Effects Of Kiwifruit Peel Polyphenols Extract On Intestinal Damage Caused By High Fat Diet

Objective:In order to investigate the effects of kiwifruit peel polyphenols extract（KPE）on intestinal damage caused by high-fat diet,so as to provide scientific basis for the mechanism of preventing or alleviating intestinal damage by KPE.Methods:Thirty-two male Sprague-Dawley（SD）rats were randomly divided into four groups of eight according to body weight:the normal diet group（Normal group）,the high-fat diet group（HFD group）,the low-dose intervention group of KPE（50mg/kg·d,HFD+LKPE group）,and the high-dose intervention group of KPE（100mg/kg·d,HFD+HKPE group）.The Normal group was fed with normal feed,the other3 groups were fed with high-fat feed,gavage once a day,the KPE intervention group was fed with a solution of the corresponding concentration,the Normal group and the HFD group were fed with distilled water,and these rats were slaughtered after being gavaged for nine consecutive weeks.1.H&E staining was used to observe the pathological changes of intestinal mucosa in rats.Immunohistochemical staining was used to observe the protein expression of tight junction proteins Claudin-1,Occludin and ZO-1.Real time-PCR was used to detect the m RNA expression levels of tight junction proteins Claudin-1,Occludin and ZO-1,as well as the kit to determine the endotoxin content in rat serum,to clarify the effect of KPE on the intestinal barrier function of rats with high-fat diet.2.The activity of T-SOD and GSH-Px,the content of GSH and MDA in intestinal tissue were detected with the kit.And Real time-PCR method detects the m RNA expression levels of phaseⅡdetoxification enzymes NQO1,antioxidant enzymes HO-1,GSH-Px,SOD1 and SOD2,transcription factor Nrf2,and Keap1 to clarify the effect of KPE on the intestinal antioxidant capacity of rats with high-fat diet.3.The Illumina Miseq PE300 high-throughput sequencing platform was used to analyze the influence of KPE on the intestinal microbial structure of high-fat diet rats,and the change in NH₃-N and SCFAS content were measured by chemical analysis and gas chromatography to clarify the effect of KPE on the gut microbes and their metabolites in rats with high-fat diet.4.The m RNA expression levels of inflammatory factors TLR-2,TLR-4,NF-κB,TNF-α,IL-1βand IL-10 were detected by Real time-PCR.Elisa kit was used to determine the levels of inflammatory factors TNF-α,IL-1βand IL-10.And Immunohistochemical staining was used to observe the expression of TLR-2 and TLR-4 proteins to clarify the effect of KPE on intestinal inflammation in rats with high-fat diet.Results:1.High-fat diet led to disordered intestinal structure and infiltration of a large number of inflammatory cells,significantly reduced the expression level of tight junction proteins（P<0.05）,and significantly increased the content of endotoxin（P<0.05）in rats.It caused intestinal damage.HKPE significantly reduced the serum endotoxin level in rats with high-fat diet（P<0.05）.LKPE and HKPE can inhibit the structural disorder of the intestinal tissues of rats with high-fat diet,significantly increase the protein expression of Claudin-1,Occludin and ZO-1（P<0.05）,and the m RNA expression level（P<0.05）in the intestinal tissue of rats with high-fat diet.2.High-fat diet significantly decreased the activities of GSH-Px and T-SOD and the content of GSH in intestinal tissues（P<0.05）,significantly increased the content of MDA（P<0.05）,and significantly down-regulated the m RNA expression level of antioxidant related enzymes（P<0.05）.HKPE significantly increased the activity of T-SOD,GSH-Px and GSH content（P<0.05）,and decreased the content of MDA（P<0.05）in the intestinal tissue.LKPE and HKPE significantly up-regulated the m RNA expression level of phase II detoxification enzyme NQO1,antioxidant enzymes HO-1,GSH-Px,SOD1 and SOD2,and transcription factor Nrf-2（P<0.05）.HKPE significantly down-regulated the m RNA expression level of Keap1（P<0.05）.3.High fat diet led to the disturbance of intestinal microbial structure and its metabolites in rats.HKPE intervention can reduce the diversity of microbial species and improve the structure of the flora in the intestinal of rats.It can improve the relative abundance of some probiotics（Lactobacillus,Allobaculum,Blautia,etc.）and inhibit the relative abundance of some harmful bacteria（Desulfovibrionaceae,Clostridiaceae,etc.）.LKPE and HKPE significantly up-regulated the m RNA expression levels of Bacteroidetes,Lactobacillus and Bifidobacterium（P<0.05）,and significantly down-regulated the m RNA expression levels of Firmicutes and C.perfringens（P<0.05）,especially the effect of HKPE was the most obvious,and HKPE significantly down-regulated the m RNA expression levels of Desulfovibrios（P<0.05）.LKPE and HKPE significantly reduced the content of NH₃-N（P<0.05）.HKPE significantly increased the content of intestinal flora metabolites acetic acid,propionic acid and butyric acid（P<0.05）.4.High-fat diet significantly increased the expression of TLR-2 and TLR-4 protein（P<0.05）and m RNA expression（P<0.05）in the intestine,significantly up-regulated the m RNA expression levels of pro-inflammatory factors NF-κB,TNF-αand IL-1β（P<0.05）,significantly increased the concentrations of pro-inflammatory factors TNF-αand IL-1β（P<0.05）,and significantly decreased the concentrations of anti-inflammatory factor IL-10（P<0.05）in rats.LKPE and HKPE significantly inhibited the expression of TLR-2 and TLR-4protein（P<0.05）and m RNA expression（P<0.05）in the intestine of rat,and significantly down-regulated the m RNA expression levels of pro-inflammatory factors NF-κB,TNF-α,and IL-1β（P<0.05）,significantly up-regulated the m RNA expression levels of the anti-inflammatory factors IL-10（P<0.05）.HKPE significantly reduced the concentration of pro-inflammatory factors TNF-αand IL-1β（P<0.05）,and significantly increased the concentration of anti-inflammatory factor IL-10（P<0.05）.Conclusions:KPE can improve the decrease of intestinal barrier function caused by high-fat diet in rats,which is related to the improvement of intestinal antioxidant capacity,the remodeling of intestinal flora structure,and the inhibition of intestinal inflammation,the effect becomes more obvious with the increase of dose.KPE can be used as a functional factor to alleviate the impaired intestinal barrier function.