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Objective To investigate the effect and mechanism of colchicine on mouse liver cancerviaHippo signaling pathway. Methods The 6-week-old male C57BL/6J mice were randomly divided into 3 groups: diethylnitrosamine(DEN)/carbon tetrachloride(CCl4)/ethanol(C2H5OH) induced mouse liver cancer model and colchicine(0.1 mg/kg) intervention were established in control group, model group and colchicine group. From week 1st to week 2nd, the model group and the colchicine group were intraperitoneally injected with 1.0% DEN once a week. From week 3rd to week 7th, 20% CCl4dissolved in olive oil solution(5 ml/kg) was intragastric administration twice a week. From week 8th to week 18th, 20% CCl4dissolved in olive oil solution(6 ml/kg) was intragastric twice a week. The colchicine group was given continuous intragastric administration for 20 weeks. The control group was given the corresponding solvent. Liver index, alanine aminotransferase(ALT) and aspartate aminotransferase(AST) serum biochemical indexes were detected. Western blot and immunofluorescence were used to detect the expression levels of MST1, pYAP, YAP, pTAZ and TAZ proteins in liver tissues of mice in each group. Results The liver surface of mice in the control group was smooth and soft, while the liver of mice in the model group was rough and hard with granular nodules. The above lesions were significantly improved in the colchicine group. HE staining showed that the liver lobular structure of mice in the control group was normal, while the liver lobular structure of mice in the model group was disordered, with a small amount of fat droplets, extensive tissue necrosis, inflammatory cell infiltration, and fat vacuoles. The degree of liver lesions of mice after colchicine intervention was significantly reduced. The results of immunofluorescence and Western blot showed that compared with the control group, the protein expression levels of pYAP and pTAZ in liver tissue of model group mice were significantly decreased, while the protein expression levels of MST1, YAP and TAZ increased. After colchicine intervention, the protein expression levels of MST1, pYAP and pTAZ were significantly up-regulated, while the protein expression levels of YAP and TAZ were down-regulated. Conclusion The therapeutic effect of colchicine on mouse liver cancer may be related to its activated Hippo signaling pathway.

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Objective To construct a stable synovial cell line MH7A from rheumatoid arthritis(RA) patients using lentiviral vectors that interfere with the expression of tumor necrosis factor receptor associated factor 2(TRAF2), and to study the role of TNF-α-TRAF2 signaling in MH7A abnormal proliferation. Methods Based on the design principles of human TRAF2 gene sequence and shRNA sequence, three pairs of TRAF2 shRNA interference sequences were designed and synthesized. The primers were annealed by PCR, and a linear vector was obtained by double enzyme digestion PLKO.1-puro. The linearized vector was connected to the annealed primers through Solution I, and the connected products were introduced into receptive cells. The plates were coated, and positive colonies were selected for sequencing. Three different recombinant plasmids of PLKO.1-TRAF2-shRNA lentivirus were constructed, and lentivirus packaging plasmids was used to package logarithmic growth phase HEK 293T cells. Virus solution was collected to infect MH7A cells. At the same time, puromycin was used to screen MH7A stable transgenic strains with low TRAF2 expression. CCK-8 method, Western blot, and qPCR were used to detect the proliferation function of MH7A induced by TNF-α and low expression of TRAF2, as well as downstream signal TRAF2, P65 protein expression and mRNA levels. Results PLKO.1-TRAF2-shRNA(1), PLKO.1-TRAF2-shRNA(2), and PLKO.1-TRAF2-shRNA(3) lentivirus vector plasmids and control group lentivirus vector plasmids PLKO.1-puro were successfully constructed. The three TRAF2-shRNA lentivirus vector plasmids and control group lentivirus vector plasmids PLKO.1-puro were respectively introduced into the lentivirus packaging plasmid of HEK 293T to obtain virus solution. After infecting MH7A cells with the virus solution, they were treated with puromycin(2.00 μ G/mL) screening and obtaining MH7A stable transgenic plants after 2 days. Through qPCR and Western blot results, it was found that the expression of TRAF2 mRNA and protein in PLKO.1-TRAF2-shRNA(1) MH7A stably transfected cells was significantly reduced compared to the negative control group. The results of CCK-8 and Western blot showed that after knocking down TRAF2 in MH7A, the proliferation of MH7A cells with low TRAF2 expression induced by TNF-α and the phosphorylation level of P65 were significantly reduced. Conclusion A stable transgenic strain of PLKO.1-TRAF2-shRNA(1) MH7A cells was successfully constructed to investigate the role of TNF-α-TRAF2 signal activation in mediating abnormal proliferation of RA synovial cells.

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Objective To investigate the effect of 4-amino-2-trifluoromethyl-phenyl retinate(ATPR) on acute liver injury induced by lipopolysaccharide(LPS) in C57BL/6 mice and its related mechanism. Methods Fifteen 6-week-old male C57BL/6 strain mice were randomly divided into normal group, model group and ATPR group, with 5 mice in each group. Mice in the ATPR group were intraperitoneally injected with ATPR(15 mg/kg·d), and normal group and model group were given solvent. After continuous administration for one week, model group and ATPR group were intraperitoneally injected with LPS(6 mg/kg), and all mice were sacrificed 6 hours later. The contents of Alanine aminotransferase(ALT) and Aspartate aminotransferase(AST) in serum of mice were detected. The mRNA levels of Interleukin-6(IL-6) and Tumor necrosis factor-alpha(TNF-α) were detected by qPCR. Hematoxylin-eosin(H&E) staining was used to observe the histopathological changes of liver in mice. The ultrastructural changes of mouse hepatocytes were observed by Transmission electron microscope(TEM). The expression levels of mitochondrial damage-related proteins FUNDC1 and OPA1 and autophagy related proteins LC3B, P62, Beclin1 and ATG5 were detected by Western blot. Results Compared with the normal group, the content of ALT and AST in serum and the mRNA levels of IL-6 and TNF-α in liver tissue increased in the model group, and the changes were reversed in the ATPR group. H&E staining showed that the hepatic lobule structure was normal in the normal group, the hepatic cords were arranged radially, there was no hyperemia and inflammatory cell infiltration, and the hepatocyte boundary was clear. In the model group, the intercellular space of liver was enlarged, the arrangement of hepatic cords was disordered, and inflammatory cells infiltrated. In the ATPR group, the intercellular space of liver and the structure of hepatic cords were restored, and the inflammatory cell infiltration was less. TEM showed that the damaged mitochondria and lipid droplet accumulation in the hepatocytes of mice in the model group were compared with that in the normal group, and the morphology and quantity of mitochondria and lipid droplet in the hepatocytes of mice in the ATPR group tended to be normal. Western blot showed that compared with the normal group, the expression of FUNDC1 protein in the liver tissues of mice in the model group increased, the expression of OPA1 protein decreased, the ratio of LC3BⅡ to LC3BⅠ decreased, the expression of P62 protein increased, the expression of Beclin1 and ATG5 protein decreased, and the above changes were reversed in the ATPR group. Conclusion ATPR alleviates acute liver injury induced by lipopolysaccharide in mice by promoting autophagy.

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Objective To study the effect of formononetin on the cell damage of glucose/oxygen deprivation/reoxygenation glyconeuronsviathe PARP1 signaling pathway, and to offer theoretical support for the use of Caragana isoflavones in the treatment of cerebral ischemia-reperfusion injury. Methods In mouse neurons(HT22), a model of Oxygen-glucose deprivation/reoxygenation(OGD/R) was created. Western blot was used to detect the expression of PARP1 and PARG in HT22 neurons at various time points of glucose-oxygen deprivation/reoxygenation, and the optimal time point of pathway modification was chosen. After OGD/R, HT22 cells were treated with formononetin, PARP1 inhibitor(PJ34), and PARG inhibitor, and six groups were developed: control group, control group+formononetin group, OGD/R group, OGD/R+ formononetin group, OGD/R+PJ34 group, OGD/R+PARG inhibitor group. HT22 cells were grown normally without OGD/R therapy in the control group. The expression levels of apoptotic factors and associated proteins in each group were determined using immunofluorescence and Western blot. Results PARP1 pathway was activated most obviously in HT22 cells after 3 hours of glucose and oxygen deprivation/reoxygenation. Under the condition of OGD/R 3 h, treatment with formononetin, PJ34 or PARG inhibitor could increase E3 ubiquitin ligase(Iduna), inhibit the expression of PARP1 and PARG pathway proteins, reduce the expression of AIF and P53, and increase the phosphorylation level of AKT protein. Conclusion Formononetin can block the PARP1/AIF/Akt signaling pathway by raising the expression of Iduna protein in the presence of OGD/R, hence decreasing the damage to HT22 mouse neurons.

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Objective To explore the effect of cannabinoid receptor 2(CB2) on orthodontic tooth movement(OTM) rate and periodontal tissue reconstruction of pressure area in mice. Methods Thirty CB2-/-male mice and thirty littermate control WT male mice were individually accepted the orthodontic appliance at their age of 6 weeks. The mice were respectively scarified at 3 days, 7 days, 14 days and 21 days after the operation. Then the tooth movement distance was examined through the stereomicroscope. Hematoxylin-eosin staining was performed to explore the biological responses of periodontium at the distal mesial root pressure area. Anti-tartrate acid phosphatase staining was performed to calculate the number and distribution of osteoclasts at the distal mesial root pressure area, and MMP-9 was evaluated by immunohistochemistry to examine the number of MMP-9(+) monocytes and multinucleated cells in the same district as the TRAP staining. Results Compared with those WT mice at 3,7,14 and 21 days, OTM distance showed a gradual increased tendency according with experimental time over 21 days. The widths of periodontal ligament on the pressure side were markedly greater in CB2-/-mice than WT mice at 7,14 and 21 days(P<0.000 1). The numbers of TRAP positive osteoclasts were significantly greater in CB2-/-mice than those in WT mice at 14 days of OTM(P<0.001). MMP-9 immunohistochemical staining showed that the number of MMP-9(+) monocytes and multinucleated cells was more in CB2-/-mice than that in WT mice at 14 days of OTM(P<0.05). Conclusion The absence of CB2 accelerates orthodontic tooth movement under orthodontic force. The absence of CB2 reinforces bone resorption in orthodontic tooth movement compressive area during orthodontic tooth movement.

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Objective To detect cell DNA damage more effectively by optimizing the influencing factors of Comet assay and establishing a positive control. Methods Based on the traditional alkaline comet assay, the gel preparation method, gel concentration, agarose solvent, lysis time, cell density and electrophoresis voltage were optimized. HL-60 cells were induced by hydrogen peroxide(H2O2) as a DNA damage inducer as a positive control. Results The optimized experimental results showed that the gel preparation method improved from three-step method to two-step method. PBS was used as agarose solvent, 0.8% normal melting point agarose was used to prepare the bottom layer, and 0.7% low melting point agarose was mixed with cell suspension as the second layer. The gel preparation effect was better, the operation was simpler, the time was saved, and the degumming problem was better solved. After lysis for 1 h and electrophoresis at 1 V/cm voltage, representative comet images were obtained. The results were easy to interpret and could effectively avoid false positive results. The positive control of comet assay was successfully established by comparing the DNA damage of HL-60 cells induced by different concentrations of H2O2. Conclusion Compared with the traditional method, the optimized comet assay method is simpler, faster, more accurate, lower cost and has better repeatability, which can quickly detect DNA damage in cells.

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Objective To investigate the effect of cinobufacini on inhibiting colorectal cancer metastasis by regulating the polarization of M2 macrophages. Methods THP-1 was induced into M0 type macrophages. The conditioned medium of HCT116 cells was collected to stimulate M0 type macrophages. The polarization of M2 type macrophages was observed by flow cytometry, real-time quantitative PCR and ELISA experiments. The conditioned medium of M0 type macrophages and HCT116-Mφ cells was collected to stimulate HCT116 cells. The ability of migration and invasion was observed by wound healing assay and Transwell assay. The effect of cinobufacini on the viability of HCT116 cells was detected by CCK-8 assay. The conditioned medium of HCT116 and HCT116+cinobufacini was collected to stimulate M0 type macrophages. The polarization of M2 type macrophages was observed by flow cytometry, real-time quantitative PCR and ELISA experiments. The conditioned media of HCT116-Mφ cells and(HCT116+cinobufacini)-Mφ cells were collected to stimulate HCT116 cells. The changes of migration and invasion ability were observed by wound healing assay and Transwell assay. Results After stimulation of M0 type macrophages in HCT116 cell conditioned medium, the morphology of M0 macrophages turned into fusiform cells, the proportion of CD11b+CD206+cells increased, and the expression of M2 macrophage markers IL-10 and TGF-β increased. The migration and invasion ability of HCT116 cells were significantly enhanced after stimulation in the conditioned medium of HCT116-Mφ cells. After the addition of cinobufacini, not only the polarization proportion of M2 macrophages decreased, but also the metastatic effect mediated by M2 macrophages was inhibited. Conclusion HCT116 cells can induce the polarization of M2 macrophages, while cinobufacini can inhibit the tumor metastasis mediated by M2 macrophages by inhibiting the polarization of M2 macrophages.

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Objective To investigate the application value of quantitative parameters MRFDGmaxand SUVmaxin the stages of hepatitis, liver fibrosis and cirrhosis in rats by whole-body dynamic18F-FDG PET/CT Patlak imaging. Methods Twenty-four SD rats were randomly divided into four groups of six rats each, which were the normal group, hepatitis group, liver fibrosis group and cirrhosis group. According to the experimental grouping, rats in each group were induced by the CCl4oil solution complex method. Whole-body dynamic18F-FDG PET/CT patlak imaging was performed on each group of rats separately at the completion of induction.After the imaging was completed, the MRFDGmax, SUVmaxand CT values of the livers of each group were analyzed; subsequently, the serum of rats in each group was extracted for the detection of liver function indexes(AST, ALT and ALP), and HE staining was performed on the livers of rats in the normal, hepatitis and cirrhosis groups, and Masson staining was performed on those in the liver fibrosis group; the α-SMA expression in the liver tissues of each group was analyzed by immunohistochemical method. The data were analyzed by one-way ANOVA, two independent samples t-test and Pearson correlation analysis. Results MRFDGmax, SUVmaxvalues were statistically significant differences among normal, hepatitis, liver fibrosis and cirrhosis groups(F=84.54, 38.35,P<0.001). The difference in CT values between liver fibrosis and cirrhosis groups was not statistically significant(t=-0.407,P=0.693), and the difference was statistically significant when compared between the rest of the groups(F=112.25,P<0.001). Compared with the normal group, AST, ALT and ALP of the experimental group showed a staged increase, and the differences were statistically significant(F=93.32, 64.63, 145.03,P<0.001). HE staining showed that hepatocytes of the normal group were neatly arranged and structurally intact; a large number of inflammatory cells infiltrated the hepatitis group with steatosis; pseudo lobe formation was observed in the cirrhosis group. Masson staining of the liver fibrosis group showed collagen fiber proliferation and thickening of the peritoneum. Immunohistochemistry test results showed that α-SMA expression increased in hepatitis group, liver fibrosis group and cirrhosis group, with a staged increase, and the difference was statistically significant(F=80.57,P<0.001). Correlation analysis showed a positive correlation between SUVmaxand MRFDGmax(r=0.967,P<0.01). α-SMA was positively correlated with AST, ALT and ALP in the hepatitis, liver fibrosis and cirrhosis groups, respectively(r=0.924, 0.756, 0.934,P<0.01). Conclusion Whole-body dynamic18F-FDG PET/CT Patlak imaging has application value in monitoring hepatitis, liver fibrosis and cirrhosis stages through quantitative parameters MRFDGmaxand SUVmaxchanges.

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Objective To explore the causal association between gut microbes and non-alcoholic fatty liver disease(NAFLD) by Mendelian randomisation analysis. Methods Genetic instrumental variables for gut microbiota were identified from a gene-wide association study of 18 340 participants, and summary statistics for NAFLD were obtained from the FinnGen database, which provided data on 894 NAFLD cases and 217 898 controls using the IVW method as the primary analysis. In order to test the robustness of the results, MR-Egger method, WM method, Simple Mode method, Weighted Mode method were used for Mendelian randomisation analysis, and heterogeneity test, sensitivity analysis, and multiplicity analysis were performed. Results class Gammaproteobacteria IVW results showed(OR=0.621, 95%CI=0.412～0.934,P=0.022); family Enterobacteriaceae IVW results showed(OR=1.481, 95%CI=1.069～2.053,P=0.018);genus Lachnospiraceae IVW results showed(OR=1.405, 95%CI=1.036～1.904,P=0.029); genus Prevotella7 IVW results showed(OR=0.834, 95%CI=0.714～0.974,P=0.021); genus Prevotella9 IVW results showed(OR=1.251, 95%CI=1.025～1.527,P=0.027); order Desulfovibrionales IVW results showed(OR=0.714, 95%CI=0.519～0.982,P=0.038); order Enterobacteriales IVW results showed(OR=1.481, 95%CI=1.069～2.053,P=0.018). And there was no heterogeneity in the heterogeneity test, and the sensitivity analyses all showed robustness and no pleiotropy was found. Conclusion This study implicates class Gammaproteobacteria, family Enterobacteriaceae, genus Lachnospiraceae, genus Prevotella7, genus Prevotella9, order Desulfovibrionales, order Enterobacteriales seven species of gut microorganisms have a causal relationship with NAFLD.

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Objective To observe the effect of different synovial cell secretions on chondrocytes after LPS-induced inflammation, and to explore the mechanism of two synovial cell secretions causing cartilage damage in the progression of KOA disease. Methods Two kinds of synovial cells were co-cultured at 1∶4 and LPS-induced inflammation. The supernatant and exocrine were extracted, and then the normal and LPS-induced inflammation were extracted. The human cartilage tissue obtained during the operation was isolated and cultured into chondrocytes, which were divided into five groups: the first group was added with FLS secretion, the second group was added with normal FLS secretion, the third group was added with secretion after co-culture of two kinds of synovial cells, the fourth group was added with inflammatory MLS secretion, and the fifth group was added with inflammatory FLS secretion. CCK-8 was used to detect the viability of chondrocytes in each group. TNF-α,IL-1β,IL-6 level in the supernatant of chondrocytes in each group was detected by ELISA. The protein expression of TLR4,NF-κB,IkK,IκB,ADAMTS5 in chondrocytes of each group was detected by Western blot method.Results CCK-8 showed that the activity of chondrocytes in the three groups of inflammatory secretions decreased compared with the secretions from normal synovial cells(P<0.05); ELISA showed TNF-α,IL-1β,IL-6 level in the supernatant of group Ⅲ, IV and V was higher than that of group Ⅰ and Ⅱ(P<0.05), TNF-α,IL-1β,IL-6 level in group Ⅲ was higher than that in group IV but lower than that in group V(P<0.05). Western blot showed the protein expression of TLR4,NF-κB,IkK,IκB,ADAMTS5 in chondrocytes of group Ⅲ, Ⅳ and Ⅴ was higher than that in group Ⅰ and Ⅱ(P<0. 05),the protein expression of TLR4,NF-κB,IkK,IκB,ADAMTS5 in group Ⅲ was higher than that in group Ⅳbut lower than that in group Ⅴ(P<0.05).Conclusion Two kinds of synovial cell-derived secretions after LPSinduced inflammation can regulate cartilage TLRs/NF-κB signal pathway, causing cartilage inflammation. The inflammatory effect of MLS secretion is stronger than that of FLS secretion, but the inflammatory effect of MLS secretion under two co-cultures is weaker than that of MLS secretion alone.

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Objective To study the effect of high mobility group box B1(HMGB1) gene knockout on alleviating acute lung injury and inhibiting toll-like receptor 4(TLR4)/nuclear factor-κB(NF-κB) pathway of sepsis mice. Methods Wild-type(WT) mice were divided into WT-Sham group and WT-model group, and HMGB1 knockout(KO) mice were divided into KO-sham group and KO-model group. Sepsis ALI model was established by cecal ligation and perforation in WT-model group and KO-model group. Sham operation was performed in WT-Sham group and KO-Sham group. 24 h after modeling, the partial pressure of arterial oxygen(PaO2) was detected, oxygenation index(OI) was calculated, pathological changes of lung tissue were detected and lung injury score was calculated, the concentrations of tumor necrosis factor-α(TNF-α), interleukin-1β(IL-1β), interleukin-6(IL-6), reactive oxygen species(ROS), malondialdehyde(MDA), superoxide dismutase(SOD), in serum and lung tissues and the expression of HMGB1, TLR4 and nuclear NF-κB in lung tissues were detected. Results The PaO2, OI and the concentration of SOD in serum and lung tissue of WT-model group were lower than those of WT-Sham group, the lung injury scores, the concentrations of TNF-α, IL-1β, IL-6, ROS and MDA in serum and lung tissue, and the expression levels of HMGB1, TLR4 and nuclear NF-κB in lung tissue were higher than those in WT-Sham group(P<0.05). HMGB1 was not expressed in lung tissue of KO-model group, and the concentrations of PaO2, OI and the concentration of SOD in serum and lung tissue of KO-model group were higher than those of WT-model group, the lung injury scores, the concentrations of TNF-α, IL-1β, IL-6, ROS and MDA in serum and lung tissue, and the expression levels of TLR4 and nuclear NF-κB in lung tissue were lower than those of the WT-model group(P<0.05). Conclusion HMGB1 gene knockout alleviates acute lung injury of sepsis mice, the related molecular mechanism may be the inhibition of TLR4/NF-κB pathway mediated inflammation and oxidative stress.

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Objective To investigate the effect and possible mechanism of microRNA-26a(miR-26a) on the synthesis of extracellular matrix(ECM) induced by high glucose(HG)in renal tubular epithelial cells(RTECs). Methods A model of diabetic kidney disease(DKD) was constructed by inducing RTECs with HG. MiR-26a was overexpressed in HG-induced RTECs, and RT-qPCR and Western blot were used to assess the effects of miR-26a on ECM synthesis and ferroptosis-related markers in HG-treated RTECs. Ferrostatin(Fer-1) was used to inhibit ferroptosis in the DKD model, and its impact on ECM synthesis was evaluated. RT-qPCR and Western blot were performed to measure ferroptosis-related markers, and fluorescence microscopy was used to observe the intensity of reactive oxygen species(ROS). Results Compared with the control group, the expression of miR-26a decreased in HG-treated cells, while the expression levels of ECM synthesis-related indexes fibronectin and collagen Ⅰ increased. After overexpressing miR-26a, the HG+miR-26a group showed a significant increase in miR-26a expression and a decrease in fibronectin and collagen Ⅰ expression compared to the HG group. In terms of ferroptosis, the protein and mRNA expression of SLC7A11 and GPX4 significantly decreased, the expression of TFR-1 and ACSL4 significantly increased, and the fluorescence intensity of ROS was significantly enhanced in the HG group compared with the control group. Inhibition of ferroptosis in the HG+Fer-1 group resulted in significant changes in ferroptosis and ECM synthesis-related indicators expression levels compared to the HG group. Furthermore, re-expression of miR-26a in the HG+miR-26a led to significant changes in ferroptosis-related indicators expression levels and decreased ROS fluorescence intensity compared to the HG group. Conclusions In HG-induced RTECs, miR-26a inhibits the occurrence of ferroptosis, thus reducing ECM synthesis.

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Objective To explore the clinical significance and mechanisms of chromatin licensing and DNA replication factor 1(CDT1) in lung adenocarcinoma). Methods The gene expression samples of lung adenocarcinoma tissue and normal lung tissue were downloaded from the TCGA database, and perform differential analysis, GO analysis, independent prognosis analysis, and correlation analysis with immunotherapy using R language. CDT1 expression in lung adenocarcinoma and normal tissues was detected by PCR in clinical samples. The changes of cell proliferation and cycle in si-CDT1 knockdown group and si-NC control group were detected by flow cytometry. The invasive ability of each group was detected by Transwell. The expressions of CDT1, TPX2 and p53 in each group were detected by Western blot. Results The TCGA data analysis revealed CDT1 as a differentially expressed gene. GO analysis indicated that CDT1 was closely associated with the cell cycle. The high expression of CDT1 in lung adenocarcinoma tissues was validated in clinical samples. CDT1 could serve as an independent factor for predicting the prognosis of lung adenocarcinoma and had predictive value for immunotherapy in lung adenocarcinoma. Knockdown of CDT1 resulted in a significant decrease in cell proliferation ability compared to the control group, and cells were noticeably arrested in the G1 phase. Transwell assay results demonstrated a significant reduction in invasive capacity in the CDT1 knockdown group. Knockdown of CDT1 led to a significant decrease in TPX2 expression and a significant increase in p53 expression, while overexpression of CDT1 yielded the opposite effect. Conclusion Results demonstrate the elevated expression of CDT1 in lung adenocarcinoma, its association with prognostic significance, and its impact on lung adenocarcinoma's occurrence and development by influencing TPX2 and p53.

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Objective To analyze the effect and molecular mechanism of phellodendron amurense in the treatment of liver injury based on network pharmacology, and to verify the relevant prediction targets and the protective effect of phellodendron amurense extract-Phellodendron amurense polysaccharide on immune liver injury through mice. Methods TCMSP and Swiss target prediction databases were used to retrieve and screen phellodendron amurenses active components and action targets, and then obtain disease-related targets on GeneCards and OMIM websites, and take compounds and disease intersection targets for protein interaction. Analysis, GO biological function and KEGG signaling pathway enrichment analysis, followed by molecular docking of compounds and key target proteins, and finally established a mouse liver injury model induced by Daodou protein A(Con A) to explore the mechanism of phellodendron amurense extract in the treatment of liver injury. Results 37 active ingredients were screened. The key targets for their treatment were tumor necrosis factor α(TNF-α), serine/threonine protein kinase 1(AKT1), signal transduction and transcription activation factor 3(STAT3), epidermal growth factor receptor(EGFR) anditin. Enzyme 3(CASP3) and other enrichment analysis showed that phellodendron amurense might play a protective role in protecting the liver through molecular mechanisms such as positive regulation of MAPK cascade reaction, oxidative stress response and regulatory PI3K-Akt signaling pathway, lipid and atherosclerosis. Animal experiments had found that the gastric treatment of phellodendron amurense polysaccharide could improve the activity of superoxide dismutase(SOD) and catalase(CAT) in liver tissue, reduce the levels of serum alkaline phosphatase(ALP), aspartate aminotransferase(AST) and malonaldehyde(MDA) in liver tissue, and regulate serum inflammatory factor while the expression of intercitin(IL)-6, IL-1β, tumor necrosis factor α(TNF-α), activated the expression of transforming growth factor β1(TGF-β1), and reduced TNF-α mRNA expression in liver tissue. Conclusion Phellodendron amurense can intervene in lipid and atherosclerosis pathways by acting on targets such as TNF-α, AKT1, STAT3, EGFR and CASP3 to reduce oxidative stress and inflammatory reactions and achieve liver protection.

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Objective To study the protective effect of lipoxin A4(LXA4) against sepsis-induced acute kidney injury(SAKI) in rats and its effect on the expression of toll-like receptor 4(TLR4),myeloid differentiation factor88(MyD88),nuclear factor kappa B(NF-κB) in the kidney. Methods Forty male specific pathogen-free C57BL/6J mice were randomly divided into SAKI group, SAKI+LXA4 group, sham procedure group, sham procedure+LXA4 group.The mice in SAKI group and SAKI+LXA4 group were given cecum ligation and puncture(CLP) to establish SAKI animal models.The mice in SAKI+LXA4 group and sham procedure+LXA4 group were given LXA4(40 ng/kg) with intraperitoneal injection 30 mins after CLP.All mice were sacrificed at the 24th hour after CLP to collect serum, urine and kidney tissues.The enzyme linked immunosorbent assay(ELISA) was used to test the serum creatinine(Scr),blood urea nitrogen(Bun),interleukin-1β(IL-1β),IL-6,tumor necrosis factor-α(TNF-α) and urine neutrophil gelatinase-associated lipocalin(NGAL),kidney injury molecule 1(KIM-1) levels of mice.The pathological changes were examined through hematoxylin-eosin(HE) staining and Periodic Acid-Schiff(PAS) staining.And the mRNA levels of TLR4,MyD88,NF-κB p65 were detected through real-time PCR(RT-PCR);the expression levels of TLR4,MyD88,NF-κB p65,phospho-NF-κB p65(p-NF-κB p65) were detected through immunohistochemistry(IHC) and Western blot assay. Results ELISA showed that the values of Scr, Bun, IL-1β, IL-6,TNF-α,NGAL,KIM-1 in SAKI group were higher than those in SAKI+LXA4 group(P<0.05),and there were no significant differences in Scr, Bun, IL-1β, IL-6, TNF-α, NGAL, KIM-1 between sham procedure group and sham procedure+LXA4 group. HE and PAS staining showed that SAKI group had severer pathological changes than SAKI+LXA4 group in the kidney structure(P<0.05), while pathological structures of the kidney were normal in sham procedure group and sham procedure+LXA4 group.RT-PCR showed that the mRNA levels of TLR4, MyD88,N F-κB p65 in SAKI group, SAKI+LXA4 group were higher than sham procedure group and sham procedure+LXA4 group(P<0.05); the mRNA levels of TLR4,MyD88,NF-κB p65 in SAKI group were higher than SAKI+LXA4 group(P<0.05);The results of IHC,Western blot assay were as follows: The expression levels of TLR4,MyD88,NF-κB p65,p-NF-κB p65 in SAKI group, SAKI+LXA4 group were higher than sham procedure group and sham procedure+LXA4 group(P<0.05); the expression levels of TLR4,MyD88,NF-κB p65, p-NF-κB p65 in SAKI group were higher than SAKI+LXA4 group(P<0.05). Conclusion TLR4/MyD88/NF-κB pathway plays an important role in the occurrence and development of SAKI, and LXA4 may reduce SAKI by inhibiting TLR4/MyD88/NF-κB signaling pathway.

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Objective To investigate the effects of long non-coding RNA 00894(LINC00894) gene on proliferation and metastasis of human gastric cancer cells, and to verify the regulatory relationship ofLINC00894,miR-205-5pandZEB1in gastric cancer. Methods The expression level ofLINC00894in gastric cancer cell lines, normal gastric lines, clinical gastric cancer and normal gastric tissue samples were determined by RT-qPCR. Through follow-up, the relationship between the expression level ofLINC00894and the prognosis of gastric cancer patients was explored.LINC00894knockdown cell lines and overexpression cell lines were constructed, and the knockdown and overexpression efficiency was detected by RT-qPCR. Cell proliferation and metastatic capacity were determined by CCK 8, clone formation and Transwell assays. Dual-luciferase reporter assays, RT-qPCR assays and Western blot assays were used to examine the targeted regulatory relationships ofLINC00894,miR-205-5pandZEB1. Results The expression ofLINC00894gene in gastric cancer tissues or cells was significantly higher than that in normal gastric tissues or cells, moreover, gastric cancer patients with highLINC00894gene expression had a worse prognosis. The knockdown ofLINC00894inhibited the viability, clonogenesis, migration and invasion ability of gastric cancer cells, and conversely, the overexpression ofLINC00894obtained the opposite results.LINC00894promotedZEB1expression by targeted downregulation ofmiR-205-5pexpression.LINC00894promoted the expression ofZEB1by targetingmiR-205-5pand down-regulating its expression. Conclusion LINC00894serves as an oncogene in gastric cancer and may promote proliferation and metastasis of gastric cancer cellsviaregulatingmiR-205-5p/ZEB1axis.

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Objective To investigate the correlation between uterine globulin associated protein 1(UGRP1) and Fas mediated apoptosis pathway in hashimoto thyroiditis(HT). Methods The expression of UGRP1 in thyroid cells of normal people and HT patients was detected by immunohistochemistry(IHC). FRTL-5 cells were transfected by plasmidsin vitro, and control group, UGRP1 group, Fas group were established respectively. Real-time fluorescent quantitative reverse transcription PCR(RT-qPCR) was used to detect the expression of Fas and UGRP1 mRNA in each group. Results UGRP1 expression was positive in thyroid cells of HT patients and negative in that of normal people. There were no significant differences between control group and UGRP1 group in Fas gene expression(1.085 0±0.124 9vs1.021 0±0.113 9). Compared with the control group, the expression of UGRP1 gene increased significantly in Fas group(P<0.000 1, 5.807 0±0.323 2vs0.752 7±0.076 0). Conclusion The high expression of UGRP1 in HT may be related to apoptosis pathway mediated by Fas.

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Objective To investigate the role of KAT7 in cartilage cell and tissue ageing by establishing an over-replicating-induced primary mouse cartilage cell ageing model and a mouse natural ageing model. Methods Chondrocytes of the mouse knee joint were obtained by type II collagenase digestion and identified by toluidine blue staining and Col II staining. The age-related proteins and KAT7 expression levels in cartilage cells from different generations of mice were discovered using Western blot and cellular immunofluorescence techniques, and the aging of the cells was assessed by SA-β-Gal coloring. The pathological alterations were examined in the joints of 22-month-old mice compared to 2-month-old mice using HE staining and safranin O-solid green staining. Additionally, immunohistochemical analysis was done to observe the expression of KAT7 and p53 in mouse joint tissue. Results Compared with the control group, the expression levels of KAT7 protein and p21 and p53 in aged mouse chondrocytes significantly increased. WM-3835, a commonly inhibitor of KAT7 that possess the capacity on halting the protein expression procedure of gene KAT7 as well as p21 in ageing chondrocytes. SA-β-Gal staining showed a significant increase in positive staining of chondrocytes in the eighth generation(P8) compared to the first generation(P1). Compared with the cartilage tissue of young mice, the cartilage tissue of elderly mice presents a near-bone distribution, with a decrease in cartilage surface integrity, a significant increase in the number of hypertrophic chondrocytes, and more KAT7 and p53 cells that were positive. Conclusion The expression of KAT7 increases in the ageing chondrocytes and the cartilage tissue of ageing mice, reveales the potential significance of KAT7 correlated to cellular aging process in cartilage.

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Objective To investigate the correlation between Yes-associated protein(YAP) nuclear expression and tumor size with prognosis of patients with epithelial ovarian cancer(EOC) and to study the role of YAP in EOC. Methods 120 patients with EOC were selected as the experimental group, including 38 patients with early stage(Ⅰ+Ⅱ) EOC and 82 patients with advanced stage(Ⅲ+Ⅳ) EOC. 30 normal ovarian tissues obtained from patients with uterine leiomyoma were enrolled as the control group. Immunohistochemical(IHC) assay was employed to determine YAP expression and sub-location. The relationship between YAP expression and the pathological parameters of the 120 patients with EOC was analyzed, so as to the prognosis of these patients. EOC cells(C13K and OV2008) were cultured with varying initial cell volumes. Ki67 expression and cell proliferation were tested by immunofluorescence and cloning assay respectively. YAP expression at mRNA and protein levels were detected by q-PCR and Western blot respectively when the cell conference of EOC cells reached to low(60%) and high(90%) cell density.Results The YAP nuclear expression was significantly higher in the EOC group compared to the control group(P<0.05). The average diameter of stage Ⅰ + Ⅱ EOC was larger than that of stage Ⅲ+ Ⅳ EOC(P<0.01). The high nuclear expression of YAP was positively associated with pathological grade,clinical stage and the level of Ca125>1 000 IU/ml, while negatively correlated with tumor size(allP<0.05).Survival analyses showed that smaller tumor size(<10 cm) and higher YAP nuclear expression were negatively associated with the 3-year overall survival rate of EOC patients(P<0.01). C13K and OV2008 cells cultured in the low density group exhibited a high number of clone formation, high Ki67 and YAP expression(P<0.01). The down-regulation of YAP expression could decrease the cell viability of EOC cells in the low-and high-density groups(P<0.05).Conclusion Higher level of YAP nuclear expression and smaller tumour size are inversely associated with the clinical prognosis of patients with EOC. Inhibiting YAP nuclear expression leads to a decrease in the proliferation capacity of EOC cells.

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Objective To investigate the effects of nuclear respiratory factor 1(NRF1) on mitochondrial and cognitive dysfunction in Alzheimer's disease(AD) model mice. Methods The 5×FAD mice were utilized as a model for Alzheimer's disease, and the sparsely labeled AAV virus overexpressing NRF1(AAV-NRF1) was administered via stereotaxic injection into the brain. The expression of NRF1 in hippocampus was determined by Western blot, the morphology of mitochondria in hippocampus was observed by transmission electron microscope, the dendritic spines of sparsely labeled neurons in the CA1 region were visualized and quantified using confocal laser microscopy, cognitive and memory functions of mice were evaluated using the Morris water maze test, while electrophysiological methods were employed to detect long-term potentiation(LTP) of synaptic efficacy. Results The expression of NRF1 in the hippocampus was significantly upregulated following stereotactic injection of AAV-NRF1(P<0.001). This intervention led to notable improvements in mitochondrial morphology within hippocampal neurons, as well as enhanced cognitive and memory functions in mice(P<0.01). Moreover, there was a significant increase in dendritic spine density among neurons located in the CA1 region of the hippocampus(P<0.001), accompanied by long-lasting and stable long-term potentiation(LTP) and a substantial elevation in fEPSP slope(P<0.01). Conclusion The overexpression of NRF1 in a 5×FAD mouse model of Alzheimer's disease(AD) initiated the restoration of mitochondrial dysfunction and enhanced synaptic plasticity, indicating that these alterations may contribute to the therapeutic efficacy of NRF1 overexpression in ameliorating cognitive dysfunction associated with AD.

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Objective To investigate the role of bufalin(BU) in inhibiting M2-type macrophage-mediated colorectal cancer metastasis. Methods Human acute leukemia mononuclear cells(THP-1) were differentiated into M0 macrophages using phorbol ester induction(PMA) for 48 hours. The M0 macrophages were then treated with IL-4 and IL-13 medium. Surface markers and morphological changes were observed through ELISA, morphology, and RT-qPCR experiments. RT-PCR and ELISA experiments were conducted to detect the surface markers TGF-β and IL-10 of M2 macrophages. The secretion level of IL-6 in the supernatant of M2 macrophages and colorectal cancer cells HCT116 was compared using ELISA. Additionally, the effect of conditioned medium on colorectal cancer cell HCT116 was assessed through Transwell, Wound healing, RT-qPCR, and Western blot experiments. Subsequently, bufalin was added to the conditioned medium and the changes in AKT/PI3K protein, migration, and epithelial-mesenchymal transition ability in HCT116 were observed using Western blot, Transwell, Wound healing and RT-qPCR experiments. Results THP-1 were successfully differentiated into M2 macrophages. The activation of AKT/PI3K protein in HCT116 cells was induced by the secretion of IL-6 from M2 macrophages, which in turn promoted the migration and epithelial-mesenchymal transition ability of the HCT116 cells. The migration and epithelial-mesenchymal transition mediated by M2 macrophages in HCT116 cells were effectively inhibited by Bufalin. Conclusion The release of IL-6 from M2 macrophages activates the AKT/PI3K signaling pathway in colorectal cancer cells, thereby promoting their migration and epithelial-mesenchymal transition capacity. Moreover, bufalin exhibits inhibitory effects on this effect.

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Objective To explore the mechanism of malignant behavior of cervical cancer(CC) cells based on AMP-activated protein kinase(AMPK)/rapamycin target protein(mTOR) signaling pathway mediated by nucleotide-binding oligomerization domain receptor 2(NOD2). Methods Bioinformatics analysis was performed to determine the expression of NOD2 in CC tissue. Plasmids targeting NOD2(shNOD2) and shRNAs negative control(shNC), NOD2 overexpression(NOD2) and vectors(Vec) were transfected into CC cells. The effect of NOD2 on the growth of CC cells was determined by cell counting kit-8 assay, colony formation and Transwell cell invasion assay. Transcriptome analysis was performed by high throughput RNA sequencing(RNA-Seq). Western blot was used to detect the expression of NOD2, AMPK/mTOR signaling pathway and autophagy protein in the cell line. 24 female BALB/c nude mice were randomly divided into four groups, with 6 mice in each group: vector group(Vec group), NOD2 overexpression group(NOD2 group), shNC group and shNOD2 group. The distant metastasis model was established in mice, and the fluorescence intensity of lung metastasis was monitored and the number of lung metastasis nodules was counted. Results On-line database analysis showed that the expression of NOD2 in CC tissues was significantly higher than that in normal tissues, and there were significant differences in the mRNA expression of NOD2 in different stages of CC(P<0.05). In addition, the high expression of NOD2 was associated with poor overall survival and disease-free survival(P<0.05). NOD2 overexpression promoted the proliferation, colony formation, migration and invasion of CC cells, while NOD2 knock-down was the opposite. Consistent with the results in vitro, the lung colonization and lung metastasis of CC cells in NOD2 group were significantly higher than those in Vec group(P<0.05), while those in shNOD2 group were significantly lower than those in shNC group(P<0.05). RNA-Seq results showed that the expression of NOD2 was significantly related to AMPK signal activation, mTOR signal inhibition, autophagy regulation pathway activation and autophagy formation. Compared with shNC group, the expression of phosphorylated AMPK and LC3 protein decreased significantly in shNOD2 group(P<0.05), and the expression levels of phosphorylated mTOR and p62 protein increased significantly(P<0.05).Compared with Vec group, the expression levels of LC3 and AMPK protein in NOD2 group increased significantly(P<0.05), and the expression levels of phosphorylated mTOR and p62 protein decreased significantly(P<0.05). Compared with shNC group, the point accumulation of GFP-mRFP-LC3 in shNOD2 group decreased significantly(P<0.05). Compared with Vec group, the point accumulation of GFP-mRFP-LC3 increased significantly(P<0.05).Conclusion NOD2 may promote the proliferation, migration and invasion of CC through AMPK/mTOR signal, and its mechanism partly involves autophagy activation.

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Objective To investigate the role of heme oxygenase-1(HO-1) on HBV replication and the antiviral effect of HO-1 combined with α-interferon(IFN-α). Methods HepG2.2.15 cells and HBV1.3-transfected HepG2 cells(HepG2-HBV1.3) were used as HBV replicating cell models; Hemin treated HepG2.2.15 and HepG2-HBV1.3 cells, to induce the expression of HO-1 molecules. CCK-8 method was used to assess the toxic effects of Hemin on HepG2 and HepG2.2.15; chemiluminescence method was used to analyze HBsAg and HBeAg in the supernatants of Hemin-treated group and si-HO-1 and other experimental groups; RT-qPCR was used to analyze HO-1, IFN-β and HBV-DNA; Western blot was used to analyze the expression of IRF-3 and the expression of related molecules in the JAK/STAT signaling pathway; Hemin combined with IFN-α treated HepG2.2.15 to monitor whether HO-1 had synergistic IFN-α antiviral effect. Results Hemin dose-dependently induced HO-1, and HO-1 was induced to exert a significant anti-HBV effect, while the expression of IFN-β, IRF-3, and IRF-9 and MxA, downstream molecules of the JAK/STAT signaling pathway, were all increased. Silencing HO-1 expression reversed the antiviral effect in the Hemin-induced group, and at the same time, type I interferon IFN-β showed low expression, and the expression of IRF-9 and MxA in the JAK/STAT signaling pathway was inhibited as well. Hemin combined with IFN-α exerted stronger antiviral effects. Conclusion HO-1 can exert an anti-HBV effect, which may be due to increased phosphorylation of IRF-3 to induce type I interferon expression and thus activate the JAK/STAT signaling pathway to exert an antiviral effect; HO-1 can synergize with IFN-α to exert an antiviral effect.

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Objective To investigate the effects of indirubatin derivative E804 on proliferation and migration of non-small cell lung cancer(NSCLC) A549 cells, and to elucidate the possible mechanism of Nrf2-HO-1/GPX4 pathway. Methods Lung cancer A549 cells were used as the cell model. The proliferation and migration of different specific inhibitors(Nec-1, CQ, Z-VAD, DFO, Fer-1 and Lip-1) in 0, 10 μmol/L E804 and 10 μmol/L E804+ groups were observed by MTT and cell scratch assay. The contents of reactive oxygen species(ROS) were detected by DCFH-DA fluorescence probe method, the contents of Fe2+were detected by colorimetric method, the contents of reduced glutathione(GSH) were detected by spectrophotometry, and the contents of malondialdehyde(MDA) were detected by micromethod. The expression levels of SLC7A11, Transferrin, GPX4, SLC40A1, Nrf2 and HO-1 were detected by Western blot in cells of 0, 2.5, 5 and 10 μmol/L E804 groups. Results Compared with the control group(0 μmol/L E804), 2.5, 5 and 10 μmol/L E804 significantly increased intracellular ROS, Fe2+and MDA levels, and decreased intracellular GSH content(P<0.01). Meanwhile, the expression levels of SLC7A11, GPX4, SLC40A1, Nrf2 and HO-1 significantly decreased(P<0.01), and the expression level of Transferrin increased(P<0.05). Compared with the 10 μmol/L E804 group alone, the apoptosis inhibitor(Z-VAD) group and the ferroptosis inhibitor(DFO, Fer-1 and Lip-1) group could significantly reverse the inhibition of proliferation and migration of A549 cells by 10 μmol/L E804(P<0.01). Conclution E804 can induce ferroptosis and inhibit the proliferation and migration of A549 cells, which may be related to the inhibition of Nrf2-HO-1/GPX4 pathway.

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Objective To understand the molecular epidemiological characteristics ofNorovirusoutbreaks and the genome evolution ofNorovirusepidemic strains in Hainan Province from 2020 to 2022. Methods The information and samples have been collected from thenorovirusoutbreaks from 2020 to 2022.Noroviruswas detected by using the real-time PCR in these samples, then the detected sequences were amplified the analyzed. TheNorovirussequences of 8 strains had been amplified and analyzed. Results From 2020 to 2022, 39 gastroenteritis outbreaks were reported, and 25 outbreaks caused byNoroviruswhich mainly occurred in childcare institutions and schools(20/25,80%). TheNorovirusoutbreaks were mainly concentrated in counties around Haikou(northeast), which including Ding′an(5 cases), Wenchang(4 cases), Chengmai(4 cases), and Lingao(3 cases); following by western regions which included Baisha(2 cases), Ledong(2 cases), and Dongfang(3 cases). 1 case was in Wanning in the southeast. Among individuals aged 2-17, the positive proportion ofNorovirusin males was higher than that in females. Among individuals aged over 55, the proportion ofNoroviruspositive in females was higher than that in males. The gender of positive samples among individuals aged 18-40 was related to their profession. According to RT-PCR typing and sequencing, GII groupNoroviruswere classified in13 outbreaks. There were 4 genotypes detected. GII.2 [P16] was the main epidemic strain with 60%(9/13), and the other three genotypes were GII.4 Sydney [P31](15.4%, 2/13) GII.4 Sydney [P16](7.7%, 1/13) and GII.3 [P12](7.7%, 1/13). Further genic analysis of 8Norovirusstrains showed that all of them were still in the same branch as the previous strain, and all exhibited a certain amount of amino acid variation. Conclusion Norovirusis the main pathogen of gastroenteritis outbreaks in Hainan province, and the main epidemic strain is GII.2 [P16]. It is necessary to continue to strengthen the monitoring that provides scientific evidence for the prevention and control ofnorovirusoutbreaks in Hainan region.

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Objective To discuss the value of clinical radiomic nomogram(CRN) and deep convolutional neural network(DCNN) in distinguishing atypical pulmonary hamartoma(APH) from atypical lung adenocarcinoma(ALA). Methods A total of 307 patients were retrospectively recruited from two institutions. Patients in institution 1 were randomly divided into the training(n=184: APH=97, ALA=87) and internal validation sets(n=79: APH=41, ALA=38) in a ratio of 7:3, and patients in institution 2 were assigned as the external validation set(n=44: APH=23, ALA=21). A CRN model and a DCNN model were established, respectively, and the performances of two models were compared by delong test and receiver operating characteristic(ROC) curves. A human-machine competition was conducted to evaluate the value of AI in the Lung-RADS classification. Results The areas under the curve(AUCs) of DCNN model were higher than those of CRN model in the training, internal and external validation sets(0.983vs0.968, 0.973vs0.953, and 0.942vs0.932, respectively), however, the differences were not statistically significant(p=0.23, 0.31 and 0.34, respectively). With a radiologist-AI competition experiment, AI tended to downgrade more Lung-RADS categories in APH and affirm more Lung-RADS categories in ALA than radiologists. Conclusion Both DCNN and CRN have higher value in distinguishing APH from ALA, with the former performing better. AI is superior to radiologists in evaluating the Lung-RADS classification of pulmonary nodules.

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Objective To investigate the correlation between serum bicarbonate level and cardiovascular events in peritoneal dialysis(PD) patients. Methods The data of PD patients who underwent PD catheterization and were followed up regularly until March 31, 2023 were retrospectively collected. The included patients were divided into low bicarbonate group and normal bicarbonate group according to the time-averaged serum bicarbonate level. The incidence of cardiovascular events(including coronary heart disease, heart failure, stroke, peripheral vascular disease, death related to cardiovascular surgery or death due to aneurysm dissection or rupture, fatal pulmonary embolism, or death from other or unknown cardiovascular causes) was compared between the two groups and the risk factors for cardiovascular events were analyzed. Results At the end of follow-up, a total of 110 PD patients were included, and 34 patients had cardiovascular events. Compared with the normal bicarbonate group, the low bicarbonate group had a higher incidence of cardiovascular events. Univariate Cox regression analysis showed that the risk of cardiovascular events in the low bicarbonate group was 4.197 times higher than that in the normal bicarbonate group(95%CI=2.115-8.331,P<0.001). After adjusting for multiple confounding factors, the risk of cardiovascular events in the low bicarbonate group was 3.506 times higher than that in the normal bicarbonate group(95%CI=1.709-7.193,P=0.001). The results of multivariate competing risk model showed that the risk of cardiovascular events in the low bicarbonate group was 3.801 times higher than that in the normal bicarbonate group(95%CI=1.920-7.525,P<0.001). Conclusion Low serum bicarbonate level is closely related to the occurrence of cardiovascular events in patients with PD, and it is an independent risk factor for cardiovascular events in patients with PD.

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Objective To investigate the diagnostic efficacy and clinical value of GNB4 and Riplet gene methylation alone and in combination in the diagnosis of primary liver cancer.Methods A total of 313 patients were selected,including 78 patients with primary liver cancer, 41 patients with other digestive system tumors, 17 patients with non-digestive system tumors, 20 patients with postoperative liver cancer, and 157 patients with benign liver diseases.The levels of GNB4 and Ripletgene methylation in plasma were detected using quantitative methylation-specific PCR(qMSP).Serum alpha-fetoprotein(AFP) levels were measured by direct chemiluminescence.Results The sensitivity and specificity of AFP in diagnosis were 51.3% and 94.3%, respectively; the sensitivity and specificity of GNB4 gene methylation in diagnosis were 83.3% and 99.4%, respectively; the sensitivity and specificity of Riplet gene methylation in diagnosis were 73.1% and 99.4%, respectively.The sensitivity and specificity of GNB4andRipletgene methylation combined diagnosis were 92.3% and 98.7%, respectively; the sensitivity and specificity of AFP,GNB4andRipletgene methylation combined diagnosis were 92.3% and 98.7%, respectively;the sensitivity and specificity of combined diagnosis including age and gender were 93.6% and 97.5%, respectively.Conclusion The sensitivity and specificity of AFP in the diagnosis of primary liver cancer are limited, while the methylation levels of GNB4 and Ripletgenes are higher, and the sensitivity and specificity of their combined detection are higher than those of AFP.The sensitivity and specificity of AFP,GNB4 and Ripletgene methylation combined diagnosis are significantly higher than those of AFP,GNB4 and Ripletgene methylation alone.

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Objective To investigate the relationship between interleukin 1 receptor a(IL-1Ra) gene polymorphism and different outcomes in asymptomatic bacterial vaginosis(aBV) patients with the aim of grouping and managing aBV patients. Methods In study on the natural attribution of aBV patients, all patients were enrolled and a sample of venous blood and vaginal lavage fluid were separately frozen. After 4 months at the end of the clinical study, patients who completed the study were divided into three groups based on clinical outcomes: self-healing, progressive, and unchanged. The IL-1Ra gene polymorphism, the concentration of IL-1β and IL-1Ra were tested, and the differences in the above indicators among three groups of patients with different outcomes were compared. Results 1 014 Chinese Han female patients were enrolled, and 984 patients completed clinical follow-up and obtained clinical outcome data. 13 specimens were unusable during testing, with a total of 971 specimens completed the test. IL-1Ra gene was detected in all patients, with three genotypes: A1/A1, A1/A2, and A2/A2. Most population had a genotype of A1/A1, with the rarest genotype being A2/A2. No rare genotype of female was found. The frequency of A2alleles in the progression group was significantly higher than that in the self-healing group(P<0.05). IL-1β and IL-1Ra were detected in all vaginal lavage fluid samples. Compared with the progression group, IL-1β in the self-healing group was significantly lower(P<0.05). When carrying the A2allele, IL-1β in progression group was relatively low, while the level of IL-1Ra was relatively high. The values of the unchanged group were middle. Conclusion The polymorphism characteristics of the IL-1Ra gene in aBV patients are related to the IL-1Ra content in vaginal secretions. Carrying allele A2is related to the elevation of IL-1Ra, the decrease of IL-1β in vaginal secretions. Carrying allele A2may affect the clinical outcome of aBV by some potential mechanism.