Objective To investigate the protective effect of astragaloside Ⅳ（AS-Ⅳ） on the lipopolysaccharide（LPS）-induced oxidative injury of osteoblasts precursor cells MC3T3-E1， and to clarify its related mechanism. Methods The MC3T3-E1 cells in logarithmic growth phase were divided into control group， LPS group and different concentrations （10，25，50 and 100 mg·L^－1） of AS-Ⅳ groups. CCK-8 method was used to detect the cell activities in various groups.The MC3T3-E1 cells in logarithmic growth phase were divided into control group， LPS group， AS-Ⅳ group， AS-Ⅳ+LY294002 ［phosphatidylinositol-3-kinase（PI3K）/protein kinase B（Akt）signaling pathway inhibitor］group. Flow cytometry was used to detect the levels of reactive oxygen species （ROS） in the cells in various groups. The levels of malondialdehyde （MDA） and glutathione（GSH） and the activities of superoxide dismutase（SOD） in the cells in various groups were determined with the related kits. The expression levels of heme oxygenase （HO-1） mRNA in the cells in various groups were detected by real-time fluorescence quantitative PCR（RT-qPCR） method. Western blotting method was used to detect the expression levels of PI3K，phosphorylated Akt（p-Akt），and nuclear factor-E2 related factor 2（Nrf2），B-cell lymphoma-2 （Bcl-2）， and Bcl-2 associated X protein（Bax） in the cells in various groups. Results The CCK-8 method results showed that compared with control group， the cell activity in LPS group was significantly decreased （P<0.01）； compared with LPS group， the activities of cells in 50 and 100 mg·L^－1 AS-Ⅳ groups were significantly increased （P<0.01）. Compared with control group， the ROS and MDA levels in the cells in LPS group were significantly increased （P<0.01）， while the SOD activity and GSH level were decreased （P<0.01）； compared with LPS group， the ROS and MDA levels in the cells in AS-Ⅳ group were significantly decreased （P<0.01）， the SOD activity and GSH level were significantly increased （P<0.01）； compared with AS-Ⅳ group， the ROS and MDA levels in the cells in AS-Ⅳ+LY294002 group were significantly increased （P<0.01）， the SOD activity and GSH level were significantly decreased （P<0.01）. Compared with control group， the expression level of HO-1 mRNA and the expression levels of p-Akt， Nrf2， HO-1， and Bcl-2 proteins in the cells in LPS group were significantly decreased（P<0.01）， while the expression level of Bax protein was significantly increased （P<0.01）； compared with LPS group， the expression level of HO-1 mRNA and the expression levels of p-Akt， Nrf2， HO-1，and Bcl-2 proteins in the cells in AS-Ⅳ group were signicantly increased （P<0.01）， while the expression level of Bax protein was significantly decreased （P<0.01）；compared with AS-Ⅳ group， the expression level of HO-1 mRNA and the expression levels of p-Akt，Nrf2 HO-1 and Bcl-2 proteins in the cells in AS-Ⅳ+ LY294002 group were significantly decreased （P<0.01），and the expression level of Bax protein was significantly increased （P<0.01）. Conclusion AS-Ⅳ can play protective effect on the LPS-induced osteoblast injury by mediating PI3K/Akt/Nrf2 signaling pathway.

Objective To investigate the ameliorative effects of sulfur dioxide （SO₂） on myocardial fibrosis in the type 2 diabetes mellitus （T2DM） rats induced by streptozocin （STZ） and high glucose and high fat diet and the effects on apoptosis and transforming growth factor-β1（TGF-β1）/Smad family member 2/3（Smad2/3） pathway， and to clarify the possible molecular mechanisms of SO₂ in ameliorating diabetic myocardial fibrosis. Methods Forty male SD rats were randomly divided into control group， T2DM group， T2DM+SO₂ group， and SO₂ group. Except for control group and SO₂ group， the rats in T2DM and T2DM+SO₂ groups were used to establish the T2DM rat models by intraperitoneal injection of STZ （35 mg·kg^－1） and high glucose and high fat diet， and the rats in T2DM+SO₂ and SO₂ groups were injected intraperitoneally with exogenous SO₂ donors ［sodium sulfite（Na₂SO₃）/sodium bisulfite（NaHSO₃），85 mg·kg^－1·d^－1］ for 8 weeks； the pathomorphology of myocardial fibrosis of the rats in various groups was observed by Masson staining， and the myocardial collagen volume fractions of the rats in various groups were calculated， and the expression levels of relevant proteins in myocardial tissue of the rats in various groups were detected by Western blotting method. Results Compared with control group， the myocardial tissue of the rats in T2DM group showed morphology and structural disorders， the myocardial collagen fiber formation was significantly increased， the myocardial collagen volume fraction was significantly increased （P<0.01）， the expression levels of α-smooth muscle actin（α-SMA） and type Ⅲ collagen（Col Ⅲ），B-cell lymphoma-2（Bcl-2） associated X protein（Bax）， TGF-β1， Smad2， and Smad3 proteins were significantly increased （P<0.01）， and the expression level of Bcl-2 protein was significantly decreased （P<0.01）. Compared with T2DM group， the morphological structure of myocardial tissue of the rats in T2DM+SO₂ group was improved， the myocardial collagen fiber formation and the myocardial collagen volume fraction were significantly decreased（P<0.01）， the expression levels of α-SMA， Col Ⅲ， Bax， TGF-β1， Smad2， and Smad3 proteins were significantly decreased（P<0.01），and the expression level of Bcl-2 protein was significantly increased （P<0.01）. Compared with control group， there were no statistically significant differences in the above indexes of the rats in SO₂ group rats（P>0.05）. Conclusion SO₂ can alleviate myocardial fibrosis in the T2DM rats by inhibiting the expressions of pro-apoptosis-related proteins and TGF-β1/Smad2/3 pathway proteins.

Objective To investigate the effect of ring finger protein 31 （RNF31） on the expression of epidermal growth factor receptor （EGFR）， and to explore the regulation mechanism of RNF31 on EGFR. Methods The interaction between RNF31 and EGFR proteins was validated by co-immunoprecipitation （Co-IP） method.The HEK293T cells were divided into empty vector group（transfcted with pcDNA3.1 emptey vector）and RNF31 group（transfected with RNF31-Flag plasmid）. The expression levels of EGFR mRNA and protein in the cells were detected by real-time fluorescence quantitative PCR （RT-qPCR） and Western blotting methods.After mutation of the ubiquitin ligase function of RNF31 （RNF31^C699/702S）and deubiquitinase binding site （RNF31^{N84A Y93A}），Western blotting method was used to detect the expression levels of EGFR protein in the cells in various groups.The HEK293T cells were divided into empty vector group，RNF31 group（transfected RNF31-Flag plasmid），RNF31+EGFR inhibitor Ggfitinib（Gefitinib） group and RNF31+EGFR inhibitor Erlotinib（Erlotinib） group； the expression levels of EGFR and its downstream sigaling pathway proteins in the cells in various groups were detected by Western blotting method. Results The Co-IP method detection results confirmed that the RNF31 was interacted with EGFR. Compared with empty vector group， the expression level of EGFR mRNA in the cells in RNF31 group was significantly decreased （P<0.05）. Compared with empty vector group， the expression levels of EGFR protein in the cells in RNF31^C699/702S group and wild-type RNF31 group were significantly increased（P<0.05）. Compared with RNF31 group，the expression level of EGFR protein in the cells in transfection RNF31^{N84A Y93A} group was decreased （P<0.05）. Compared with empty vector group，the expression levels of EGFR and nuclear transcription factor-κB（NF-κB）， phosphorylated signal transducer and activiator of transcription 3（p-STAT3），mitogen-activated protein kinase（MAPK） and phosporylated MAPK（p-MAPK） proteins in its downstream signling pathways in the cells in RNF31 group were significantly increased（P<0.05）；the expression levels of protein kinase B（Akt），phosphorylated Akt（p-Akt），STAT3 and MAPK proteins in the cells in RNF31+Gefitinib group and RNF31+Erlotinib group were significantly decreased（P<0.05）. Conclusion RNF31 exerts deubiquitinating through the deubiquitinase enzyme-binding struatural domain， reduces the ubiquitination level of the EGFR protein molecule surface， and slows down the degradation rate of EGFR protein， while highly expressed EGFR can activate the expressions of its downstream proteins related to cell proliferation and division.

Objective To investigate the protective effect of rosaminic acid （RA） on astrocyte injury induced by β-amyloid protein 1-42 （Aβ_1-42） and the inhibitory effect of the inflammatory factor release， and to clarify the related mechanism. Methods After primary culture，the astrocytes were divided into control group， Aβ_1-42 group （5 μmol?L^－1）， different concentrations（20，50，100 μmol?L^－1） of RA+Aβ_1-42 groups， nuclear factor E2-related factor 2（Nrf2） inhibitor ML385 group，and ML385+RA+Aβ_1-42 group.The astrocytes were pretreated with different concentrations （20，50，100 μmol·L^－1） of RA for 24 h and then treated with Aβ_1-42 at a final concentration of 5 μmol·L^－1 for 24 h，and the final concentration of 10 μmol·L^－1 ML385 was pretreated for 6 h before added RA in ML385 group. The cell activities in various groups were detected by MTT method， the lactate dehydrogenase（LDH）release rates of the cells in various groups were detected by LDH kits， the levels of interleukin-1β（IL-1β） and tumor necrosis factor-α（TNF-α） in the cells in various groups were detected by enzyme linked immunosorbent assay （ELISA） kits， the levels of nitric oxide （NO） in culture medium in various groups were detected by Griess method， the expressions of glial fibrillary acidic protein （GFAP） and Nrf2 in the cells in various groups were detected by immunofluorescence method，and the expression levels of GFAP， Nrf2 and heme oxygenase-1 （HO-1） proteins in the cells in various groups were detected by Western blotting method. Results Compared with control group， the cell activity in Aβ_1-42 group was significantly decreased（P<0.01）， the LDH release rate and the levels of IL-1β， TNF-α， and NO were significantly increased（P<0.01）， the fluorescence intensity of GFAP was significantly enhanced，the fluorescence intensity of Nrf2 was significantly reduced，the expression level of GFAP protein in the cells was significantly increased（P<0.01），and the expression levels of Nrf2 and HO-1 proteins were significantly decreased （P<0.01）. Compared with Aβ_1-42 group， the cell activities in different concentrations of RA+Aβ_1-42 groups were significantly increased （P<0.01）， the LDH release rates and the levels of IL-1β，TNF-α， and NO were significantly decreased （P<0.01）， the fluorescence intensities of GFAP were significantly decreased，the fluorescence intensities of Nrf2 were significantly enhanced，the expression levels of GFAP protein were significantly decreased（P<0.01）， and the expression levels of Nrf2 and HO-1 proteins were significantly increased （P<0.01）. Compared with RA （50 μmol?L^－1） +Aβ_1-42 group， the cell activity in ML385+Aβ_1-42+RA group was significantly decreased（P<0.01）， the LDH release rate and the levels of IL-1β， TNF-α， and NO were significantly increased（P<0.01）， the fluorescence intensity of GFAP was significantly enhanced，the fluorescence intensity of Nrf2 was significantly，the expression level of GFAP protein was significantly increased （P<0.01），and the expression levels of Nrf2 and HO-1 proteins were significantly decreased （P<0.01）. Conclusion RA can inhibit the release of the inflammatory factors and NO， the mechanism of protecting astrocytes from damage may be related to the activation of Nrf2 / HO-1 pathway.

Methods Forty male SD rats were randomly divided into control group， diabetic periodontitis model group（model group）， low dose （100 mg·kg^－1） of GSPE group and high dose （200 mg·kg^－1） of GSPE group； there were 10 rats in each group. Except for control group， the rats in other groups were used to establish the diabetic periodontitis models by periodontal ligation combined with intraperitoneal injection of streptozotocin （STZ）. After the successful establishment of the models， the rats in low and high doses of GSPE groups were given 100 and 200 mg·kg^－1 GSPE， respectively， while the rats in control group and model group were given the same amount of normal saline once a day for 8 weeks. The general situation of the rats and the changes of blood glucose levels were observed， the pathomorphology of periodontal tissue of the rats in various groups were observed by HE staining， the alveolar bone absorption of the rats in various groups was measured， the activities of superoxide dismutase （SOD） and the catalase （CAT） and the levels of reactive oxygen species （ROS） and malondialdehyde （MDA） in periodontal tissue of the rats in various groups were detected by related kits.Enzyme-linked immunosorbent assay（ELISA） method was used to detect the levels of tumor necrosis factor-α （TNF-α）， interleukin-1β（IL-1β） and interleukin-6（IL-6） in serum of the rats in various groups. The expression levels of Toll-like receptor 4（TLR4） and nuclear factor-κB（NF-κB） proteins in periodontal tissue of the rats in various groups were detected by Western blotting method. Results Compared with control group， the rats in model group showed obvious symptoms of “three more and one less”；the level of blood glucose of the rats was increased obviously（P<0.05）， the inflammatory cells were infiltrated in periodontal connective tissue， periodontal tissue was damaged， and the alveolar bone absorption amount was increased（P<0.05）；the activities of SOD and CAT in periodontal tissue were significantly decreased （P<0.05）， the levels of ROS and MDA were significantly increased （P<0.05）， the levels of TNF-α， IL-1β and IL-6 in serum were significantly increased（P<0.05），and the expression levels of TLR4 and NF-κB proteins were significantly increased （P<0.05）. Compared with model group， the general condition of the rats in low and high doses of GSPE groups was obviously improved，the levels of blood glucose were reduced（P<0.05）， periodontal tissue damage was obviously reduced，the alveolar bone absorption amounts were reduced（P<0.05）， the activities of SOD and CAT in periodontal tissue were significantly increased （P<0.05）， the levels of ROS and MDA were significantly decreased （P<0.05）， the levels of TNF-α， IL-1β and IL-6 in serum were significantly decreased（P<0.05），and the expression levels of TLR4 and NF-κB proteins were significantly decreased （P<0.05）. Compared with low dose of GSPE group， the activities of SOD and CAT in periodontal tissue of the rats in high dose of GSPE group were significantly increased （P<0.05）， the levels of ROS and MDA were significantly decreased （P<0.05）， the levels of TNF-α， IL-1β and IL-6 in serum were significantly decreased（P<0.05），and the expression levels of TLR4 and NF-κB proteins were significantly decreased （P<0.05）. Conclusion GSPE can improve the periodontal inflammation in the diabetic periodontitis rats， and its mechanism may be related to regulating the TLR4 and NF-κB signaling pathway in periodontal tissue. Objective To explore the improvement effects of grape seed proanthocyanidin extract （GSPE） on the periodontal inflammation in the diabetic periodontitis rats， and to clarify its mechanism.

Objective To investigate the effects of up-regulation of protein phosphatase Mg²⁺/Mn²⁺-dependent 1F （PPM1F） on the proliferation and migration of nasopharyngeal carcinoma HONE-1 cells， and to clarify their mechanisms. Methods The nasopharyngeal carcinoma HONE-1 cells were divided into pcDNA3.1 group （transfected with pcDNA3.1 plasmid） and pcDNA3.1-Flag-PPM1F group （transfected with pcDNA3.1-Flag-PPM1F plasmid）. The expression levels of PPM1F and E-cadherin mRNA and proteins in the cells in two groups were detected by real-time fluorescence quantitative PCR（RT-qPCR） and Western blotting methods； the activities of proliferation and clone formation rates of the cells in two groups were detected by CCK-8 method and clone formation assay； the scratch healing rates of the cells in two groups were detected by cell scratch assay，and the numbers of migation cells in two groups were detected by Transwell experiment. Results Compared with pcDNA3.1 group， the expression level of PPM1F mRNA in the cells in pcDNA3.1-Flag-PPM1F group was significantly increased（P<0.01），the expression amount of PPM1F protein was significantly increased，the expression levels of E-cadherin mRNA and protein in the cells were significantly increased（P<0.01），the proliferation activity，the clone formation rate and the scratch healing rate were significantly decreased （P<0.05 or P<0.01）， and the number of migration cells was significantly reduced（P<0.05）. Conclusion Up-regulation of PPM1F expression can inhibit the proliferation and migration of nasopharyngeal carcinoma HONE-1 cells， and its mechanism may be related to intercellular adhesion.

Objective To explore the improvement effect of propofol postconditioning on neurological function in the rats with cerebral ischemia-reperfusion injury and its protective effect on brain mitochondrial injury，and to clarify its mechanism. Methods Twenty-four SD rats were randomly divided into sham operation group， model group and propofol postconditioning group， with 8 rats in each group. The rats in model group and propofol postconditioning group were used to establish the focal middle cerebral artery obstruction focal cerebral ischemia-reperfusion models by carotid artery ligation. The rats in propofol postconditioning group were infused with 20 mg·kg^－1·h^－1 propofol for 2 h in the femoral vein immediately after reperfusion； the rats in sham operation group and model group were given the same amount of normal saline.The neurological deficit scoring in the rats in various groups 24 h after reperfusion were performed； HE staining method was used to observe the pathomorphology of the hippocampus tissue of the rats in various groups； the activities and levels of oxidative stress-related factors in hippocampus tissue of the rats in various groups were detected by related kits； fluorescence probes of reactive oxygen species （ROS） were used to detect the levels of ROS in hippocampus tissue of the rats in various groups； TUNEL method was used to detect the TUNEL positive cell rates in hippocampus tissue of the rats in various groups. Western blotting method was used to detect the expression levels of mitochondrial fission， fusion and biogenesis-related proteins and nicotinamide adenine dinucleotide phoshate oxidase 4（Nox4） and nuclear factor E2-related factor 2（Nrf2） proteins； the expression levels of Nrf2 in hippocampus tissue of the rats in various groups were detected by immunofluorescence method. Results Compared with sham operation group， the neurological deficit score of the rats in model group was significantly increased （P<0.05）， the pathological damage of hippocampus tissue of the rats was obvious， the activities of superoxide dismutase（SOD） and glutathion peroxidase（GSH-Px） and the level of total antioxidant capacity（T-AOC） in hippocampus tissue were significantly decreased（P<0.05）， the levels of malonadehyde（MDA） and ROS and the TUNEL positive cell rate were significantly increased（P<0.05），the expression levels of dynamin-related protein 1 （DRP1）， fission protein 1（Fis1）， Nox4， and Nrf2 proteins were significantly increased （P<0.05），and the expression levels of optic atrophy 1（OPA1）， mitofusin 2（Mfn2）， peroxisome proliferator-activated receptor γ co-activator 1α（PGC-1α），and mitochondrial transcription factor A（TFAM） proteins were significantly decreased （P<0.05）. Compared with model group， the neurological deficit score of the rats in propofol postconditioning group was significantly decreased （P<0.05）， the pathological damage of hippocampus tissue of the rats was significantly improved， the levels of MDA and ROS and the TUNEL positive cell rate were signficantly increased（P<0.05）， the expression levels of DRP1， Fis1 and Nox4 proteins were significantly decreased （P<0.05）， and the expression levels of OPA1， Mfn2， PGC-1α， TFAM and Nrf2 proteins were significantly increased （P<0.05）. Conclusion Postconditioning of propofol can inhibit apoptosis and oxidative stress in hippocampus tissue of the rats with cerebral ischemia-reperfusion injury， promote mitochondrial fusion and biogenesis and inhibit mitochondrial fission， and improve the neurological damage of rats； its mechanism may be related to the regulation of Nox4/Nrf2 signaling pathway.

Objective：To clarify the possible mechanism of suboptimal efficacy of enhancer of zeste homolog 2 （EZH2） inhibitors in pancreatic cancer， and to provide basis for exploring a new treatment plan for pancreatic cancer. Methods The pancreatic cancer PANC-1 cells， CFPAC-1 cells and pancreatic metastasis cancer SW1990 cells were treated with different concentrations（0－50 μmol·L^－1）of EZH2 inhibitor GSK126. CCK8 method was used to detect the survival rates of the cells in various groups. Then the cells were divided into control group and GSK126 group. After 48 h of GSK126 incubation， the apoptotic rates of the cells in various groups were detected by flow cytometry，the lipid droplet formation in the cells in various groups were observed with oil red O staining，and the levels of triglyceride（TG） in the cells in various groups were detected.The expression levels of apoptosis genes cysteine-aspartic acid protease（1，3，7 and 9） （Caspase-1， Caspase-3， Caspase-7， Caspase-9）， vascular endothelial growth factor A （VEGFA）， E-cadherin and stemness markers CD133， CD24 and CD44 mRNA and the expression levels of fat acid synthase （FASN）， acetyl CoA carboxylase alpha （ACACA）， citrate synthase （CS）， ATP citrate lyase （ACLY）， stearoyl-CoA desaturase （SCD） and acyl-CoA synthetase short chain family member 2（ACSS2） mRNA were detected by real-time fluorescence quantitative PCR（RT-qPCR） method. The three kinds of cells were divided into control group， GSK126 group， FASN inhibitor Orlistat （Orlistat） group and GSK126+Orlistat group，respectively. After incubated for 48 h， the survival rates of cells in various groups were detected by CCK-8 method， and the combination index （CI） value of the two drugs was calculated.The lipid droplet formation in the cells in various groups were observed with oil red O staining，and the levels of TG in the cells in various groups were detected； the apoptotic rates of cells in various groups were detected by flow cytometry，and the expression levels of Caspase-1， Caspase-3， Caspase-7 and Caspase-9 and CD133， CD24 and CD44 mRNA in the cells in various groups were detected by RT-qPCR method. Results GSK126 inhibited the proliferation of pancreatic cancer cells in a concentration dependent manner. There were no significant differences in the apoptotic rates and apoptosis gene mRNA expression levels in the three kinds of cells between control group and GSK126 group（P>0.05）. Compared with control group， the expression levels of VEGFA mRNA in the SW1900 cells and E-cadherin mRNA in the PANC-1 cells in GSK126 group were significantly decreased （P<0.05）；the expression levels of CD133 in three kinds of cells and the expression levels of CD24 mRNA in the CFPAC-1 cells and SW1990 cells were signicantly increased （P<0.05）.Compared with control group， the number of lipid droplets in three kinds of cells in GSK126 group were significantly increased，the TG levels and the expression levels of FASN mRNA were significantly increased （P<0.05 or P<0.01）， and the expression levels of other lipid metabolism-related genes were also increased to varying degrees.Compared with control group， GSK126 group and Orlistat group， the survival rates of the cells in GSK126+Orlistat group were significantly decreased（P<0.05 or P<0.01）； the CI of two drugs was less than 1， which meaned synergistic effect of GSK126 and Orlistat.Compared with control group，GSK126 group and the number of lipid droplets in the cells in GSK126+Orlistat group was signifieantly decreased；compared with control group and GSK126 group，the TG level in the cells in GSK126+Orlistat group was significantly decreased（P<0.01），and the expression levels of apoptosis-related gene and stemness gene mRNA were significantly increased（P<0.05 or P<0.01）；compared with control group，GSK126 group and Orlistat group，the apoptotic rates of the cells in GSK126+Orlistat group was significantly increased（P<0.01）. Conclusion GSK126 inhibits the proliferation of pancreatic cancer cells and induces reprogramming of lipid metabolism， which weakens its anticancer effect. The combined use of GSK126 and FASN inhibitor Orlistat can partially reverse this effect， synergistically inhibit the cell proliferation， enhance apoptosis and reduce the expression of stemness genes.

Objective To investigate the improvement effect of tetramethylpyrazine（TMP） on oxidative damage of kidney tissue in the nephrolithiasis model rats， and to clarify its possible mechanism. Methods A total of 40 healthy male SD rats were randomly divided into control group， model group， TMP group and positive control group. After pre-experiment，except control groups，the rats in other groups were intraperitoneally injected with 80 mg·kg^－1 glyoxylate stock solution to establish the rat models of kidney stones； the rats in TMP group were intraperitoneally injected with TMP hydrochloride injection 100 mg·kg^－1，the rats in positive control group were intragastrically perfused with 3.12 g·kg^－1 of Shenshitong granules， and the rats in control group were intraperitoneally injected with 2.5 mL·kg^－1 of 0.9% sodium chloride injection，lasted for 10 d.The rat kidney tissue sections were taken， Von Kossa staining and HE staining were performed to observe the calcium deposition and the pathomorphology of kidney tissue， and the levels of malondialdehyde（MDA） and the activities of superoxide dismutase （SOD） and glutathione peroxidase （GSH-Px） in kidney tissue of the rats in various groups were detected. Western blotting method was used to detect the expression levels of nuclear protein nuclear factor E2-related factor 2 （Nrf2）， holoprotein heme oxygenase-1 （HO-1） and NAD（P）H quinone oxidoreductase 1 （NQO1） proteins in kidney tissue of the rats in various groups. Results Compared with control group，obvious calcium deposition was seen in the Von Kossa staining of kidney tissue of the rats in model group，and the quantitative grading score of crystal was significantly increased （P<0.01）；the HE staining results showed obvious pathological damage in kidney tissue cells，the level of MDA in kidney tissue was significantly increased （P<0.01）， the activities of SOD and GSH-Px were significantly decreased （P<0.01），and the expression levels of Nrf2， HO-1 and NQO1 proteins in kidney tissue were significantly decreased （P<0.01）. Compared with model group， calcium deposition was seen in the Von Kossa staining of kidney tissue of the rats in TMP group and positive control group， and the quantitative grading scores of crystal were decreased （P<0.05）；the HE staining results showed that the pathological damage of kidney tissue cells was alleviated， the levels of MDA were significantly decreased （P<0.05），the activities of SOD and GSH-Px were significantly increased （P<0.05）， and the expression levels of Nrf2， HO-1 and NQO1 proteins in kidney tissue were significantly increased （P<0.05）. Conclusion Kidney stones can cause oxidative stress injury in the rat kidney tissue， resulting in the changes in oxidative stress indicators and Nrf2/antioxidant response element（ARE） signaling pathway protein expression levels in kidney tissue. TMP can activate this signaling pathway after intervention， so as to improve the oxidative stress injury of kidney tissue.

Objective To explore the effect of Physalis Calyx seu Fructus （PCF） on the occurrence and development of leukemia based on network pharmacology， and to clarify the main targets and signaling pathway-related mechanism of its bioactive components. Methods Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform （TCMSP） database was used to search its bioactive components and corresponding potential targets with “Physalis Calyx seu Fructus” as the keyword； the potential targets were evaluated and predicted with Protein Data Bank （PDB） and Swiss Target Prediction website； GeneCards database， Online Mendelian Inheritance in Man （OMIM） database and DrugBank database were used to search the relevant targets with “leukemia” as the keyword. The bioactive component targets of PCF and leukemia target were intersected by Draw Veen Diagram online software， and the intersected targets were the targets of PCF acting on leukemia； the network information of protein-protein interaction （PPI） of intersection target proteins was obtained and visualized with STRING database； DAVID database was used for Gene Ontology （GO） biological function enrichment analysis and Kyoto Encyclopedia of Genes and Genomes （KEGG） signaling pathway enrichment analysis of intersection genes； topological analysis on PPI network was performed with Cytoscape 3 7.2 to screen the core targets and the core active components； the molecular docking verification was performed with AutoDock and PyMol. Results After screening the retrieval results of TCMSP database， 6 active components of PCF were obtained； 29 common targets of PCF and leukemia were collected； GO enrichment analysis mainly involved the development of hematopoietic and lymphoid organs， protein metabolism regulation， apoptosis and other biological processes； KEGG enrichment analysis mainly included cancer pathway， tumor necrosis factor （TNF）， T cell receptor （TCR） and other signal pathways. Ten core targets such as protein kinase B1 （AKT1） and cysteinyl aspartate specific proteinase-3（Caspase-3） were obtained by topological network analysis， combined with KEGG signaling pathway， the core active components Sitosterol and Gramisterol were obtained. The results of molecular docking showed that the docking energy of Sitosterol with proto-oncogene （JUN）， Gramisterol with AKT1 and Caspase-3 were －6.0 kcal·mol^－1，indicating the docking was good. Conclusion The main active components of PCF， such as Sitosterol and Gramisterol， may affect the occurrence and development of leukemia by regulating JUN， AKT1， Caspase-3 and other genes and participating in cancer pathway， TCR and other signaling pathways.

Objective To print the porous titanium alloy scaffolds by 3D printing technology，and to study the effect of micro arc oxidation （MAO）/alkali treatment and arginine-glycine-aspartic acid（RGD） coating on the biological behavior of osteoblasts. Methods The 3D porous titanium alloy scaffolds were designed and printed， and they were divided into MAO group， MAO/alkali treatment （MN） group， MAO/alkali treatment loaded with RGD peptide coating （MNR） group after different surface treatments，another blank control group was set up.The elastic modulus of the scaffolds in various groups was detected.The microstructures of the scaffold surface in various groups were observed by scanning electron microscope （SEM）， the elemental compositions of the scaffold surface were detected by energy dispersive spectroscopy （EDS）， and the contact angles of water droplets on the scaffold surface were measured by contact angle measuring instrument.The mouse embryonic osteoblast precursor cells （MC3T3-E1 cells） were co-cultured with the scaffolds in various groups.CCK-8 assay was used to detect the cell proliferation activities，Live/Dead cell staining was used to detect the biocompatibility of the scaffolds in various groups， cell adhesion test was used to observe the adhesion of cells on the material surface， and alkaline phosphatase （ALP） kit was used to detect the ALP activities.Real-time fluorescence quantitative PCR （RT-qPCR） method was used to detect the expression levels of Runt-related transcription factor 2（Runx2）， osteopontin（OPN） and osteocalcin（OCN） mRNA in the cells in various groups. Results The elastic modulus of 3D printed scaffolds was （1.17±0.62） GPa. The SEM observation results showed that the surface of the scaffolds after MAO treatment showed crater-like morphology. After alkali treatment， small cracks and nano scale structure appeared.The scattered RGD particles were found on the surface of the scaffolds loaded with RGD peptide coating.The ERS detection showed that the RGD peptide coating was successfully loaded on the surface of scaffolds. The contact angle measuring instrument detection results showed that the surface contact angles were MAO group>MN group>MNR group. The cell proliferation activities in three groups showed an increasing trend on the 1st， 3rd and 5th days detected by CCK-8 assay， and there were statistically significant differences in the cell proliferation activities between various groups on the 3rd and 5th days of culture（P<0.05 or P<0.01）. The results of Live/Dead cell staining showed that the scaffolds in three groups had good in vitro compatibility. In cell adhesion test， after 48 h of co-culture， the number and morphological extension of cells in MNR group were better than those in MAO group and MN group. On the 7th day of culture， compared with MAO group， the activity of ALP in the cells in MNR group was significantly increased （P<0.01）； on the 14th day of culture， there were significant differences in the ALP activities between three groups （P<0.05 or P<0.01）.The results of RT-qPCR method showed that the expression levels of Runx2 and OPN mRNA in MN group and MNR group were higher than those in blank control group and MAO group on the 7th day of culture （P<0.01）， and the expression level of OCN mRNA in MNR group was higher than those in blank control group and MAO group （P<0.05）； on the 14th day of culture，there were significant differences in the expression levels of Runx2 and OPN mRNA between three groups（P<0.05 or P<0.01）. Compared with blank control group， the expression levels of OCN mRNA in MAO group，MN group，and MNR group were significantly increased （P<0.01）. Conclusion The 3D printed porous titanium alloy scaffolds hve the elastic modulus matching with the human bone tissue. The surface MAO/alkali treatment and loading with RGD peptide coating are non-toxic to the MC3T3-E1 cells and can promote their osteogenic differentiation.

Objective To prepare the oxaliplatin （OXA）-loaded elastin-like polypeptides （ELPs） hydrogels， and to investigate its in vitro drug release and the killing effect in the CT26 colon cancer cells. Methods The prokaryotic proteins ELPs were produced in Escherichia coli （E.coli） system and purified via the inverse transition cycling （ITC）.The ELPs hydrogel was prepared by mixing the ELPs protein solution with the cross-linking agent tetrakis hydroxymethyl phosphonium chloride（THPC）.The microstructures of hydrogels were observed by scanning electron microscope （SEM）； CCK-8 assay was used to determine the survival rates of the CT26 cells and NIH3T3 cells after hydrogel treatment and Live/Dead cell staining was used to observe the survival of the CT26 cells after treated with 30 and 40 g?L^－1 hyrogels for 48 h. The OXA-loaded ELPs hydrogels were prepared， and the drug release of OXA-loaded hydrogels was detected at different pH values （pH 6.5 and pH 7.4） in the sustained release liquid. The survival rates of CT26 cells after added with the ELPs hydrogels soak solution loading different concentrations （0.25， 1.00， 4.00， 16.00， and 64.00 mg?L^－1） of OXA were detected and the survival rates of CT26 cells after treated for different time in various groups were detected by CCK-8 assay. Results The gel electrophoresis analysis showed that an obvious bold protein band was observed in the region with expected molecular weight 38 000. After protein purification using ITC， almost no other protein was found. The ELPs protein solution was transited from transparent solution into an opaque white hydrogel after the addition of THPC. The hydrogel showed uniformed pores and three-dimensional structure under SEM.The CCK-8 assay results indicated that the survival rates of CT26 cells and NIH3T3 cells were close to 100%. The Live/Dead cell staining results showed the green fluorescence in the CT26 cells under fluorescence microscope. The OXA-loaded hydrogels showed sustained drug release profile in vitro，and a slightly faster drug release in pH 6.5 was found than that in pH 7.4.The CCK-8 assay results showed that the OXA-loaded hydrogels could significantly inhibit the proliferation of CT26 cells in a dose-dependent manner. With the prolongation of the incubation time， the survival rates of CT26 cells treated with drug-loaded hydrogel were decreased gradually. Conclusion The OXA-loaded ELPs hydrogels have sustained release profile in vitro and can kill efficiently and consistenly the colon cancer CT26 cells of the mice. ELPs hydrogels may be used as the safe and effective drug delivery system for cancer therapy.

Objective To investigate the effect of human circular RNA_0114428 （circ_0114428） on the lipopolysaccharide （LPS）-induced injury of human renal tubular epithelial HK-2 cells， and to clarify the possible molecular mechanism of circ_0114428 in improving the injury of HK-2 cells by regulating microRNA-139-5p （miR-139-5p）/transforming growth factor β receptor 2 （TGFBR2） axis. Methods The HK-2 cells were cultured in vitro， the dual luciferase reporter gene experiment was used to verify the targeted regulation relationship between miR-139-5p and circ_0114428 or TGFBR2， and the CCK-8 method was used to screen the appropriate LPS intervention time and concentration. The HK-2 cell injury model was established by administering 10 mg·L^－1 LPS for 12 h， and the HK-2 cells were divided into control group， LPS group， LPS+circ_0114428 silence control （LPS+si-NC） group， LPS+circ_0114428 silence （LPS+si-circ_0114428） group， LPS+circ_0114428 silence+miR-139-5p inhibitor control （LPS+si-circ_0114428+in-NC） group， and LPS+circ_0114428 silence+miR-139-5p inhibitor（LPS+si-circ_0114428+in-miR-139-5p） group. Real-time fluorescence quantitative PCR （RT-qPCR） method was used to detect the expression levels of circ_0114428， miR-139-5p and TGFBR2 mRNA in the cells in various groups， flow cytometry was used to detect the apoptotic rates of the cells in various groups， and Western blotting method was used to detect the expression levels of TGFBR2， B-cell lymphoma-2 （Bcl-2） and Bcl-2 associated X protein （Bax） proteins in the cells in various groups；enzyme linked immunosorbent assay（ELISA） was used to detect the levels of interleukin-6（IL-6）， tumor necrosis factor-α （TNF-α） and interleukin-1β （IL-1β） in the cell supernatant in various groups. Results The results of dual luciferase reporter gene experiment showed that miR-139-5p had a targeted regulatory relationship with circ_0114428 and TGFBR2 （t=10.171， P<0.05；t=7.682， P<0.05）.The CCK-8 method results showed that the concentration of LPS induction was 10 mg·L^－1 and the optimal time was 12 h.Compared with control group， the expression levels of circ_0114428， expression levels of TGFBR2 mRNA and protein， and expression level of Bax protein， the apoptotic rate， and levels of IL-6， TNF-α and IL-1β in cell supernatant of the HK-2 cells in LPS group were significantly increased （P<0.05），and the expression levels of miR-139-5p and Bcl-2 protein were significantly decreased （P<0.05）. Compared with LPS+si-NC group， the expression levels of circ_0114428， expression levels of TGFBR2 mRNA and protein， and expression level of Bax protein， the apoptotic rate， and the levels of IL-6， TNF-α and IL-1β in cell supernatant of the HK-2 cells in LPS+si-circ_0114428 group were significantly decreased （P<0.05），and the expression levels of miR-139-5p and Bcl-2 protein were significantly increased （P<0.05）. Compared with LPS+si-circ_0114428+in-NC group， the expression levels of circ_0114428， expression levels of TGFBR2 mRNA and protein， and expression level of Bax protein， the apoptotic rate， and the levels of IL-6， TNF-α and IL-1β in cell supernatant of the HK-2 cells in LPS+si-circ_0114428+in-miR-139-5p group were significantly increased （P<0.05）， and the expression levels of miR-139-5p and Bcl-2 protein were significantly decreased （ P<0.05）. Conclusion Circ_0114428 can alleviate the LPS-induced HK-2 cell injury， and its mechanism may be related to the regulation of the miR-139-5p/TGFBR2 axis.

Objective To investigate the effects of Dendrobium huoshanense polysaccharide （DHP） on the levels of dopamine（DA） in striatum， levels of inflammatory factors and activities of antioxidant enzymes in substantia nigra tissue of the Parkinson’s disease（PD） mice induced by 1-methyl-4-phenyl-1，2，3，6-tetrahydropyridine（MPTP），and to elucidate the ameliorative effect of DHP on PD. Methods The C57BL/6J mice were randomly divided into control group，model group， and different doses（25， 50 and 100 mg·kg^－1） of DHP groups.Except for control group，the mice in other groups were used to establish the PD models by intraperitoneal injection of MPTP.The mice in different doses of DHP groups were given by gavage with different doses of DHP， and the mice in control and model groups were given the equal amount of saline by gavage. The behaviors of mice in various groups were examined by the method of turning sticks. The levels of DA in striatum of the mice in various groups were determined by high performance liquid chromatography（HPLC）； the levels of malondialdehyde（MDA） and the activities of superoxide dismutase （SOD） and glutathione peroxidase （GSH-Px） in substantia nigra tissue of the mice in various groups were determined by chemical colorimetry.The levels of interleukin-1β （IL-1β） and tumor necrosis factor-α （TNF-α） in substantia nigra tissue of the mice in various groups were determined by enzyme linked immunosorbent assay（ELISA）. Results Compared with control group， the drop latency of the mice in model group was significantly shortened（P<0.05）； compared with model group， the drop latencies of the mice in 50 and 100 mg·kg^－1 DHP groups were significantly prolonged （P<0.05）. Compared with control group， the level of DA in striatum of the mice in model group was significantly decreased （P<0.05）；compared with model group， the levels of DA in the striatum of the mice in 50 and 100 mg·kg^－1 DHP groups were significantly increased （P<0.05）. Compared with control group， the level of MDA in substantia nigra of the mice in model group was significantly increased （P<0.05）， and the activities of SOD and GSH-Px were significantly decreased （P<0.05）；compared with model group，the levels of MDA in substantia nigra of the mice in 50 and 100 mg·kg^－1 DHP groups were significantly decreased（P<0.05），and the activities of SOD and GSH-Px in substantia nigra of the mice were significantly increased（P<0.05）. Compared with control group， the levels of IL-1β and TNF-α in substantia nigra tissue of the mice in model group were significantly increased （P<0.05）；compared with model group， the levels of IL-1β and TNF-α in substantia nigra tissue of the mice in 50 and 100 mg·kg^－1 DHP groups were significantly decreased （P<0.05）. Conclusion DHP has a certain protective effect on nerve function in the MPTP-induced PD mice， and its mechanism may be related to inhibition of oxidative damage and reduction of neuroinflammatory response.

Objective To investigate the effect of miR-216b-5p on the proliferation， migration，invasion and apoptosis of laryngeal squamous cell cancer（LSCC） TU686 cells，and to analyze the target genes in laryngeal cancer cells and its possible mechanism. Methods The TU686 cells were divided into control group and miR-216b-5p group；the cells in control group were infected with the nonsense sequence through lentivirus，and the cells in miR-216b-5p group were infected with overexpressed miR-216b-5p by lentivirus.The expression levels of miR-216b-5p in the cells in two groups were detected by real-time fluorescence quantitative PCR（RT-qPCR） method， the clone formation rates of the cells in two groups were detected by clone formation assay，the scratch healing rates of cells in two groups were detected by cell scratch assay，the number of invasion cells in two groups was detected by Transwell assay，and the apoptotic rates of cells in two groups were detected by flow cytometry. The TargetScan website was used to predict the potential target genes of miR-216b-5p. The expression levels of target gene mRNA were detected by RT-qPCR method.The targeting relationship between miR-216b-5p and the target gene was detected by dual-luciferase assay. Results After lentivirus infection， compared with control group， the expression level of miR-216b-5p in the cells in miR-216b-5p group was significantly increased（P<0.01）.Compared with control group，the clone formation rate and the scratch healing rate of the cells in miR-216b-5p group were significantly decreased（P<0.01），the number of invasion cells was significantly decreased（P<0.01）， and the apoptotic rate was significantly increased（P<0.01）. Targetscan website predicted that autophagy related gene 5 （ATG5） was the potential target gene of miR-216b-5p；compared with control group，the expression level of ATG5 mRNA in the cells in miR-216b-5p group was significantly decreased（P<0.01）.The dual-luciferase assay proved that there was a targeting correlation between ATG5 and miR-216b-5p. Conclusion MiR-216b-5p can inhibit the proliferation， migration， and invasion and induce apoptosis of laryngeal cancer cells to exert a tumor suppressor effect， and its mechanism may be related to the targeted regulation of the expression level of ATG5.

Objective To explore the targeting relationship between miR-150-5p and non-SMC condensate Ⅰ complex subunit G （NCAPG） and its effect on the biological function of hepatocellular carcinoma （HCC） cells， and to provide the potential targets for the clinical diagnosis and treatment of HCC. Methods The targeting relationship between miR-150-5p and NCAPG was verified by dual-luciferase reporter gene experiment. The Huh7 cells were divided into mimic negative control （mimic NC）group， miR-150-5p mimic group， miR-150-5p mimic+overexpression negative control （miR-150-5p mimic+ovNC）group and miR-150-5p mimic+overexpression NCAPG （miR-150-5p mimic+ovNCAPG） group. The expression levels of NCAPG mRNA in the cells in various groups were detected by real-time fluorescence quantitative PCR （RT-qPCR） method， and the expression levels of NCAPG protein in the cells in various groups were detected by Western blotting method； 5-ethynyl-2'-deoxyuridine （EdU） assay was used to detect the EdU positive rates of cells in various groups， flow cytometry was used to detect the apoptotic rates， and the numbers of migration and invasion cells were detected by Transwell assay. The nude mice were divided into agomiR-NC group， agomiR-150-5p group， agomiR-150-5p+overexpression negative control（agomiR-150-5p+ovNC） group and agomiR-150-5p+overexpression NCAPG （agomiR-150-5p+ovNCAPG） group. The Huh7 cells formed subcutaneous tumors， and the tumor volumes and tumor weights of nude mice in various groups were measured. Results The dual luciferase reporter gene assay results confirmed that NCAPG was the target gene of miR-150-5p. Compared with mimic NC group， the expression levels of NCAPG mRNA and protein and the EdU positive rate in the Huh7 cells in miR-150-5p mimic group were significantly decreased （P<0.05），the apoptotic rate was significantly increased （P<0.05），and the numbers of migration and invasion cells were significantly decreased（P<0.05）； compared with miR-150-5p mimic +ovNC group， the expression levels of NCAPG mRNA and protein and the EdU positive rate in the Huh7 cells in miR-150-5p mimic +ovNCAPG group were significantly increased （P<0.05），the apoptotic rate was significantly decreased （P<0.05），and the numbers of migration and invasion cells were significantly increased（P<0.05）. Compared with agomiR-NC group， the transplanted tumor volume and tumor weight of the nude mice in agomiR-150-5p group were significantly reduced （P<0.05）； compared with agomiR-150-5p+ovNC group， the transplanted tumor volume and tumor weight of the nude mice in agomiR-150-5p ovNCAPG group were significantly increased （P<0.05）. Conclusion MiR-150-5p inhibits the HCC cell proliferation， migration， invasion and tumor growth through targeting binding and inhibitiing the NCAPG expression， and promotes the apoptosis.

Objective To explore the replication of rotavirus SA11 strain in vivo and in vitro after adding Enterococcus faecalis （E. faecalis），and to provide the evidence for clarifying the effect of E. faecalis on rotavirus infection. Methods In in vitro experiment， 10⁸－10¹² CFU ·mL^－1E. faecalis were diluted with 10 times concentration gradient and co-cultured with Caco-2 cells for 12， 18， 24， 30，and 48 h，and control group （Caco-2 cells infacted with rotavius without the addttion of E. faecalis） was set up at the same time. The survival rates ofCaco-2 cellsin various groups after treatment of different concentrations of E. faecalis were determined by CCK-8 method.One hour after the Caco-2 cells were treated with rotavirus with multiplicity of infection （MIO） value of 0.1，they were cultured with 10⁸－10¹² CFU·mL^－1E.faecalis for 18 h. The rotavirus VP6 gene copy numbers of Caco-2 cells in various groups were detected by real-time fluorescence quantitative PCR（RT-qPCR） method and the rotavirus titers of Caco-2 cells in various groups were detected by immunofluorescence focus method.In in vivo experniment， 18 litters of 4－5 d old SPF Kunming suckling mice were randomly divided into control group， antibiotic treatment group and E. faecalis transplantation group. The suckling mice in antibiotic treatment group and E. faecalis transplantation group were gavaged with quadruple antibiotics for 3 d to deplete their intestinal flora， and the suckling mice in three groups received 10⁷ FFU·mL^－1 rotavirus 100 μL by gavage daily for 8 d， and the suckling mice in E. faecalis transplantation group were given 10¹⁰ CFU·mL^－1E. faecalis 100 μL at the same time.The feces were collected on the 2nd，4th，6th，and 8th days after virus infection， and the copy numbers of rotavirus in feces were measured by RT-qPCR method. Results In in vitro experiment，the CCK-8 method results showed that compared with control group，the survival rates of Caco-2 cells in various groups in 48 h of treatment of different concentrations of E. faecalis were increased（P<0.05）， indicating that E. faecalis was non-toxic to the Caco-2 cells within this concentration range.The RT-qPCR results showed that compared with control group，the number of rotavirus gene copies in the Caco-2 cells in 10¹⁰， 10¹¹，and 10¹² CFU·mL^－1E. faecalis groups weredecreased by 75.8%， 99.9%， and 99.9%，respectively（P<0.05）. The results of immunofluorescence focus detection showed that compared with control group，the rotavirus titer in 10⁸，10¹⁰，and 10¹² CFU·mL^－1E. faecalis groups were decreased by 13%，77%， and 100%， respectively （P<0.05）.In in vivo experiment， compared with control group， the copy number of fecal virus gene of the suckling mice in E. faecalis transplantation group was significantly decreased（P<0.05）. Conclusion In in vitro cell model and rotavirus infection model of suckling mice， E. faecalis can inhibit the replication of rotavirus.

Objective To screen the survival-associated alternative splicing （AS） events in the lung adenocarcinoma （LUAD） patients using bioinformatics method and construct the splicing factor （SF）-AS regulatory network， and to provide a new approach for the prognostic evaluation of the LUAD patients. Methods The transcriptome data and clinical data of the LUAD patients were downloaded from The Cancer Genome Atlas （TCGA） database， and the AS data of the LUAD patients were downloaded from the SF expression data and RNA target motifs（SpliceAid2）database. The survival-associated AS events of the LUAD patients were analyzed by univariate Cox regression analysis. The prognostic risk model of the LUAD patients was established by LASSO regression analysis and multivariate Cox regression analysis，and the risk score of each AS event was calculated based on the model. Kaplan-Meier （K-M） survival analysis and receiver operating characteristic （ROC） curve were used to evaluate the reliability of the model. The relationships between risk model or clinical parameters and independent prognosis of the LUAD patients were analyzed by using Cox regression analysis. The Pearson test was used to analyze the correlation of SF with AS events associated with prognosis. Cytoscape software was utilized to construct the SF-AS interaction network. Results A total of 43 948 cases of survival-associated AS events were screened， including 7 types of events. Exon skip （ES） events and alternate terminator （AT） events were the main events. The prognostic risk model of AS events was constructed， and the LUAD patients were divided into high and low risk groups based on the risk scores of the model.The K-M survival analysis results showed that the overall survival （OS） rate of the LUAD patients in high risk group was lower than that in low risk group （P<0.05）.The ROC curve analysis indicated that prognostic risk model of the LUAD patients had a good predictive effect，and the area under curve （AUC） was 0.824. The SF-AS regulatory network showed that 12 prognosis-related SFs modulated AS events positively or negatively，and they could predict the poor prognosis of the LUAD patients. Conclusion The prognostis-related AS events of LUAD patients and their upstream regulatory factor SF are screened，the SF-AS network is constructed，and the study provides the theoretical basis for further research on the correlation of AS events with prognosis of the LUAD patients.

Objective To analyze the expression of microRNA-223 （miR-223） in the ovarian cancer tissue and explore the effect of miR-223 on the proliferation and invasion of ovarian cancer OVCAR3 cells， and to clarify the molecular regulatory mechanism. Methods The expression levels of miR-223 in tissue samples taken during surgical removal of 45 patients with ovarian cancer and different ovarian cancer cells were detected by real-time fluorescence quantitative PCR （RT-qPCR） method， and the expression levels of miR-223 in cancer tissue of the ovarian cancer patients with different clinicopathological features were analyzed. The ovarian cancer OVCAR3 cells were cultured in vitro； bioinformatics， dual luciferase reporter gene experiment and Western blotting method were used to analyze the targeted regulation relationship of miR-223 to N-myc downstream regulatory gene 1 （NDRG1）. The OVCAR3 cells were transfected with negative control mimic（NC group），miR-223 inhibitor（MiR in group） and miR-223 inhibitor and siRNA-NDRG1 plasmid at the same time（MiR in+si-NDRG1 group）. The number of clone formation was detected by colony formation test， the apoptotic rate was measured by flow cytometry， and the number of invasion cells was measured by Transwell chamber assay. Results Compared with adjacent normal tissue the expression level of miR-223 in cancer tissue was significantly increased（P<0.01）；the miR-223 expression level was closely related to the degree of histological differentiation， FIGO stage and lymph node metastasis of ovarian cancer patients （P<0.01）. Meanwhile， the expression levels of miR-223 in the different ovarian cancer cells were higher than that in normal ovarian epithelial cells（P<0.01）. MiR-223 could target NDRG1 and inhibit the expression of NDRG1， and the expression level of NDRG1 mRNA in ovarian cancer tissue was negatively correlated with the expression level of miR-223 （r=－0.291， P<0.01）. Compared with NC group， the number of clone formation and the number of invasion cells in MiR in group were decreased （P<0.05 or P<0.01）， while the apoptotic rate was significantly increased（P<0.01）. Compared with MiR in group， the number of clone formation and the number of invasion cells in MiR in + si-NDRG1 group were increased （P<0.05 or P<0.01）， while the apoptotic rate was decreased （P<0.01）. Conclusion MiR-223 is highly expressed in the ovarian cancer tissue， which is positively correlated with the severity of ovarian cancer. MiR-223 can promote the proliferation and invasion of ovarian cancer cells and inhibit apoptosis by targeted inhibition of NDRG1 expression.

Objective To analyze the influencing factors of ectopic pregnancy in the patients with in vitro fertilization-embryo transfer （IVF-ET），and to establish the nomogram model to predict the risk of ectopic pregrancy in the patients with IVF-ET. Methods The inclusion and exclusion criteria of the subjects were developed， the clinical informations of 522 patients with IVF-ET and successful pregnancy were analyzed retrospectively for modeling and internal validation； according to the above inclusion and exclusion criteria， the clinical informations of 1 126 patients with IVF-ET and successful pregnancy who were not in our hospital were screened for external verification. The baseline data of the patients were recorded， according to the occurrence of ectopic pregnancy in the IVF-ET patients， they were divided into occurrence group and non-occurrence group， Logistic regression was used to analyze the influencing factors of ectopic pregnancy in the patients with IVF-ET； based on the above influencing factors， the nomogram model for predicting the occurrence of ectopic pregnancy in the IVF-ET patients was established. The Bootstrap method was used for model verification， and the consistency index （C-index） was calculated to test the accuracy of the model； the area under the curve （AUC） of receiver operating characteristics （ROC） and calibration curve were used to evaluate the discrimination and calibration of the nomogram model. Results Among 522 patients in the internal validation group， 45 patients had ectopic pregnancy， the incidence rate was 8.62%； after the baseline data of the patients in two groups were compared， the Logistic regression analysis results showed that the history of intrauterine operation， smoking，pelvic inflammatory disease， previous ectopic pregnancy and elevated serum estradiol （E2） level on the day of human chorionic gonadotrophin （hCG） injection were the influencing factors of ectopic pregnancy in the patients with IVF-ET （OR>1，P<0.05）； the nomogram model for predicting the ectopic pregnancy in the IVF-ET patients was constructed； Bootstrap internal verification method was used to verify the nomogram model， the calibration degree of the model was good， and the C-index was 0.702； it showed that the model had good discrimination. The nomogram model was internally verified， the ROC curve was drawn and the AUC of nomogram model in predicting the risk of ectopic pregnancy in the patients with IVF-ET was 0.735（>0.70）， which had certain predictive value； the nomogram model was externally verified， the ROC curve was drawn and the AUC of nomogram model for predicting the risk of ectopic pregnancy in the patients with IVF-ET was 0.862（>0.80）， which had better predictive value. Conclusion Ectopic pregnancy in the patients with IVF-ET is affected by many factors such as history of intrauterine operation， smoking， pelvic inflammatory disease， previous ectopic pregnancy and the E2 level on the day of hCG injection， and the nomogram model based on multiple influencing factors provides an effective method for reasonably predicting the risk of ectopic pregnancy in the patients with IVF-ET.

Objective To investigate the changes of follicular helper T lymphocytes （Tfh） in peripheral blood of the patients with systemic lupus erythematosus （SLE） during pregnancy， and to clarify the immune effects of Tfh cells duing pregnancy. Methods The SLE patients from Qingdao Municipal Hospital and Qingdao Women and Children Hospital were divided into SLE with pregnancy group （n=32） and SLE without pregnancy group （n=30）；at the same time， healthy pregnancy group （healthy pregnancy women，n = 30） and healthy non-pregnancy group（healthy non-pregnancy women，n=25） were used as control. According to the changes of lupus during pregnancy and after termination of pregnancy within 3 months， the 32 cases of SLE patients in SLE with pregnancy group （2 cases were lost during follow-up） were divided into poor prognosis group（n=8） and clinical stabilization group（n=22）. The clinical data and laboratory examination indexes of the patients were collected. The percentages of CD4+inducible T-cell costimulatory factor （ICOS）+ chemokine C-X-C-motif receptor 5（CXCR5）+Tfh cells in the CD4+ T lymphocytes in peripheral blood of the patients in various groups（ percentages of Tfh cells ） were detected by flow cytometry. The relationships between the percentages of Tfh cells in peripheral blood of the patients in SLE with pregnancy group and the laboratory examination indexes were analyzed by Spearman correlation analysis.The levels of interleukin-6 （IL-6）， interleukin-21 （IL-21） and interferon-γ （IFN- γ） in peripheral blood of the subjects in various groups were detected by enzyme linked immunosorbent assay（ELISA）method. Results Compared with healthy pregnancy group， the percentage of Tfh cells in peripheral blood of the patients in SLE with pregnancy group was significantly increased（P<0.01）， and the percentage of Tfh cells in peripheral blood of the subjects in healthy non-pregnancy group was significantly decreased（P<0.05）. Compared with SLE without pregnancy group， the percentage of Tfh cells in peripheral blood of the patients in SLE with pregnancy group was significantly increased（P<0.05）. During the follow-up， compared with clinical stabilization group， the percentage of Tfh cells in peripheral blood of the patients in poor prognosis group was significantly increased（P<0.05）. The percentage of Tfh cells in peripheral blood of the patients in SLE with pregnancy group was positively correlated with the levels of anticardiolipin antibody （ACA） and anti-β2 glycoprotein Ⅰ（β2GPⅠ） antibody（r=0.743 1，P<0.05； r=0.830 7，P<0.01）. Compared with healthy pregnancy group， the levels of IL-6 and IL-21 in peripheral blood of the patients in SLE with pregnancy group were significantly increased（P<0.05）；compared with SLE without pregnancy group，the level of IL-21 in peripheral blood of the patients in SLE with pregnancy group was significantly increased（P<0.05）. Conclusion The increase of the percentage of Tfh cells in peripheral blood of the patients with SLE in early pregnancy may be related to the poor pregnancy prognosis in the patients with SLE.

Objective To analyze the influencing factors of the debilitation status of elderly hospitalized patients with comorbid diseases， and to provide reference for its early intervention. Methods A total of 124 elderly patients with comorbid diseases were selected from April 2017 to June 2021 in the Department of Geriatrics （Jinan Shande Nursing Home） in the Southern District of the Second Hospital of Shandong University. The clinical data and laboratory indexes were collected， including age， gender， body mass index（BMI）， education level， albumin（ALB） level， white blood cell （WBC） count， C-reactive protein （CRP） level， hemoglobin（Hb） level， serum creatinine （SCr） level，and blood urea nitrogen （BUN） level；the comprehensive assessment of the elderly was carried out and FRAIL scale was used to evaluate the debilitation of the patients. According to the score of FRAIL scale， the patients were divided into debilitation group and non-debilitation group. Univariate analysis and binary multivariate Logistic regression analysis were used to analyze the influencing factors of the debilitating status of elderly inpatients with comorbid diseases. Results In 124 patients with comorbid diseases， there were 68 cases （54.84%） in debilitation group and 56 cases （45.16%） in non-debilitation group. The differences in BMI level， mininutritional assessment short form （MNA-SF） score， Charlson comorbidity index （CCI）score， sleep status， cognitive function status， Hb level， SCr level and BUN level between two groups were statistically significant （P<0.05）.The binary multivariate Logistic regression analysis showed that MNA-SF score （OR=0.511，95%CI： 0.373－0.700，P<0.01） was the protective factor for debilitation in the elderly hospitalized comorbid patients， and CCI score （OR=2.612， 95%CI： 1.627－4.191， P<0.01） was the independent risk factor for debilitation in the elderly hospitalized comorbid patients （P<0.05）. Conclusion The incidence of debilitation in the elderly hospitalized patients with comorbid diseases is high， and nutritional status and comorbidity degree are the independent influencing factors for debilitation.

Objective To collect the clinical data of a Sotos syndrome patient with epilepsy and necrotic enterocolitis as performance， and to analyze the importance of diagnosis， antiepileptic therapy and complication follow-up of this disease. Methods The male patient was 21 months of age.The clinical manifestion of the patient was developmental delay and recurrent seizures，and the physical examination of patient showed excessive growth and special face.The patient had a previous history of necrotizing enterocolitis in the neonatal period.Combined with the clinical and genetic data of the patient， the etiology was confirmed and the characteristics of diagnosis and treatment were analyzed，and the relevant literatures were reviewed and summarized. Results Combined with the clinical features， auxiliary examination and genetic testing，the patient was diagnosed as Sotos syndrome and epilepsy.After 1 year and 3 months of rehabilitation，the developmental delay of the patient was improved and the rehabilitation was continued and the patient was followed-up；after 1 month of levetiracetam monotherapy， the seizure was completely controlled；there were no more seizures，and the reexamination electroencephalogram was normal.The patient appeared vomiting and bloody stools during the neonatal period，and was diagnosed as necrotic enterocolitis and colonic perforation combined with abdominal ultrasound and X-ray.After more than 20 d of non-operative conservative treatment，the symptoms of patient were gradually improved and the patient recovered. Conclusion In clinical practice，genetic testing is recommended for the patients with suspected Sotos syndrome to confirm diagnosis and symptomatic treatment.Active rehabilitation therapy is recommended to improve the developmental delay of the patient；epileptic seizures in the patients with Sotos syndrome should be actively treated with antiepileptic drugs.Neonatal necrotic enterocolitis is the first case of this syndrome，which may provide reference and treatment experience for the study of the relationship between genotype and phenotype of this syndrome.

Objective To analyze the clinical manifestations， diagnostic methods and treatment process of the patients with non-Hodgkin’s lymphoma complicated with human coronavirus（HCoV）-HKU1 pneumonia and improve the clinical medical staff’s awareness of the disease， and to reduce the occurrence of clinical adverse events. Methods The clinical data of a patient with non-Hodgkin’s lymphoma complicated with HCoV-HKU1 pneumonia with hot flashes and night sweats， dry cough and dry throat as the main clinical features who were hospitalized in the hospital in January 2021 were analyzed， and the relevant literatures were reviewed and the clinical manifestations and diagnosis of HCoV-HKU1 were analyzed. Results The female patient was admitted to the hospital due to diagnosed non-Hodgkin’s lymphoma for more than 2 months. The physical examination results showed Karnofsky score was 90 points； there was no palpable enlargement of systemic superfical lymph nodes； mild tenderness in the right lower abdomen， no rebound tenderness，and slightly thicker breath sounds in both lungs were found， and a few moist rales were heard in both lower lungs. The chest CT results showed diffuse exudative foci in both lungs，and the number of white blood cells in the urine analysis was 158 μL^－1； next generation sequencing technique（NGS） was used the detect the bronchoalveolar lavage fluid，and HCoV-HKU1 pneumonia was diagnosed. At admission， the patient had symptoms such as dull pain in the right lower abdomen， nighttime cough， and night sweats； antiviral treatment with oseltamivir was ineffective. After treatment with Compound Sulfamethoxazole Tablets and Lianhua Qingwen Granules， the respiratory symptoms of the patient disappeared. The re-examination chest CT results showed the exudation was absorbed. Conclusion The clinical symptoms of the patients with non-Hodgkin’s lymphoma complicated with HCoV-HKU1 pneumonia are non-specific. When the diffuse shadow changes in the lungs are found in clinic， and the new coronavirus nucleic acid test is negative， attention should still be paid to the possibility of other HCoV infections. The NGS can efficiently screen the infectious pathogens， which is beneficial to guide the diagnosis and treatment of pulmonary infectious diseases more accurately.

Objective： To investigate the clinical characteristics， diagnosis procedure and treatment method of the patient with recurrent vulvar angiomyofibroblastoma （AMFB），and to improve the understanding of clinical doctors to this disease. Methods The clinical data， imaging examination results， pathological examination results and immunohistochemical examination results of 1 patient with recurrent vulvar AMFB were collected； the above clinical data were analyzed， and the relevant literatures were reviewed. Results A 50-year-old female patient underwent excision of vulvar mass 5 years ago in the local hospital for a vulvar mass，the postoperative pathology suggested AMFB.The patient was now admitted to the hospital due to a 6-month self-reported vaginal mass and a high degree of suspicion of recurrence of AMFB.The gynecological examination suggested a swelling under the labia major on the left side， with complete skin surface， deep protrusion towards the rectum and unclear boundary with the bowel；the size of tumor was about 5.0 cm×4.0 cm×4.0 cm，with regular shape， clear boundary，and poor activity，and there was no obvious tenderness，ulceration and skin lesion. Superficial ultrasound showed a slightly low anechoic light mass with a range of 5.0 cm×4.0 cm×4.0 cm at the left labia major， in which multiple mass hyperechoics were observed. Color Doppler flow imaging （CDFI） showed visible blood flow signals in and around the labia major. Conclusion Although recurrence and malignancy of AMFB are rare， postoperative follow-up of AMFB should be given high priority to reduce the recurrence and increase the cure rate.

Objective To investigate the application of arthroscope in the surgical treatment of the patients with temporomandibular joint synovial osteoma disease， and to clarify its value in improving the clinical diagnosis and treatment efficiency and prognosis of the patients with the disease. Methods The clinical data of two patients with temporomandibular synovial osteoma disease treated with arthroscope were collected.The opening degree， maximum anterior extension and lateral movements， and pain improvement of the patients were compared before and 6 months after surgery， and the operation procedure was summarized，and the significance of arthroscope application was analyzed combined with literature review. Results Two patients with temporomandibular synovial osteoma disease both chose the method of open surgery combined with arthroscope.With the assistance of arthroscope， the open surgery completely removed the chondroma bodies， and the removal of inflammatory synovial tissue was carried out.The opening degrees of the patients reached 33 and 35 mm， respectively， 6 months after operation， and the maximum anterior extension and lateral movement were ≥7 mm，the joint function was gradually restored. Conclusion Arthroscope has obvious advantages in the surgical treatment of the patients with temporomandibular synovial osteoma disease， but fully arthroscopic surgery is limited by the complex structure of the joint， and if the tumor is too large or not confined to the supra-articular cavity， open surgery combined with arthroscope is required when necessary.

Objective To explore the feasibility of the treatment strategy of no treatment and only regular follow-up for the patient with asymptomatic postmenopausal pulmonary benign metastasizing leiomyomat（PBML）， and to provide a reference for the diagnosis， treatment， and prognosis of the disease. Methods The clinical characteristics， examination results， and follow-up results of a patient with PBML were analyzed retrospectively， and the pathogenesis and treatment measures were discussed in combination with the literature review. Results The female patient， 52 years old，was admitted to the hospital due to the physical examination of multiple nodules in both lungs， and previously underwent total hysterectomy for multiple uterine leiomyomas. There were no obvious pulmonary signs，and imaging examination showed multiple round nodular hyperdense shadows in both lungs. Percutaneous biopsy of pulmonary nodules was performed under the guidance of computed tomography （CT） in the left lung. The pathological diagnosis was PBML， and estrogen receptor（ER） and progesterone receptor（PR） were positive. The patient’s sex hormone level was in menopause，so no treatment was given， and regular imaging examinations and follow-up were carried out. The CT follow-up at 5， 11， 20， and 34 months after percutaneous lung puncture showed that no obvious enlargement of bilateral pulmonary nodules was found， and no cough， wheezing and other discomfort symptoms were found at 41 months. Conclusion The asymptomatic female PBML patients with hormone levels in menopause may not be treated and only need the regular imaging follow-up.

Methods The clinical materials of a patient with CNSL complicated with SS were collected. The diagnosis process was summarized and the relevant literatures were reviewed. Results The female patient，51 year old，was admitted to the hospital for dry mouth and dry eyes with nausea and vomiting for more than one month.The physical examination results showed the tongue surface was dry，the patient moved restlessly at times，and the limb strength was level 4；the left calcaneal and tibial tests were not stable， and the Romberg’s sign were positive and bilateral pathological signs were positive.The results of laboratory tests showed anti-Sjogren’s syndrome A antibody（anti-SSA antibody） and anti-Sjogren’s syndrome B antibody（anti-SSB antibody） were positive.The results of tear secretion test showed left side 5 mm； the corneal staining was positive，and salivary gland isotope test was positive. The enhanced magnetic resonance imaging（MRI） results showed multiple abnormal enhancement signals in bilateral medial cerebellar hemispheres， optic chiasm， pituitary stalk and pineal gland；the magnetic resonance spectrum（MRS） results of the head revealed that the N-acetyl-L-aspartic acid（NAA） peak and creatine（Cr） peak of L-aspartic acid were decreased， while the choline（Cho） peak was increased， NAA/Cr=1.74， Cho/Cr=3.52， Cho/NAA=2.02；the lactate（Lac） peak and lipid（Lip） peak were also seen， single-voxel ¹H-MRS and multi-voxel ¹H-MRS were found in bilateral cerebellar hemispheres. The pathological examination showed that high-grade B-cell lymphoma with hemorrhage and the clinical diagnosis was CNSL complicated with SS. Radiotherapy and chemotherapy were given after discharge in local hospital. Conclusion The incidence of CNSL with SS is low， and it is easy to be misdiagnosed as central nervous system demyelinating disease. CNSL should be considered when the symptoms and signs of central nervous system are present in the SS patients. Objective To analyze the clinical manifestation， diagnosis and treatment process of a patient with central nervous system lymphoma （CNSL） complicated with Sj?gren’s syndrome（SS） and explore its pathogenesis， and to provide reference for the diagnosis of this rare disease.

Objective To establish the rat models of intrauterine adhesion （IUA） with the chemical and heat conduction dual injury method， and to clarify its characteristics and advantages as an improved approach of chemical injury in constructing the animal model with similar pathological changes of IUA. Methods Six female SD rats aged 6－8 weeks were randomly divided into chemical injury group and chemical and heat conduction dual injury group （dual injury group），with 3 rats in each group.The left uterine horns of rats were used as experimental group and the right uterine horns were used as self-control group. The left uterine horns of the rats in chemical injury group， which were defined as the chemical injury experiment group， were treated with 95% ethanol for 3 min； the right uterine horns of the rats， which were defined as chemical injury self-control group，didn’t received any treatment. The left uterine horns of the rats in dual injury group， which were defined as dual injury experiment group， were treated with 95% ethanol in the left uterus for 3 min， and then treated with 100 ℃ hot water for 1 min； the right uterine horns of the rats， which were defined as dual injury self-control group， didn’t receive any treatment.On the 14 th day after modeling， the thickness of endometrial， the number of glands， the area of endometrial fibrosis and the proliferation index （PI） of glandular epithelial cells and stromal cells of the rats in various groups were detected by hematoxylin and eosin （HE） staining， Masson staining， and Ki67 immunohistochemical staining method. Results The HE staining results showed that compared with self-control group，the thickness of endometrium of the rats and the number of glands in chemical injury experiment group and dual injury experiment group were significantly reduced（P<0.05）.Compared with chemical injury experiment group， the uterine cavity of the rats in dual injury experiment group was closed， the endometrial thickness was reduced （P<0.05）， and the number of endometrial glands was decreased（P<0.05）. The Masson staining results showed that compared with self-control group， the fibrotic hyperplasia in endometrial layers of the rats in chemical injury experiment group and dual injury experiment group was seen， and the ratios of endometrial fibrosis area in exdometrium tissue were apparently increased （P<0.05）. Compared with chemical injury experiment group， fibrotic adhesion and intact myometrium of the rats in dual injury experiment group were found， and the ratio of endometrial fibrosis area was apparently increased （P<0.05）. The Ki67 immunohistochemical staining results showed that compared with self-control group， the nuclear shape and arrangement of glandular epithelial cells appeared the changes in endometrium tissue of the rats in chemical injury experiment group and dual injury experiment group， and the PI was significantly increased （P<0.05）. Conclusion Chemical and heat conduction dual injury method is an easy and reproducible operation that can establish the IUA model with fibrotic adhesion and intact muscle layer for the safety evaluation， mechanism research and clinical transformation of IUA， which possesses a good application perspect.

Objective To establish a method of nucleic acid colloidal gold strip for rapid detection of respiratory tract fungal infection with polymerase chain reaction （PCR） technique and colloidal gold technique， and to evaluate the detection effect. Methods The fungal internal transcribed spacer（ITS） gene fragment was selected as the target gene according to GenBank database， and the fungal ITS gene fragment was analyzed by DNAMAN software， and labeled with Biotin and fluorescein isothiocyanate （FITC） at the 5' end of fungal universal primers， respectively； the PCR system was established and the optimal conditions of the systerm were optimized. The best labeling amount of colloidal gold test strips to label streptavidin and the best coating concentration of quality control line and detection line were comfirmed， the nucleic acid colloidal gold test strip was assembled， and the specificity， sensitivity， repeatability and stability of the colloidal gold test strip were verified. Results The DNA of Candida albicans， Cryptococcus neoformans and Mucor was extracted by boiling method， the concentrations were more than 180 μg·L^－1，and the purities were 1.60－2.10. Under the condition of pH 7.0， the optimal labeling amount of colloidal gold- labeled streptavidin was 3.3 μg in 100 μL of colloidal gold-solution， the concentration of biotinylated bovine serum albumin （BSA-Biotin） coated by quality control line was 2.00 g·L^－1， and the concentration of anti-FITC antibody coated by detection line was 1.00 g·L^－1. The specificity of nucleic acid test strip was consistent with the results of electrophoresis， and only Candida albicans， Cryptococcus neoformans and Mucor showed positive results. There were no cross reactions with Staphylococcus aureus， Streptococcus pneumoniae， Type B Hemolytic streptococcus， Pseudomonas aeruginosa， Klebsiella pneumoniae， Acinetobacter baumannii， Escherichia coli and other common respiratory tract infection bacteria. The sensitivity detection results showed that the nucleic acid test strips could still accurately detect the DNA of Candida albicans， Cryptococcus neoformans and Mucor when the DNA concentrations were of 10^－4， 10^－2 and 10^－3 μg·L^－1， respectively. The results of ordinary PCR electrophoresis showed that the lowest detection concentrations of Candida albicans， Cryptococcus neoformans and Mucor were 0.01， 1.00，and 0.10^{μg·L－1， respectively.In repeatability test， nucleic acid test strips were verified by different operators in different laboratories， the results were consistent， and the reproducibility was good； in stability test， nucleic acid test strips were tested in 6， 9 and 12 months and the results were as expected with good stability. Conclusion The established method of nucleic acid colloidal gold test strip for respiratory tract fungal infection in this study can be used to detect Candida albicans， Cryptococcus neoformans， Mucor and other fungi with high specificity， sensitivity， simple and rapid operation.}