BACKGROUND Primary sclerosing cholangitis (PSC) is characterized by chronic inflammation and it predisposes to cholangiocarcinoma due to lack of effective treatment options. Recombinant adeno-associated virus (rAAV) provides a promising platform for gene therapy on such kinds of diseases. A microRNA (miRNA) let-7a has been reported to be associated with the progress of PSC but the potential therapeutic implication of inhibition of let-7a on PSC has not been evaluated. AIM To investigate the therapeutic effects of inhibition of a miRNA let-7a transferred by recombinant adeno-associated virus 8 (rAAV8) on a xenobiotic-induced mouse model of sclerosing cholangitis. METHODS A xenobiotic-induced mouse model of sclerosing cholangitis was induced by 0.1% 3,5-Diethoxycarbonyl-1,4-Dihydrocollidine (DDC) feeding for 2 wk or 6 wk. A single dose of rAAV8-mediated anti-let-7a-5p sponges or scramble control was injected in vivo into mice onset of DDC feeding. Upon sacrifice, the liver and the serum were collected from each mouse. The hepatobiliary injuries, hepatic inflammation and fibrosis were evaluated. The targets of let-7a-5p and downstream molecule NF-κB were detected using Western blot. RESULTS rAAV8-mediated anti-let-7a-5p sponges can depress the expression of let-7a-5p in mice after DDC feeding for 2 wk or 6 wk. The reduced expression of let-7a-5p can alleviate hepato-biliary injuries indicated by serum markers, and prevent the proliferation of cholangiocytes and biliary fibrosis. Furthermore, inhibition of let-7a mediated by rAAV8 can increase the expression of potential target molecules such as suppressor of cytokine signaling 1 and Dectin1, which consequently inhibit of NF-κB-mediated hepatic inflammation. CONCLUSION Our study demonstrates that a rAAV8 vector designed for liver-specific inhibition of let-7a-5p can potently ameliorate symptoms in a xenobiotic-induced mouse model of sclerosing cholangitis, which provides a possible clinical translation of PSC of human.

BACKGROUND:Primary sclerosing cholangitis (PSC) is a chronic cholestatic liver disease characterized by inflammatory fibrosis usually involving the whole biliary tree. However, there are very limited treatment options to treat this disease. Our previous study found a lipid-protein rCsHscB from a liver fluke - Clonorchis sinensis, which had full capacities of immune regulation. Therefore, we investigated the role of rCsHscB in a mouse model of sclerosing cholangitis induced by xenobiotic 3,5- diethoxycarbonyl-1,4-dihydrocollidine (DDC) to explore whether this protein had potential therapeutic value for PSC.METHODS:Mice were fed 0.1% DDC for 4 weeks and treated with CsHscB (30 μg/mouse, intraperitoneal injection, once every 3 days); the control group was given an equal amount of PBS or CsHscB under normal diet conditions. All the mice were sacrificed at 4 weeks for the evaluation of biliary proliferation, fibrosis, and inflammation.RESULTS:rCsHscB treatment attenuated DDC-induced liver congestion and enlargement and significantly decreased the upregulation of serum AST and ALT levels. The administration of rCsHscB to DDC-fed mice significantly decreased cholangiocyte proliferation and pro-inflammatory cytokine production compared to mice fed with DDC alone. Also, rCsHscB treatment showed a decreased expression of α-SMA in the liver and other markers of liver fibrosis (Masson staining, Hydroxyproline content, and collagen deposit). More interestingly, DDC-fed mice treated with rCsHscB showed a significant up-regulation of PPAR-γ expression, which was similar to control mice, indicating the involvement of PPAR-γ signaling in the protective action of rCsHscB.CONCLUSION:Overall, our data show that rCsHscB attenuates the progression of cholestatic fibrosis induced by DDC and supports the potential for manipulating the parasite-derived molecule to treat certain immune-mediated disorders.

目的 研究华支睾吸虫来源的分子伴侣(molecular chaperones HscB,CsHscB)对氧化偶氮甲烷(azoxymethane,AOM)联合葡聚糖硫酸钠(dextran sodium sulfate,DSS)诱导的小鼠炎症性结肠癌的治疗效果及潜在的分子机制.方法 C57BL/6 小鼠随机分为正常对照组(NC,n=10),rCsHscB组(n=10),模型组(AOM+DSS,n=15),干预组(AOM+DSS+rCsHscB,n=15).模型组和干预组按8 mg/kg体重的剂量经腹腔注射AOM,1 次/d,连续7 d.然后给予7 d 2％DSS饮水和14 d正常饮水,共4个循环.在给予DSS饮水的第4 d、7 d,rCsHscB组和干预组小鼠按照1.25 mg/kg体重的剂量经腹腔注射rCsHscB蛋白.记录小鼠体重和生存状况,第84 d剖杀小鼠,测量结肠长度,记录结肠肿瘤个数,HE和Masson染色观察结肠病变,ELISA检测结肠组织中IL-6、MCP-1 和IL-10 含量,Western blot检测结肠组织中α-SMA、MMP2、Occludin表达水平和ERK1/2、JNK、P38、AKT、STAT3 的磷酸化及非磷酸化水平.结果 与正常对照组相比,模型组71.4％的小鼠结肠中出现肿瘤,小鼠体重和结肠长度显著降低缩短(均P＜0.05),结肠组织中有大量炎症细胞浸润和纤维化形成.与模型组相比,干预组小鼠的生存率显著升高(P＜0.01),体重、结肠长度和炎症纤维化均有所改善,仅有10％小鼠出现结肠肿瘤(P＜0.01).与模型组相比,干预组小鼠结肠组织中IL-6 和MCP-1 显著降低(均P＜0.05),IL-10 显著升高(P＜0.0501),α-SMA 和MMP2 蛋白表达水平降低(均P＜0.05),Occludin蛋白表达水平升高(P＜0.05),ERK1/2、JNK、P38、AKT和STAT3蛋白的磷酸化和非磷酸化水平均显著降低(均P＜0.05).结论 rCsHscB可能通过抑制MAPK和AKT/STAT3 信号通路的活化改善AO M/DSS诱导的小鼠炎症性结肠癌.

Clonorchiasis caused by Clonorchis sinensis is a mainly foodborne parasitic disease. It can lead to hepatobiliary duct inflammation, fibrosis, obstructive jaundice, liver cirrhosis, and even cholangiocarcinoma. Interleukin (IL)-10 is an immune-regulatory cytokine which plays an immunosuppressive role during infection. Our previous study found that IL-10 was increased in mice with C. sinensis infection. However, the role and mechanism of IL-10 playing in hepatobiliary injury induced by C. sinensis infection remain unknown. Herein, Il10(+/+) mice and Il10(+/-) C57BL/6J mice were infected with C. sinensis. It was found that IL-10 deficiency aggravated biliary hyperplasia and exacerbated periductal fibrosis induced by C. sinensis infection. Moreover, IL-10 deficiency increased CD4(+)T cells and CD8(+)T cells but not macrophages in the liver of mice with infection. There were no apparent differences in Th1 and Treg cells between Il10(+/+) and Il10(+/-) mice infected with C. sinensis. However, the proportion of Th17 cells in CD4(+)T cells in Il10(+/-) infected mice was significantly higher than that in Il10(+/+) infected mice. IL-10 deficiency also enhanced the increase of Th17 cells induced by ESPs stimulation in vitro. Taken together, our results suggest that IL-10 plays a protective role in hepatobiliary injury in C57BL/6J mice induced by C. sinensis infection via inhibiting Th17 cells, which could deepen our understanding of the immunopathology of clonorchiasis.

Background Chinese medical postgraduates (CMPs) are a special subpopulation that has a great risk of mental health due to high workload, and heavy academic and clinical pressure during the COVID-19 pandemic. Physical activity has been demonstrated to be positive for the mental health of human being. However, little is known about the risks to mental health among CMPs as well as the potential effects of physical activity on mental health in CMPs during the COVID-19 pandemic. Aims In the present study, the aims are to (1) investigate the levels of PA and mental health that CMPs had; (2) to identify the potential factors that contribute to mental health among CMPs; (3) to explore the potential effects of PA on the mental health in CMPs. Methods The survey was conducted online across 25 provinces of mainland China in 28 medical colleges or universities with structured questionnaires. Physical Activity Rating Scale-3 (PARS-3), Depression, Anxiety, Stress Scales (DASS-21), and General Self-Efficacy Scale (GSES) were employed to assess the levels of PA, Negative emotional states, and self-efficacy of the participants, respectively. Resilience was evaluated using Connor-Davidson Resilience Scale (CD-RISC). The chi-square and Wilcoxon rank-sum tests were used to compare inter-group differences in demographic data and mental health conditions. Spearman's rank correlation test and partial correlation analysis were used to assess associations between exercise scores and mental health. Results We found some socio-demographic variables such as the location, education levels of parents and the levels of degrees they are training had potential effects on outcomes of mental health among 2,217 CMPs ( P < 0.05); furthermore, we also found that PA was negatively correlated with the negative emotion ( r = −0.045, P < 0.05) such as depression ( r = −0.052, P < 0.05), anxiety and stress, ( r = −0.051, P < 0.05) but positively correlated with the self-efficacy ( r = 0.143, P < 0.001) and resilience ( r = 0.192, P < 0.001) among Chinese postgraduate medical students. Conclusion We concluded that for Chinese postgraduate medical students, taking part in physical activity would depress negative emotions such as depression, anxiety, and stress, but improve their self-efficacy and resilience, which will benefit them in completing their studies and training.

Clonorchis sinensis (C. sinensis) infection induces severe hepatobiliary injuries, which can cause inflammation, periductal fibrosis, and even cholangiocarcinoma. Sphingolipid metabolic pathways responsible for the generation of sphingosine-1-phosphate (S1P) and its receptor S1P receptors (S1PRs) have been implicated in many liver-related diseases. However, the role of S1PRs in C. sinensis-mediated biliary epithelial cells (BECs) proliferation and hepatobiliary injury has not been elucidated. In the present study, we found that C. sinensis infection resulted in alteration of bioactive lipids and sphingolipid metabolic pathways in mice liver. Furthermore, S1PR2 was predominantly activated among these S1PRs in BECs both in vivo and in vitro. Using JTE-013, a specific antagonist of S1PR2, we found that the hepatobiliary pathological injuries, inflammation, bile duct hyperplasia, and periductal fibrosis can be significantly inhibited in C. sinensis-infected mice. In addition, both C. sinensis excretory-secretory products (CsESPs)- and S1P-induced activation of AKT and ERK1/2 were inhibited by JTE-013 in BECs. Therefore, the sphingolipid metabolism pathway and S1PR2 play an important role, and may serve as potential therapeutic targets in hepatobiliary injury caused by C. sinensis-infection.

Clonorchiasis caused by Clonorchis sinensis is a zoonotic parasitic disease characterized by cholangitis, biliary proliferation, biliary fibrosis, and even cholangiocarcinoma. Our previous study showed that the expression of interleukin (IL)-33 is increased in both humans and mice infected by C. sinensis, suggesting that IL-33 is potentially involved in the pathogenesis of clonorchiasis. However, the roles and potential mechanism of IL-33 underlying remain unknown. Wild-type (WT) and IL-33 knockout (KO) mice (BALB/c female mice) were orally infected with 45 metacercariae of C. sinensis for 8 weeks. Biliary injuries and fibrosis were extensively evaluated. Hepatic type II cytokines (IL-4, IL-13, and IL-10) were detected by ELISA. For wild-type mice, we found that the mice infected with C. sinensis showed severe biliary injuries and fibrosis compared with the normal mice that were free from worm infection. In addition, the levels of type II cytokines such as IL-4, IL-13, and IL-10 in infected wild-type mice were significantly higher than in the control mice without infection (P < 0.05). However, IL-33 deficiency (IL-33 KO) prevents the augmentation of biliary injuries and fibrosis caused by C. sinensis infection. Furthermore, the increased levels of these type II cytokines induced by worm infection were also reversed in IL-33 KO mice. Our present study demonstrates that IL-33 contributes to the pathogenesis of C. sinensis-induced biliary injuries and repair, which can potentially orchestrate type 2 responses. These findings highlight the pathophysiological role of IL-33 in the progression of clonorchiasis.

The autonomic nervous system has been studied for its involvement in the control of macrophages; however, the mechanisms underlying the interaction between the adrenergic receptors and alternatively activated macrophages (M2) remain obscure. Using FVB wild-type and beta 2 adrenergic receptors knockout, we found that β2-AR deficiency alleviates hepatobiliary damage in mice infected with C. sinensis . Moreover, β2-AR-deficient mice decrease the activation and infiltration of M2 macrophages and decrease the production of type 2 cytokines, which are associated with a significant decrease in liver fibrosis in infected mice. Our in vitro results on bone marrow–derived macrophages revealed that macrophages from Adrb2 −/− mice significantly decrease M2 markers and the phosphorylation of ERK/mTORC1 induced by IL-4 compared to that observed in M2 macrophages from Adrb2 +/+ . This study provides a better understanding of the mechanisms by which the β2-AR enhances type 2 immune response through the ERK/mTORC1 signaling pathway in macrophages and their role in liver fibrosis.