To investigate the protective effect of melatonin on the hydrogen peroxide（H₂O₂）-induced oxidative stress injury of human neuroblastoma SH-SY5Y cells， and to explore its mechanism.

The SH-SY5Y cells were cultured in vitro and divided into control group，H₂O₂ group， melatonin groups， and N-acetyl-2-benzyltryptamine（Luzindole） group. The medium containing 200 μmol·L^－1 H₂O₂ was added in H₂O₂ group；different concentrations （1， 5 and 10 μmol·L^－1） of melatonin and medium containing 200 μmol·L^－1 H₂O₂ were added in melatonin groups； 50 μmol·L^－1 Luzindole， 10 μmol·L^－1 melatonin and the medium containing 200 μmol·L^－1 H₂O₂ were added in Luzindole group. The survival rates of human neuroblastoma SH-SY5Y cells were measured by CCK-8 method， the apoptotic rates of SH-SY5Y cells in various groups were detected by flow cytometry， and DCFH-DA fluorescence probe was used to detect the levels of reactive oxygen species（ROS） in the cells in various groups. The expression levels of microtubule-associated protein light chain 3-Ⅱ（LC3-Ⅱ）protein in the cells in various groups were detected by Western blotting method， and the fluorescence intensities of autophagic vesicles in various groups were observed by fluorescence microscope.

Compared with control group， the survival rate of SH-SY5Y cells in H₂O₂ group was decreased （P<0.01），the ROS level and the apoptotic rate were increased （P<0.01）. Compared with H₂O₂ group， the survival rates of SH-SY5Y cells in different concentrations of melatonin groups were increased（P<0.05），the ROS levels and the apoptotic rates were significantly decreased （P<0.01），the LC3-Ⅱ protein expression levels were significantly increased （P<0.01）， and the fluorescence intensity of autophagic vesicles in 10 μmol·L^－1 melatonin group was increased（P<0.05）. Compared with 10 μmol·L^－１ melatonin group， the survival rates of SH-SY5Y cells in 1 μmol·L^－１ melatonin group and Luzindole group were significantly decreased（P<0.05）；the ROS levels and the apoptotic rates in the SH-SY5Y cells in 1 and 5 μmol·L^－1 melatonin groups and Luzindole group were significantly increased（P<0.05），and the LC3-Ⅱ protein expression levels in the SH-SY5Y cells were decreased（P<0.05 or （P<0.01）， and the fluorescence intensities of autophagic vessicles in the SH-SY5Y cells in 1 μmol·L^－1 melatonin group and Luzindole group were significantly decreased （P<0.05）.

Melatonin can inhibit the H₂O₂-induced oxidative stress injury of SH-SY5Y cells and has a neuroprotective effect，and its mechanism may be related to reducing the ROS levels and enhancing the autophagy of cells.

To investigate the inhibitory effects of endoplasmic reticulum stress（ERS） and endoplasmic reticulum autophagy on the growth of transplanted melanoma model in amyloid precursor protein（APP）/presenilin 1（PS1）mice， and to clarify the possible mechanism of its inhibitory effect of protein kinase R-like endoplasmic reticulum（PERK）-eukaryotic translation initiation factor 2α（eIF2α）-activating transcription factor 4（RATF4） pathway.

The C57BL/6J（C57） and APP/PS1 mice were transplanted with melanoma B16 cells to establish the C57 transplanted tumor and APP/PS1 transplanted tumor models of male mice），and used as C57 transplanted tumor group （n=7） and APP/PS1 transplanted tumor group （n=7）. The tumor appearance time was observed and the tumor volume was calculated of the mice in two groups. The expression levels of glucose regulatory protein 78（GRP78），PERK and lysosomal cathepsin L （cathepsin L） mRNA in the transplanted tumor tissue of the mice in two groups were detected by real-time fluorescence quantitative PCR（RT-PCR）.The expression levels of GRP78， PERK， phos-PERK， eukaryotic translation initiation factor 2α （eIF2α），p-eIF2α， ATF4， and FAM134B in the transplanted tumor tissue of the mice in two groups were detected by Western blotting method. The expression levels of protein disulfide isomerase（PDI） protein in two groups were detected by immunohistochemistry and the adenosine triphoshate（ATP） levels in the transplanted tumor tissue of the mice were determined by fluorescence assay.

Compared with C57 transplanted tumor group， the tumor appearance time of the mice in APP/PS1 transplanted tumor group was late， and the tumor volume was decreased（P<0.05）；the expression levels of GRP78 and PERK mRNA in the transplanted tumor tissue were increased （P<0.05）， and the expression levels of GRP78， p-PERK/PERK， p-eIF2α/eIF2α and ATF4 proteins were increased（P<0.05）. In C57 transplanted tumor group， melanoma granules were found in the cytoplasm of the tumor cells， which was the morphological characteristic of transplanted tumor tissue， and a small amount of brown granules were found， which were PDI positive granules. In APP/PS1 transplanted tumor group， a small amount of melanoma granules and widely expressed brown granules were found in the cytoplasm of tumor cells. Compared with C57 transplanted tumor group， the expression level of FAM 134B protein in the transplanted tumor tissue of the mice in APP/PS1 transplanted tumor group was increased（P<0.05）， the expression level of cathepsin L mRNA was increased（P<0.05），and the ATP level was decreased（P<0.05）.

The growth of tumor in the APP/PS1 transplanted tumor model mice is slow， and its mechanism may be related to the activation of endoplasmic reticulum stress PERK-eIF2α-ATF4 signaling pathway and regulated endoplasmic reticulum autophagy.

To investigate the expression levels of Wnt5a protein in ovary tissue of the mice at different development stages， and to clarify its effect on oocyte autophagy.

The ovary tissue of mice at different developmental stages were collected， and Western blotting method was used to detect the expression levels of Wnt5a protein in perinatal ovary tissue of the mice. The ovarian tissue of the 1 d after partum （1 dpp） mice were selected for immunofluorescence staining. The oocyte cytoplasmic marker DDX4 and granulosa cell marker FOXL2 （red） were co-stained with Wnt5a （green） and nuclear marker Hoechst （blue）， respectively，and the morphology and the number of oocytes were observed. The ovary tissue of the mice at 17.5 d of embryonic period（17.5 dpc）were divided into control group， 1 mol·L^－1 overexpressing Wnt5a group （rWnt5a group）， 1 mol·L^－1 inhibitor IWP2 group and 1 mol·L^－1 inhibitor BOX5 group. The expression levels of Wnt5a， LC3 and P62 proteins in ovary tissue of the mice in various groups were detected by Western blotting method， and the number of ovarian oocytes was observed by immunofluorescence method.

Wnt5a was expressed in ovary tissue of the mice at different developmental stages. Compared with control group， the expression levels of Wnt5a protein in 17.5 dpc and 1 dpp groups were significantly increased （P<0.05）. The fluorescence localization revealed that the target protein Wnt5a overlapped with the markers DDX4 and FOXL2. The Western blotting results showed that compared with control group， the expression level of Wnt5a protein in rWnt5a group was significantly increased （P<0.01）， the expression levels of Wnt5a and LC3 proteins in inhibitor IWP2 group were significantly decreased （P<0.01）， and the expression level of Wnt5a protein in inhibitor BOX5 group was significantly increased （P<0.01）. Compared with rWnt5a group， the expression level of LC3 protein in inhibitor IWP2 group was significantly decreased（P<0.01），and the expression level of P62 protein was significantly increased （P<0.01）. The immunofluorescence staining results showed that the number of oocytes in IWP2 inhibitor group was reduced compared with control group.

Wnt5a may regulate the oocyte survival by affecting the level of autophagy in ovary of the mice.

To establish a zebrafish xenograft model of human nasopharyngeal carcinoma and explore the inhibitory effect of curcumin on nasopharyngeal carcinoma in vivo and in vitro， and to provide the basis for elucidating the molecular mechanism of anti-nasopharyngeal carcinoma of curcumin.

The nasopharyngeal carcinoma CNE-2 cells were divided into control group and CM-Dil group. The cells were counted for 5 consecutive days， and the fluorescence intensity changes were observed. The zebrafish xenograft model of nasopharyngeal carcinoma was established by microinjection. The zebrafishes were divided into control group， PBS group and model group. The zebrafishes in control group were not treated， the zebrafishes in PBS group were injected with 10 nL PBS buffer， and the zebrafishes in model group were injected with 10 nL cell suspension. The survival number of zebrafishes in each group was counted， and the fluorescence intensity of transplanted tumor of the zebrafishes in model group was observed and quantitatively analyzed by macro stereomicroscope. The morphology of zebrafish transplanted tumors in control group and model group were observed by HE staining. The zebrafishes with the days postfertilization（dpf） was 3 were exposed to feeding water containing different concentrations （0， 0.625， 1.250， 2.500， 5.000，and 7.500， 10.000 μmol·L^－1） of curcumin， the death and deformity were observed；the death rates of zebrafishes were calculated，and the concentration of curcumin in vivo was selected. The zebrafish models were treated with differnent concentrations （0，0.625， 1.250， 2.500， and 5.000 μmol·L^－1） of curcumin for 48 h， the growth and metastasis of the transplanted tumor were detected. The CNE-2 cells were treated with different concentrations（0， 10， and 20 μmol·L^－1） of curcumin for 24 h， the proliferation rates and migration rates were detected by CCK-8 method and wound healing assay.

Compared with control group， there was no significant difference in the number of CNE-2 cells in CM-Dil group （P>0.05）. On the 5th day of the experiment， compared with control group， there was no significant difference in the survival number of zebrafishes in PBS group （P>0.05）； compared with control group and PBS group， the survival number of zebrafishes in model group was decreased significantly （P<0.01）. The red fluorescence intensity in zebrafishes in model group was increased with the prolongation of time. Compared with the first day after injection， the red fluorescence intensity of zebrafishes in model group was increased significantly on the fifth day after injection （P<0.05）.The HE staining results showed that the CNE-2 cells were found in the yolk sac of the zebrafishes in model group， indicating that the models were successfully constructed. The zebrafishes in 0.625， 1.250， 2.500， and 5.000 μmol·L^－1 concentrations of curcumin groups had no death and obvious toxicity. Compared with control group（0 μmol·L^－1 cureumin group），the fluorescence intensities in the zebrafish xenograft tissue in 2.500 and 5.000 μmol·L^－1 cureumin groups were signifricantly decreased（P<0.05 or P<0.01）， and the number of zebrafishes with head and tail metastasis in 5.00 μmol·L^－1 curcumin group was decreased （P<0.05）. The results of CCK-8 method and wound healing assay showed that compared with control group （0 μmol·L^－1 curcumin group），the proliferation rates and migration rates of CNE-2 cells in 10 and 20 μmol·L^－1 curcumin groups were significantly decreased （P<0.05）.

The CNE-2 cells can be successfully implanted in the zebrafishes and to estabish the zebrafish models of nasopharyngeal carcinoma. Curcumin can inhibit the proliferation and metastasis of nasopharyngeal carcinoma in vivo and in vitro， and it has a good prospect of cell anti-tumor.

To investigate the effect of human umbilical cord mesenchymal stem cells （MSCs） on the proliferation and apoptosis of cervical cancer HeLa cells， and to elucidate its possible mechanism.

The human umbilical cord MSCs were isolated and cultured. The morphology of cells were observed by light microscope. The expressions of surface antigen markers CD90， CD105， CD34 and CD45 were identified by flow cytometry. The MSCs were added with second antibody （without first antibody） were used as parallel control group.The conditioned medium containing 20% and 60% MSCs and cervical cancer HeLa cells were co-cultured for 72 h， colony formation experiment was used to observe the colony forming ability of HeLa cells after treated with of MSCs conditioned medium.CCK-8 experiment was used to detect the inhibitory rates of proliferation of the HeLa cells. Flow cytometry was used to detect the apoptotic rates of HeLa cells. Western blotting method was used to detect the expression levels of proliferation- and apoptosis-related gene proteins in the HeLa cellsn.

After 72 h of inoculation， a small number of cells adhered to the wall， and gradually became flat monolayer cells， which grew in clusters and vortices after one week. With the increase of cell density， the cell body became slender， similar to that of fibroblasts.The MSCs showed positive expressions of CD90 and CD105， but negative expressions of CD34 and CD45， which conformed to the phenotype of stem cells.The results of colony formation test showed that the inhibitory rates of colony formation of HeLa cells were 18.56% and 37.64% respectively when the conditioned medium containing 20% and 60% MSCs were added.Compared with control group，the early apoptotic rate of HeLa cells after treated with MSCs for 48 h was increased （P<0.05）. After the HeLa cells were treated with MSCs for 0， 24 and 48 h， the expression levels of cysteinyl aspartate-specific proteinase-3 （Caspase-3） and P53 proteins were increased continuously （P<0.05）， and the expression levels of B-cell lymphoma-2 （Bcl-2） protein were decreased continuously （P<0.05）.

The MSCs can inhibit the proliferation of HeLa cells and induce the apoptosis of HeLa cells in vitro，and its mechanism may be related to up-regulating the expression of Caspase-3 and P53 and down-regulating the expression of anti-apoptotic factor Bcl-2.

To explore the effect of rutin on the colorectal cancer （CRC） SW480 cells， and to clarify the effect of rutin in the treatment of CRC and its possible molecular mechanism.

The Comparative Toxicogenomics（CTD） and GeneCards databases were used to predict the intersection mRNA related to rutin； Gene Ontdogy（GO） and Kyoto Encyclopedia of Genes and Genones（KEGG） databases were used to predict the biological function and action pathways of the intersection mRNA； String database was used to find the Notch-1 related proteins. The human CRC SW480 cells were used as the subjects， 0.1， 1.0， and 10.0 μmol·L^－1 DAPT were given for preliminary screening， then 1， 2， 4， 8 and 10 μmol·L^－1 DAPT were given to intervene， and the optimal concentrations of rutin and DAPT were screened out. The human CRC SW480 cells were divided into blank control group， rutin group （22 μmol·L^－1）， DAPT group （1 μmol·L^－1） and rutin+DAPT group（22 μmol·L^－1 rutin+1 μmol·L^－1 DAPT）.MTT method was used to detect the cell proliferation activities； Hoechst33258 nuclear staining was used to observe the apopotic morphology of cells，and flow cytometry was used to detect the apoptotic rates；real-time fluorescence quantitative PCR（RT-qPCR） method was performed to detect the expression levels of Notch-1 mRNA in the SW480 cells in various groups； Western blotting method was used to determine the protein expression levels of Notch-1，caspase-3，caspase-9，B-cell lymphoma 2（Bcl-2），and Bcl-2 associated X protein in the SW480 cells in various groups.

In CTD and GeneCards databases， there were a total of 34 intersection mRNA；the GO and KEGG databases predicted that the intersection mRNA was related to the process of colon cancer apoptosis； String database verified 16 Notch-1 related proteins （score>0.42）； the optimal administration concentrations of rutin and DAPT were 22 μmol·L^－1 and 1 μmol·L^－1； compared with blank control group， the apoptotic rates in rutin group， DAPT group and rutin+DAPT group were increased significantly （P<0.05）， the expression levels of Notch1 mRNA and protein were significantly reduced （P<0.05）， the expression levels of caspase-3， caspase-9， and Bax proteins in the cells were increased significantly（P<0.05）， while the expression levels of Bcl-2 protein were decreased significantly （P<0.05）. There was no significant difference in the apoptotic rate between rutin group and DAPT group （P>0.05）. Compared with rutin group and DAPT group， the apoptotic rate in rutin+DAPT group was significantly increased（P<0.05）， the expression levels of caspase-3， caspase-9， and Bax proteins were increased significantly（P<0.05），while the expression level of Bcl-2 protein was decreased significantly（P<0.05）.

The results of bioinformatics analysis and experiment show that rutin can promote the apoptosis of CRC cells， which may be achieved by targeting Notch-1 and regulating MAPK signal pathway.

To investigate the effect of atorvastatin（ATO） on the vascular endothelial cell injury induced by oxidized low-density lipoprotein （Ox-LDL） /β2-glycoprotein Ⅰ （β2GPⅠ）/anti-β2-glycoprotein Ⅰ antibody （anti-β2GPⅠ） complex and its effect on the TRAF3 interacting protein 2 （TRAF3IP2）/ Ⅰ kappa B kinase （IKKγ）/ nuclear factor kappa-B （NF-κB） signaling pathway， and to elucidate the possible molecular mechanism of improvement effect of ATO on the endothelial cell injury in the context of antiphospholipid syndrome （APS）.

The endothelial cell injury models were established by treating the human umbilical vein endothelial cells （HUVECs） with β2GPⅠ（100 mg·L^―1），Ox-LDL（50 mg·L^―1），β2GPⅠ/anti-β2GPⅠ（100 mg·L^―1），Ox-LDL/β2GPⅠ and Ox-LDL/β2GPⅠ/anti-β2GPⅠ complex.The HUVECs were divided into control group，β2GPⅠ group，Ox-LDL group，β2GPⅠ/anti-β2GPⅠ group，Ox-LDL/β2GPⅠ group，Ox-LDL/β2GPⅠ/anti-β2GPⅠ group，ATO（10 μmol·L^―1）+ Ox-LDL/β2GPⅠ group and ATO+ Ox-LDL/β2GPⅠ/anti-β2GPⅠ group. The survival rates of HUVECs in various groups were detected by MTT assay， the fluorescence intensities of intracellular reactive oxygen species （ROS） in various groups were detected by chemical fluorescence probe， and the endothelin-1（ET-1） levels in various groups were detected by ELISA. The expression levels of TRAF3IP2， p-IKKγ and p-NF-κB p65 in the HUVECs in various groups were detected by Western blotting method.

Compared with control group， the survival rates of HUVECs in Ox-LDL/β2GPⅠ group and Ox-LDL/β2GPⅠ/anti-β2GPⅠ group were significantly decreased （P<0.01）；compared with Ox-LDL/β2GPⅠ group and Ox-LDL/β2GPⅠ/anti-β2GPⅠ group，after treatment for 1 h，the survival rates in ATO+Ox-LDL/β2GPⅠ group and ATO+Ox-LDL/β2GPⅠ/anti-β2GPⅠ group were significantly increased （P<0.05）. Compared with control group， the fluorescence intensities of ROS and the levels of ET-1 in the HUVECs in Ox-LDL/β2GPⅠ group and Ox-LDL/β2GPⅠ/anti-β2GPⅠ group were significantly increased （P<0.01）； the expression levels of TRAF3IP2， p-IKKγ and p-NF-κB P65 proteins in the HUVECs in were significantly increased （P<0.05）. Compared with Ox-LDL/β2GPⅠ group and Ox-LDL/β2GPⅠ/anti-β2GPⅠ group，the fluorescence intensties of ROS and the levels of ET-1， and the expression levels of TRAF3IP2， p-IKKγ and p-NF-κB P65 in ATO+Ox-LDL/β2GPⅠ group and ATO Ox-LDL/β2GPⅠ/anti-β2GPⅠ group were significantly decreased （P<0.05）.

ATO could alleviate the vascular endothelial dysfunction induced by Ox-LDL/β2GPⅠ/anti-β2GPⅠ complex，and its mechanism may be related to down-regulating the TRAF3IP2/IKK/NF-κB signaling pathway.

To investigate the effect of silencing the Bloom’s syndrome helicase（BLM） expression on the chemotherapy sensitivity of irinotecan （CPT-11） in the colorectal cancer （CRC） cells， and to elucidate its related mechanism.

The HT-29， Lovo， HCT-116， RKO， and DLD1 cells were cultured in vitro. The RKO and DLD1 cells with high expressions of BLM mRNA and protein were screened by real-time fluorescence quantitative PCR（RT-qPCR） and Western blotting methods. The cells were infected with lentivirus vector carrying shRNA to silence the expression of BLM in the RKO and DLD1 cells. The survival rates of the CRC cells and the half inhibitory concentration （IC₅₀） values of CPT-11 in various groups after transfection were detected by CCK-8 assay. The RKO or DLD1 cells were divided into LV-shNC group （infected with LV-shNC）， CPT-11+LV-shNC group （ infected with LV-shNC and treated with CPT-11） and CPT-11+LV-shBLM group （infected with LV-shBLM and treated with CPT-11）. The percentages of the cells in various groups at different cell cycles were detected by flow cytometry. The expression levels of BCL2 associated death promoter（Bad），cleaved of cysteinyl aspartate specific proteinase（cleaved caspase-3）， cyclin-dependent protein kinase 4（CDK4），cyclin-dependent protein kinase 6 （CDK6），and cyclin-dependent kinase inhibitor 1A（p21） proteins in the CRC cells in various groups were detected by Western blotting method； the apoptotic rates of the cells in various groups were detected by flow cytometry.The tumor volume and positive expression rates of BLM， Ki-67，and TUNEL in tumor tissue of the CRC-bearing mice were detected by constructing the allograft tumor model.

Compared with the HT-29， Lovo or HCT-116 cells， the expression levels of BLM mRNA and protein in the RKO and DLD1 cells were increased （P<0.05）；compared with RKO or DLD1 cells infected with LV-shNC， the expression levels of BLM mRNA in the RKO or DLD1 cells infected with LV-shBLM lentivirus were decreased（P<0.05）. Compared with LV-shNC group， the survival rates and IC₅₀ values of the cells in CPT-11+LV-shNC group and CPT-11+LV-shBLM group were decreased（P<0.05）.Compared with LV-shNC group， the percentages of the cells at G₀/ G₁ phase and S phase and the apoptotic rates of the cells in CPT-11+LV-shNC group and CPT-11+LV-shBLM group were increased （P<0.05）， while the percentages of the cells at G₂ / M phase were decreased （P<0.05），and the expression levels of Cyclin D1， CDK4， CDK6 and Bcl-2 proteins were decreased（P<0.05）， while the expression levels of p21， Bax， Bad，and cleaved caspase-3 proteins were increased （P<0.05）；compared with CPT-11+LV-shNC group， the percentages of the cells at G₀/G₁ phase and S phase and the apoptotic rate of the cells in CPT-11+LV-shBLM group were increased （P<0.05）， while the percentage of the cells at G₂/M phase was decreased （P<0.05），and the expression levels of Cyclin D1，CDK4， CDK6，and Bcl-2 proteins were decreased（P<0.05）， while the expression levels of p21， Bax， Bad，and cleaved caspase-3 proteins were increased （P<0.05）. The nude mice transplanted tumor experiment results showed that compared with LV- shNC group， the tumor volumes of the nude mice and the positive expression rates of BLM and Ki-67 in the cells in CPT-11+LV-shNC group and CPT-11+LV-shBLM group were decreased （P<0.05）， and the positive expression rates of TUNEL were increased（P<0.05）； compared with CPT-11+LV-shNC group， the tumor volume of the nude mice in CPT-11+LV -shBLM group was decreased （P<0.05）， and the positive expression rates of BLM and Ki-67 in tumor tissue were decreased （P<0.05）， and the positive expression rate of TUNEL was increased（P<0.05）.

Silencing BLM expression in the CRC cells may promote CPT-11-induced apoptosis by inhibiting the cycle progression of tumor cells， and ultimately improve chemotherapy resistance of CRC cells in vivo and in vitro.

To compare the inhibitory effects of juglone on the proliferation of human papilloma virus（HPV）-positive cervical cancer Caski cells， negative C33A cells and Caski cells knocked down the Pin1 gene（shCaski cells），and to explore its possible mechanism.

A lentiviral vector expressing Pin1 shRNA was constructed and used to infect the cells， and shCaski cells were obtained. The Caski， shCaski and C33A cells in the logarithmic growth phase were selected and divided into normal control group and 10， 20， 50 and 100 μmol·L^－1 juglone groups. After juglone treatment for 24 h， the cell proliferation activities in various groups were detected by MTT method. The Caski cells and C33A cells in the logarithmic growth phase were selected and divided into control group and 20 μmol·L^－1 juglone group. After 24 h of treatment of the cells in each group， the percentages of cells in different cell cycles and the early apoptotic rates in various groups were detected by flow cytometry. Hoechst33258 fluorescence staining was used to observe the morphology of nucleus； Western blotting method was used to detect the expressions of cell cycle and apoptosis-related proteins.

The MTT experiment showed that the proliferation activities of the Caski cells in different concentrations of juglone groups were significantly decreased （P<0.05or P<0.01） compared with normal control group， while the proliferation activities of C33A and shCaski cells in 50 and 100 μmol·L^－1 juglone groups were significantly decreased（P<0.05）.The results of Hoechst 33258 staining showed that compared with control group， both the Caski cells and C33A cells in 20 μmol·L^－1juglone group showed increased nuclear fragmentation， and the number of nuclei fragmentation of Caski cells was more than that of C33A cells. The results of flow cytometry showed that compared with control group， the percentage of Caski cells in G₂ phase in 20μmol·L^－1 juglone group was increased significantly （P<0.01）； while there was no significant difference in the percentage of C33A cells in G₂ phase in 20 μmol·L^－1 juglone group（P>0.05）；compared with control group，the early apoptotic rate of Caski cells in 20 μmol·L^－1 juglone group was increased significantly （P<0.01）； while the early apoptotic rate of C33A cells in 20 μmol·L^－1 juglone group was also increased， but there was no significantly difference（P>0.05）. The Western blotting results showed that compared with the C33A cells，the expression amount of Pin1 protein in the Caski cells was more higher； and the expression amount of Pin1 protein in Caski and shCaski cells in 20 μmol·L^－1 juglone group were decreased. Compared with control group， the expression amounts of cell cycle-related proteins in the C33A cells in 20 μmol·L^－1 juglone group did not change significantly； the expression amount of pATM， pChk2， pCdc25c Ser216 and pCdc25c Tyr216 in the Caski cells in 20 μmol·L^－1 juglone group were increased， while the expression amount of pCdc25c protein was decreased in 20 μmol·L^－1 juglone group；the expression amount of Cdk1 protein didn’t change obviously. Compared with control group， the amounts of Bcl-2 and mitochondrial CytC Caski cells in 20 μmol·L^－1 juglone group were decreased， but the expression amounts of cytoplasmic CytC，Bax，Cleaved Caspase-3 and Cleaved PARP proteins in the Caski cells in 20 μmol·L^－1 juglone group were increased.

Juglone has stronger inhibitory and pro-apoptotic effects in the HPV-positive cervical cancer cells than negative cells； its mechanism may be related to the specific inhibition of Pin1 gene by juglone.

The expressions of YTHDF2 protein in 113 cases of ESCC tissue and 95 cases of normal tissue adjacent to esophageal carcinoma were detected by immunohistochemical method. The relationships between the expression of YTHDF2 protein and the clinicopathological features and prognosis of the ESCC patients were analyzed.The Eca109 and EC9706 cells cultured in vitro were divided into control group （transfected with si-NC） and si-YTHDF2 groups （transfected with si-YTHDF2#1 and si-YTHDF2#2）. The expressions of YTHDF2 protein in the cells in various groups were detected by Western blotting method. The proliferation abilities of cells in various groups were detected by CCK-8 method， the number of colony formation in various groups was detected by colony formation experiment， and the number of migration cells in various groups were detected by Transwell chamber assay.

The YTHDF2 protein mainly expressed in the cytoplasm of ESCC cells and a small amount of YTHDF2 protein expressed in the nucleus.The expression amount of YTHDF2 protein in ESCC tissue was significantly higher than that in paracancerous normal tissue （P<0.01）. There were significant differences in the expression intensities of YTHDF2 protein in the ESCC patients with different ages of onset and TNM stages （P=0.008，P=0.041）. The Kaplan-Meier survival analysis showed that compared with YTHDF2 low expression group，the survival time of the patients in YTHDF2 high expression group was significantly shortened（P<0.05）. The CCK-8 method and colony formation experiment results showed that the proliferation activities and the number of colony formation of esophageal carcinoma cells in si-YTHDF2 transfection groups were significantly lower than those in control group （P<0.01）. The Transwell chamber assay results showed that the number of migration esophageal carcinoma cells in si-YTHDF2 transfection groups was significantly lower than that in control group （P<0.01）.

The expression of YTHDF2 in ESCC tissue is significantly higher than that in the normal tissue. YTHDF2 knockdown contributes to inhibiting the proliferation， colony formation and migration of esophageal carcinoma cells.

To observe the inhibitory effect of valproic acid （VPA）， a histone deacetylase inhibitor （HDACi）， on the proliferation of the human triple negative breast cancer （TNBC）MDA-MB-231 cells alone or combined with X-ray irradiation， and to analyze the effect of combined irradiation on the apoptosis and autophage of the MDA-MB-231 cells.

The MDA-MB-231 cells at logarithmic growth phase were divided into control group， different concentrations（2.5，5.0，10.0，20.0， and 40.0 mmol·L^－1） of VPA groups 4 Gy， X-ray irradiation groups，and different concentrations （2.5，5.0，10.0，20.0， and 40.0 mmol·L^－1） of VPA combined with 4 Gy X-ray irradiation groups. The proliferation rates of the cells in various groups were detected by CCK-8 method after treated for 24， 48， and 72 h.The cells were divided into control group， 4 Gy X-ray irradiation group， 5 mmol·L^－1 VPA group， 5 mmol·L^－1 VPA combined with 4 Gy X-ray irradiation group， the apoptotic rates and autophage rates of the cells in various groups were analyzed by fiow cytometry after treated for 24 and 48 h.

Compared with control group， the proliferation rates of the MDA-MB-231 cells in VPA group and VPA combined with 4 Gy X-ray irradiation groups were decreased significantly in a concentration and time-dependent manner （P<0.01）. Compared with 4 Gy X-ray irradiation group， the proliferation rates of the MDA-MB-231cells in VPA combined with 4 Gy X-ray irradiation groups were significantly decreased （P<0.01）. Compared with 2.5 mmol·L^－1 VPA group， the proliferation rate of the MDA-MB-231 cells in 2.5 mmol·L^－1 VPA combined with 4 Gy X-ray irradiation group after treated for 24 h was decreased（P<0.05）. After treated for 72 h， the proliferation rates of the MDA-MB-231 cells in 2.5 and 10.0 mmol·L^－1 VPA combined with 4 Gy X-ray irradiation groups were lower than those in 2.5 and 10.0 mmol·L^－1 VPA groups（P<0.05），respectively. After treated for 24 and 48 h， the apoptotic rates and autophagy rates of the MDA-MB-231 cells in 4 Gy X-ray group， 5 mmol·L^－1 VPA group and 5 mmol·L^－1 VPA combined with 4 Gy X-ray group were significantly higher than those in control group （P<0.05 or P<0.01）. Compared with 4 Gy X-ray irradiation group， the apoptotic rate and autophagy rate of the MDA-MB-231 cells in 5 mmol·L^－1 VPA combined with 4 Gy X-ray irradiation group were significantly increased （P<0.01）. After treated for 48 h，compared with 5 mmol·L^－1 VPA group，the autophagy rate of the MDA-MB-231 cells in 5 mmol·L^－1 VPA combined with 4 Gy X-ray irradiation group was increased （P<0.05）.

VPA can inhibit the proliferation of MDA-MB-231 cells in a concentration and time-dependent manner， and the combined inhibitory effect of VPA and X-ray is better， and the mechanism may be involved in the increasing of apoptosis and autophage of the MDA-MB-231 cells induced by the combination of VPA and X-ray.

To explore the anti-fatigue effect of ginseng adventitious root protein （GARP） in the mice and to clarify its mechanism， and provide the experimental basis for the development and utilization of ginseng adventitious root resources.

Forty kunming mice of clean grade were used to establish fatigue animal models by the weight-bearing swimming test. They were randomly divided into control group， low， medium and high doses （0.25， 0.50 and 1.00 g·kg^－1） of GARP groups， and there were 10 mice in each group. The swimming time of the mice in various groups was recorded through the exhaustive swimming experiment. The mice were killed after swimming experiment. The spectrophotometric method was used to detect the blood lactic acid （BLA） and blood urea nitrogen （BUN） levels in the serum of the mice in various groups. The spectrophometric method was used to detect the glutathione （GSH） levels， the superoxide dismutase （SOD） activities and the liver glycogen levels in liver tissue of the mice in various groups， and the spectrophometric method was used to detect the muscle glycogen level in the nuscle tissue of the mice in various groups. The C2C12 myoblasts were divided into control group and different doses （5， 10 and 20 mg·L^－1） of GARP groups.The spectrophotometric method was used to detect the GSH levels， SOD activities， and the glycogen levels in the cells in various groups， and the glucose uptakes of the cells in various groups were determined with phenol sulphuric acid process.The phosphorylation levels of adenosine 5'-monophosphate （AMP）-activated protein kinase （AMPK） （p-AMPK/AMPK） and the expression level of glucose transporter 4 （GLUT4） protein were detected by Western blotting method.

Compared with control group， the swimming time of mice in medium and high doses of GARP groups was increased （P<0.05 or P<0.01）； the BLA and BUN levels in the serum of the mice were reduced （P<0.05 or P<0.01）， the GSH levels and SOD activities in different doses of GAPP groups were increased （P<0.05 or P<0.01）； the liver glycogen and muscle glycogen levels were increased （P<0.05 or P<0.01）. Compared with control group the GSH levels，the SOD activities and the glycogen levels in the C2C12 myoblasts in 5，10 and 20 μg·mL^－1 GARP groups were increased （P<0.05 or P<0.01）， the glucose uptakes were increased （P<0.05 or P<0.01），and the ratios of p-AMPK AMPK and the expression levels of GLUT4 protein were increased （P<0.05 or P<0.01）.

GARP has anti-fatigue effect，and its mechanism may be achieved by activating AMPK/GLUT4 signaling pathway to promote glucose uptake.

To investigate the effect of p38 mitogen-activated protein kinase （p38 MAPK） inhibitor SB303580 （SB） on the progression of chronic obstructive pulmonary disease （COPD） in the mice， and to elucidate its possible mechanism.

A total of 48 C57BL/6 mice were divided into control group，SB group， COPD group， and COPD+SB group， and there were 12 mice in each group. The mice in COPD group and COPD+SB group were given cigarette smoke and lipopolysaccharide （LPS） to establish the COPD models. The airway resistance after inhalation of methacholine （Mch） was observed to evaluate the lung function of the mice in various groups， and the pathomophology of lung tissue of the mice in various groups was observed by HE staining. The total number of white blood cells， neutrophils， macrophages and lymphocytes， the levels of tumor necrosis factor-α （TNF-α）， interleukin-6 （IL-6）， interleukin-1β （IL-1β）， and interleukin-18 （IL-18） in pulmonary alveolar lavage fluid （BALF） of the mice in various groups were detected by cell counting and ELISA method.The mouse macrophage RAW264.7 cells were stimulated by cigaritte smoke extract （CSE） in vitro， and the cells were divided into control group， SB group， CSE group and SB+CSE group. Cell immunofluorescence staining was used to observe the expressions of NOD-like receptor protein 3（NLRP3） and cleaved caspase-3 in the macrophages in various groups， and flow cytometry was used to detect the pyroptotic rates and early apoptotic rates of the cells in various groups，and Western blotting method was used to detect the expression levels of phosphorylated p-38（p-p-38），pyroptosis-related and nuclear factor κB（NF-κB） signaling pathway related poteins in the macrophages in various groups.

The COPD mouse models were successfully established.Compared with control group， the airway resistance of the mice in COPD group and COPD+SB group， the total number of white blood cells， neutrophils， macrophages and lymphocytes in BALF， the levels of TNF-α， IL-6， IL-1β ，IL-18 in BALF， and the expression levels of p-p-38， NLRP3，cysteine protease 1（caspase-1），apoptosis-associated spotted protein（ASC），NF-κB p65，and Toll-like receptor 4 （TLR4） proteins in lung tissue of the mice were increased （P<0.05 or P<0.01）；the lung tissue was obviously damaged， the airway epithelial cells were exfoliated， and some adjacent alveoli fused to bullae.Compared with control group， the airway resistance of the mice in SB group， the total number of white blood cells， neutrophils， macrophages and lymphocytes in BALF， the levels of TNF-α， IL-6， IL-1β ，and IL-18 in BALF， and the expression levels of NLRP3， caspase-1， ASC， NF-κB p65， and TLR4 proteins in lung tissue had no significant differences（P>0.05）； the lung tissue was normal， but the expression level of p-p-38 in lung tissue was significantly decreased （P<0.05）. Compared with COPD group， the airway resistance of the mice in COPD+SB group， the total number of white blood cells， neutrophils， macrophages and lymphocytes in BALF， the levels of TNF-α， IL-6， IL-1β， and IL-18 in BALF， and the expression levels of NLRP3， caspase-1， ASC， NF-κB p65，and TLR4 proteins were decreased（P<0.05）；the pathological injury of lung tissue of the mice in COPD+SB group was alleviated（P<0.05）. In vitro experiments， compared with control group， the expression amounts of NLRP3 and cleaved caspase-3， pyroptotic rates and early apoptotic rates of the cells， the expression levels of p-p-38， ASC， NF-κB p65， TLR4， and cleaved caspas-3 proteins in the macrophages in CSE group and CSE+SB group were increased （P<0.05 or P<0.01）；the pyroptotic rate and early apoptotic rate of the cells， and expression levels of NLRP3， cleaved caspase-3， ASC， NF-κB p65， TLR4， and cleaved caspase-3 proteins in the macrophages in SB group had no significant differences（P>0.05）， and the expression level of p-p-38 in SB group was decreased （P<0.05）. Compared with CSE group， the early apoptotic rate of the macrophages and the expression level of cleaved caspase-3 in the cells in CSE+SB group were increased （P<0.05），and the pyroptotic rate， the expression levels of p-p-38， NLRP3， ASC， NF-κB p65，TLR4， and cleaved caspase-3 proteins were decreased （P<0.05）.

SB may improve the lung injury and inflammatory response of COPD by inhibiting the pyroptosis of macrophages mediated by the NLRP3 pathway.

To explore the effect of palmitic acid （PA） on the lipid deposition in the skeletal muscle cells，and to discuss the optimum action concentration and time.

The C2C12 cells were divided into control group and different concentrations （50，75，100，and 125 μmol·L^－1） of PA groups when the C2C12 cells grew to 70%－80%. The C2C12 cells in different concentrations of PA groups were given growth media containing different concentrations of PA， while the C2C12 cells in control group were not treated with PA（0 μmol·L^－1 PA）.CCK-8 method was used to detect the cell proliferation activities of cells in various groups，the triglyceride （TG） levels in the C2C12 cells in various groups were detected， Oil Red O staining was used to observe the formation of lipid droplets in the C2C12 cells in various groups， and 2% horse serum was used to induce the cell differentiation and evaluate the differentiation of C2C12 cells in various groups.

Compared with control group， there were no significant differences in the proliferation activities of the C2C12 cells in 50， 75， and 100 μmol·L^－1 PA groups（P>0.05）， the proliferation activity of the C2C12 cells in 125 μmol·L^－1 PA group was decreased（P<0.01），the TG levels in the C2C12 cells in 50， 75 and 100 μmol·L^－1 PA groups were increased （P<0.01）， and the TG level in the C2C12 cells in 100 μmol·L^－1 PA group was increased significantly （P<0.01）.Compared with cultured for 24 h， the TG level in the C2C12 cells after treated with 100 μmol·L^－1 PA for 48 h was increased （P<0.01）. After differentiation and culture， more myotubes were formed in the C2C12 cells in PA groups and control group.

The lipid deposition in the C2C12 cells could be effectively promoted after cultured with 100 μmol·L^－1 PA for 48 h.

To discuss the effect of different concentrations of peiminine （PMI） on the apoptosis of the human lung cancer A549 cells， and to clarify its possible molecular mechanism.

The A549 cells were treated with different concentrations （0.025， 0.050， 0.100， 0.200 and 0.400 mmol·L^－1） of PMI for 24， 48 and 72 h，respectively. The proliferation activity of the A549 cells was detected by MTT method， and the half inhibitory concentration （IC₅₀） was calculated.The A549 cells were treated with different concentrations （0.050，0.100，0.200 mmol·L^－1）of PMI or 0.200 mmol·L^－1 PMI combined with p38 MAPK inhibitor SB203580 for 48 h. The cells were divided into blank control group， different concentrations（0.050， 0.100 and 0.200 mmol·L^－1） of PMI groups and combination group （0.200 mmol·L^－1 PMI + 20 μmol·L^－1 SB203580）.The apoptotic rates of the A549 cells in various groups were detected by Annexin Ⅴ-FITC/PI method； the expression levels of apoptosis-related proteins B-cell lymphoma-2 （Bcl-2），Bcl-2 associated X protein（Bax），and cleaved-caspase-3 and mitogen-activated protein kinases （p38MAPK）/p53 signaling pathway related proteins phosphorylaed p38MAPK （p-p38 MAPK） （Thr180/Thr182）， p-p53 （ser15）， p53， PUMA， and murine double minute 2（MDM2） in the A549 cells in various groups were detected by Western blotting method the localization of p53 protein of the A549 cells in various groups was observed by immunofluorescence assay.

Compared with control group， the survival rates of the A549 cells in 0.10 and 0.20 mmol·L^－1 PMI groups were decreased （P<0.05）， the apoptotic rates were increased （P<0.05）， and the expression levels of Bax， cleaved-caspase-3， p-p38 MAPK （Thr180/Thr182）， p-p53 （ser15）， p53 and PUMA proteins were increased （P<0.05），and the expression levels of Bcl-2 and MDM2 proteins were decreased （P<0.05）.Compared with 0.20 mmol·L^－1 PMI group， the survival rate of the A549 cells in combination group was increased （P<0.05）， the apoptotic rate was decreased （P<0.05）， the expression levels of Bax， cleaved-caspase-3， p-p38MAPK （Thr180/Thr182）， p-p53 （ser15）， p53， and PUMA proteins were decreased （P<0.05）， while the expression levels of Bcl-2 and MDM2 proteins were increased （P<0.05）.The immunofluorescence results showed that the phenomenon of p53 protein transferring from cytoplasm to nucleus could be seen， and the p53 protein was expressed in both the cytoplasm and nucleus.

Peiminine can induce the apoptosis of the A549 cells， and its molecular mechanism may be related to the p38 MAPK/ P53 signaling pathway mediated apoptosis.

To explore the effects of histone deacetylase inhibitor （HDACi） sodium butyrate （NABT） alone or combined with X-ray irradiation on the apoptosis of human non-small cell lung cancer A549 cells， and to clarify its mechanism.

The A549 cells in logarithmic growth phase were divided into control group， NaBt group， 4 Gy X-ray irradiation group and NaBt combined with 4 Gy X-ray irradiation group. Flow cytometry was used to detect the apoptotic rates at 24 and 48 h after treatment. After 6 h of irradiation， the mitochondrial membrane potential （Δψm） and ROS （reative oxygen species） levels in the cells of various groups were analyzed by flow cytometry. After 24 h of irradiation， the expression levels of caspase-3 and p21 mRNA in the cells in various groups were detected by real-time quantitative PCR （RT-qPCR）.

After 4 Gy irriadiation for 24 and 48 h， compared with control group， 4 Gy X-ray irradiation group and NaBt group， the apoptotic rate of cells in NaBt combined with 4 Gy X-ray irradiation group was significantly increased （P<0.05 or P<0.01）. Compared with control group， the apoptotic rate of cells in NaBt group was significantly increased after 48 h of irradiation （P<0.01）. After treated with NaBt alone or combined with 4 Gy X-ray for 6 h， compared with control group， 4 Gy X-ray irradiation and NaBt group， the Δψm of A549 cells in NaBt combined with 4 Gy X-ray irradition group was decreased （P<0.05）. Compared with control group， the ROS levels in NaBt group and NaBt combined with 4 Gy X-ray irradiation group were increased （P<0.05）. Compared with 4 Gy X-ray irradiation group， the ROS level in NaBt combined with 4 Gy X-ray irradiation group was increased （P<0.05）. After treated with NaBt alone or combined with 4 Gy X-ray irradiation for 24 h， the mRNA levels of caspase-3 and p21 in the cells in NaBt combined with 4 Gy X-ray irradiation group were significantly higher than those in control group， 4 Gy X-ray irriadiation group and NaBt group （P<0.01）. Compared with control group， the p21 mRNA levels in NaBt group and 4 Gy X-ray irriadiation group were significantly increased （P<0.01）.

NaBt can induce the apoptosis of A549 cells， and its mechanisms may be related to the regulation of mitochondrial apoptosis pathway.

To explore the effects of tumor-associated macrophages （TAM） polarization state on the self-renewal ability and vasculogenic mimicry of prostate cancer （Pca） stem cells， and to clarify the effect of TAM in the development of PCa.

The PCa stem cells were selected from human PCa cell line DU145， and the human mononuclear leukemia cell strain THP-1 was induced into the M2 TAM， which was used as induction group， and the uninduced THP-1 cells were used as control group；real-time fluorescence quantitative PCR（RT-qPCR） method was used to detect the expression levels of inducible nitric oxide synthase （iNOS） mRNA and recombinant human arginase-1 （Arg-1） mRNA in the cells. The experiment was divided into PCa-cancer stem cells（CSC） group （PCa stem cells were cultured in the conventional culture medium）， THP-1+PCa-CSC group （PCa stem cells were cultured in the culture supernatant of uninduced THP-1 cells） and TAM+PCa-CSC group （PCa stem cells were cultured in TAM-conditioned medium of induced THP-1 cells）； a co-culture system of TAM and PCa stem cells was established by simulating the microenvironment.After 48 h of culture， the spheroidization of PCa stem cells in various groups was detected by cell spheroidization experiment， the number of cell colonies of PCa stem cells in various groups was detected by cell clone formation experiment，cell scratch test was used detect the scratch healing rates of cells in various groups， and Transwell chamber test was used to detect the number of invasion PCa stem cells in various groups； vasculogenic mimicry formation experiment was used to detect the number of tubular structures of PCa stem cells in various groups.

The human PCa DU145 cells were sorted to obtain the CD44+CD133+ labeled prostate cancer stem cells. Compared with control group， the THP-1 cells in induction group showed irregular polygonal shapes， the expression level of iNOS mRNA was decreased （t=21.021， P<0.05）， the expression level of Arg-1 mRNA was increased （t=26.153， P<0.05）， the expression level of CD86 protein was decreased（t=34.556，P<0.05），and the expression level of CD206 protein was increased（t=31.095，P<0.05）. Compared with PCa-CSC group， the volumes of cell spheroids in THP-1+Pca-CSC group and TAM+Pca-CSC group were larger， the number of cell colonies was increased（P<0.05）， the scratch healing rates were increased（P<0.05）， the number of invasion cells was increased（P<0.05）， and the number of tubular structures of cells was also increased（P<0.05）. Compared with THP-1+PCa-CSC group， the volume of the cell spheroids in TAM+PCa-CSC group was further increased（P<0.05）， the number of cell colonies and the number of invasion cells were increased（P<0.05）， the scratch healing rate was increased（P<0.05），and the number of cell tubular structures was increased（P<0.05）.

Regulating the polarization state of TAM can induce the metastasis of PCa stem cells， and further promote their self-renewal and vasculogenic mimicry formation.

To investigate the effect of Tongguan Xiaozheng Decoction （TGXZ） in the rats with tubal inflammatory infertility， and to explore its possible molecular mechanism.

A total of 92 SD female rats aged 6－8 weeks were selected， and the rat models of tubal inflammatory infertility were constructed by vaginal inoculation with mixed bacteria. The 60 model rats were randomly divided into model group， western medicine （metronidazole tablets and levofloxacin hydrochloride capsules） group and low （5.33 g·kg^－1）， medium （10.67 g·kg^－1）， and high （21.33 g·kg^－1） doses of TGXZ groups， with 12 rats in each group， and another 12 normal rats were taken as normal control group. All rats were intragastrically administered twice a day， once in the morning and evening， for a total of 1 dose， for 30 d. The mental state， activity levels， water consumptions， paw color， etc. of the rats in various groups were observed and recorded； ELISA method was used to measure the levels of serum interleukin-1β （IL-1β）， interleukin-6 （IL-6）， interleukin-10 （IL-10）， tumor necrosis factor-α （TNF-α） of the rats in various groups； Hematoxylin eosin （HE） staining was used to observe the pathomorphology of fallopian tube tissue of the rats in various groups； Western blotting method was used to detect the expression levels of Toll-like receptor 4/nuclear factor-kB （TLR4/NF-κB） signaling pathway related proteins in the fallopian tube tissue of the rats in various groups； the conception rates of rats in various groups were calculated.

The rats in normal control group had good mental state， normal activity and drinking water， and white claw color. The mental state of the rats in model group was depressed， the amount of activity and drinking water were decreased， and the claw color was dark red. The general situation of rats in medium dose of TGXZ group， high dose of TGXZ group and Western medicine group was significantly improved， the mental state of rats recovered， the amounts of activity and drinking water were increased， and the color of claw became lighter. Compared with normal control group， the serum IL-1β， IL-6， IL-10， and TNF-α levels of the rats in model group were significantly increased （P<0.05）， and the inflammatory cell infiltration score of fallopian tube tissue was significantly increased （P<0.05）， the expression levels of TLR4， p-IκBα and NF-κB p65 proteins in fallopian tube tissue were significantly increased （P<0.05）， the IκBα protein expression level was significantly reduced （P<0.05）， and the conception rate of rats was significantly reduced （P<0.05）. Compared with model group， the levels of serum IL-1β， IL-6， IL-10， and TNF-α in medium and high doses of TGXZ groups were significantly reduced （P<0.05）， and the inflammatory cell infiltration scores of fallopian tube tissue were significantly reduced （P<0.05）， the expression levels of TLR4， p-IκBα and NF-κB p65 in fallopian tube tissue were significantly decreased （P<0.05）， the IκBα protein expression level was significantly increased （P<0.05）， and the conception rate was significantly increased （P<0.05）； but there were no significant differences in the above indicators in low dose of TGXZ group （P>0.05）. Compared with low dose of TGXZ group， the serum IL-1β， IL-6， IL-10， and TNF-α levels of the rats in medium and high dose of TGXZ groups， the inflammatory cell infiltration scores of fallopian tube tissue， the expression levels of TLR4， p-IκBα and NF-κB p65 proteins in fallopian tube tissue were decreased significantly （P<0.05），and the conception rates and the IκBα expression levels were increased significantly （P<0.05）.

TGXZ can effectively reduce the levels of serum inflammatory factors in the rats with tubal inflammatory infertility， and its mechanism may be related to the inhibition of the activity of TLR4/NF-κB signaling pathway.

To construct the lentiviral vector of lymphocyte activation gene 3 （LAG3）-mCherry red fluorescent protein （LAG3-mCherry） and establish the cell line stably expressed LAG3-mCherry after transfection of the HEK293T cells， and to investigate the cellular location of LAG3-mCherry fusion protein in the HEK293T cells.

The LAG3 plasmid and lentiviral vector including mCherry gene were digested by endonucleases EcoRⅠ and NotⅠ. The insert and vector fragments were extracted and ligated to construct the pEZ-LAG3-mCherry plasmid. The pEZ-LAG3-mCherry plasmid was sequenced and transfected into the HEK293T cells using Lipofectamine 3000. The expression location of LAG3-mCherry in the HEK293T cells was observed with fluorescence microscope and the expression of LAG3-mCherry fusion proteins in the cells were detected by Western blotting method.

The results of double digestion of recombinant plasmid showed that two DNA bands of about 8 012 and 2 066 bp were found in gel electrophoresis， which were consistent with the size of pEZ-Lv-mCherry vector and LAG3 gene fragment. The DNA sequencing results showed that the LAG-3 gene was successfully inserted into the lentiviral expression vector. The fluorescence microscope results demonstrated that most of LAG3-mCherry proteins were located on the cell membrane of HEK293T cells， while a few fusion proteins were found in the cytoplasm area. The results of Western blotting method showed a specific protein band of LAG3 in the HEK293T cells transfected with pEZ-LAG3-mCherry plasmid.

The pEZ-LAG3-mCherry plasmid including LAG3-mCherry DNA sequence is successfully constructed and the cell line stably expressing LAG3-mCherry is established. The LAG3-mCherry fusion protein is mainly distributed on the cell membrane of HEK293T cells transfected with pEZ-LAG3-mCherry plasmid.

To explore the effect of sufentanil on the apoptosis of myocardial cells and the long non-coding RNA-MALAT1 （LncRNA-MALAT1） targeted extracellular signal-regulated kinase 1/2 （ERK1/2） signaling pathway in the myocardial ischemia-reperfusion injury（MIRI） rats， and to clarify the possible mechanism of sufentanil in the MIRI rats.

Thirty male SD rats were randomly divided into sham operation group， model group and sufentanil group， with 10 rats in each group. The MIRI model was prepared by ligation of the anterior descending branch of the left coronary artery （LAD）. In sham operation group， only thorax of the rats was opened without ligation.The rats in sufentanil group were injected with sufentanil 1 μg?kg^－1 through the tail veins 10 min before reperfusion，and the rats in sham operation group and model group were injected with the same amount of normal saline through the tail veins.The expression levels of LncRNA-MALAT1 in myocardium tissue of the rats in various groups were detected by real-time fluorescence quantitative PCR （RT-qPCR） and fluorescence in situ hybridization （FISH） methods； the apoptotic rates of cardiomyocytes of the rats in various groups were determined by TUNEL method； the expression levels of cysteine protease protein-3 （caspase-3），cleaved cysteine protease protein-3 （cleaved-caspase-3），B-cell lymphoma-2 （Bcl-2）， Bcl-2 associated X protein（Bax），ERK1/2 and phosphorylated ERK1/2 （p-ERK1/2） proteins in myocardium tissue of the rats in various groups were detected by Western blotting method. The hypoxia-reoxygenation model of cardiomyocytes was constructed to simulate the MIRI in vivo， and the experiment was divided into control group， model group， sufentanil group， pcDNA-MALAT1 group （MALAT1 group）， sufentanil+pcDNA-MALAT1 group （sufentanil+MALAT1 group）. The expression levels of caspase-3， cleaved-caspase-3， Bax，Bcl-2， ERK1/2 and p-ERK1/2 in the H9C2 cells were detected by Western blotting method.

The animal experiment results showed that compared with sham operation group， the LncRNA-MALAT1 expression level in myocardium tissue of the rats in model group， the apoptotic rate of cardiomyocytes， the expression levels of cleaved-caspase-3， and the ratio of Bax/Bcl-2 were increased significantly （P<0.05）； the expression levels of p-ERK1/2 proteins were decreased significantly （P<0.05）； compared with model group， the LncRNA-MALAT1 expression level in the myocardium tissue of the rats in sufentanil group， the apoptotic rate of cardiomyocytes， the expression level cleaved-caspase-3，and the ratio of Bax/Bcl-2 were decreased significantly （P<0.05）； the expression levels of p-ERK1/2 proteins were increased significantly （P<0.05）. The cell experiment results showed that compared with control group， the expression level of cleaved-caspase-3 and the ratio of Bax/Bcl-2 in the HPC2 cells in model group were increased significantly （P<0.05）， and the expression levels of p-ERK1/2 proetins were decreased significantly （P<0.05）； compared with model group， the expression level of cleaved-caspase-3 and the ratio of Bax/Bcl-2 in the HPC2 cells in the sufentanil group were decreased significantly （P<0.05）， and the expression levels of p-ERK1/2 proteins were increased significantly （P<0.05）； compared with MALAT1 group， the expression level of cleaved-caspase-3 and the ratio of Bax/Bcl-2 in the HPC2 cells in sufentanil+MALAT1 group were decreased significantly （P<0.05），and the expression levels of p-ERK1/2 proteins were increased significantly （P<0.05）.

Sufentanil can attenuate the apoptosis of cardiomyocytes in the MIRI rats， and its mechanism may be related to the inhibition of LncRNA-MALAT1 to activate the ERK1/2 signaling pathway.

To construct an immundeficient mouse model with low density lipoprotein receptor （LDLR） gene knockout by CRISPR/cas9 technology and analyze and evaluate the blood cholesterol level，and to provide a new method for constructing a humanized mouse model of immune system with hyperlipidemia.

Using CRISPR/cas9 technology， sgRNA/cas9 mRNA effectively identifying exons 2 and 18 of LDLR gene was injected into the fertilized eggs of NOD SCID mice. The gene knockout F0 positive （LDLR^+/+， AA） mice were screened by genotyping of neonatal mice， and the mice were bred with NOD SCID （LDLR^+/+， AA） mice to identify the F1 generation （AA） with stable genotypes mice. The F1 positive heterozygous mice and NOD SCID mice were bred to obtain a large number of F2 generation （AA） mice with exactly the same gene sequence. Large scale breeding was carried out between F2 generations.The F3 generation mice were grouped according to genotype and gender， respectively： male AA， male Aa， female AA and female Aa.The body weights were monitored， and the peripheral blood was collected for blood cholesterol level detection.

The NOD SCID LDLR^+/－ （AA） mice were obtained by the above construction method. After 8 weeks of weight detection， it was found that Aa heterozygous genotype did not affect the body weight of mice during growth and development process. The cholesterol level of female Aa was （100.80±4.42） mg·dL^－1 and male Aa was （120.56±11.16） mg·dL^－1， compared with negative control （AA mice），the cholesterol levels were significantly increased（P<0.05）；the cholesterol levels of female AA and male AA were（60.78±2.11） and （75.43±10.06） mg·dL^－1，and the difference between them was statistically significant （P<0.05）.

Without affecting the body weight of mice， a mouse model with spontaneous increase of cholesterol level is successfully constructed by using CRISPR/cas9 technology under the background of NOD SCID， which can be used as the basis for humanization.