Background:Collagen triple helix repeat containing protein-1(Cthrc1)has been reported playing an important role in liver diseases,especially in liver fibrosis,however,its effect on hepatic stellate cells proliferation is not fully clear. Aims:To investigate the effect of Cthrc1 on proliferation of hepatic stellate cells. Methods:Recombinant adenovirus vector of Cthrc1 was constructed. After injecting Ad-Cthrc1 through tail vein,mRNA and protein expressions of Cthrc1 were determined by real-time PCR and Western blotting,respectively. Liver fibrosis model was established by bile duct ligation and fed with 3,5-diethoxycarbonyl-1,4-dihydrocollidine(DDC)in mice,respectively. The liver fibrosis mice were injected with Ad-Cthrc1 or Ad-GFP through tail vein. Immunofluorescence was used to determine number of hepatic stellate cells. Results:Recombinant Cthrc1-adenovirus vector was successfully constructed. Real-time PCR and Western blotting showed that mRNA and protein expressions of Cthrc1 were increased in Ad-Cthrc1 group than in control group. HE and Masson staining demonstrated that mice model of liver fibrosis was successfully established. Immunofluorescence showed that overexpression of Cthrc1 inhibited significantly the proliferation of hepatic stellate cells. Conclusions:Recombinant adenovirus vector of Ad-Cthrc1 constructed can express stably in vivo,and inhibit the proliferation of hepatic stellate cells. Therefore,Cthrc1 may become a potential target for treatment of liver fibrosis.

Background:Primary biliary cirrhosis( PBC ) is an autoimmune liver disease,the cause of the disease remain incompletely understood. In addition to genetic and environmental factors,autoantibodies,multiple immunocytes and cytokines are considered to be involved in the development of PBC. Recent studies indicated that interleukin-9(IL-9)had pleiotropic functions in inflammatory regulation in allergic and autoimmune diseases. Aims:To investigate the expression and clinical significance of IL-9 in patients with PBC. Methods:A total of 30 specimens of peripheral blood and 20 specimens of liver tissue from PBC patients were collected. Ten specimens of peripheral blood from healthy subjects and 4 specimens of normal liver tissue were served as controls. Level of serum IL-9 was determined by ELISA,and expression of IL-9 in liver tissue was detected by immunohistochemistry. Correlations between IL-9 and serum biochemical indicators, immune indicators and histologic stage of liver tissue were analyzed. Results:Level of serum IL-9 was significantly increased in PBC patients than in normal control group(P<0. 05),and was positively correlated with level of serum IgG (r=0. 681,P<0. 01). Amount of IL-9 positive cells in liver tissue was significantly increased in PBC patients than in normal control group(P <0. 01),and was positively correlated with histologic stage of liver tissue(rs =0. 465,P <0. 05). Conclusions:Expression of IL-9 is significantly increased in peripheral blood and liver tissue in patients with PBC and is positively correlated with level of serum IgG and histologic stage of liver tissue,which suggests an important role of IL-9 in the pathogenesis of PBC.

Background:Activation of hepatic stellate cells( HSCs)plays a pivotal role in development of liver fibrosis. Interleukin-17(IL-17)is the most important effector of T helper 17(Th17)cells that causes inflammatory cell infiltration and tissue damage. Preliminary studies showed that the number of IL-17-positive cells in liver tissue was positively correlated with the severity of liver fibrosis in patients with chronic hepatitis B and autoimmune hepatitis. However,the mechanism of IL-17 in liver fibrosis is not yet clarified. Aims:To investigate the effect of IL-17 on activation of human HSC cell line LX2 and collagen expression. Methods:Human HSC cell line LX2 was treated with different concentrations of IL-17. Viability of LX2 cells was measured by CCK-8 assay. mRNA expressions of α-smooth muscle actin(α-SMA), type Ⅰ collagen(Col-Ⅰ)and Col-Ⅲ were determined by real-time PCR. Protein expressions of α-SMA、Col-Ⅰ and Col-Ⅲ were detected by immunofluorescence. Results: Viability of LX2 cells increased with the increase of IL-17 concentration,but no significant differences were seen between any two groups(P > 0. 05). mRNA expressions of α-SMA, Col-Ⅰ and Col-Ⅲ in IL-17 treatment group(100 ng/ mL)were significantly higher than those in blank control group(P <0. 05). With the increase of IL-17 concentration,protein expressions of α-SMA,Col-Ⅰ and Col-Ⅲ gradually increased. Conclusions:IL-17 can promote the activation of HSCs and expressions of Col-Ⅰ and Col-Ⅲ,thereby contributing to the development of liver fibrosis.

Drug-induced liver injury ( DILI) and autoimmune hepatitis ( AIH) have many similar clinical and histological manifestations , which brings difficulties to clinicians in differential diagnosis .This article elaborates on the association between DILI and AIH and their simi-larities and differences in clinical and histological manifestations , in order to help clinicians with the differential diagnosis of DILI and AIH . This article reviews the selection of therapeutic regimens for DILI and AIH and introduces how to select therapeutic strategies based on patient conditions and make a definite diagnosis when there are difficulties in differential diagnosis .

Objective To investigate the effects of interleukin(IL)-34 on the polarization and migration ability of macrophages derived from human peripheral blood monocytes.Methods The CD14+monocytes were isolated from human peripheral blood monocytes by immunomagnetic bead sorting,and the purity of CD14+monocytes was detected by flow cytometry.The CD14+monocytes were divided into M1 type group,IL-34-M1 type group,M2 type group and IL-34-M2 type group.The cells in the M1 type group and IL-34-M1 type group were added granulocyte-macrophage colony stimulating factor(GM-CSF)to induce for 5 days,and half of the fluid was changed,and then the interferon-γ,lipopolysaccharides,IL-6 and GM-CSF were added for another 4-day induction;the cells in the M2 type group and IL-34-M2 type group were added macrophage colony stimulating factor(M-CSF)to induce for 5 days,and half of the fluid was changed,and M-CSF,IL-4,IL-6,and IL-13 were added for another 4-day induction.The cells in IL-34-M1 group and IL-34-M2 group were co-induced with IL-34 at the beginning of induction and on the 5th day of induction.On the 9th day of induction,the proportion of CD14+CD86+cells(M1 type macrophages)and CD14+CD163+cells(M2 type macrophages)in each group was detected by flow cytometry,and the migration ability of cells in the M2 type group and IL-34-M2 type group was detected by the Transwell chamber experiments.Results High purity CD14+monocytes were obtained through magnetic bead sorting,with a CD14 positive rate of(96.77± 2.72)％,which could be used for macrophage induction.The proportion of CD14+CD86+cells in the M1 type group and IL-34-M1 type group was(43.20±7.59)％and(27.87±2.06)％,respectively.The proportion of CD14+CD163+cells in the M2 type group and IL-34-M2 type group was(47.70±4.49)％and(58.95±3.65)％,respectively;the proportion of CD14+CD86+cells in the IL-34-M1 type group was significantly lower than that in the M1 type group(P＜0.05),while the proportion of CD14+CD163+cells in the IL-34-M2 type group was significantly higher than that in the M2 type group(P＜0.05).The number of migrating cells of macrophages in the M2 type group and IL-34-M2 type group was 97.8±9.0 and 205.6±21.9,respectively;the number of migrating cells of macrophages in the IL-34-M2 type group was significantly higher than that in the M2 type group(P＜0.05).Conclusion IL-34 can inhibit the polarization of macrophages derived from human monocytes cells towards M1 type,promote the polarization of macrophages towards M2 type,and enhance the migration ability of M2 type macrophages.

Objective:To investigate the correlation between the expression of programmed death ligand-1 (PD-L1) and clinicopathological parameters and prognosis in hepatocellular carcinoma (HCC) tissues.Methods:From January 2008 to December 2016, 344 HCC patients underwent surgery in Nantong Third Hospital Affiliated to Nantong University were enrolled. The expression of PD-L1 in paraffin HCC tissues was detected by tissue microarray and immunohistochemistry. The correlation between the expression of PD-L1 in tumor cells, tumor-infiltrating immune cells and the clinicopathological parameters and prognosis of HCC patients were analyzed. And the related factors affecting the prognosis of patients were explored. Chi-square test, log-rank test and univariate and multivariate Cox regression analysis were used for statistical analysis.Results:Positive PD-L1 located in the membrane and/or cytoplasm of HCC tumor cells and tumor-infiltrating immune cells. The positive rate of PD-L1 expression in tumor cells was 21.8%(75/344). The expression of PD-L1 in tumor cells was related to histological grade and microvascular invasion, the positive rates of PD-L1 expression of patients with histological gradeⅠ, Ⅱ and Ⅲ were 7.7% (2/26), 16.5% (19/115) and 26.6% (54/203), respectively, the positive rates of PD-L1 expression of patients with or without microvascular invasion were 29.3% (34/116) and 18.0% (41/228), respectively, and the differences were statistically significant ( χ2=7.659 and 5.787, P=0.022 and 0.016). The positive expression rate of PD-L1 in tumor-infiltrating immune cells was 47.1% (162/344). The expression of PD-L1 in tumor-infiltrating immune cells was related with microvascular invasion, the positive rates of PD-L1 expression in patients with or without microvascular invasion were 56.9% (66/116) and 42.1% (96/228), respectively, and the differences were statistically significant ( χ2=6.751, P=0.009). The median survival time of patients with negative expression of PD-L1 in HCC tumor cells was 61 months (30 months, 92 months), while that of patients with positive PD-L1 expression was 16 months(6 months, 44 months), and the difference was statistically significant ( χ2=55.722, P<0.01). The expression of PD-L1 in HCC tumor cells was an independent risk factor affecting overall survival time (hazard ratio=3.090, P<0.01). Conclusions:The expression of PD-L1 in HCC tumor cells may be related to the malignancy and invasion of HCC, and may be a potential risk factor for prognosis.

Objective: To explore the changes of the peripheral invariant natural killer T (iNKT) cells in patients with human immunodeficiency virus (HIV) infection. Methods: A total of 101 patients with HIV infection including 52 asymptomatic patients and 49 acquired immunodeficiency syndrome (AIDS) patients were enrolled in the study from June 2016 to July 2017. Flow cytometry was used to detect iNKT cells, CD4⁺ T cells and CD8⁺ T cells, and the relationship among them and HIV RNA was studied. At same time, 12 healthy persons were enrolled as control group. T test or variance analysis, rank sum test, Chi-square test and Fisher exact test were used for statistical analysis. Results: In HIV infected asymptomatic patients, AIDS patients and healthy controls, iNKT cells were 0.135% (0.066%, 0.228%), 0.058% (0.034%, 0.100%) and 0.385% (0.205%, 0.600%), respectively, and the difference was statistical significant (Z=40.113, P<0.01). CD4⁺ T cell counts in the three groups were (340.82±119.26) cells/μL, (72.73±61.84) cells/μL and (555.17±229.43) cells/μL, respectively, and the difference was statistical significant (t=113.79, P<0.01); CD8⁺ T cell counts in the three groups were (842.29±423.68) cells/μL, (540.43±257.85) cells/μL and (875.92±516.45) cells/μL, respectively, and the difference was statistical significant (t=9.423, P<0.01). Ratios of CD4⁺ /CD8⁺ T cells in the three groups were 0.490 (0.240, 0.695), 0.120 (0.030, 0.210) and 0.600 (0.475, 0.895), respectively, and the difference was statistical significant (Z=53.603, P<0.01). iNKT cell counts in patients with or without hepatitis B virus infection, pneumocystis pneumonia, oral mold infection, treponema pallidum, latent tuberculosis or EB virus infection were not significantly different (Z=0.244, 2.325, 2.393, 0.168, 1.183 and 0.454, respectively, all P>0.05). There were correlations between iNKT cells and CD4⁺ T cells, CD4⁺ /CD8⁺ T cells (r=0.513 and 0.261, respectively, both P<0.05), and no relationship was found between iNKT cells and CD8⁺ T cells (r=0.155, P=0.126). In HIV infected asymptomatic patients and AIDS patients, iNKT cells was not associated with HIV RNA (r=-0.113 and -0.111, respectively, both P>0.05). Conclusions The level of peripheral iNKT cells in HIV infected patients decreases with the disease progression. To certain extent, iNKT cells can reflect the severity of immune damaging in HIV infected patients.

Background: Hepatic encephalopathy (HE) is one of the main causes of death in patients with end-stage liver disease. Early evaluation of the condition of patients with liver cirrhosis complicated with HE is the key for improving the prognosis. Aims: To investigate the effects of controlling nutritional status (CONUT) and CTP score on predicting the short term prognosis of liver cirrhosis patients complicated with HE. Methods: The clinical data of 168 liver cirrhosis patients complicated with HE initially diagnosed in Nantong Third People's Hospital from January 2018 to December 2021 were enrolled and analyzed retrospectively. HE patients were divided into survival group and death group. Cox regression analysis was used to evaluate the risk factors affecting the prognosis. A new prediction model was established, and ROC curve was used to evaluate the values of different models for short⁃term prognosis of HE patients. Survival was evaluated by Kaplan⁃Meier method. Results: Compared with survival group, ALT, AST, TBIL, INR, white blood cell count, neutrophil were significantly increased in death group (P<0.05), while ALB, total cholesterol, serum Na, fibrinogen, lymphocyte were significantly decreased (P<0.05). CONUT (OR=1.499, 95% CI: 1.092⁃2.057, P=0.012), CTP score (OR=1.474, 95% CI: 1.178⁃1.844, P=0.001) were independent risk factors for 90⁃day mortality in liver cirrhosis patients complicated with HE. AUC of CTP, CONUT and CONUT⁃CTP scoring models were 0.90, 0.94 and 0.95, respectively. Kaplan⁃Meier analysis showed that the survival rate in CONUT⁃CTP≥7.50 group was significantly lower than that in <7.50 group (P<0.000 1). Conclusions: CONUT⁃CTP score has good predictive value for the prognosis of liver cirrhosis patients complicated with HE, CONUT⁃CTP ≥7.50 indicates poor prognosis.

Background: Interleukin-34 (IL-34) is an important immunomodulatory factor that plays a crucial role in a variety of autoimmune diseases. Aims: To investigate the expression of IL-34 in primary biliary cholangitis (PBC) and its influence on intrahepatic inflammation and bile duct damage. Methods: Liver tissues were obtained from 26 PBC patients and 10 hepatic hemangioma patients without PBC. Expression and localization of IL-34 were detected by immunohistochemistry. In animal experiment, Poly I:C intraperitoneal injection was used to construct PBC model in wild-type and IL-34-knockout C57BL/6 mice (WT-PBC group and IL-34KO-PBC group). Subsequently, the intrahepatic inflammation and bile duct damage were evaluated pathologically, and the expressions of IL-34 and associated cytokines in liver tissues were detected by real-time PCR and Western blotting. Results: Expression of IL-34 in liver tissues of PBC patients and PBC model mice was significantly higher as compared with those of the controls (all P<0.05). No morphological changes in hepatic pathological evaluation were observed in IL-34KO mice receiving intraperitoneal saline injection. In IL-34KO-PBC mice, the portal area inflammation and biliary epithelial cell damage were more severe than those in WT-PBC mice (all P<0.05). Expressions of proinflammatory cytokine interleukin-1β (IL-1β) and monocyte chemotactic protein-1 (MCP-1) in liver tissues of IL-34KO-PBC mice were significantly increased than those of WT-PBC mice, whereas expressions of antiinflammatory cytokine IL-10 and CD163, the surface marker of M2 macrophages, were significantly reduced (all P<0.05). Conclusions: IL-34 expression is increased in liver tissues of PBC patients and animals. It might reduce the portal area inflammation and bile duct damage via modulating cytokines expression and driving macrophages polarization to the M2 phenotype. IL-34 might act as a self-rescue factor which negatively regulates hepatic immune microenvironment and prevents disease progression.

INTRODUCTION: Paraquat (PQ) intoxication is frequently associated with a high mortality rate. No specific treatment has been shown to reduce mortality in victims within the first 72 hours. We investigated the protective effects of rapamycin (Rapa) against PQ-induced toxicity in a zebrafish model. METHODS: To determine the maximum nonlethal concentration (MNLC) and lethal concentration 50 (LC50) of Rapa, zebrafish were treated at 2-5 days post fertilisation (dpf) and their mortality was recorded every 24 hours. At 5 dpf, the zebrafish were treated with PQ 100 µg/mL or PQ+Rapa (MNLC, 1/3 MNLC or 1/9 MNLC) for 72 hours, and the rate of survival was recorded every 24 hours. Reverse transcription-polymerase chain reaction was used to test the signalling pathway of mTOR (mammalian target of rapamycin). RESULTS: MNLC and LC50 of Rapa were determined to be 6.7 µg/mL and 28.9 µg/mL, respectively. At 48 hours, the PQ+Rapa groups had much lower mortality than the PQ group. The rates of survival of the PQ+Rapa groups were 43.33% (MNLC), 53.89% (1/3 MNLC) and 44.45% (1/9 MLNC), as compared to 19.45% in the PQ group, with the 1/3 MNLC group showing the highest rate of survival (p < 0.001). atg1 was slightly activated in the PQ group. In the PQ+Rapa groups, the expression of atg1 was markedly increased, suggesting strengthening of the autophagy process. CONCLUSION Rapa can increase the rate of survival of PQ-intoxicated zebrafish by inhibiting mTOR complex 1 and activating autophagy. Rapa could be an alternative first-line drug in the treatment of PQ poisoning.