The effect of Massa Medicata Fermentata on the cytokine secretion of colonic mucosa and visceral sensitivity in rats with IBS-D

Objective To study the effect of MMF(Massa Medicata Fermentata)on the secretion of cytokines of colon mucosa and visceral sensitivity in rats with IBS-D(diarrhea-predominant irritable bowel syndrome). Methods 45 adult male SD rats (Spragua-dawley) were randomly divided into Normal control group (NC). IBS-D model group (Model); MMF intervention group (MMF). qPCR (quantitative real-time polymerase chain reaction) was used to detect the quantity of fecal Bifidobacterium , Lactobacillus , and Escherichia coli ; FITC-Dextran (Fluorescein Isothiocyanate-Dextran) was used to detect intestinal permeability; Immunofluorescence was used to detect the localization of FliC (Flagellin main component) and TLR5 (Toll-like receptor 5) in the intestinal mucosa; qPCR and WB (Western blot) were used to detect the mRNA and protein expression of TLR5, TRIF (Toll/Interleukin-1 Receptor Domain-Containing Adapter Inducing Interferon-β), EPK1/2 (Extracellular signal-regulated kinase 1/2) in LPDCs (Lamina Propria Dendritic Cells); CCK8 (Cell Counting Kit-8) was used to detect the proliferation of CD4+ T lymphocytes stimulated by LPDCs; ELISA (Enzyme-Linked Immunosorbent Assay) was used to detect the secretion of IL-12, IL-4, IL-6, IL-9, IL-17A, IL-10 (Interleukin-12, 4, 6, 9, 10, 17A), IFN-γ (interferon-gamma), TGF-β (Transforming Growth Factor - beta), from CD4+ T lymphocytes. Results After IBS-D modeling, When compared with the NC group, the visceral sensitivity of rats in model group and MMF group were increased; The quantities of fecal Lactobacillus and Bifidobacterium decreased, The quantities of fecal Escherichia coli increased; The permeability of colonic mucosa was enhanced, accompanied with Flagellin and TLR5 protein upregulated; The expression of TLR5, TRIF, EPK1/2 signals inside LPDCs were increased; CD4+ T lymphocytes proliferation ability was hyperfunction, followed by the excessive secretion of IL-6, IL-17A, TGF-β, IL-12, IFN-γ, and IL-4 from CD4+ T lymphocytes, in contrast IL-10 was hyposecretion (p