SK-OV-3 cells in si-TIM+oe-PD-L1/T group was significantly decreased （P<0.01）. Compared with BC group， the positive expression rate of PD-L1 on surface of the SK-OV-3 cells in si-TIM group was significantly decreased （P<0.01）. The ELISA results showed that compared with BC/T group， the levels of IFN-γ and TNF-α in the culture supernatant in si-TIM/T group were significantly increased （P<0.01）； compared with si-TIM/T group， the levels of IFN-γ and TNF-α in the supernatant in si-TIM+oe-PD-L1/T group were significantly decreased （P<0.01）. The LDH release assay results showed that compared with BC/T group， the cytotoxicity of the CD8+T lymphocytes in si-TIM/T group was significantly increased （P<0.01）； compared with si-TIM/T group， the cytotoxicity of the CD8+T lymphocytes in si-TIM+oe-PD-L1/T group was significantly weakened （P<0.01）. The RT-qPCR and Western blotting results showed that compared with BC group， the expression levels of PD-L1 mRNA and protein in the SK-OV-3 cells in si-TIM group were significantly decreased （P<0.01）； compared with si-TIM group， the expression level of PD-L1 protein in the cells in si-TIM+oe-PD-L1 group was significantly increased （P<0.01）. Conclusion TIM gene silencing enhances the cytotoxic effect of CD8+T lymphocytes on ovarian cancer SK-OV-3 cells and inhibits immune escape， and its mechanism may be related to the regulation of PD-L1 protein expression.

Objective To discuss the effects of microRNA （miR）-199a-5p overexpression on cell migration and apoptosis in the glioblastoma U251 cells， and to clarify the targeting regulatory relationship between miR-199a-5p and caveolin-1 （CAV-1）. Methods The glioblastoma U251 cells and oligodendroglioma Hs683 cells were cultured in vitro. Western blotting method was used to detect the expression levels of CAV-1 protein in 2 kinds of cells； real-time fluorescence quantitative PCR （RT-qPCR） method was used to detect the expression levels of miR-199a-5p in 2 kinds of cells. The U251 cells were divided into blank group （non-transfection）， mimics NC group （transfected with empty vector）， and miR-199a-5p mimics group （transfected with miR-199a-5p mimics）. The Hs683 cells were divided into blank group （no transfection）， inhibitor NC group （transfected with empty vector）， and miR-199a-5p inhibitor group （transfected with miR-199a-5p inhibitor）. RT-qPCR method was used to detect the transfection efficiency of the cells in various groups； Western blotting method was used to detect the expression levels of CAV-1 protein in the cells in various groups. TargetScan database was used to predict the binding sites between miR-199a-5p and CAV-1 in the 3' untrans lated region（3'UTR）； psiCHECKTM-2-CAV-1-WT and psiCHECKTM-2-CAV-1-Mut were co-transfected with miR-199a-5p mimics and mimics NC into the U251 cells， respectively， forming psiCHECKTM-2-CAV-1-WT+mimics NC group， psiCHECKTM-2-CAV-1-WT+miR-199a-5p mimics group， psiCHECKTM-2-CAV-1-Mut+mimics NC group， and psiCHECKTM-2-CAV1-Mut+miR-199a-5p mimics group； dual-luciferase reporter gene assay was used to verify the targeting relationship between miR-199a-5p and CAV-1； cell scratch assay was used to detect the scratch healing rates of the U251 cells in various groups； flow cytometry was used to detect the apoptotic rates of the U251 cells in various groups. Results The Western blotting and RT-qPCR results showed that compared with Hs683 cells， the expression level of CAV-1 protein in the U251 cells was significantly decreased （P<0.05）； compared with U251 cells， the expression level of miR-199a-5p in the Hs683 cells was significantly increased （P<0.01）. Compared with blank group and mimics NC group， the expression level of miR-199a-5p in the U251 cells in miR-199a-5p mimics group was significantly increased （P<0.01）， the expression level of CAV-1 protein in the U251 cells in miR-199a-5p mimics group was significantly decreased （P<0.05）. Compared with blank group， the expression levels of miR-199a-5p in the Hs683 cells in inhibitor NC group and miR-199a-5p inhibitor group were significantly decreased （P<0.01）. No significant differences were observed in the expression levels of CAV-1 protein in the Hs683 cells among various groups （P>0.05）. The dual-luciferase reporter gene assay results showed that psiCHECKTM-2-CAV-1-wild type （WT） and psiCHECKTM-2-CAV-1-mutant （Mut） expression vectors were successfully constructed； compared with psiCHECKTM-2-CAV-1-WT-mimics NC group， the relative luciferase activity of WT CAV-1 in the U251 cells in psiCHECKTM-2-CAV-1-WT-miR-199a-5p mimics group was significantly decreased （P<0.01）. The cell scratch assay results showed that at 12， 24， and 48 h after transfection， compared with blank group， the scratch healing rate of the U251 cells in miR-199a-5p mimics group was significantly decreased （P<0.05 or P<0.01）. The flow cytometry results showed that compared with blank group and mimics NC group， the apoptotic rate of the U251 cells in miR-199a-5p mimics group was significantly increased （P<0.01）. Conclusion Transfection of mature miR-199a-5p mimics into the glioblastoma U251 cells can reduce the expression of CAV-1 protein， inhibit glioma cell migration， promote apoptosis， and suppress tumorigenesis and development. The targeting relationship between miR-199a-5p and CAV-1 may represent a potential mechanism for glioma development and could serve as a potential diagnostic and therapeutic target for glioma.

Objective To discuss the effect of hirudin on post-stroke depression （PSD） in the mice， and to clarify its potential mechanism. Methods Sixty male C57BL/6 mice were randomly divided into control group， PSD group， PSD+low dose of hirudin group， PSD+medium dose of hirudin group， and PSD+high dose of hirudin group， and there were 12 mice in each group. The stroke model was established by middle cerebral artery occlusion （MCAO）， and the depression model was induced by chronic unpredictable mild stress （CUMS） combined with solitary housing. The mice in PSD+low dose of hirudin， PSD+medium dose of hirudin， and PSD+high dose of hirudin groups were intravenously injected with 10， 20， and 40 U·kg^-1 hirudin， respectively， while the mice in control and PSD groups received equal volumes of saline. The body weights of the mice were recorded on days 0， 7， 14， and 21 of CUMS. The LONGA neurological function score was calculated. Sucrose preference test， tail suspension test， and forced swimming test were used to detect the sucrose preference rate， immobility time in tail suspension， and forced swimming in various groups， respectively； HE staining was used to observe the histopathological changes in the medial prefrontal cortex （mPFC）； biochemical kits were used to detect the levels of malondialdehyde （MDA） and reduced glutathione （GSH） as well as superoxide dismutase （SOD） activity in mPFC tissue of the mice in various groups； 2'，7'-dichlorodihydrofluorescein diacetate （DCFH-DA） fluorescence probe method was used to detect the reactive oxygen species （ROS） positive rate in mPFC tissue of the mice in various groups； real-time fluorescence quantitative PCR （RT-qPCR） and Western blotting method were used to detect the expression levels of nucleoredoxin （NXN） mRNA and protein in mPFC tissue of the mice in various groups. Results Compared with control group， the body weight of the mice in PSD group was significantly decreased on days 0， 7， 14， and 21 of CUMS （P<0.05 or P<0.01）. Compared with PSD group， the body weights of the mice in PSD+low dose of hirudin， PSD+medium dose of hirudin， and PSD+high dose of hirudin groups were significantly increased on days 14 and 21 of CUMS （P<0.05 or P<0.01）， and the neurological function scores were significantly decreased （P<0.05 or P<0.01）. The sucrose preference test， tail suspension test， and forced swimming test results showed that compared with control group， the sucrose preference rate of the mice in PSD group was significantly decreased （P<0.01）， while the immobility times in tail suspension and forced swimming were significantly increased （P<0.01）. Compared with PSD group， the sucrose preference rates of the mice in PSD+low dose of hirudin， PSD+medium dose of hirudin， and PSD+high dose of hirudin groups were significantly increased （P<0.05 or P<0.01）， and the immobility times were significantly decreased （P<0.05 or P<0.01）. The HE staining showed normal cell morphology， clear structure， and uniform size distribution in mPFC tissue in control group. In PSD and PSD+low dose of hirudin groups， the number of the cells in mPFC tissue was significantly reduced， with severe vacuolar degeneration and pyknotic nuclei. Compared with PSD group， the numbers of the cells in PSD+medium dose of hirudin and PSD+high dose of hirudin groups were significantly increased， and the vacuolar degeneration and nuclear pyknosis were alleviated. The Biochemical and DCFH-DA fluorescence probe assays results showed that compared with control group， the GSH level and SOD activity in mPFC tissue of the mice in PSD group were significantly decreased （P<0.01）， while the MDA level and ROS positive rate were significantly increased （P<0.01）. Compared with PSD group， the GSH levels and SOD activities of the mice in PSD+low dose of hirudin， PSD+medium dose of hirudin， and PSD+high dose of hirudin groups were significantly increased （P<0.05 or P<0.01）， while the MDA levels and ROS positive rates were significantly decreased （P<0.05 or P<0.01）. The RT-qPCR and Western blotting results showed that compared with control group， the expression levels of NXN mRNA and protein in mPFC tissue of the mice in PSD group were significantly decreased （P<0.01）. Compared with PSD group， the expression levels of NXN mRNA and protein in mPFC tissue of the mice in PSD+low dose of hirudin， PSD+medium dose of hirudin， and PSD+high dose of hirudin groups were significantly increased （P<0.05 or P<0.01）. Conclusion Hirudin promotes redox balance in mPFC of the PSD mice， repairs neurological damage， and improves PSD.

Objective To discuss the inhibitory effects of combined application of chlorogenic acid （CA）， procyanidin （PC）， and paeoniflorin （PF）， the active components of Paeoniae Radix Rubra， on Enterococcus faecalis （E.faecalis） and its biofilm， and to clarify the mechanism. Methods The minimal inhibitory concentration （MIC） and minimal bactericidal concentration （MBC） of CA， PC， and PF against E.faecalis were detected by microdilution method； the fractional inhibitory concentration index （FICI） and fractional bactericidal concentration index （FBCI） of the three active components of Paeoniae Radix Rubra in combination were detected by checkerboard dilution method. The experiment was divided into control group， high concentration of single-drug groups （PF-10 group， PC-6 group， and CA-10 group）， and drug combination groups （CA-2+PC-1 group， CA-2+PC-2 group， PF-4+PC-2 group， PF-6+PC-2 group， PF-4+CA-4 group， and PF-6+CA-4 group）. Crystal violet staining was used to detect the biofilm formation of E.faecalis in various groups after treated with three active components in combination； scanning electron microscope （SEM） was used to observe the morphology of E.faecalis biofilm in various groups after treated with three active components in combination； spot assay was used to detect the inhibitory effects of three active components in combination on E.faecalis planktonic bacteria and biofilm in various groups； SEM was used to observe the damage to E.faecalis cell membrane in various groups after treated with three active components in combination； kit was used to detect the adenosine triphosphate （ATP） levels in E.faecalis planktonic bacteria and biofilm in various groups after treated with three active components in combination. Results Among the three active components of Paeoniae Radix Rubra, the MIC of PC was 4 g·L?1 and the MBC was 6 g·L?1； the MIC of CA was 8 g·L?1 and the MBC was 10 g·L?1； the MIC and MBC of PF were both >10 g·L?1, and the concentration of PF was selected as 10 g·L?1. The combination of PC and CA showed synergistic effects, the combination of PC and PF showed additive effects, and the combination of CA and PF showed additive effects. The crystal violet staining results showed that compared with control group, the biofilm formations of E.faecalis in PF-10 group, PC-6 group, CA-10 group, and drug combination groups were significantly decreased （P<0.05）； compared with PF-10 group, the biofilm formations of E.faecalis in PC-6 group， CA-10 group， CA-2+PC-1 group， CA-2+PC-2 group， PF-4+PC-2 group， PF-6+PC-2 group， and PF-6+CA-4 group were significantly decreased （P<0.05 or P<0.01）. The SEM results showed that in control group, the E.faecalis biofilm was thick, with tightly connected bacteria, regular morphology, and intact cell membranes; in PF-10 group, PC-6 group, and CA-10 group, the thickness of E.faecalis biofilm was significantly reduced, and the arrangement of bacteria became relatively loose; in all drug combination groups, the E.faecalis biofilm was significantly reduced or even completely disappeared, and under high magnification, the biofilm structure was completely absent, with bacterial fragments adhering and aggregating, losing their original bacterial morphology. The spot assay results showed that compared with control group, the colonies of E.faecalis planktonic bacteria in PF-10 group, PC-6 group, and CA-10 group were significantly reduced after treated for 5, 10, and 30 min, indicating gradually enhanced bactericidal effects; among drug combination groups, the combination of CA and PC significantly reduced the colonies of E.faecalis planktonic bacteria within 5 min, showing strong bactericidal effects. Compared with CA group and PC group, the colonies of E.faecalis planktonic bacteria in all drug combination groups showed no significant reduction after treated for 5, 10, and 30 min； compared with control group, the colonies of E.faecalis biofilm in PF-10 group, PC-6 group, and CA-10 group were gradually decreased after the treated for 30 and 60 min, suggesting that the high concentration of single-drug groups exhibited gradually enhanced bactericidal effects on E.faecalis in biofilm. Among them, the biofilm-killing effect of PC-6 group was the most significant, with no colony formation observed after treated for 30 min； in drug combination groups, only a few colonies of E.faecalis biofilm were observed in CA-2+PC-2 group after treated for 30 min, indicating effective killing of bacteria in biofilm; compared with PC-6 group and CA-10 group, all drug combination groups achieved the bactericidal effects of high concentration of single-drug groups at low concentrations. The SEM results showed that in control group, E.faecalis exhibited an oval shape with intact cell membranes; in PF group, bacterial morphology was altered, and cell membrane integrity was damaged； in CA group, most bacterial cell membranes remained relatively intact, but the bacterial surface showed shrinkage and depression, with a few bacteria exhibiting disrupted cell membrane integrity； in PC group, the integrity of bacterial cell membranes was most severely damaged, leading to leakage of cellular contents and aggregation of cell fragments into flocculent structures; in all drug combination groups, E.faecalis exhibited ruptured cell membranes, leakage of contents, and aggregation of bacterial debris, especially in the combination of CA and PC, where the most severe disruption of bacterial cell membrane integrity and complete leakage of contents were observed; in the combination of PF and CA, bacterial surface pits and shrinkage were observed, with occasional cell membrane rupture. The kit results showed that compared with control group, the ATP levels in E.faecalis planktonic bacteria and biofilm in various groups were significantly decreased （P<0.01）； compared with PF-10 group, the ATP levels in E.faecalis planktonic bacteria in CA-10 group, CA-2+PC-2 group， PF-4+CA-4 group, and PF-6+CA-4 group were significantly decreased （P<0.05 or P<0.01）， and the ATP levels in E.faecalis biofilm in CA-10 group and CA-2+PC-2 group were significantly decreased （P<0.05 or P<0.01）. Conclusion The combined application of PF， PC， and CA， the active components of Paeoniae Radix Rubra， exhibits significant inhibitory effects on E.faecalis and its biofilm formation. The pairwise combinations of three active components show synergistic or additive effects， with the combination of CA and PC demonstrating the most significant synergistic effect. The underlying mechanism may be related to the disruption of E.faecalis cell membrane integrity and inhibition of bacterial ATP levels.

Objective To investigate the active ingredients and targets of Compound Gastritis Mixture （CGM） in the treatment of chronic atrophic gastritis （CAG） by network pharmacology method， and to validate the potential mechanism combined with molecular docking technology and cellular experiments. Methods The Traditional Chinese Medicine System Analysis Platform （TCMSP） and Swiss Target Prediction databases were used to select the herbal ingredients of CGM and the corresponding targets； the GeneCards and Online Mendelian Inheritance in Man （OMIM） database were used to screen the targets of CAG； the common targets of CGM and CAG were analyzed from the Venny2.1.0 platform； STRING online platform was used to construct protein-protein interaction （PPI） networks for common drug-disease targets and screen the core targets. Cytoscape 3.9.1 software was used to construct the drug-disease-target network and screen the drug core components； Gene Ontology （GO） fuctional， Kyoto Encyclopedia of Genes and Genomes （KEGG） signaling pathway enrichment analysis were used to analyze the common targets of CGM and CAG； and AutoDock analysis software was used to perform molecular docking analysis of predicted main components of the drugs and core targets. The gastric mucosal epithelial cells GES-1 were induced by lipopolysaccharide （LPS） to construct CAG cell model.The GES-1 cells were divided into blank group （10% serum complete medium），model group （10 mg·L^-1 LPS）， and different concentrations of CGM groups （50， 100， 200， 400， 800 and 1 600 g·L^-1 CGM+10 mg·L^-1 LPS）， and cells were incubated for 12， 24， and 48 h. The cell counting kit-8 （CCK-8） assay was used to detect the proliferation activities of GES-1 cells. The GES-1 cells were divided into blank group （10% serum complete medium）， model group （10 mg·L^-1 LPS） and CGM group（1 600 g·L^-1 CGM+10 mg·L^-1 LPS）. Real-time fluorescence quantitative PCR （RT-qPCR） method was used to detect the expression levels of interleukin（IL）-6， tumor necrosis factor （TNF）， serine/threonine protein kinase 1 （AKT1），IL-1β， and epidermal growth factor receptor （EGFR） mRNA in the cells in various groups. Results A total of 198 ingredients of CGM were screened， and 128 common targets with CAG were identified. The main herbal ingredients of CGM in treatment of CAG were quercetin， kaempferol， and lluteolin， which mainly acted on the core targets of IL-6， TNF， AKT1， IL-1β， and EGFR. The GO function enrichment analysis results showed that the top 15 targets mainly focused on biological processes（BP） such as apoptosis， inflammatory response and cell proliferation， mainly included cellular components （CC） such as cytoplasm， cell surface and macromolecular complexes， and mainly exerted molecular functions （MF）such as proteins， enzymes and ubiquitin-protein ligases. A total of 158 pathways were obtained from KEGG signaling pathway enrichment analysis， mainly involved cancer-related pathways， TNF signaling pathways， viral infection， programmed cell death-ligand 1（PD-L1）/ programmed cell death protein-1 （PD-1） pathways， apoptosis， NOD-like receptor signaling pathways， Toll-like receptor signaling pathways， EGFR， and IL-17 signaling pathways.The binding energies of the core targets IL-6， TNF， IL-1β， AKT1， and EGFR with main herbal ingredients quercetin， kaempferol， and luteolin were<-5 kcal·mol^-1. The CCK-8 assay results showed that compared with blank group， after 24 and 48 h of cell culture， the proliferation activities of the cells in model group were significantly decreased （P<0.01）， and the inhibition of the proliferation activity was more obvious after 48 h； therefore， 48 h was selected for the modeling time； compared with model group， the proliferation activities of cells in 800 and 1 600g·L^-1 GCM groups were significantly decreased （P<0.01）， and the promotion of cell proliferation activity was more obvious in 1 600g·L^-1 GCM group， so the intervening concentration of this drug was selected for the subsequent experiments. The RT-qPCR method results showed that compared with blank group， the expression levels of IL-6， TNF， IL-1β， AKT1， and EGFR mRNA in the cells in model group were significantly increased （P<0.01）； compared with model group， the expression levels of IL-6， IL-1β， AKT1 and EGFR mRNA in the cells in CGM group were significantly decreased （P<0.01）. Conclusion CGM may play a role in the prevention and treatment of CAG through multiple ingredients such as quercetin， kaempferol and lignocerol， acting on the multiple target proteins such as IL-6， TNF， AKT1， IL-1β， and EGFR， as well as involving a variety of “inflammatory-cancer-related” pathways.

Objective To discuss the regulatory role of complement alternative pathway in mouse colorectal cancer （CRC） liver metastasis model based on bioinformatics methods， and to clarify its mechanism through experimental verification. Methods Using “CRC liver metastasis” as the keyword， the GSE81558 dataset was retrieved from Gene Expression Omnibus （GEO） database， including normal colon tissue samples， CRC tissue samples and CRC liver metastasis tissue samples. Bioinformatics methods were used to analyze and screen differentially expressed genes （DEGs）. Gene Ontology （GO） functional enrichment analysis and Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment analysis were performed using R and Cytoscape software， and the results were visualized. Search Tool for the Retrieval of Interacting Genes/Proteins （STRING） database was used to evaluate protein-protein interactions （PPIs） of DEGs and construct PPI network. Twelve C57BL/6 mice were injected with SL4 tumor cells into spleen， and the liver tissues were collected at 0， 7 and 14 d. Real-time fluorescence quantitative PCR （RT-qPCR） method was used to detect the expression levels of complement pathway-related genes in liver metastatic foci. The CRC liver metastasis mouse model was used to verify the complement signaling pathway. The mice were divided into control group， factor B knockout group （FB^-/-） and C4 factor knockout group （C4^-/-）， and there were 6 mice in each group. The liver weights of the mice were measured； HE staining was used to detect the percentage of metastatic area in liver tissue in control group and FB^-/- group； immunohistochemistry was used to detect macrophage infiltration in liver tissue in control group and FB^-/- group， and the percentage of macrophage infiltration was calculated. Results The distances between normal colon tissue samples and CRC tissue samples， as well as between CRC tissue samples and CRC liver metastasis tissue samples were far， indicating significant differences between samples， allowing subsequent analysis of DEGs. A total of 1 908 DEGs were screened in the dataset comparing normal colon tissue samples and CRC tissue samples， including 771 up-regulated DEGs and 1 137 down-regulated DEGs. Twenty-three up-regulated DEGs and 100 down-regulated DEGs were identified in the dataset comparing CRC and CRC liver metastasis. The GO functional enrichment analysis results showed that compared with normal colon tissue samples， DEGs in CRC samples were mainly enriched in biological processes （BP） related to cell cycle and mitosis， including mitotic cell cycle process， cell division， response to hormone， mitotic nuclear division and response to lipid. Compared with CRC samples， the DEGs in CRC liver metastasis samples were mainly enriched in coagulation-related BP， including platelet degranulation， blood coagulation regulation， acute-phase response， hemostasis regulation and coagulation regulation. The KEGG pathway enrichment analysis results showed that compared with normal colon tissue samples， the DEGs in CRC tissue samples were mainly enriched in cell cycle and p53 signaling pathways. Compared with CRC tissue samples， the DEGs in CRC liver metastasis tissue samples were mainly enriched in complement， coagulation cascade and metabolism-related signaling pathways. The Hub genes identified in PPI network were related to blood proteins. The RT-qPCR results showed that compared with 0 d group， the mRNA expression level of complement related genes complement 1q （C1q） in liver metastatic foci tissue sampres in 7 d group was significantly decreased （P<0.05）， the mRNA expression levels of complement 3 （C3）， complement 5 （C5）， FB， and factor D （FD） were significantly increased （P<0.05 or P<0.01）， the mRNA expression levels of complement pathway-related genes C1q， complement 2 （C2）， C3， complement fragment 3a receptor （C3aR）， C5， complement fragment 5a receptor （C5aR）， decay-accelerating factor （DAF）， FB and FD in liver metastatic foci tissue sampres in 14 d group were significantly increased （P<0.05 or P<0.01）. Compared with control group， the liver weight of the mice in FB^-/- group was significantly decreased （P<0.01）， while there was no significant difference was observed in C4^-/- group （P>0.05）. The HE staining results showed that compared with control group， the liver metastatic foci in FB^-/- mice were significantly decreased， and the percentage of metastatic area was decreased （P<0.01）. The immunohistochemistry results showed that compared with control group， the macrophage infiltration in liver metastatic foci of the mice in FB^-/- group was reduced， and the percentage of macrophage infiltration was decreased （P<0.01）. Conclusion Complement cascade is associated with CRC liver metastasis， and the alternative complement pathway regulates CRC liver metastasis， suggesting this pathway may serve as a potential therapeutic target for CRC liver metastasis.

Objective To discuss the expression and biological function of gap junction protein β2 （GJB2） in the lung adenocarcinoma （LUAD） A549 cells， and to procide the basis for the treatment of LUAD. Methods The TIMER database was used to analyze the differential expression of GJB2 gene in various tumors； the GEPIA database was used to detect the mRNA expression of GJB2 in LUAD and analyze its correlation with different clinical stages of LUAD； the Kaplan-Meier plotter database was used to analyze the correlation between GJB2 protein and the prognosis of the LUAD patients. A total of 111 pairs of cancer tissues and adjacent normal lung tissues of LUAD patients were collected， and immunohistochemical staining was used to observe the expression of GJB2 protein in LUAD tissues and adjacent normal lung tissues. The human LUAD A549 cells were cultured in vitro and divided into GJB2 overexpression group （OE-GJB2 group， transfected with GJB2 overexpression plasmid） and its negative control group （OE-NC group， transfected with GJB2 empty vector plasmid）， small interfering RNA （siRNA）-GJB2 group （si-GJB2 group， transfected with GJB2-siRNA） and its negative control group （si-NC group， transfected with control siRNA）. Western blotting method was used to verify the transfection efficiency of the cells； cell couting kit-8 （CCK-8） method was used to detect the proliferation activities of the A549 cells in various groups； 5-ethynyl-2'-deoxyuridine （EdU） staining was used to detect the positive expression rates of the A549 cells in various groups； colony formation assay was used to detect the number of colony formation of the A549 cells in various groups； cell scratch assay was used to detect the migration rate of the A549 cells in various groups； Transwell chamber assay was used to detect the number of the invasion A549 cells in various groups； Western blotting method was used to detect the expression levels of GJB2 protein and epithelial-mesenchymal transition （EMT）-related proteins in the A549 cells in various groups. Results The RT-PCR results showed that GJB2 gene was abnormally expressed in various tumors； compared with adjacent normal lung tissue， the mRNA expression level of GJB2 in cancer tissue of LUAD patients was significantly increased （P<0.05）， and the high expression of GJB2 was correlated with the clinicopathological stage of LUAD （P<0.05）. The Kaplan-Meier plotter results showed that compared with patients with low GJB2 expression， the overall survival of the patients with high GJB2 expression was significantly decreased ［hazard ratio （HR）=1.71， 95% CI： 1.34-2.18， P<0.05］. GJB2 protein was negatively expressed in adjacent normal alveolar and bronchial epithelial cells； compared with adjacent normal lung tissue， the expression of GJB2 protein in cancer tissue of LUAD patients was significantly enhanced. In LUAD， the positive expression of GJB2 protein was significantly associated with lymphnode metastasis （P<0.05） but not with gender （P=0.626）， age （P=0.639）， or TNM stage （P=0.837） （P>0.05）. The CCK-8 results showed that compared with OE-NC group， the proliferation activity of the A549 cells in OE-GJB2 group was significantly increased （P<0.05 or P<0.01）； compared with si-NC group， the proliferation activity of the A549 cells in si-GJB2 group was significantly decreased （P<0.05 or P<0.01）. The EdU staining results showed that compared with OE-NC group， the positive expression rate of EdU in the A549 cells in OE-GJB2 group was significantly increased （P<0.01）； compared with si-NC group， the positive expression rate of EdU in the A549 cells in si-GJB2 group was significantly decreased （P<0.01）. The colony formation assay results showed that compared with OE-NC group， the number of colony formation of the A549 cells in OE-GJB2 group was significantly increased （P<0.01）； compared with si-NC group， the number of colony formation of the A549 cells in si-GJB2 group was significantly decreased （P<0.01）. The Transwell chamber assay results showed that compared with OE-NC group， the number of invasion A549 cells in OE-GJB2 group was significantly increased （P<0.01）； compared with si-NC group， the number of invasion A549 cells in si-GJB2 group was significantly decreased （P<0.01）. The cell scratch assay results showed that 24 h after scratching， compared with OE-NC group， the migration rate of the A549 cells in OE-GJB2 group was significantly increased （P<0.01）； compared with si-NC group， the migration rate of the A549 cells in si-GJB2 group was significantly decreased （P<0.01）. The Western blotting results showed that compared with OE-NC group， the expression levels of GJB2 and N-cadherin proteins in the A549 cells in OE-GJB2 group were significantly increased （P<0.01）， while the expression level of E-cadherin protein was significantly decreased （P<0.05）； compared with si-NC group， the expression levels of GJB2 and N-cadherin proteins in the A549 cells in si-GJB2 group were significantly decreased （P<0.01）， while the expression level of E-cadherin protein was significantly increased （P<0.01）. Conclusion GJB2 is highly expressed in cancer tissue of LUAD patients and is associated with the poor prognosis in LUAD patients. GJB2 promotes the proliferation， invasion， migration， and EMT of the A549 cells.

Objective To discuss the effect of KIAA1522 on the proliferation， migration， and invasion of lung cancer cells， and to clarify its signaling mechanism. Methods Bioinformatics analysis was used to detect the expression levels of KIAA1522 mRNA and protein in 75 cases of human non-small cell lung cancer （NSCLC） tissues and adjacent normal lung tissues； immunohistochemical staining was used to detect the expression of KIAA1522 protein in NSCLC tissue and adjacent normal lung tissues； Western blotting method was used to detect the expression level of KIAA1522 protein in various lung cancer cell lines. KIAA1522-small interfering（siRNA） and over-expression plasmids were transfected into the lung cancer H1299 and A549 cells， respectively. The KIAA1522-siRNA experiment was divided into blank group， negative control group （si-NC group）， KIAA1522-siRNA#1 group， and KIAA1522-siRNA#2 group. The KIAA1522 over-expression experiment was divided into control group， empty vector control group （OE-NC group， transfected with KIAA1522 over-expression empty vector plasmid）， KIAA1522 overexpression group （OE-KIAA1522 group， transfected with KIAA1522 over-expression plasmid）， KIAA1522 over-expression+MK2206 group ［OE-KIAA1522+MK2206 group， co-transfected with KIAA1522 over-expression plasmid and protein kinase B （AKT） signaling pathway inhibitor MK2206］， and MK2206 group （transfected with MK2206）. Western blotting method was used to verify the transfection efficiencies of the cells in various groups； MTT assay was used to detect the proliferation activities of the lung cancer cells in various groups； cell scratch assay was used to detect the migration rates of lung cancer cells in various groups； Transwell chamber assay was used to detect the numbers of invasion lung cancer cells in various groups； Western blotting method was used to detect the expression levels of phosphorylated AKT （p-AKT）， total AKT （t-AKT）， cyclin D1 （Cyclin D1）， vascular endothelial growth factor （VEGF）， and epithelial-mesenchymal transition （EMT）-related proteins ［vimentin （Vimentin）， N-cadherin （N-cadherin）， and E-cadherin （E-cadherin）］ proteins in the cells in various groups. Results The bioinformatics analysis results showed that compared with adjacent normal lung tissue， the expression levels of KIAA1522 mRNA and protein in NSCLC tissue were significantly increased （P<0.05 or P<0.01）. The immunohistochemistry staining results showed that compared with adjacent normal lung tissue， the positive expression rate of KIAA1522 protein in NSCLC tissue was significantly increased （P<0.05） and was associated with TNM stage （P<0.01）. The Western blotting results showed that compared with normal lung epithelial cells BEAS-2B， the expression levels of KIAA1522 protein in lung cancer cell lines PC9， H1299， H460， A549， H1975， and H226 were significantly increased （P<0.05 or P<0.01）. Compared with si-NC group， the expression levels of KIAA1522 protein in the H1299 cells in KIAA1522-siRNA#1 group and KIAA1522-siRNA#2 group were significantly decreased （P<0.01）； compared with OE-NC group， the expression level of KIAA1522 protein in the A549 cells in OE-KIAA1522 group was significantly increased （P<0.01）. The MTT results showed that at 24， 48， and 72 h of cell culture， compared with si-NC group， the proliferation activities of the H1299 cells in KIAA1522-siRNA#1 group and KIAA1522-siRNA#2 group were significantly decreased （P<0.01）； compared with OE-NC group， the proliferation activity of the A549 cells in OE-KIAA1522 group was significantly increased （P<0.05）； compared with OE-KIAA1522 group， the proliferation activity of the A549 cells in OE-KIAA1522+MK2206 group was significantly decreased （P<0.01）； compared with OE-KIAA1522+MK2206 group， the proliferation activity of the A549 cells in MK2206 group was significantly decreased （P<0.05）. The cell scratch assay results showed that compared with si-NC group， the migration rates of the H1299 cells in KIAA1522-siRNA#1 group and KIAA1522-siRNA#2 group were significantly decreased （P<0.01）； compared with OE-NC group， the migration rate of the A549 cells in OE-KIAA1522 group was significantly increased （P<0.01）； compared with OE-KIAA1522 group， the migration rate of the A549 cells in OE-KIAA1522+MK2206 group was significantly decreased （P<0.05）； compared with OE-KIAA1522+MK2206 group， the migration rate of the A549 cells in MK2206 group was significantly decreased （P<0.05）. The Transwell chamber assay results showed that compared with si-NC group， the numbers of invasion H1299 cells in KIAA1522-siRNA#1 group and KIAA1522-siRNA#2 group were significantly decreased （P<0.01）； compared with OE-NC group， the number of invasion A549 cells in OE-KIAA1522 group was significantly increased （P<0.01）； compared with OE-KIAA1522 group， the number of invasion A549 cells in OE-KIAA1522+MK2206 group was significantly decreased （P<0.01）； compared with OE-KIAA1522+MK2206 group， the number of invasion A549 cells in MK2206 group was significantly decreased （P<0.01）. The Western blotting results showed that compared with si-NC group， the expression levels of p-AKT， Cyclin D1， Vimentin， N-cadherin， and VEGF proteins in the H1299 cells in KIAA1522-siRNA#1 group and KIAA1522-siRNA#2 group were significantly decreased （P<0.05 or P<0.01）， while the expression level of E-cadherin protein was significantly increased （P<0.01）； compared with OE-NC group， the expression levels of p-AKT， Cyclin D1， Vimentin， N-cadherin， and VEGF proteins in the A549 cells in OE-KIAA1522 group were significantly increased （P<0.05 or P<0.01）， while the expression level of E-cadherin protein was significantly decreased （P<0.05）； compared with OE-KIAA1522 group， the expression levels of p-AKT， Cyclin D1， Vimentin， N-cadherin， and VEGF proteins in the A549 cells in OE-KIAA1522+MK2206 group were significantly decreased （P<0.05 or P<0.01）， while the expression level of E-cadherin protein was significantly increased （P<0.05）； compared with OE-KIAA1522+MK2206 group， the expression levels of Cyclin D1， Vimentin， N-cadherin， and VEGF proteins in the A549 cells in MK2206 group were significantly decreased （P<0.05 or P<0.01）， while the expression level of E-cadherin protein was significantly increased （P<0.05）. Conclusion The KIAA1522 protein upregulates the expression of Cyclin D1， EMT-related proteins， and VEGF protein in lung cancer cells， promoting the proliferation， migration， and invasion of lung cancer cells， and its mechanism is related to the activation of the AKT signaling pathway.

Objective To observe the clinical effect of tracheal tube with an attached laryngeal pillow cushion （LPC） in the patients under general anesthesia， and to provide new methods for the preventing arytenoid dislocation during tracheal intubation. Methods Forty-eight patients scheduled for elective oral tracheal intubation under general anesthesia and meeting the inclusion criteria were selected. Based on the head and neck positions， the patients were divided into supine without pillow （SWOP） group， supine with pillow （SWP） group， trendelenburg position （TP） group， and head side position （HSP） group， each group consisted of 12 patients. The patient in the following groups underwent two sequential treatments after tracheal intubation： intervention group （LPC-inflated） and control group （LPC uninflated， representing the current method of tracheal tube use）. One patient in TP group and two patients in HSP group rminated the experiment， so a total of 45 patients were successfully included in this study. Electronic fiber laryngoscopy was used to observe and record the positional relationship between the endotracheal tube LPC and the posterior commissure arytenoid joint area under different head and neck positions after two treatments. The evaluation indicators were whether the tracheal tube was lifted from the posterior commissure arytenoid joint area and the degree of compression of the tracheal tube on the arytenoid joint. The incidence of tracheal tube lift-off and the percentage of compression degree on cricoarytenoid joint of the patients in various groups were calculated. Results Within the same head and neck position group， compared with control group， the incidence of endotracheal tube lift-off of the patients in intervention group was significantly increased （P<0.05）， and the percentage of compression degree of endotracheal tube on the arytenoid joint was significantly decreased （P<0.05）. In control and intervention groups， there were no statistically significant differences in the incidence of endotracheal tube lift-off and the percentage of compression degree on arytenoid joint of the patients in various head and neck positions groups （P>0.05）. Conclusion Under the four head and neck positions， inflating the LPC of the tracheal tube can lift the tracheal tube from the posterior commissure arytenoid joint area， effectively relieving or reducing the compression and mechanical friction injuries to the arytenoid joint.

Objective To discuss the relationship between Msh homeobox 1 （MSX1） gene methylation and the clinicopathological characteristics and prognosis of the patients with cervical cancer （CC）， and to analyze the application prospect of MSX1 gene methylation in the diagnosis and treatment of CC. Methods The clinical data， cancer tissues， and adjacent normal tissues were collected from 120 CC patients who underwent surgery between January 2019 and June 2020， and all CC patients were followed up for 3 years； methylation-specific PCR （MSP） was used to detect the methylation status of the MSX1 gene in cancer tissues of the CC patients， who were divided into MSX1 gene hypo-methylation group （n=50） and MSX1 gene hyper-methylation group （n=70）； MSP was used to detect the methylation level of the MSX1 gene in cancer tissues of the CC patients， and the relationship between MSX1 gene methylation status and clinicopathological characteristics and prognosis of the CC patients was analyzed； Western blotting method was used to detect the expression level of MSX1 protein in cancer tissues and adjacent normal tissues of the CC patients； small interfering RNA （siRNA） targeting MSX1 mRNA was designed； the CC cells were divided into control group （transfected with blank vector） and siMSX1 group （transfected with MSX1 siRNA）； real-time fluorescence quantitative PCR （RT-qPCR） method was used to verify the cell transfection efficiency； cell scratch assay was used to detect the migration activity of the cells in two groups； Logistic regression was used to analyze the influencing factors of prognosis in the CC patients. Results The methylation rate in cancer tissues of the CC patients was 58.33%， which was significantly higher than that in adjacent normal tissues （11.67%， χ2=42.725， P<0.01）； the Western blotting results showed that compared with adjacent normal tissue， the expression level of MSX1 protein in cancer tissue of the CC patients was significantly decreased （P<0.01）； high methylation status of the MSX1 gene in cancer tissue of the CC patients was associated with high-risk human papillomavirus （HR-HPV） DNA， lymph node metastasis， and TNM stage （P<0.05）， but not associated with patient age， pathological type， and tumor size （P>0.05）； the cell scratch assay results showed that compared with control group， the expression level of MSX1 mRNA in the cells in siMSX1 group was significantly decreased （P<0.01）， suggesting successful knockout of the MSX1 gene； the cell scratch assay results showed that compared with control group， the migration activity of the cells in siMSX1 group was significantly increased （P<0.01）； the 3-year cumulative survival rate of patients with MSX1 gene methylation was 54.29%， which was significantly lower than that of patients without MSX1 gene methylation （80.00%， χ2=9.717， P=0.002）； the Logistic regression results showed that positive HR-HPV DNA， lymph node metastasis， TNM stage Ⅲ-Ⅳ and MSX1 gene high methylation were independent risk factors for prognosis in CC patients （P<0.05）. Conclusion MSX1 gene methylation is associated with poorer clinicopathological characteristics and adverse prognosis in the CC patients， suggesting its potential as a biomarker for cervical cancer.

Objective To discuss the characteristics of age， antral follicle count （AFC）， anti-Müllerian hormone （AMH）， sex hormones， and glycolipid metabolism in the infertile patients with different phenotypes of polycystic ovarian syndrome （PCOS）， and to improve the outcomes of assisted reproductive technology （ART）. Methods A total of 11 660 infertile female patients treated in our hospital were selected as the research subjects， including 3 110 PCOS patients and 8 550 non-PCOS patients. According to the Rotterdam criteria and inclusion/exclusion criteria， the subjects were divided into PCOS group （2 261 PCOS patients） and control group （1 871 non-PCOS patients）. The PCOS group was further divided into four phenotypes： type A （345 cases， oligo-ovulation or anovulation （OA）+hyperandrogenemia or clinical hyperandrogenism （HA）+polycystic ovary morphology （PCO））， type B （204 cases， OA+HA）， type C （102 cases， HA+PCO）， and type D （1 610 cases， OA+PCO）. Chemiluminescent immunoassay was used to detect the serum AMH levels of the subjects in various groups； glucose oxidase method and biochemical method were used to detect the serum levels of triglycerides （TG）， total cholesterol （TCHO）， fasting blood glucose （FBG）， and fasting insulin （FINS） of the subjects in various groups； chemiluminescence method was used to detect the serum basal sex hormone levels of the subjects in various groups； transvaginal ultrasound was used to detect the AFC of the subjects in various groups. Results Compared with control group， the age and serum basal follicle-stimulating hormone （bFSH） levels of the subjects in different PCOS phenotype groups were significantly decreased （P<0.01）， while AFC and serum levels of AMH， total testosterone （TESTO）， and basal luteinizing hormone （bLH） of the subjects were significantly increased （P<0.01）. Compared with type A PCOS group， the AFC and serum levels of AMH and bLH of the subjects in type B， C， and D PCOS groups were significantly decreased （P<0.01）. Compared with control group， the serum levels of TG， TCHO， FBG， and FINS， as well as homeostasis model assessment of insulin resistance （HOMA-IR） of the subjects in type A and D PCOS groups were significantly increased （P<0.01）； the serum levels of FBG and FINS， as well as HOMA-IR of the subjects in type B PCOS group were significantly increased （P<0.01）； the serum TG level of the subjects in type C PCOS group was significantly increased （P<0.01）. Compared with type A PCOS group， the serum levels of TG and FINS， as well as HOMA-IR of the subjects in type B， C， and D PCOS groups were significantly decreased （P<0.01）. Conclusion The patients with different PCOS phenotypes exhibit distinct basal sex hormone levels and glycolipid metabolism characteristics. Phenotypic classification of PCOS infertile patients helps predict disease severity， and personalized pretreatment should be performed for different PCOS phenotypes before ART.

Objective To explore the risk factors associated with aneurysmal subarachnoid haemorrhage （aSAH） complicated with acute hydrocephalus （aHCP）， and to provide the clinical reference for the early identification and intervention of these patients. Methods The clinical data and laboratory indexes of 175 patients with aSAH were retrospectively analysed， and the patients were divided into aHCP group （n=56） and non-aHCP group （n=119） according to whether they presented with aHCP after the onset of the disease. Univariate analysis and binary logistic regression were applied to identify the risk factors for the aHCP in aSAH patients， and receiver operating characteristic （ROC） curve analysis with area under the curve （AUC） was used to evaluate the predictive value of these factors. Results A total of 56 （32.0%） out of 175 aSAH patients included developed aHCP after the onset of the disease. Compared with non-aHCP group，the levels of neutrophil count， blood glucose， neutrophil-albumin ratio （NAR）， platelet-lymphocyte ratio （PLR）， neutrophil-lymphocyte ratio （NLR）， monocyte-lymphocyte ratio （MLR）， systemic immune-inflammatory index （SII）， systemic inflammatory response index （SIRI）， and systemic inflammation composite index （AISI） of the patients in aHCP group were significantly increased（P<0.05）， the level of lymphocyte count was significantly decreased （P<0.05）， the Hunt-Hess grade and modified Fisher grade were higher （P<0.05）， and the incidence of ventricular haematochezia was more high （P<0.05）. The binary Logistic regression analysis results showed that the elevated levels of NAR （OR=2.237， 95%CI： 1.063-4.708， P=0.034） and NLR （OR=1.210， 95%CI： 1.095-1.337， P<0.01） were the independent risk factors for the development of aHCP after aSAH. The ROC curve analysis showed that the AUC of NAR was 0.812 （95%CI： 0.745-0.878， P<0.001）， the AUC of NLR was 0.844 （95%CI： 0.785-0.903， P<0.001）， and the combined AUC of NAR and NLR was 0.854 （95%CI： 0.798-0.910， P<0.001）. Conclusion NAR and NLR are independent risk factors for the development of aHCP in aSAH patients.

Objective To discuss the efficacy of temporomandibular joint disc anchoring （TMJDA） in the treatment of anterior disc displacement without reduction （ADDWoR） from clinical and imaging perspectives， in order to improve the clinicians’ understanding of this surgical approach. Methods Twenty-one ADDWoR patients who underwent TMJDA were retrospectively collected， involving a total of 25 temporomandibular joints （TMJs）. The maximum mouth opening， visual analogue scale （VAS） score for pain， and Helkimo index were measured preoperatively， 1 month postoperatively， and 6 months postoperatively in all the patients. Postoperative complications and satisfaction questionnaires were designed for the patients to self-evaluate the efficacy， and their magnetic resonance imaging （MRI） findings were assessed. Results Compared with preoperative period， the maximum mouth opening at 1 month and 6 months postoperatively was significantly increased （P<0.05）； compared with 1 month postoperatively， the maximum mouth opening at 6 months postoperatively was significantly increased （P<0.05）. Compared with preoperative period， the VAS scores at 1 month and 6 months postoperatively were significantly decreased （P<0.05）； compared with 1 month postoperatively， the VAS score at 6 months postoperatively was significantly decreased （P<0.05）. Compared with preoperative period， the percentages of the patients with Di 0 and DiⅠ scores were significantly increased （P<0.05）， while those with DiⅡ and DiⅢ scores were significantly decreased （P<0.05）， indicating significant improvement in TMJ function after surgery. Among the 21 ADDWoR patients， 14 （66.67%） were satisfied and 7 （33.33%） were basically satisfied. Compared with preoperative period， the disc length was significantly increased postoperatively （P<0.01）， while no significant difference was observed in condylar height （P>0.05）； all displaced discs were repositioned postoperatively； among the 25 joints， 23 （92.00%） were evaluated as “excellent” and 2 （8.00%） were evaluated as “good”. No patients experienced postoperative facial nerve injury， local alopecia， surgical area depression， salivary fistula， or Frey syndrome； 3 patients （3 joints） developed numbness in the preauricular area within 24 hours postoperatively， which resolved by the 6-month follow-up. Conclusion TMJDA for the treatment of ADDWoR can stably reposition the disc， significantly improve mouth opening and pain levels， with a low incidence of postoperative complications.

Objective To discuss the diagnostic value of levels of growth stimulation expressed gene 2 protein （ST2）， carbohydrate antigen 125 （CA125）， and human epididymis protein 4 （HE4） in serum of the ovarian cancer patients， and to clarify their relationships with clinicopathological parameters of the ovarian cancer patients. Methods A total of 136 patients with ovarian benign lesions or primary ovarian malignancies in our hospital confirmed by postoperative histopathology were randomly selected， including 53 cases in ovarian benign ovarian disease group and 83 cases in ovarian cancer group. Additionally， 55 healthy female volunteers during the same period were enrolled as healthy control group. The fasting venous blood from all the subjects was collected on the first day of admission， and serum was retained； cyclic enhanced fluorescence immunoassay was used to detect ST2 levels of the subjects in various groups； chemiluminescence method was used to detect serum CA125 and HE4 levels of the subjects in various groups； receiver operating characteristic （ROC） curve was used to assess the diagnostic performance of each indicator； the Cut-off value and area under the ROC curve （AUC） were calculated， with AUC representing the diagnostic performance； Kendall’s method was used to analyze the correlations between serum ST2 levels and TNM stage， maximum tumor diameter， distant metastasis， lymphnode metastasis， peritoneal metastasis， carcinoembryonic antigen （CEA）， CA125， carbohydrate antigen 199 （CA199）， HE4， P53， and Kiel-67（Ki67） in the ovarian cancer patients. Results Compared with healthy control group， the serum CA125 level of the patients in ovarian benign ovarian disease group was significantly increased （P<0.05）. Compared with healthy control group and ovarian benign group， the serum levels of ST2， CA125， and HE4 of the patients in ovarian cancer group were significantly increased （P<0.05）. The AUC values were 0.719 （95%CI： 0.616-0.822） for ST2， 0.868 （95%CI： 0.794-0.942） for CA125， and 0.867 （95%CI： 0.793-0.942） for HE4. The combined detection of ST2+CA125+HE4 yielded an AUC of 0.894 （95%CI： 0.832-0.955）. Significant differences were observed in serum ST2， CA125， and HE4 levels among ovarian cancer patients with different TNM stages， lymph node metastasis， distant metastasis， and peritoneal metastasis （P<0.05）， while there were no significant differences in the patients with different ages or maximum tumor diameters （P>0.05）. The ST2 level in serm of the ovarian cancer patients was positively correlated with TNM stage， distant metastasis， lymph node metastasis， peritoneal metastasis， CA125 level， HE4 level， and Ki67 level （P<0.05）， but was not correlated with maximum tumor diameter， CEA， CA199 level， or P53 level （P>0.05）. Conclusion ST2， CA125， and HE4 are highly expressed in the serum of the ovarian cancer patients. Combined detection of serum ST2， CA125， and HE4 exhibits good sensitivity and specificity for ovarian cancer screening and improves diagnostic efficacy. ST2 is associated with advanced TNM stage and metastasis in ovarian cancer and may participate in the occurrence and progression of ovarian cancer.

Crigler-Najjar syndrome （CNS） is an autosomal recessive genetic disorder caused by mutations in the UGT1A1 gene. This author retrospectively analyzes the clinical data of one patient presenting with postnatal hyperbilirubinemia， who was genetically diagnosed with CNS. Her clinical course， genetic characteristics， diagnostic and therapeutic approaches are discussed to enhance clinicians’ understanding of this condition. The patient exhibited recurrent jaundice due to elevated unconjugated bilirubin （UCB） levels since birth. During this period， the gallstones were identified and surgically removed， yet her bilirubin levels did not improve. In March 2024， the patient presented to our hospital and underwent genetic testing via a third-party facility. The results revealed two risk variants in the UGT1A1 gene. After one week of oral phenobarbital administration， her UCB levels decreased by approximately 47%， confirming a diagnosis of CNS type Ⅱ. The patient is currently under active follow-up with regular monitoring of bilirubin levels. For the infants and children presenting with persistent， fluctuating， non-hemolytic， and non-hepatitic hyperbilirubinemia since birth， early genetic testing should be prioritized to establish the definitive diagnosis.

Adult-onset Still’s disease （AOSD） is a rare autoinflammatory disease characterized by fever， rash， arthritis， liver， spleen and lymph node enlargement， increased total number of peripheral white blood cells and neutrophil ratio. This paper reported a case of AOSD with dermal lymphadenitis （DL） complicated with hemophagocytic syndrome （HPS）， in order to improve the clinicians’ understanding for this complicated complication. The patient was a 48-year-old female who was admitted to the hospital with the complant of “intermittent fever with rash for 15 d”. After 10 d of active anti-infection treatment， the symptoms were not improved， and there were new large congestive edematous erythema on the face and trunk， muscle pain in limbs and joints， and spleen enlargement. Laboratory tests showed increased white blood cell count， significantly decreased platelet count， hypofibrinogenemia， elevated serum ferritin， and elevated soluble interleukin-2 receptor sCD25； DL was pathologically diagnosed by axillary lymph node biopsy. After excluding other diseases， the diagnosis was confirmed as AOSD with DL complicated with HPS. After diagnosis of HPS， the patient was treated with hemophagocytic lymphohistiocytosis（HLH）- 1994 regimen combined with rucotinib for 6 weeks， and the symptoms were improved； the patrent was discharged. The diagnosis of AOSD is particularly complex when complicated with the complications such as HPS， which requires carefully differential diagnosis， especially to exclude lymphoma. The cases of AOSD with DL are rare， and its etiology and pathogenesis need further study； early diagnosis and multidisciplinary collaboration are essential to improve the patient’s prognosis.

Objective To clarify the changes in incidence and mortality of various cancers based on analysis on registration data of malignant tumor incidence and mortality from Dehui City and Yanji City in Jilin Province. Methods The incidence and mortality data of malignant tumors from 2009 to 2016 in Dehui City and Yanji City in Jilin Province， were collected from the Chinese Cancer Registry Annual Report published by the National Cancer Center. The number of cases， deaths， crude incidence rate， crude mortality rate， age-standardized incidence rate （ASIR）， age-standardized mortality rate （ASMR）， and annual percentage change （APC） of the malignant tumors were analyzed by cancer sites and genders. Results From 2009 to 2016， the CIR of malignant tumors in Dehui City （APC=1.2%， P=0.019） and Yanji City （APC=3.6%， P=0.058） showed an increasing trend. After standard population age adjustment， the ASIR in males in Dehui City showed a significant decline （APC=-5.7%， P=0.021）， while the ASIR in females exhibited an overall downward trend， but the difference was not significant （APC=-2.2%， P=0.111）. In Yanji City， the ASIR in males （APC=-1.4%， P=0.535） and females （APC=0.0%， P=0.988） showed no significant changes. The CMR of malignant tumors in Dehui City （APC=1.9%， P=0.001） and Yanji City （APC=5.9%， P=0.001） showed a continuous upward trend. After age-standardization， the ASMR in males （APC=-3.1%， P=0.100） and females （APC=-4.2%， P=0.053） in Dehui City， as well as in males （APC=-1.3%， P=0.438） in Yanji City， showed a slight downward trend. Although the ASMR in females in Yanji City showed a slight increase， the difference was not statistically significant （APC=0.5%， P=0.838）. In 2016， the most common malignant tumor in terms of both incidence and mortality in Dehui City was lung cancer， with a CIR of 60.76/100 000 and a CMR of 46.96/100 000. In Yanji City， the most common malignant tumor was liver cancer， with a CIR of 49.04/100 000 and a CMR of 51.09/100 000. Conclusion Lung cancer， liver cancer， and gastric cancer are the major malignant tumors threatening residents in Dehui City， Yanji City， and even the entire Jilin Province， and should be prioritized in cancer prevention and control efforts. Early diagnosis and treatment should be strengthened.

Objective To discuss the differences in depression symptoms between genders and between only-children and non-only-children among adolescents through symptom network analysis of adolescent depression. Methods A total of 650 adolescents were randomly selected from Chaoyang District， Changchun City， Jilin Province，and they were as the study samples. The Patient Health Questionnaire-9 （PHQ-9） was used to assess depression symptoms in the adolescents； network analysis was performed on the survey results to identify the core symptoms of adolescent depression and to compare gender differences and differences between only-children and non-only-children in depression symptoms. Results The core symptoms of the adolescent depression were depressed mood ［Closeness（Clo）=1.700 5， Strength（Str）=1.722 8］ and lack of energy ［Betweenness（Bet）=1.875 1］. No significant differences in core symptoms were observed between males and females， but significant differences were found between only-children and non-only-children. Depression manifestations in the only-children also included psychomotor agitation or retardation. In the depression symptom network of the adolescents， the strongest edge connection was between anhedonia and depressed mood ［Edge strength（Edge strength）=0.322 5］， and significant differences were observed between genders and between only-children and non-only-children. The symptom network in male adolescents was consistent with the overall pattern， whereas in female adolescents， the strongest connection was the biggest between psychomotor agitation or retardation and suicidal ideation or self-harm （Edge strength=0.320 5）. The symptom network in only-children was consistent with the overall pattern， whereas in non-only-children， the strongest connection was between depressed mood and feelings of worthlessness （Edge strength=0.287 4）. Conclusion The core symptoms of the adolescent depression exhibit differences between only-children and non-only-children， and the edge strengths in the symptom network show variations by gender and between only-children and non-only-children.

Objective To synthesize a hydrolysis-resistant urushiol-modified monomer （UMM） to improve the hydrolysis resistance of light-cured composite resin， while reducing the volume shrinkage rate （VS）， increasing the double bond conversion rate （DC）， and mitigating the potential biosafety concerns of bisphenol A glycidyl methacrylate （Bis-GMA） monomer. Methods UMM was synthesized by modifying urushiol via an acyl chloride reaction， and its structure was analyzed and characterized using Fourier transform infrared spectroscopy （FT-IR）. Control group was consisted of Bis-GMA/triethylene glycol dimethacrylate （TEGDMA） without UMM， while 10%UMM， 15%UMM， and 20%UMM groups were prepared by partially replacing Bis-GMA with UMM at mass fractions of 10%， 15%， and 20%， respectively. The viscosity of UMM was measured using a rheometer. The DC of light-cured composite resin in various groups was detected by FT-IR spectroscopy， and the VS was calculated. The contact angle of light-cured composite resin in various groups was measured using the sessile drop method， and the water sorption and solubility values were calculated. The mechanical properties of light-cured composite resin in various groups were tested. The in vitro cytotoxicity of light-cured composite resin in various groups was evaluated using the cell counting kit-8 （CCK-8） assay. Results The FT-IR spectra results showed that the absorption peak of the hydroxyl group at 3 402 cm^-1 disappeared， while characteristic absorption peaks of -C=O and -C=C appeared at 1 745 and 1 637 cm^-1， indicating that urushiol successfully reacted with acryloyl chloride to form UMM. The viscosity of UMM ranged from 25.14 to 29.43 Pa·s. Compared with control group， the DC of light-cured composite resin in 10%UMM， 15%UMM， and 20%UMM groups was significantly increased （P<0.05）， while the VS was significantly decreased （P<0.05）， both in a dose-dependent manner. Compared with control group， the contact angle of light-cured composite resin in 10%UMM， 15%UMM， and 20%UMM groups was significantly increased （P<0.05）. Compared with 10%UMM group， the contact angle of light-cured composite resin in 15%UMM and 20%UMM groups was further increased （P<0.05）. Compared with control group， the water sorption and solubility values of light-cured composite resin in 10%UMM， 15%UMM， and 20%UMM groups were significantly decreased （P<0.05）， showing a dose-dependent trend. After 24 h of water immersion， compared with control group， the flexural strength （FS） and elastic modulus （EM） of light-cured composite resin in 10%UMM， 15%UMM， and 20%UMM groups were significantly decreased （P<0.05）， also in a dose-dependent manner. After 7 d of water immersion， compared with control group， the FS of light-cured composite resin in 10%UMM group was significantly increased （P<0.05）， while that in 20%UMM group was significantly decreased （P<0.05）. Compared with 10%UMM group， the FS of light-cured composite resin in 15%UMM and 20%UMM groups was significantly decreased （P<0.05）， exhibiting a dose-dependent trend. Compared with control group， the EM of light-cured composite resin in 15%UMM and 20%UMM groups was significantly decreased （P<0.05）， also in a dose-dependent manner. The relative growth rate （RGR） of the L929 cells in control， 10%UMM， 15%UMM， and 20%UMM groups was above 90%， with no statistically significant differences among groups （P>0.05）， and all cytotoxicity results were qualified. Conclusion A novel low-viscosity monomer UMM is successfully synthesized in this study. All UMM-containing light-cured composite resin formulations exhibit higher DC， lower VS， reduced water sorption and solubility values， improved hydrolysis resistance， and low cytotoxicity. UMM can serve as a potential resin monomer to enhance the hydrolysis resistance of light-cured composite resin.

Protein phosphatase 1 （PP1） is a widely expressed and highly conserved serine/threonine phosphatase in organisms. It regulates cellular signaling pathways by catalyzing the dephosphorylation of various proteins， thereby influencing biological processes such as cell proliferation， apoptosis， migration， and transcription. In vivo， PP1 does not exist as a free catalytic subunit but instead forms distinct PP1 holoenzymes by binding with at least one PP1-interacting protein （PIP）. The interaction between PP1’s catalytic subunit and its specific regulatory proteins is central to PP1’s function. Under normal conditions， PP1 stably performs its dephosphorylation role in vivo； however， in tumors， PP1 function is aberrantly regulated， leading to either increased or decreased PP1 activity. PP1 exerts a dual influence on tumorigenesis and progression， acting as a suppressor in some cancers while promoting oncogenesis in others. Based on domestic and international research findings on PP1， this review summarizes the structure and biological functions of PP1， as well as the impact of its various subunits on the development and progression of different cancers， including breast cancer， lung cancer， ovarian cancer， pancreatic adenocarcinoma （PAAD）， liver cancer， endometrial cancer， esophageal cancer （EC）， colorectal cancer， and glioblastoma （GBM）. This review aims to provide the insights for developing highly efficient and environmentally friendly anticancer drugs and therapeutic approaches targeting PP1 holoenzymes.

Human immunodeficiency virus type 1 （HIV-1） latent reservoirs pose the primary obstacle to curing acquired immunodeficiency syndrome （AIDS）. Transactivator（Tat）， a regulatory protein encoded by HIV-1， influences the establishment and reactivation of latent reservoirs by promoting viral transcription. Inhibitors targeting Tat protein suppress viral rebound by reducing Tat protein levels and disrupting its transcription-promoting functions. Among the two strategies for treating HIV-1 latent reservoirs， the “block-and-lock” strategy， which is based on Tat protein inhibitors， aims to target HIV-1 proteins or host factors while interfering with histone epigenetic modifications， thereby permanently silencing proviral DNA and maintaining HIV-1 latency even after treatment discontinuation. Tat-related inhibitors such as didehydrocortistatin A （dCA）， triptolide， and apalutamide-derived Q308 regulate HIV-1 latency through distinct mechanisms. This review summarizes the regulatory roles of Tat in HIV-1 latency， Tat protein inhibitor-based therapeutic strategies for targeting latent reservoirs， and the mechanisms of action of related inhibitors， with the goal of providing insights for the development of drugs toward achieving functional HIV-1 cure.

Female reproductive system diseases， such as premature ovarian insufficiency （POI）， premature ovarian failure （POF）， polycystic ovary syndrome （PCOS）， intrauterine adhesions （IUA）， ulterus scar diverticulum， salpingitis， and tubal obstruction， may induce infertility， severely impacting patients’ physical and mental health and quality of life. Currently， the umbilical cord mesenchymal stem cells （UCMSCs） have emerged as a research focus in gynecological and obstetric fields， demonstrating significant therapeutic potential for female reproductive system disorders. UCMSCs secrete various cytokines， activate relevant signaling pathways and key molecules， reduce inflammation mediators and oxidative stress， and prevent excessive cellular damage and apoptosis， thereby achieving therapeutic effects. In recent years， extensive studies have explored the therapeutic effects of UCMSCs on female reproductive system diseases. This article review the current in vitro and in vivo research progress in UCMSCs for treating female reproductive system diseases， aiming to provide the novel strategies and directions for future research and clinical applications.

Compared with traditional obesity evaluation indexes， the weight-adjusted waist index （WWI） is a combination of waist circumference （WC） and the square root of body mass.WWI can accurately reflect the relationship between abdominal fat accumulation and body mass changes in the obese patients， and it can predict the risk of obesity-related diseases， such as type 2 diabetes mellitus （T2DM）， hyperuricemia， heart failure， abdominal aortic calcification （AAC）， erectile dysfunction （ED） in the males，osteoarthritis（OA）， asthma， dementia and stroke， and may be an independent determinant of left ventricular hypertrophy （LVH） and an independent risk factor for stroke in Chinese adult hypertensive patients. The correlations between WWI and the risk assessment and prediction of obesity-related diseases， such as endocrine metabolic diseases， cardiovascular diseases（CVD）， reproductive diseases， neurological diseases， asthma， osteoarthritis， and non-alcoholic liver fibrosis （NAFLD）， were now analysed in cornbination of the recent domestic and international related studies， with the aim of providing references for the further study of the assessment and prediction of obesity-related diseases.