The Underlying Mechanism Of Modified Citrus Pectin On Myocardial Fibrosis Based On Gal-3/TLR4/MyD88/NF-κB Signaling Pathway

Objective: The purpose of this study was to investigate the effect of modified citrus pectin(MCP),an active ingredient of Citri Reticulatae Pericarpium,on cardiac function,cardiac inflammation,myocardial fibrosis and TLR4/My D88/NF-κB signaling pathway of the model of myocardial fibrosis in rat,which was induced by isoprenaline(ISO),so as to clarify the occurrence and development of myocardial fibrosis and provide guidance for clinical treatment.Methods: 80 Wistar male rats were randomly divided into 5 groups,including control group(Control),model group(ISO),low-dose MCP intervention group(ISO + MCP 100 mg/kg/d),high-dose MCP intervention group(ISO + MCP 100 mg/kg/d)and spironolactone(SPI)intervention group(ISO + SPI 20mg/kg/d).Except the control group,the other groups were injected subcutaneously with ISO(5 mg/kg/d)for 7 days to establish a model of myocardial fibrosis in rats,and were administered with the intervention drugs by gavage for 14 or 21 days.Cardiac function was analyzed by echocardiography and electrocardiogram.cardiac morphology and pathological changes were observed by HE and Masson staining.Immunohistochemical staining was used to observe changes in protein levels of collagen Ⅰ,collagen Ⅲand galectin-3;western blot was used to analysis the protein expression levels of collagen Ⅰ,collagen Ⅲ,galectin-3,TLR4,My D88,NF-κB-p65 and the phosphorylation level changes of NF-κB-p65 in cardiac tissue;q RT-PCR was used to analysis the m RNA levels of collagen Ⅰ,collagen Ⅲ,galectin-3,IL-1β,IL-18 and TNF-α in cardiac tissue.ELISA was used to detect the protein expression levels of IL-1β,IL-18 and TNF-α in serum,and RIA was used to detect serum aldosterone levels.Results:1.MCP prevents cardiac dysfunction caused by ISOThe results of echocardiogram and electrocardiogram showed that ISO induced myocardial infarction and caused cardiac dysfunction mainly with reduced cardiac output,while MCP intervention obviously prevented myocardial infarction and improved cardiac dysfunction.2.MCP improves histopathological changes induced by ISOThe results of HE staining and Masson staining showed that ISO induced myocardial cell necrosis,myocardial fiber structural disorder,inflammatory cell infiltration and large amount of collagen fibers deposition,leading to cardiac remodeling;while MCP intervention significantly reduced myocardial cell necrosis,myocardial fiber structural disorder,inflammatory cell infiltration and collagen fibers deposition,and prevented cardiac remodeling.3.MCP ameliorates ISO-induced MFThe results of immunohistochemistry,western blot and q RT-PCR showed that ISO induced a large amount of collagen Ⅰ and collagen Ⅲ;while MCP intervention significantly reduced the m RNA and protein expression levels of collagen Ⅰ and collagen Ⅲ,and reduced myocardial fibrosis.4.MCP downregulates ISO-induced Gal-3 expressionThe results of immunohistochemistry,western blot and q RT-PCR showed that galectin-3 m RNA and protein expression levels were increased in the model of myocardial fibrosis induced by ISO,while MCP intervention reduced the m RNA and protein expression levels of galectin-3.5.MCP inhibits TLR4/My D88/NF-κB signaling pathway induced by ISOWestern blot results showed that the TLR4/My D88/NF-κB signaling pathway was activated in the rat model of myocardial fibrosis induced by ISO,while the MCP intervention blocked the activation of the TLR4/My D88/NF-κB signaling pathway.6.MCP decreases ISO-induced pro-inflammatory cytokine levelsThe results of ELISA and RIA showed that ISO increased the levels of IL-1β,IL-18,TNF-α and aldosterone,while MCP intervention reduced the levels of IL-1β,IL-18,TNF-α and aldosterone.Conclusion: MCP inhibits the activation of TLR4/My D88/NF-κB signaling pathway,cardiac inflammation and myocardial fibrosis by downregulating galectin-3 expression,thereby improving cardiac dysfunction.