Differences in the effects and action modes of gut commensals against dextran sulfate sodium-induced intestinal inflammation

Inflammatory bowel disease(IBD)is a complex relapsing inflammatory disease in the gut and is driven by complicated host-gut microbiome interactions.Gut commensals have shown different functions in IBD prevention and treatment.To gain a mechanistic understanding of how different commensals affect intestinal inflammation,we compared the protective effects of 6 probiotics(belonging to the genera Akkermansia,Bifidobacterium,Clostridium,and Enterococcus)on dextran sulfate sodium(DSS)-induced colitis in mice with or without gut microbiota.Anti-inflammatory properties(ratio of interleukin(IL)-10 and IL-12)of these strains were also evaluated in an in vitro mesenteric lymph nodes(MLN)co-culture system.Results showed that 4 probiotics(belonging to the species Bifidobacterium breve,Bifidobacterium bifidum,and Enterococcus faecalis)can alleviate colitis in normal mice.The probiotic strains differed in regulating the intestinal microbiota,cytokines(IL-10,IL-1βand interferon(IFN)-γ),and tight junction function(Zonulin-1 and Occludin).By constrast,Akkermansia muciniphila AH39 and Clostridium butyricum FHuNHHMY49T1 were not protective.Interestingly,B.breve JSNJJNM2 with high anti-inflammatory potential in the MLN model could relieve colitis symptoms in antibiotic cocktail(Abx)-treated mice.Meanwhile,E.faecalis FJSWX25M1induced low levels of cytokines in vitro and showed no beneficial effects.Therefore,we provided insight into the clinical application of probiotics in IBD treatment.

Alkaline sphingomyelinase deficiency impairs intestinal mucosal barrier integrity and reduces antioxidant capacity in dextran sulfate sodium-induced colitis

BACKGROUND Ulcerative colitis is a chronic inflammatory disease of the colon with an unknown etiology.Alkaline sphingomyelinase(alk-SMase)is specifically expressed by intestinal epithelial cells,and has been reported to play an anti-inflammatory role.However,the underlying mechanism is still unclear.AIM To explore the mechanism of alk-SMase anti-inflammatory effects on intestinal barrier function and oxidative stress in dextran sulfate sodium(DSS)-induced colitis.METHODS Mice were administered 3%DSS drinking water,and disease activity index was determined to evaluate the status of colitis.Intestinal permeability was evaluated by gavage administration of fluorescein isothiocyanate dextran,and bacterial translocation was evaluated by measuring serum lipopolysaccharide.Intestinal epithelial cell ultrastructure was observed by electron microscopy.Western blotting and quantitative real-time reverse transcription-polymerase chain reaction were used to detect the expression of intestinal barrier proteins and mRNA,respectively.Serum oxidant and antioxidant marker levels were analyzed using commercial kits to assess oxidative stress levels.RESULTS Compared to wild-type(WT)mice,inflammation and intestinal permeability in alk-SMase knockout(KO)mice were more severe beginning 4 d after DSS induction.The mRNA and protein levels of intestinal barrier proteins,including zonula occludens-1,occludin,claudin-3,claudin-5,claudin-8,mucin 2,and secretory immunoglobulin A,were significantly reduced on 4 d after DSS treatment.Ultrastructural observations revealed progressive damage to the tight junctions of intestinal epithelial cells.Furthermore,by day 4,mitochondria appeared swollen and degenerated.Additionally,compared to WT mice,serum malondialdehyde levels in KO mice were higher,and the antioxidant capacity was significantly lower.The expression of the transcription factor nuclear factor erythroid 2-related factor 2(Nrf2)in the colonic mucosal tissue of KO mice was significantly decreased after DSS treatment.mRNA levels of Nrf2-regulated downstream antioxidant enzymes were also decreased.Finally,colitis in KO mice could be effectively relieved by the injection of tertiary butylhydroquinone,which is an Nrf2 activator.CONCLUSION Alk-SMase regulates the stability of the intestinal mucosal barrier and enhances antioxidant activity through the Nrf2 signaling pathway.

Resveratrol alleviates intestinal mucosal barrier dysfunction in dextran sulfate sodium-induced colitis mice by enhancing autophagy

BACKGROUND Intestinal mucosal barrier dysfunction plays an important role in the pathogenesis of ulcerative colitis(UC).Recent studies have revealed that impaired autophagy is associated with intestinal mucosal dysfunction in the mucosa of colitis mice.Resveratrol exerts anti-inflammatory functions by regulating autophagy.AIM To investigate the effect and mechanism of resveratrol on protecting the integrity of the intestinal mucosal barrier and anti-inflammation in dextran sulfate sodium(DSS)-induced ulcerative colitis mice.METHODS Male C57BL/6 mice were divided into four groups:negative control group,DSS model group,DSS+resveratrol group,and DSS+5-aminosalicylic acid group.The severity of colitis was assessed by the disease activity index,serum inflammatory cytokines were detected by enzyme-linked immunosorbent assay.Colon tissues were stained with haematoxylin and eosin,and mucosal damage was evaluated by mean histological score.The expression of occludin and ZO-1 in colon tissue was evaluated using immunohistochemical analysis.In addition,the expression of autophagy-related genes was determined using reverse transcription-polymerase chain reaction and Western-blot,and morphology of autophagy was observed by transmission electron microscopy.RESULTS The resveratrol treatment group showed a 1.72-fold decrease in disease activity index scores and 1.42,3.81,and 1.65-fold decrease in the production of the inflammatory cytokine tumor necrosis factor-α,interleukin-6 and interleukin-1β,respectively,in DSS-induced colitis mice compared with DSS group(P<0.05).The expressions of the tight junction proteins occludin and ZO-1 in DSS model group were decreased,and were increased in resveratrol-treated colitis group.Resveratrol also increased the levels of LC3B(by 1.39-fold compared with DSS group)and Beclin-1(by 1.49-fold compared with DSS group)(P<0.05),as well as the number of autophagosomes,which implies that the resveratrol may alleviate intestinal mucosal barrier dysfunction in DSS-induced UC mice by enhancing autophagy.CONCLUSION Resveratrol treatment decreased the expression of inflammatory factors,increased the expression of tight junction proteins and alleviated UC intestinal mucosal barrier dysfunction;this effect may be achieved by enhancing autophagy in intestinal epithelial cells.

Sodium selenite ameliorates dextran sulfate sodiuminduced chronic colitis in mice by decreasing Th1, Th17, and γδT and increasing CD4(+)CD25(+) regulatory T-cell responses

AIM To assess the effect of sodium selenite on the severity of dextran sulfate sodium(DSS)-induced colitis in C57BL/6 mice.METHODS Mice were randomly divided into four groups(n = 10/group): normal group, selenium(Se) group, chronic colitis group, and Se + chronic colitis group. The mice were sacrificed on day 26. Survival rates, clinical symptoms, colon length, and histological changes were determined. The percentages and absolute numbers of immune system cells in the lamina propria lymphocytes(LPL) of the colon, the expression of m RNA in colon tissue, and the concentrations of Th1, Th17, and Treg cytokines in LPL from the large intestine, were measured.RESULTS Se significantly ameliorated the symptoms of colitis and histological injury(P < 0.05 each), increasing the proportions of neutrophils and CD4+ CD25+ T cells(P < 0.05 each) and decreasing the proportions of γδT cells, CD4+, CD4+CD44+, and CD4+ CD69+ T cells in LPL(P < 0.05 each). Moreover, Se reduced the expression of IL-6, IFN-γ, IL-17 A, IL-21, T-bet, and RORγt(P < 0.05 each), but enhanced the expression of IL-10 and Foxp3(P < 0.05 each). CONCLUSION These results suggest that Se protects against DSSinduced chronic colitis perhaps by increasing the number of CD4(+)CD25(+) Tregs that suppress the secretion of proinflammatory cytokines and populations of Th1, Th17, and γδT cells.

Differential analysis of intestinal microbiota and metabolites in mice with dextran sulfate sodium-induced colitis

BACKGROUND Intestinal micro-ecological imbalances impair the intestinal barrier and induce intestinal inflammation,for example,ulcerative colitis(UC).According to the latest research,abnormalities in intestinal microbiota structure and their metabolites play a dominant role in UC progression;in addition,they could affect the mucus barrier based on different factors.Although numerous studies have confirmed the important role of intestinal microbiota in UC pathogenesis,the intricate connection between microbiota and metabolites and mucus barrier in UC occurrence remains unclear,and correlation analyses of differential microbiota and their metabolites under UC are relatively scarce.AIM To reveal the differential intestinal microbiota and metabolites in UC pathogenesis and explore more sensitive biomarker compositions.METHODS We used the antibiotic combination method to establish intestinal pseudo-aseptic mice;afterward,dextran sulfate sodium(DSS)was applied to establish an acute experimental colitis mice model.Colitis severity,assessed based on disease activity index,colorectal length,colorectal wet weight,and histological lesions,and mucus-related staining(mucopolysaccharide alcian blue and immunofluorescence of mucin),was compared between the pseudo-aseptic and bacterial colitis mice.Finally,differential intestinal microbiota,metabolites,and their association and correlations,were analyzed by 16s rDNA sequencing in combination with non-targeted metabolomics,through gas chromatography-mass spectrometry.RESULTS Compared with the pseudo-aseptic mice,intestinal bacteria positive mice were more severely ill and their intestinal mucus loss was more pronounced in DSS-induced colitis(P<0.05),suggesting that different microbiota and metabolites could cause the different degrees of colitis.Subsequently,we observed that in addition to Klebsiella,and Bacteroides,which were widely associated with colitis,Candidatus Stoquefichus,Anaerobiospirillum,Muribaculum,and Negativibacillus may be involved in protection against colitis.Furthermore,differential metabolites of the microbiota were mainly enriched in the synthesis-related pathways of key structural sequences of mucin.In combination with the mucin-related staining and immunofluorescence results,the findings indicate that the differential microbiota and their metabolites potentially regulate the composition and function of mucus under colitis.CONCLUSION Microbiota and their metabolites are major factors regulating the composition and function of mucus,in turn influencing the function and structure of intestinal mucus barrier under colitis.The different microbiota and metabolites identified in the present study could be novel biomarkers for colitis.

Dramatic Changes of Matrix Metalloproteinases-7 and Lysozyme in the Ulcerative Colitis of Mice Induced by Dextran Sulfate Sodium

Ulcerative colitis （UC） is a lifelong illness with profound emotional and social impacts, and could cause serious damage to large intestine, especially in colon. However, the pathogenesis of UC remained unclear. The present study attempts to find out the role of matrix metalloproteinases-7 （MMP-7） and lysozyme in the pathogenesis of UC through a mice model induced by dextran sulfate sodium （DSS）. The UC model was evaluated both by disease activity index （DAI） and the intestinal histopathology. The results show that there is a high correlation between the DAI score and the pathological changes of colon. Interleukin-6 （IL-6） serum levels and large intestinal fluids levels in UC mice are always higher than that of the control groups, which might be associated with the degree of the inflammation damage in the colon. The change tendency of the MMP-7 mRNA and protein expressions are both up-regulated firstly and then down-regulated from 1 to 5 d in the colon, but only the MMP-7 protein is up-regulated at 7 d again. The up-regulated MMP-7 levels in the early stage of UC may play a protective role through the activated defensins, while the down-regulated levels in the mid-later stage of UC may be connected with the severe lesions in the colon. However, the up-regulated MMP-7 levels in the later stage of UC in the colon may also contribute to the tissue repair or be served as a marker to CRC （colorectal cancer）. The distribution of lysozyme protein indicates that there may be Paneth-like cells in the colon. Both the changes of MMP-7 and lysozyme in the small intestine may play a protective role for the safe environment of the whole gut, especially to the colon of UC.

The Involvement of Ca^2＋ Signal Pathways in Distal Colonic Myocytes in a Rat Model of Dextran Sulfate Sodium-induced Colitis

Background： Disrupted Ca2＋ homeostasis contributes to the development of colonic dysmotility in ulcerative colitis （UC）, but the underlying mechanisms are unknown. This study aimed to examine the alteration of colonic smooth muscle （SM） Ca2＋ signaling and Ca2＋ handling proteins in a rat model of dextran sulfate sodium （DSS）-induced UC. Methods： Male Sprague-Dawley rats were randomly divided into control （n = 18） and DSS （n = 17） groups. Acute colitis was induced by 5% DSS in the drinking water for 7 days. Contractility of colonic SM strips （controls, n = 8 and DSS, n = 7） was measured in an organ bath. Cytosolic resting Ca2＋ levels （n = 3 in each group） and Ca2＋ transients （n = 3 in each group） were measured in single colonic SM cells. Ca2＋ handling protein expression was determined by Western blotting （n = 4 in each group）. Differences between control and DSS groups were analyzed by a two-sample independent t-test. Results： Average tension and amplitude of spontaneous contractions of colonic muscle strips were significantly enhanced in DSS-treated rats compared with controls （1.25 ± 0.08 g vs. 0.96 - 0.05 g, P = 0.007; and 2.67 - 0.62 g vs. 0.52 ±0.10 g, P= 0.013）. Average tensions of carbachol-evoked contractions were much weaker in the DSS group （1.08 ±0.10 g vs. 1.80 ±0.19 g, P = 0.006）. Spontaneous Ca2＋ transients were observed in more SM cells from DSS-treated rats （15/30 cells） than from controls （5/36 cells）. Peak caffeine-induced intracellular Ca2＋ release was lower in SM cells of DSS-treated rats than controls （0.413 ±0.046 vs. 0.548 ±0.041, P = 0.033）. Finally, several Ca2＋ handling proteins in colonic SM were altered by DSS treatment, including sarcoplasmic reticulum calcium-transporting ATPase 2a downregulation and phospholamban and inositol 1,4,5-trisphosphate receptor 1 upregulation. Conclusions： Impaired intracellular Ca2＋ signaling of colonic SM, caused by alteration of Ca2＋ handing proteins, contribute to colonic dysmotility in DSS-induced UC.

New approach of medicinal herbs and sulfasalazine mixture on ulcerative colitis induced by dextran sodium sulfate

BACKGROUND Sulfasalazine has been used as a standard-of-care in ulcerative colitis for decades,however,it results in severe adverse symptoms,such as hepatotoxicity,blood disorders,male infertility,and hypospermia.Accordingly,the new treatment strategy has to enhance pharmacological efficacy and stimultaneously minimize side effects.AIM To compare the anti-inflammatory action of sulfasalazine alone or in combination with herbal medicine for ulcerative colitis in a dextran sodium sulfate(DSS)-induced colitis mouse model.METHODS To induce ulcerative colitis,mice received 5%DSS in drinking water for 7 d.Animals were divided into five groups(n=9 each)for use as normal(non-DSS),DSS controls,DSS+sulfasalazine(30 mg/kg)-treatment experimentals,DSS+sulfasalazine(60 mg/kg)-treatment experimentals,DSS+sulfasalazine(30 mg/kg)+Citrus unshiu peel and Bupleuri radix mixture(30 mg/kg)(SCPB)-treatment experimentals.RESULTS The SCPB treatment showed an outstanding effectiveness in counteracting the ulcerative colitis,as evidenced by reduction in body weight,improvement in crypt morphology,increase in antioxidant defenses,down-regulation of proinflammatory proteins and cytokines,and inhibition of proteins related to apoptosis.CONCLUSIONSCPB may represent a promising alternative therapeutic against ulcerative colitis,without inducing adverse effects.

Protective Effect of Total Alkaloids of Sophora Alopecuroides on Dextran Sulfate Sodium-Induced Chronic Colitis

Objective：To investigate the effect of total alkaloids of Sophora alopecuroides（TASA） on dextran sulfate sodium（DSS）-induced colitis in mice.Methods：Chronic experimental colitis was induced by administration of 4 cycles of 4%DSS.Fifty mice were randomly distributed into 4 groups（normal,DSS,DSS/high-dose TASA, and DSS/low-dose TASA groups） by a random number table with body weight stratification.Mice in the normal group（n=11） and DSS-induced colitis control group（n=15） received control treatment of 20 mL/kg distilled water; DSS plus TASA high- and low-dose groups（n=12 each） were treated with TASA solution（20 mL/kg） at the doses of 60 mg/kg and 30 mg/kg,respectively.The severity of colitis was assessed on the basis of clinical signs, colon length,and histology scores.Moreover,secretory immunoglobulin A（slgA） and haptoglobin（HP） were analyzed by enzyme linked immunosorbent assay;intercellular adhesion molecule 1（ICAM-1） and macrophage-migration inhibitory factor（MIF） gene expressions were analyzed by quantitative reverse transcriptase realtime polymerase chain reaction（qRT-PCR） using SYBA greenⅠ;and nuclear factorκB（NF-κB） expression and activation and p65 interaction with the promoter of ICAM-1 gene were assessed by Western blotting and chromatin immunoprecipitation assay.Results：TASA administration significantly attenuated the damage and substantially reduced HP elevation and maintained the level of cecum slgA.TASA inhibited the ICAM-1 gene expression and had no effect on MIF gene expression.Also,TASA was able to reduce phospho-lκBα（p-lκBα） protein expression;however,it had no effect on the activation of IκB kinaseα（IKKα） and inhibitor of NF-κBα（IκBα）.Moreover,TASA inhibited the p65 recruitment to the ICAM-1 gene promoter.Conclusions：TASA had a protective effect on DSS-induced colitis.Such effect may be associated with its inhibition of NF-κB activation and blockade of NF-κB-regulated transcription activation of proinflammatory mediator gene.

Inhibitory Effect of Recombinant IL-25 on the Development of Dextran Sulfate Sodium-Induced Experimental Colitis in Mice

The role of interleukin 25 （IL-25） in a number of human diseases still has not been extensively studied, here we attempt to evaluate the role of recombinant IL-25 （rIL-25） in the development of dextran sulfate sodium （DSS）- induced experimental colitis. Acute colitis was induced in female C57BL/6 mice by oral administration of 2.5% DSS in drinking water ad libitum. At the same time as the start of DSS exposure, mice were injected intraperitoneally with 0.4 ＋tg of rIL-25 or PBS. Then disease activity index （DAI）, histological changes and survival rate were observed. The levels of IL-17, IL-23, and TGF-β1 in colon tissues were determined by ELISA, and the production of IL-17 by CD4＋/CD8＋ T cells was detected by intracellular flow cytometry. In contrast to the DSS treated mice, DSS ＋ rIL-25 treated mice displayed a lower DAI, limited histological changes and prolonged survival. The levels of IL-23 and TGF-β1 were significantly elevated in the DSS ＋ rIL-25 treated mice compared to the DSS treated mice. There was no significant difference in the production of IL-17 in colon tissues and CD4＋/CD8＋ T cells between the DSS ＋ rIL-25 treated mice and DSS treated mice. Our findings suggest the role of IL-25 in inhibiting development and progression of acute colitis in DSS-induced mouse colitis model. Cellular ＆ Molecular Immunology. 2008;5（6）：425-431.

Changes of CD8＋ T cells in dextran sulfate sodium-induced colitis mice pretreated with oral immune regulation

Background It has been reported that CD8＋ regulatory cells could be induced upon oral tolerance. The purpose of this study was to investigate the changes of CD8a＋ T cells in dextran sulfate sodium （DSS）-induced colitis mice pretreated by oral immune regulation. Methods The effects of five low oral doses of colitis-extracted proteins （CEP） on colitis were evaluated by clinical manifestation and histological lesions. The percentages of CD8a＋ T cells gating on CD3＋ T cells were evaluated in the gut-associated lymphoid tissues （GALT） and the spleens by flow cytometry. Differences between the two groups were compared by Student＇s t test or Mann-Whitney U test. Results Compared to bovine serum albumin （BSA）-fed control mice, administration of CEP resulted in marked alleviation of colitis. The proportion of CDSa＋ T ceils, not only in intraepithelial lymphocytes （IELs） and lamina propria lymphocytes （LPLs） of the large intestine （LI） but also in spleen from CEP-fed colitis mice, was significantly higher than that from BSA-fed colitis mice （LI-IELs： （71.5±5.4）% vs. （60.1±4.3）%, P 〈0.01; LI-LPLs： （60.7±5.2）% vs. （51.9±4.7）%, P 〈0.01; spleen： （24.1±3.6）% vs. （20.3±4.1）%, P 〈0.05; n=8）. Mucosal repair in repair-period mice five days after termination of DSS treatment was also accompanied by an increase of CD8a＋ T cells in large intestinal mucosal lymphocytes （LI-IELs： （72.1±3.7）% vs. （61.5±4.5）%, P 〈0.01; LI-LPLs： （62.1±5.7）% vs. （52.7±3.6）%, P 〈0.01; n=8）. The proportion of CD3＋ T cells increased in Peyer＇s patches （PPs） and decreased in mesenteric lymph nodes （MLNs） from colitis mice compared to untreated mice, whereas the change pattern of CD3＋T cells in PPs and MLNs from CEP-fed colitis mice was just on the contrary. Conclusion Improvement of DSS-induced colitis resulted from oral immune regulation is associated with an increase in CD8a＋T cells in spleen and large intestinal mucosa.

Dextran sodium sulfate colitis murine model: An indispensable tool for advancing our understanding of inflammatory bowel diseases pathogenesis

Inflammatory bowel diseases(IBD),including Crohn's disease and ulcerative colitis,are complex diseases that result from the chronic dysregulated immune response in the gastrointestinal tract. The exact etiology is not fully understood,but it is accepted that it occurs when an inappropriate aggressive inflammatory respon-se in a genetically susceptible host due to inciting environmental factors occurs. To investigate the path-ogenesis and etiology of human IBD,various animal models of IBD have been developed that provided indispensable insights into the histopathological and morphological changes as well as factors associated with the pathogenesis of IBD and evaluation of therapeutic options in the last few decades. The most widely used experimental model employs dextran sodium sulfate(DSS) to induce epithelial damage. The DSS colitis model in IBD research has advantages over other various chemically induced experimental models due to its rapidity,simplicity,reproducibility and controllability. In this manuscript,we review the newer publicized advances of research in murine colitis models that focus upon the disruption of the barrier function of the intestine,effects of mucin on the development of colitis,alterations found in microbial balance and resultant changes in the metabolome specifically in the DSS colitis murine model and its relation to the pathogenesis of IBD.

Fluoxetine Ameliorates the Aggravation of UC Symptoms in C57BL/6 Mice Induced by CUMS

Objective Patients with chronic ulcerative colitis(UC)often have mental symptoms such as depression and anxiety,and stress can lead to gastrointestinal diseases.However,the correlation between mental stress and UC is unclear.In this paper,chronic unpredictable mild stress(CUMS)was utilized to evaluate the involvement of mental factors in the pathogenesis of UC.Methods The CUMS model was used to evaluate the direct/indirect involvement of mental factors in the pathogenesis of UC.The behavior was evaluated by the open field,forced swimming,and tail suspension tests.Body weight,the disease activity index(DAI)score,colon length,and HE staining of colon tissue were used to evaluate the action of CUMS and fluoxetine.Results The results showed that weight loss and the DAI score increased in CUMS mice,but they had no meaningful effect on colon length and morphological structure of colon tissue.However,CUMS aggravated dextran sulfate sodium(DSS)-induced colon length shortening and colon morphological structure damage.Fluoxetine significantly improved the DAI score,shortened colon length,and damaged morphology and structure of the colons induced by CUMS combined with DSS in mice.Fluoxetine also decreased the level of IL-6 in the serum and the TNF-αand IFN-γlevels of colon tissue.Fluoxetine simultaneously improved behavioral abnormalities induced by CUMS combined with DSS in mice.Conclusion CUMS aggravated the UC symptoms induced by DSS,and fluoxetine could improve the UC symptoms due to its improvement in the inflammatory level and behavioral abnormalities.

Intravenous vs intraperitoneal mesenchymal stem cells administration:What is the best route for treating experimental colitis?

AIM:To investigate the therapeutic effects of mesenchymal stem cells(MSCs)transplanted intraperitoneally andintravenously in a murine model of colitis.METHODS:MSCs were isolated from C57BL/6 mouse adipose tissue.MSC cultures were analyzed according to morphology,cellular differentiation potential,and surface molecular markers.Experimental acute colitis was induced in C57BL/6 mice by oral administration of2%dextran sulfate sodium(DSS)in drinking water ad libitum from days 0 to 7.Colitis mice were treated with1×106 MSCs via intraperitoneal or intravenous injection on days 2 and 5.The disease activity index was determined daily based on the following parameters:weight loss,stool consistency and presence of blood in the feces and anus.To compare morphological and functional differences in tissue regeneration between different MSC injection modalities,mice were euthanized on day 8,and their colons were examined for length,weight,and histopathological changes.Inflammatory responses were determined by measuring the levels of different serum cytokines using a CBA Th1/Th2/Th17 kit.Apoptotic rates were evaluated by terminal deoxynucleotidyl transferase-mediated d UDPbiotin nick end labeling assay.RESULTS:Intravenous infusion of MSCs was more effective than intraperitoneal treatment(P<0.001)in reducing the clinical and histopathologic severity of colitis,which includes weight loss,diarrhea and inflammation.An histological evaluation demonstrated decreased colonic inflammation based on reduced crypt loss and reduced infiltration of inflammatory cells.This therapeutic effect was most likely mediated by the down-regulation of pro-inflammatory cytokines[interleukin(IL)-6 and tumor necrosis factor(TNF)];and by the up-regulation of anti-inflammatory cytokines(IL-10 and IL-4).Intravenous transplantation alsoinduced high levels of IFN that lead to activation of the immunosuppressive activity of the MSCs,which did not occur with intraperitoneal transplantation(P=0.006).An increase in apoptotic T cells was observed after intravenous,but not intraperitoneal,MSC infusion,suggesting that MSCs can induce apoptosis in resistant T cells in colonic inflammation(P=0.027).CONCLUSION:Our results demonstrate that intravenous treatment is a superior method for reducing colon inflammation compared with intraperitoneal therapy.

Anti-inflammatory activity of Alpinia officinarum hance on rat colon inflammation and tissue damage in DSS induced acute and chronic colitis models

The purpose of this study was to investigate the possible prophylactic effects of Alpinia officinarum hance on experimentally induced acute and chronic colitis models,in-vivo and in-vitro.Acute and chronic colitis were induced in Male Wistar rats by administration of Dextran Sulfate Sodium(DSS)in drinking water.DSS induction exhibited colon shrinkage,increased the Disease Activity Index(DAI)score,increased the levels of inflammatory markers and caused severe anemia.DSS induced animals,co-treated with the hexane extract of Alpinia officinarum(HEAO)(200 mg/kg body wt),effectively suppressed colonic injury that was evidenced by the reduced DAI score,colon weight/length ratio,histological damage,proinflammatory markers and MPO activity.Further,it restored the colonic antioxidants near to normal levels by regulating the oxidative stress via attenuation of lipid peroxidation.Our results revealed that the degree of colitis caused by the administration of DSS was significantly attenuated by HEAO.In addition,the in-vitro study showed that HEAO treatment inhibited the proliferation of HT-29 cells and down regulated the mRNA expression of NF-B and COX-2.Taken together,these results suggest that HEAO is a promising anti-oxidant and anti-inflammatory agent that support its possible therapeutic role in the treatment of colitis.

Lactobacillus acidophilus alleviates pouchitis after ileal pouch-anal anastomosis in rats

AIM To assess the therapeutic potential of Lactobacillus acidophilus(LA) for the treatment of pouchitis in a rat model.METHODS Sprague Dawley rats underwent proctocolectomy and ileal pouch-anal anastomosis followed by administration of dextran sulfate sodium(DSS) to induce pouchitis. Rats with pouchitis were randomly divided into three groups: no intervention(NI), normal saline(NS, 3 m L/d normal saline for 7 d), and LA(3 m L/d LA at 1× 1010 colony-forming units for 7 d). General body condition was recorded and pouch specimens were obtained for histological examination. m RNA expression levels of interleukin(IL)-1β, IL-6, IL-10, and tumor necrosis factor-α were determined by RT-PCR. Zonula occludens protein 1(ZO-1) levels were measured by immunohistochemistry. RESULTS LA reduced weight loss associated with pouchitis(P < 0.05) and improved the symptoms of pouchitis in rats. Compared with the NI and NS groups, rats in the LAgroup showed earlier disappearance of hematochezia(6.17 ± 0.75, 6.50 ± 0.55, 3.17 ± 0.75, P < 0.05) and higher fecal scores(2.67 ± 0.48, 2.50 ± 0.51, 4.42 ± 0.50, respectively, P < 0.05). Histological scores were also lower in the LA group compared with the other two groups(7.17 ± 0.98, 8.00 ± 0.89, 4.00 ± 0.89, respectively, P < 0.05). m RNA expression levels of IL-1β, IL-6, and tumor necrosis factor-α were significantly reduced, while IL-10 m RNA levels were significantly increased in the LA group(P < 0.05, respectively). ZO-1 protein levels were also significantly increased after administration of LA(P < 0.05). CONCLUSION LA alleviates pouchitis induced by DSS after ileal pouchanal anastomosis by decreasing pro-inflammatory factors and increasing anti-inflammatory factors, and restoring ZO-1 expression in the mucosa.

Lysophosphatidic acid increases SLC26A3 expression in inflamed intestine and reduces diarrheal severity in C57BL/6 mice with dextran-sodium-sulfate-induced colitis

Background Diarrhea is a common clinical feature of ulcerative colitis resulting from unbalanced intestinal fluid and salt absorption and secretion.The Cl-/HCO3-exchanger SLC26A3 is strongly expressed in the mid-distal colon and plays an essential role in colonic Cl-absorption and HCO3-secretion.Sic26a3 expression is up-regulated by lysophosphatidic acid （LPA） in vitro.Our study was designed to investigate the effects of LPA on SLC26A3 expression and the diarrheal phenotype in a mouse colitis model.Methods Colitis was induced in C57BL/6 mice by adding 4％ of dextran sodium sulfate （DSS） to the drinking water.The mice were assigned to LPA treatment DSS group,phosphate-buffered saline （PBS） treatment DSS group,DSS only group and untreated mice with a completely randomized design.Diarrhea severity was evaluated by measuring mice weight,disease activity index （DAI）,stool water content and macroscopic evaluation of colonic damage.The effect of LPA treatment on Sic26a3 mRNA level and protein expression in the different groups of mice was investigated by quantitative PCR and Western blotting.Results All mice treated with DSS lost weight,but the onset and severity of weight loss was attenuated in the LPA treatment DSS group.The increases in stool water content and the macroscopic inflammation score in LPA treatment DSS group were significantly lower compared to DSS control group or PBS treatment DSS group （（18.89±8.67）％ vs.（28.97±6.95）％ or （29.48±6.71）％,P=0.049,P=0.041,respectively and 2.67±0.81 vs.4.5±0.83 or 4.5±0.54,P=0.020,P=0.006,respectively）,as well as the increase in DAI （P=0.004,P=0.008,respectively）.LPA enema resulted in higher Slc26a3 mRNA and protein expression levels compared to PBS-treated and untreated DSS colitis mice.Conclusion LPA increases Slc26a3 expression in the inflamed intestine and reduces diarrhea severity in DSS-induced colitis,suggesting LPA might be a therapeutic strategy in the treatment of colitis associated diarrhea.

3,6-dichlorobenzo[b]thiophene-2-carboxylic acid alleviates ulcerative colitis by suppressing mammalian target of rapamycin complex 1 activation and regulating intestinal microbiota

BACKGROUND 3,6-dichlorobenzo[b]thiophene-2-carboxylic acid(BT2)is a benzothiophene carboxylate derivative that can suppress the catabolism of branched-chain amino acid(BCAA)-associated mammalian target of rapamycin complex 1(mTORC1)activation.Previous studies have demonstrated the therapeutic effects of BT2 on arthritis,liver cancer,and kidney injury.However,the effects of BT2 on ulcerative colitis(UC)are unknown.AIM To investigate the anti-UC effects of BT2 and the underlying mechanism.METHODS Mouse UC models were created through the administration of 3.5%dextran sodium sulfate(DSS)for 7 d.The mice in the treated groups were administered salazosulfapyridine(300 mg/kg)or BT2(20 mg/kg)orally from day 1 to day 7.At the end of the study,all of the mice were sacrificed,and colon tissues were removed for hematoxylin and eosin staining,immunoblot analyses,and immunohistochemical assays.Cytokine levels were measured by flow cytometry.The contents of BCAAs including valine,leucine,and isoleucine,in mouse serum were detected by liquid chromatography-tandem mass spectrometry,and the abundance of intestinal flora was analyzed by 16S ribosomal DNA sequencing.RESULTS Our results revealed that BT2 significantly ameliorated the inflammatory symptoms and pathological damage induced by DSS in mice.BT2 also reduced the production of the proinflammatory cytokines interleukin 6(IL-6),IL-9,and IL-2 and increased the anti-inflammatory cytokine IL-10 level.In addition,BT2 notably improved BCAA catabolism and suppressed mTORC1 activation and cyclooxygenase-2 expression in the colon tissues of UC mice.Furthermore,highthroughput sequencing revealed that BT2 restored the gut microbial abundance and diversity in mice with colitis.Compared with the DSS group,BT2 treatment increased the ratio of Firmicutes to Bacteroidetes and decreased the abundance of Enterobacteriaceae and Escherichia-Shigella.CONCLUSION Our results indicated that BT2 significantly ameliorated DSS-induced UC and that the latent mechanism involved the suppression of BCAA-associated mTORC1 activation and modulation of the intestinal flora.

Engineering microparticles based on solidified stem cell secretome with an augmented pro-angiogenic factor portfolio for therapeutic angiogenesis

Tissue (re)vascularization strategies face various challenges, as therapeutic cells do not survive long enough in situ, while the administration of pro-angiogenic factors is hampered by fast clearance and insufficient ability to emulate complex spatiotemporal signaling. Here, we propose to address these limitations by engineering a functional biomaterial capable of capturing and concentrating the pro-angiogenic activities of mesenchymal stem cells (MSCs). In particular, dextran sulfate, a high molecular weight sulfated glucose polymer, supplemented to MSC cul-tures, interacts with MSC-derived extracellular matrix (ECM) components and facilitates their co-assembly and accumulation in the pericellular space. Upon decellularization, the resulting dextran sulfate-ECM hybrid material can be processed into MIcroparticles of SOlidified Secretome (MIPSOS). The insoluble format of MIPSOS protects protein components from degradation, while facilitating their sustained release. Proteomic analysis demonstrates that MIPSOS are highly enriched in pro-angiogenic factors, resulting in an enhanced pro-angiogenic bioactivity when compared to naïve MSC-derived ECM (cECM). Consequently, intravital microscopy of full-thickness skin wounds treated with MIPSOS demonstrates accelerated revascularization and healing, far superior to the ther-apeutic potential of cECM. Hence, the microparticle-based solidified stem cell secretome provides a promising platform to address major limitations of current therapeutic angiogenesis approaches.