The ubiquitin ligase Stub1 negatively modulates regulatory T cell suppressive activity by promoting degradation of the transcription factor Foxp3.

Regulatory T (Treg) cells suppress inflammatory immune responses and autoimmunity caused by self-reactive T cells. The key Treg cell transcription factor Foxp3 is downregulated during inflammation to allow for the acquisition of effector T cell-like functions. Here, we demonstrate that stress signals elicited by proinflammatory cytokines and lipopolysaccharides lead to the degradation of Foxp3 through the action of the E3 ubiquitin ligase Stub1. Stub1 interacted with Foxp3 to promote its K48-linked polyubiquitination in an Hsp70-dependent manner. Knockdown of endogenous Stub1 or Hsp70 prevented Foxp3 degradation. Furthermore, the overexpression of Stub1 in Treg cells abrogated their ability to suppress inflammatory immune responses in vitro and in vivo and conferred a T-helper-1-cell-like phenotype. Our results demonstrate the critical role of the stress-activated Stub1-Hsp70 complex in promoting Treg cell inactivation, thus providing a potential therapeutic target for the intervention against autoimmune disease, infection, and cancer.

Clinical value of circular RNAs and autophagy-related miRNAs in the diagnosis and treatment of pancreatic cancer.

BACKGROUND: Circular RNAs (circRNAs) are a special group of long-chain and non-coding RNAs characterized by a closed-loop structure without 3' and 5' polarity. In recent years, studies have demonstrated that circRNAs act as microRNA (miRNA) sponges to regulate the function of miRNAs. Increasing evidence indicates that circRNAs and targeted miRNAs are involved in the development, progression and metastasis of various cancers and drug resistance. A number of miRNAs are known to be associated with the pathogenesis, development and treatment of pancreatic cancer by regulating the autophagic activity. DATA SOURCES: A comprehensive literature search was executed in PubMed and EMBASE using the medical subject headings (MeSH) terms "Pancreatic Neoplasms", "autophagy", "RNA, circular" and "microRNA". We also used text terms such as "diagnosis", "prognosis" and "biomarker" to supplement the results. RESULTS: Autophagy-related miRNAs is closely related to pancreatic cancer. On basis of the retrieval results, we summarized the synthesis, features and functions of circRNAs and analyzed the association between autophagy-related miRNAs and pancreatic cancer. CONCLUSIONS: circRNAs act as the miRNA sponges and there is an association between miRNAs and autophagy, which provides a new concept to broaden the knowledge about the mechanisms underlying the development, progression and metastasis of pancreatic cancer. Additionally, clinical value of circRNAs and autophagy-related miRNAs in the diagnosis and treatment of pancreatic cancer would be further verified with in-depth researches.

Suppressive effect of microRNA-29b on hepatic stellate cell activation and its crosstalk with TGF-ß1/Smad3.

The microRNA (miR)-29 family is closely associated with fibrotic processes by virtue of its low expression in many tissues during organ fibrosis. The present study investigated whether miR-29b overexpression suppressed hepatic stellate cell (HSC) activation and its interactions with transforming growth factor (TGF)-ß1/mothers against decapentaplegic homolog 3 (Smad3), a classical signal transduction pathway contributing to the activation of HSCs. The results showed that transfection of LX-2 (human HSC) cells with miR-29b mimic or pSUPER-Smad3 silencing (si)RNA resulted in significantly increased expression of miR-29b and decreased expression of Smad3. miR-29b overexpression inhibited proliferation of LX-2 cells 24 h after transfection. Both miR-29b overexpression and Smad3 silencing antagonized the effects of TGF-ß1 on the expression of α-smooth muscle actin (α-SMA) and collagen type I (col-1). Furthermore, infection with miR-29b mimics suppressed Smad3 and TGF-ß1 expression, suggesting that miR-29b inhibited LX-2 activation mediated by both Smad3 and TGF-ß1. Nevertheless, primary miR-29a/b1, miR-29b2/c and mature miR-29b were downregulated by TGF-ß1 and stimulated by Smad3 silencing, suggesting that TGF-ß1/Smad3 signalling pathway regulate not just mature miR-29b but also its transcription. In summary, our results show overwhelming evidence corroborating the suppressive effect of miR-29b on TGF-ß1-induced LX-2 cell activation. The results also revealed the existence of crosstalk between miR-29b and TGF-ß1/Smad3 during LX-2 activation, suggesting a feedback loop between miR-29b and TGF-ß1/Smad3 signalling that promotes liver fibrosis. Copyright © 2016 The Authors. Cell Biochemistry and Function published by John Wiley & Sons, Ltd.

Therapeutic effects of curcumin on constipation-predominant irritable bowel syndrome is associated with modulating gut microbiota and neurotransmitters.

Introduction: Constipation-predominant irritable bowel syndrome (IBS-C) is a functional bowel disease that affects 10-20% of the population worldwide. Curcumin (CUR) is widely used in traditional Chinese medicine to treat IBS, but its mechanism of action needs further investigation. Methods: In this study, we used mosapride (MOS) as a positive control to evaluate the changes in gut microbiota in IBS-C rat models after treatment with CUR or MOS by analyzing 16S rDNA variation. In addition, we used enzyme immunoassay kits and immunohistochemical analysis to investigate whether CUR or MOS influenced serotonin (5-HT), substance P (SP), and vasoactive intestinal peptide (VIP) levels in the serum and colon of IBS-C rats. Results: The study showed that rats supplemented with CUR showed significantly increased fecal weight, fecal water content, small intestine transit rate and significantly decreased serum levels of 5-HT, VIP and SP compared to the IBS group (p < 0.05). In addition, treatment with CUR changed the relative abundance of Blautia, Sutterella, Acetanaerobacterium and Ruminococcus2 in the gut microbiota. Discussion: This study showed that the efficacy of CUR on IBS-C was possibly by modulating the microbiota and lowering the serum levels of HT, SP, and VIP.

The construction and analysis of a ferroptosis-related gene prognostic signature for pancreatic cancer.

Ferroptosis is a regulated cell death nexus linking metabolism, redox biology and diseases including cancer. The aim of the present study was to identify a ferroptosis-related gene prognostic signature for pancreatic cancer (PCa) by systematic analysis of transcriptional profiles from Cancer Genome Atlas (TCGA) and Genotype-Tissue Expression (GTEx). Altogether 14 ferroptosis-relevant genes with potential prognostic values were identified, based on which a risk score formula was constructed. According to the risk scores, we classified the patients into a high- and a low-risk score group. It was verified in Gene Expression Omnibus (GEO) and ICGC (International Cancer Genome Consortium) datasets. The Kaplan-Meier survival curves demonstrated that patients with lower risk scores had significantly favorable overall survival (OS) (P < 0.0001). The area under the receiver operating curve (ROC) for 12, 18 and 24 months was about 0.8 in all patients. The result of immune status analysis revealed that the signature significantly associated with the immune infiltration and immune checkpoint blockade (ICB) proteins. In addition, we used quantitative real time PCR (q-rtPCR) and Human Protein Atlas (HPA) to validate the expression of the key genes. Collectively, the signature is valuable for survival prediction of PCa patients. As the signature also has relevance with the immune characteristics, it may help improve the efficacy of personalized immunotherapy.

The Therapeutic Efficacy of Curcumin vs. Metformin in Modulating the Gut Microbiota in NAFLD Rats: A Comparative Study.

Structural disruption of gut microbiota is closely related to the occurrence of non-alcoholic fatty liver disease (NAFLD). Previous research has demonstrated that both curcumin (CUR) and metformin (MET) have a therapeutic effect against NAFLD and play a role in modulating the gut microbiota. However, there is a lack of direct comparison between the two medications in terms of the therapeutic efficacy and the regulatory effect on gut microbiota. In this study, we administered either CUR or MET to rats with high-fat diet (HFD)-induced obesity to observe changes in body parameters, biochemical parameters, liver, and ileum pathology and gut microbiota, and used next generation sequencing and multivariate analysis to evaluate the structural changes of gut microbiota in a NAFLD rat model before and after CUR and MET intervention. It was found that both CUR and MET attenuated hepatic ectopic fat deposition, alleviated inflammatory factors, and improved intestinal barrier integrity in HFD-fed rats. More importantly, CUR and MET reduced the Firmicutes/Bacteroidetes ratio and reverted the composition of the HFD-disrupted gut microbiota. Both CUR and MET treatments effectively modified the gut microbiome, enriched the abundance of beneficial bacteria and reduced opportunistic pathogens in obese rats. The abundance of Butyricicoccus was increased while the abundance of Dorea was decreased in HFD + CUR group. Besides, some beneficial bacteria such as Prevotella were increased in MET-treated animals. Spearman's correlation analysis showed that Helicobacter, Akkermansia, Desulfovibrio, Romboutsia, Corynebacterium, Lactobacillus, Ruminococcaceae_unclassified, Lachnospiraceae_unclassified, and Clostridiales_unclassified showed significantly positive correlations with TG, TC, LDL-C, GLU, IL-6, IL-1ß, and TNF-α, and negative correlations with HDL-C (both p < 0.05). However, Prevotella and Stomatobaculum showed an opposite trend. In summary, CUR and MET showed similar effects in alleviating hepatic steatosis, improving intestinal barrier integrity and modulating gut microbiota in HFD-induced obesity rats, and therefore may prove to be a novel adjunctive therapy for NAFLD.

Epidemiological and Molecular Investigations on Salmonella Responsible for Gastrointestinal Infections in the Southwest of Shanghai From 1998 to 2017.

PURPOSE: To investigate the characteristics of gastrointestinal infections in Southwest Shanghai. METHODS: Clinical and epidemiological characteristics of patients with Salmonella infections between 1998 and 2017 admitted to the Jinshan Hospital in the Southwest of Shanghai were retrospectively analyzed. A total of 565 isolated Salmonella strains were classified by serotyping and pulsed field gel electrophoresis (PFGE). RESULTS: From 1998 to 2006, diarrhea was mainly caused by Vibrio parahaemolyticus followed by Shigella and Salmonella. From 2007 to 2010, Vibrio parahaemolyticus infection was the major cause of diarrhea followed by Salmonella and Shigella. From 2011 to 2017, Salmonella infections became the main cause of diarrhea after Vibrio parahaemolyticus. Salmonella infections increased from 2006 on and peaked between May and October, accounting for 82.48% of yearly infections. Patients with Salmonella infections (90.5%) had a history of eating unclean food, abdominal pain (58.05%), diarrhea ≥5 times a day (50.44%), moderate fever (24.96%) and increased fecal leukocytes (41.42%). From 1998 to 2017, infected specimens from clinical cases were dominated by Salmonella enterica serovar Typhimurium (S. Typhimurium) (21.59%) followed by Salmonella enterica serovar Enteritis (S. Enteritidis) (16.81%), Salmonella enterica serotype London (6.55%) and Salmonella group B (13.10%). Other species included Salmonella enterica serovar Thompson, Salmonella enterica serovar Saintpaul, Salmonella group D, Salmonella group C, Salmonella enterica serovar Choleraesuis and Salmonella enterica serovar Aberdeen. The PFGE classification of Salmonella serovars in 2008-2017 demonstrated that S. Enteritidis had 9 PFGE banding patterns and S. Typhimurium 16 with varying degrees of similarity among S. Enteritidis and S. Typhimurium. The results of antibiotic susceptibility tests for the 330 Salmonella strains revealed that fosfomycin had the highest sensitivity rate (97.5%) followed by levofloxacin and ceftriaxone (81%), and ampicillin/sulbactam (78.2%). The resistance to piperacillin and ciprofloxacin was 60.9 and 50.61%, respectively. CONCLUSION: The features of onset, epidemiological characteristics and molecular subtyping of Salmonella were conducive to clinical diagnosis, rational use of antibiotics and improved therapeutic efficacy.

Role and mechanism of AT1-AA in the pathogenesis of HELLP syndrome.

HELLP syndrome remains a leading cause of maternal and neonatal mortality and morbidity worldwide, which symptoms include hemolysis, elevated liver enzymes and low platelet count. The objective of this study was to determine whether HELLP is associated with AT1-AA. The positive rate and titer of AT1-AA in plasma from pregnant women were determined, and the correlation of AT1-AA titer with the grade of HELLP was analyzed. A HELLP rat model established by intravenous injection of AT1-AA. Our experimental results show the AT1-AA titer and positive rate were significantly higher in HELLP group, and AT1-AA titer were positively correlated with the level of TNF-α and ET-1 in plasma and the grade of HELLP syndrome. The results of animal experiments showed that the typical features of HELLP in the pregnant rats after AT1-AA injection. The levels of TNF-α and ET-1 in plasma and liver tissue were significantly increased in AT1-AA-treated rats compared with control rats. The HELLP syndrome induced by AT1-AA was attenuated markedly after administration of losartan. These data support the hypothesis that one the potential pathway that AT1-AA induce damage to capillary endothelial cells and liver during pregnancy is through activation of TNF-α and ET-1.

Curcumin Induces Autophagy, Apoptosis, and Cell Cycle Arrest in Human Pancreatic Cancer Cells.

OBJECTIVE: Curcumin is an active extract from turmeric. The aim of this study was to identify the underlying mechanism of curcumin on PCa cells and the role of autophagy in this process. METHODS: The inhibitory effect of curcumin on the growth of PANC1 and BxPC3 cell lines was detected by CCK-8 assay. Cell cycle distribution and apoptosis were tested by flow cytometry. Autophagosomes were tested by cell immunofluorescence assay. The protein expression was detected by Western blot. The correlation between LC3II/Bax and cell viability was analyzed. RESULTS: Curcumin inhibited the cell proliferation in a dose- and time-dependent manner. Curcumin could induce cell cycle arrest at G2/M phase and apoptosis of PCa cells. The autophagosomes were detected in the dosing groups. Protein expression of Bax and LC3II was upregulated, while Bcl2 was downregulated in the high dosing groups of curcumin. There was a significant negative correlation between LC3II/Bax and cell viability. CONCLUSIONS: Autophagy could be triggered by curcumin in the treatment of PCa. Apoptosis and cell cycle arrest also participated in this process. These findings imply that curcumin is a multitargeted agent for PCa cells. In addition, autophagic cell death may predominate in the high concentration groups of curcumin.

tRNASer(CGA) differentially regulates expression of wild-type and codon-modified papillomavirus L1 genes.

Exogenous transfer RNAs (tRNAs) favor translation of bovine papillomavirus 1 wild-type (wt) L1 mRNA in in vitro translation systems (Zhou et al. 1999, J. Virol., 73, 4972-4982). We, therefore, investigated whether papillomavirus (PV) wt L1 protein expression could be enhanced in eukaryotic cells following exogenous tRNA supplementation. Both Chinese hamster ovary (CHO) and Cos1 cells, transfected with PV1 wt L1 genes, effectively transcribed the genes but did not translate them. However, L1 protein translation was demonstrated following co-transfection with the L1 gene and a gene expressing tRNA(Ser)(CGA). Cell lines, stably transfected with a bovine papillomavirus 1 (BPV1) wt L1 expression construct, produced L1 protein after the transfection of the tRNA(Ser)(CGA) gene, but not following the transfection with basal vectors, suggesting that tRNA(Ser)(CGA) gene enhanced wt L1 translation as a result of endogenous tRNA alterations and phosphorylation of translation initiation factors elF4E and elF2alpha in the tRNA(Ser)(CGA) transfected L1 cell lines. The tRNA(Ser)(CGA) gene expression significantly reduced translation of L1 proteins expressed from codon-modified (HB) PV L1 genes utilizing mammalian preferred codons, but had variable effects on translation of green fluorescent proteins (GFPs) expressed from six serine GFP variants. The changes of tRNA pools appear to match the codon composition of PV wt and HB L1 genes and serine GFP variants to regulate translation of their mRNAs. These findings demonstrate for the first time in eukaryotic cells that translation of the target genes can be differentially influenced by the provision of a single tRNA expression construct.