α7 nicotinic acetylcholine receptor agonist GTS-21 attenuates DSS-induced intestinal colitis by improving intestinal mucosal barrier function.

BACKGROUND AND AIMS: Cholinergic output, which could modulate innate immune responses through stimulation of α7 nicotinic acetylcholine receptor (α7nAChR), might be a target to minimize tissue damage in autoimmune disease. GTS-21, a selective α7nAChR agonist, has previously demonstrated to inhibit synovium inflammation in rheumatoid arthritis. In this study, we investigated the effect of GTS-21 on dextran sulfate sodium (DSS)-induced colitis model and its potential mechanism. METHODS: Male BABL/c mice (n = 32) were randomly divided into four groups: normal control group, DSS-induced colitis group, GTS-21 treatment with or without α7nAChR antagonist α-BGT treatment group. Disease activity index (DAI), histological activity index (HAI) and colonic macroscopic damage were evaluated. Fluorescein isothiocyanate (FITC)-dextran assay was applied to measure intestinal permeability. The expressions of tight junction (TJ) proteins and NF-κB associated proteins were detected by Western blot. RESULTS: GTS-21 could decrease DAI scores, HAI scores, intestinal permeability and reduce the intestinal bacterial translocation in DSS-induced colitis group, whereas α7nAChR antagonist α-BGT could impair this protective influence. The expressions of TJ proteins were increased with administration of GTS-21 both in vivo and in vitro. Furthermore, GTS-21 also inhibited the NF-қB activation in intestinal epithelial cells and colitis model, while α-BGT reversed the inhibitory effect. CONCLUSION: The α7nAChR agonist GTS-21 attenuated DSS-induced colitis through increasing expressions of TJ proteins in colon tissues and improved intestinal barrier function, which might be due to  modulating NF-қB activation in intestinal epithelial cells.

TNF-α-induced lncRNA LOC105374902 promotes the malignant behavior of cervical cancer cells by acting as a sponge of miR-1285-3p.

Long noncoding RNAs (lncRNAs) regulate gene expression at epigenetic, transcriptional, post-transcriptional levels and play important roles in tumorigenesis and inflammation. In order to explore the effects of lncRNAs on the malignant behavior of cervical cancer (CC) which may be involved in mechanism stimulated by inflammatory factors, we screened a differential expression profile of lncRNAs in CC cells stimulated by TNF-α by deep sequencing. We characterized a significantly upregulated lncRNA LOC105374902 induced by TNF-α. Then, we found that TNF-α accelerated the binding of STAT3 to the promoter region of lncRNA LOC105374902 and promoted its expression. Mechanistically, lncRNA LOC105374902 directly bond to miR-1285-3p as a competing endogenous RNA (ceRNA) to derepress RPL14; functional analysis indicated that both lncRNA LOC105374902 and RPL14 promoted migration, invasion and epithelial-mesenchymal transition (EMT) of CC cells. Taken together, TNF-α-induced lncRNA LOC105374902 may function as a ceRNA for miR-1285-3p to promote the expression of RPL14, promoting the migration, invasion and EMT of CC cells. These findings may provide new insights into the molecular pathogenesis of CC.

Probiotic Mixture Protects Dextran Sulfate Sodium-Induced Colitis by Altering Tight Junction Protein Expressions and Increasing Tregs.

Bifico is a probiotic mixture containing Bifidobacterium, Lactobacillus acidophilus, and Enterococcus. Studies support that Bifico has a protective effect in experimental colitis (IL-10-deficient and TNBS) models and in patients with inflammatory bowel disease (IBD). However, the mechanism underlying the protective effects of this mixture of probiotic bacteria remains incompletely clear. Here, we investigated the effect of Bifico on intestinal inflammation. In an in vivo experiment, dextran sulfate sodium was used to induce colitis. Bifico treatment significantly attenuated the severity of colitis in this model. Bifico increased the expression of tight junction proteins (TJs). In addition, Bifico increased the number of Tregs, but reduced the number of total CD4+ T cells in the peripheral blood. Furthermore, the expression of colonic CD4 protein was decreased while the level of forkhead box P3 (Foxp3) was upregulated. These results suggested that Bifico exerts beneficial effects on experimental colitis by increasing the expressions of TJs, upregulating the number of Tregs, and reducing the total CD4+ T cell number in both colon and peripheral blood. The intestinal damage in the pretreated + treated-Bifico-colitis group was more severe than that in only the pretreated-Bifico-colitis group. This suggested that Bifico might aggravate intestinal damage when the mucosal barrier is impaired.

Novel T-cell epitopes on Schistosoma japonicum SjP40 protein and their preventive effect on allergic asthma in mice.

Allergic asthma is a chronic inflammatory disease mediated by Th2 cell immune responses. Currently, immunotherapies based on immune deviation are attractive, preventive, and therapeutic strategies for asthma. Many studies have shown that intracellular bacterial infections such as mycobacteria and their components can suppress asthmatic reactions by enhancing Th1 responses, while helminth infections and their proteins can inhibit allergic asthma via immune regulation. However, some helminth proteins such as SmP40, the major egg antigen of Schistosoma mansoni, are found as Th1 type antigens. Using a panel of overlapping peptides, we identified T-cell epitopes on SjP40 protein of Schistosoma japonicum, which can induce Th1 cytokine and inhibit the production of Th2 cytokines and airway inflammation in a mouse model of allergic asthma. These results reveal a novel form of immune protective mechanism, which may play an important role in the modulating effect of helminth infection on allergic asthmatic reactions.

Phospholipase Cε deficiency delays the early stage of cutaneous wound healing and attenuates scar formation in mice.

This study aimed to investigate the role of phospholipase Cε (PLCε) in the skin wound healing process. PLCε, an effect factor of Ras/Rap small G protein, plays a crucial role in skin inflammation by regulating inflammatory cytokines. Inflammatory responses are closely associated with wound healing. Full-thickness skin wounds were made in the PLCε knockout (KO) and wild-type (WT) mice, and the healing process was analyzed. The macroscopic wound closure rate declined in the PLCε KO mice on days 3, 4, and 5 after wounding, following the decreased expression of interleukin (IL)-6, chemokine (C-X-C motif) ligand (Cxcl)-1, Cxcl-2, and chemokine (C-C motif) ligand (Ccl) 20. The proliferation rate of epidermal keratinocytes was not affected by PLCε, but silencing of PLCε resulted in the delayed migration of keratinocytes. Moreover, the scars were found to be much smaller in the PLCε KO mice than in the WT mice. The mRNA expression of Ccl20, collagen (Col) 6a1, and Col17a1 decreased in the PLCε KO mice. These results were in agreement with a previous hypothesis that PLCε might delay the early stage of cutaneous wound healing by inhibiting the migration of keratinocytes, and decrease the expression of Col6a1, Col17a1, and Ccl20 by inhibiting the inflammatory response to reduce scar formation. This study shed light on a novel role of PLCε in wound healing and provided new therapeutic approaches to target PLCε for diminishing scar formation after injury.

The Arf GAP AGAP2 interacts with ß-arrestin2 and regulates ß2-adrenergic receptor recycling and ERK activation.

AGAP2 [Arf (ADP-ribosylation factor) GAP (GTPase-activating protein) with GTP-binding-protein-like, ankyrin repeat and PH (pleckstrin homology) domains] is a multidomain Arf GAP that was shown to promote the fast recycling of transferrin receptors. In the present study we tested the hypothesis that AGAP2 regulates the trafficking of ß2-adrenergic receptors. We found that AGAP2 formed a complex with ß-arrestin1 and ß-arrestin2, proteins that are known to regulate ß2-adrenergic receptor signalling and trafficking. AGAP2 co-localized with ß-arrestin2 on the plasma membrane, and knockdown of AGAP2 expression reduced plasma membrane association of ß-arrestin2 upon ß2-adrenergic receptor activation. AGAP2 also co-localized with internalized ß2-adrenergic receptors on endosomes, and overexpression of AGAP2 slowed accumulation of ß2-adrenergic receptor in the perinuclear recycling endosomes. In contrast, knockdown of AGAP2 expression prevented the recycling of the ß2-adrenergic receptor back to the plasma membrane. In addition, AGAP2 formed a complex with endogenous ERK (extracellular-signal-regulated kinase) and overexpression of AGAP2 potentiated ERK phosphorylation induced by ß2-adrenergic receptors. Taken together, these results support the hypothesis that AGAP2 plays a role in the signalling and recycling of ß2-adrenergic receptors.

Efficacy and safety of rectal 5-aminosalicylic acid versus corticosteroids in active distal ulcerative colitis: a systematic review and network meta-analysis.

Topical 5-aminosalicylic acid (5-ASA) and corticosteroids are used frequently in the treatment of active distal ulcerative colitis (UC). Our study aimed to determine the efficacy and safety of different topical drugs used to treat active distal UC. A random-effects model within a Bayesian framework was utilized to compare treatment effects and safety as odds ratios (ORs) with corresponding 95% credible intervals (CrI). The surface under the cumulative ranking area (SUCRA) and median rank (MR) with corresponding 95% CrI were calculated to rank the treatment outcomes. In the induction of clinical and endoscopic remission, most regimens showed significant advantages over placebo except topical budesonide 0.5 mg/d and hydrocortisone 100 mg/d. According to SUCRA and MR values, rectal 5-ASA 1.5 to 2.0 g/d + Beclomethasone dipropionate (BDP) 3 mg/d rendered the highest probability of being the best regimen to achieve clinical and endoscopic remission, followed by the separate use of 5-ASA 4 g/d and BDP 3 mg/d. The occurrence of adverse events was not significantly different between each treatments and placebo. In conclusion, the combined use of topical 5-ASA and BDP proved to be the best choice for active distal UC and further well-designed researches are warranted to assess its efficacy and safety.

Efficacy and safety of medical therapy for low bone mineral density in patients with Crohn disease: A systematic review with network meta-analysis.

BACKGROUND: Low bone mineral density (BMD) is a frequent complication of inflammatory bowel disease (IBD), particularly in patients with Crohn disease (CD). The aim of our study is to determine the efficacy and safety of different drugs used to treat low BMD in patients with CD. METHODS: PUBMED/MEDLINE, EMBASE, and Cochrane Central Register of Controlled Trials were searched for eligible studies. A random-effects model within a Bayesian framework was applied to compare treatment effects as standardized mean difference (SMD) with their corresponding 95% credible interval (CrI), while odds ratio (OR) was applied to compare adverse events with 95% CrI. The surface under the cumulative ranking area (SUCRA) was calculated to make the ranking of the treatments for outcomes. RESULTS: Twelve randomized controlled trials (RCTs) were eligible. Compared with placebo, zoledronate (SMDs 2.74, 95% CrI 1.36-4.11) and sodium-fluoride (SMDs 1.23, 95% CrI 0.19-2.26) revealed statistical significance in increasing lumbar spine BMD (LSBMD). According to SUCRA ranking, zoledronate (SUCRA = 2.5%) might have the highest probability to be the best treatment for increasing LSBMD in CD patients among all agents, followed by sodium-fluoride (27%). For safety assessment, the incidence of adverse events (AEs) demonstrated no statistical difference between agents and placebo. The corresponding SUCRA values indicated that risedronate (SUCRA = 77%) might be the most safe medicine for low BMD in CD patients and alendronate ranked the worst (SUCRA = 16%). CONCLUSIONS: Zoledronate might have the highest probability to be the best therapeutic strategy for increasing LSBMD. For the safety assessment, risedronate showed the greatest trend to decrease the risk of AEs. In the future, more RCTs with higher qualities are needed to make head-to-head comparison between 2 or more treatments.

Temporal and spatial distributions of PBDEs in atmosphere at Shanghai rural regions, China.

Atmospheric samples were collected using polyurethane foam (PUF) passive air sampling device for every 3 months from June 2012 to May 2013 in Shanghai rural regions in order to investigate the concentrations, profiles, spatial distributions, and seasonal variations of polybrominated diphenyl ethers (PBDEs). Twelve PBDE congeners (BDE-17, BDE-28, BDE-47, BDE-49, BDE-66, BDE-85, BDE-99, BDE-100, BDE-138, BDE-153, BDE-154, and BDE-183) were measured and analyzed by GC-MS. The results showed that detectable PBDEs were examined in all air samples, which indicated that these pollutants are widespread in the research areas. The ∑12PBDE concentrations in Shanghai rural air ranged from 4.49 to 77.5 pg m-3, with mean value up to 26.7 pg m-3. The highest concentration was found at Jinshan sampling site in summer (from June to August in 2012). Furthermore, among the PBDE compounds investigated, the most frequently detected and the major congeners were BDE-17, BDE-28, BDE-47, and BDE-99. And the lower brominated diphenyl ethers (accounting for 75.0%) were the majority of the PBDE congeners. Finally, the result of principal component analysis (PCA) revealed that the lower and higher brominated diphenyl ethers in Shanghai rural regions were emitted from different pollutant sources.

Arf GTPase-activating protein AGAP2 regulates focal adhesion kinase activity and focal adhesion remodeling.

Focal adhesions are specialized sites of cell attachment to the extracellular matrix where integrin receptors link extracellular matrix to the actin cytoskeleton, and they are constantly remodeled during cell migration. Focal adhesion kinase (FAK) is an important regulator of focal adhesion remodeling. AGAP2 is an Arf GTPase-activating protein that regulates endosomal trafficking and is overexpressed in different human cancers. Here we examined the regulation of the FAK activity and the focal adhesion remodeling by AGAP2. Our results show that FAK binds the pleckstrin homology domain of AGAP2, and the binding is independent of FAK activation following epidermal growth factor receptor stimulation. Overexpression of AGAP2 augments the activity of FAK, and concordantly, the knockdown of AGAP2 expression with RNA interference attenuates the FAK activity stimulated by epidermal growth factor or platelet-derived growth factor receptors. AGAP2 is localized to the focal adhesions, and its overexpression results in dissolution of the focal adhesions, whereas knockdown of its expression stabilizes them. The AGAP2-induced dissolution of the focal adhesions is independent of its GTPase-activating protein activity but may involve its N-terminal G protein-like domain. Our results indicate that AGAP2 regulates the FAK activity and the focal adhesion disassembly during cell migration.

[Construction and expression of the eukaryotic expression plasmids of human TSHR extracellular domain].

AIM: To construct the eukaryotic expression plasmids of hTSHR extracellular domain and study their expression in CHO cells. METHODS: The human TSHR extracellular domain cDNAs, which were 188-403 bp and 407-904 bp, were amplified from human normal thyroid by RT-PCR. Two fragments were inserted into pcDNA3.1(D)/V5-His-TOPO.Then the recombinant plasmids pcDNA3.1-hTSHRf and pcDNA3.1-hTSHRe were transfected into CHO cells by Lipofectin after they were identified by restricting enzyme HindIII digestion analysis, PCR amplifying and DNA sequencing. RT-PCR and Western blot analysis were used to analyse hTSHR expression on mRNA and at protein levels. RESULTS: Two bands of 220 bp and 540 bp were amplified from CHO cells transfected by the recombinant plasmids pcDNA3.1-hTSHRf and pcDNA3.1-hTSHRe, respectively. Western blot analysis revealed that CHO cells transfected by pcDNA3.1-hTSHRf and pcDNA3.1-hTSHRe had strong bands with molecular weight of about 11 900 and 23 600, respectively. CONCLUSION: The recombinant plasmids have been successfully constructed. The transcription on CHO cells transfected by the recombinant plasmids has been proved by RT-PCR and eukaryotic expression has been confirmed by Western blot analysis. Our research will contribute to further study on gene expression in vivo and the establishment of animal models of Graves' disease.