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The human gut microbiota is a complex and dynamic community of microorganisms living in our intestines and has emerged as an important factor for colorectal adenocarcinoma (CRC). The purpose of our study was to investigate the microbiota composition in Brazilian CRC patients compared with a local control population (CTL) to find out which changes could be considered universal or regional features in CRC microbiota. Fecal samples were obtained from 28 CRC and 23 CTL individuals. The 16S rRNA gene was used for metagenomic analysis. In addition to the anthropometric variables, the clinical stage (TNM 2018) was considered. Patients with CRC had a significant increase in alpha diversity and a higher percentage of genus Prevotella and a decreased proportion of Megamonas and Ruminococcus. Additionally, the proportion of Faecalibacterium prausnitzii was associated with a better prognosis in the first stages of CRC, and Fusobacterium nucleatum proved to be an important marker of colorectal carcinogenesis and tumor aggressiveness. Although regional differences influence the composition of the microbiota, in the case of CRC, the microhabitat created by the tumor seems to be a major factor. Our results contribute to a better understanding of the carcinogenic process, and even in different environments, some factors appear to be characteristic of the microbiota of patients with CRC.
ABSTRACT
It has been widely accepted that Faecalibacterium prausnitzii (Fp) induces the differentiation of Treg cells.Lymphocyte function-associated antigen-1 (LFA-1) is also involved in the differentiation of Treg cells.Aims: To investigate the effect of Fp on Treg cells and cytokines in colitis mice with LFA-1 knockout (LFA-1-/-).Methods: Twenty wild type mice and twenty LFA-1-/-mice with same genetic background were randomly divided into wild type control group, wild type treatment group, LFA-1-/-control group and LFA-1-/-treatment group.Colitis model was induced by drinking DSS solution.Mice in the two treatment groups were intragastrically administrated with Fp.General status and histopathological score were assessed.Percentages of Treg cells in spleen and mesenteric lymph nodes were measured by flow cytometry.Serum levels of IL-10 and TGF-β1 were measured by ELISA.mRNA expressions of IL-10 and TGF-β1 in colonic tissue were detected by real time PCR.Results: Compared with corresponding control groups, histopathological score was significantly decreased in wild type treatment group (P<0.05);percentages of Treg cells in spleen and mesenteric lymph nodes were significantly increased (P<0.05), serum levels of IL-10 and TGF-β1 were significantly increased (P<0.01), expression of TGF-β1 mRNA was significantly increased in wild type treatment group and LFA-1-/-treatment group (P<0.05);expression of IL-10 mRNA was significantly decreased in LFA-1-/-treatment group (P<0.01).Compared with wild type treatment group, serum level of TGF-β1 was significantly decreased (P<0.05), and mRNA expressions of IL-10 and TGF-β1 were significantly decreased in LFA-1-/-treatment group (P<0.05).Conclusions: Fp can up-regulate the percentages of Treg cells and enhance the secretion of anti-inflammatory cytokines IL-10 and TGF-β1 in LFA-1-/-mice.The therapeutic efficacy for colitis in wild type mice is superior to that in LFA-1-/-mice, which may be related to the inhibition of function of Treg cells and secretion of cytokines due to LFA-1 knockout.
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Background:The abundance of Faecalibacterium prausnitzii (Fp) is reduced in patients with inflammatory bowel disease (IBD).Previous studies demonstrated that the supernatant of Fp exerts an obvious anti-inflammatory effect in experimental colitis.Aims:To investigate the effect of Fp supernatant on monocytes/macrophages and colonic inflammation in dextran sulfate sodium (DSS)-induced colitis in mice.Methods:Thirty male C57BL/6J mice were randomly divided into control group,colitis group and Fp treatment group.Acute colitis was induced by drinking 4.5％ DSS in distilled water for seven days in colitis and Fp treatment groups.Mice in Fp treatment group were also given gastric infusion of Fp supernatant during the process of colitis induction.Weight loss,defecation and histopathological damage of colon tissue were observed.The percentages of monocytes and macrophages of different phenotypes in spleen and colonic lamina propria,as well as the serum levels of a series of inflammatory cytokines were detected by flow cytometry.Results:Compared with colitis group,Fp treatment showed an ameliorating effect on weight loss,shortening of colon length and histopathological damage (P ＜ 0.05).The percentages of total amount of monocytes and proinflammatory monocytes in spleen and colonic lamina propria,and the percentage of M1 macrophages in colonic lamina propria in Fp treatment group were significantly lower than those in colitis group;the percentage of M2 macrophages in colonic lamina propria in Fp treatment group was significantly higher than that in colitis group (P all ＜ 0.05).The serum levels of interleukin (IL)-10 and IL-4 in Fp treatment group were significantly higher than those in colitis group,and the levels of IL-6,interferon (IFN)-γ and chemokine CCL2 were significantly lower than those in colitis group (P all ＜ 0.05).Conclusions:Fp supernatant may exert anti-inflammatory effect in experimental colitis in mice via reducing proinflammatory monocytes,inducing M2 polarization of macrophages in inflamed colon,promoting secretion of anti-inflammatory cytokines,and inhibiting secretion of proinflammatory cytokines and chemokines.
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Background:NLRP3 inflammasome attracts widespread attention in study of inflammatory bowel disease. Faecalibacterium prausnitzii(Fp)is an anti-inflammatory commensal bacterium that has preventive and therapeutic effects on rat colitis. Aims:To explore the underlying mechanism of Fp in treating experimental colitis in rats. Methods:Fifty rats were randomly divided into two groups,10 in control group and 40 in model group. Rats in model group were administered intrarectally with 5% TNBS and dehydrated alcohol to induce experimental colitis. Twenty-four hours afterwards,the model rats were further divided into four groups and administered intragastrically with PBS,culture medium,live Fp and Fp supernatant 1 mL per day,respectively,for 7 days. On day 8,all the rats were sacrificed for evaluation of colonic inflammation. Expressions of the constituents of NLRP3 inflammasome(NLRP3,ASC,and caspase-1)were assessed by Western blotting and real-time PCR;levels of IL-1β and IL-18,the downstream effectors of NLRP3 inflammasome,in colon and plasma were measured by real-time PCR and ELISA,respectively. Results:Weight loss, reduced colon length and colonic inflammatory injury were observed in model rats. These manifestations were ameliorated in live Fp and Fp supernatant groups than those in PBS and culture medium groups. In PBS and culture medium groups, expressions of NLRP3,ASC,and caspase-1 protein and mRNA in colonic tissue were significantly higher than those in control group(P < 0. 05),the colonic and plasma levels of IL-1β were increased(P < 0. 05),and IL-18 levels were decreased(P < 0. 05). In live Fp and Fp supernatant groups,IL-18 level showed a further reduction as compared with PBS and culture medium groups( P < 0. 05),but the increasing trend for other parameters was reduced( P < 0. 05). Conclusions:NLRP3 inflammasome participates in the development of TNBS-induced colitis in rats. Fp might alleviate colonic inflammation by inhibiting NLRP3 inflammasome and its downstream effectors.
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Background:Faecalibacterium prausnitzii( Fp) is one of the most abundant bacterium in human intestinal microbiota,and is closely correlated with the process of colitis-associated colorectal cancer(CAC). Aims:To observe the effect of Fp on CAC,and investigate the possible mechanism. Methods:The model of CAC was induced by azoxymethane (AOM)and dextran sodium sulfate( DSS). Fifty-two C57BL/ 6J mice were randomly divided into 4 groups:group A (AOM + DSS),group B(AOM + DSS + Fp),group C(AOM + DSS + Fp supernatant)and group D(control group). All the mice were sacrificed on day 92. DAI was assessed,serum levels of TNF-α and IL-10 were determined by ELISA. HE staining was used to examine the grade of tumor. Expressions of VEGF,COX-2,NF-κB in tumor tissue were measured by immunohistochemistry. Results:The tumorigenesis rates of group A,B,C were 100% ,100% and 77. 8% ,respectively;mainly were high-grade intraepithelial neoplasia. The tumor load in group A was significantly higher than that in group B (P < 0. 01),and the spleen index in group B was significantly higher than that in group C(P < 0. 01). Serum level of TNF-α was significantly lower(P < 0. 05)and IL-10 was significantly higher(P < 0. 05)in group A than that in group B. No significant differences in expressions of VEGF,COX-2,NF-κB were found among group A,B and C. Conclusions:Fp had no obvious effect on the occurrence rate of CAC,and Fp supernatant could decrease the incidence of CAC in mice. Fp and its supernatant could reduce the tumor load via regulating the expressions of TNF-α,IL-10.
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Background:Faecalibacterium prausnitzii(Fp)is a commensal intestinal bacterium that exhibits anti-inflammatory and immunomodulatory capacity in vivo and in vitro. It has been reported that Fp in intestinal lumen was reduced in patients with colorectal cancer,which might be a factor associated with cancer development. Aims:To investigate the effect and immunological mechanism of Fp and its genomic DNA(fDNA)on the killing activity of peripheral blood mononuclear cells (PBMCs)against human colon cancer LoVo cells. Methods:PBMCs derived from healthy adults were co-cultured in vitro with Fp,fDNA,or the digested fDNA(d-fDNA),respectively. Killing activity of PBMCs against LoVo cells was measured by MTT assay;concentrations of interferon-gamma(INF-γ),a Th1-type cytokine and interleukin-4(IL-4),a Th2-type cytokine in culture supernatant of PBMCs were determined by ELISA;and expressions of T-bet and GATA3,the transcription factors specific for Th1 and Th2 cells,were measured by real-time PCR. Results:Compared with the PBMCs not treated,fDNA could significantly enhance the killing activity of PBMCs against LoVo cells(P < 0. 05);meanwhile,it promoted IFN-γ secretion,up-regulated T-bet mRNA expression and inhibited IL-4 secretion and GATA3 mRNA expression in PBMCs(P < 0. 05). Similar effects were not observed in PBMCs treated with Fp and d-fDNA. Conclusions:fDNA enhances the killing activity of PBMCs against human colon cancer cells by up-regulating Th1 immune response.