| Objective:To establish a model of NAFLD in rats, explore the regulation of multiple soluble extract of tea (pu-erh tea, green tea, black tea and oolong tea)on intestinal long chain fatty acids absorption and tight junction proteins in NAFLD rats by studying the gene expression trend of relative proteins in rats'intestine, andtheir intervention effect on body weight, liver weight and lipid level in NAFLD rats.Methods:96 male SD rats were randomly divided into two groups:normal control group(NCG) and model group(MG).24 rats in NCG fed a normal dietand 72 rats in MG fed a high-fat diet.12 rats from each groups were randomly chosen and killed at the end of the 4th weeks, the body weight, liver weight, blood lipid parameter were measured and hepatic pathology were observed to ensure the building result. MG randomly devided into five new groups with 12 rats in each from the 5th week:NAFLD model group(NMG), pu-erh tea treating group(PTG), green tea treating group(GTG), black tea treating group(BTG) and oolong tea treating group(OTG). Rats in four treatment groups fed a high-fat diet along with gavaging soluble extract of tea at the dose of 1.0 g/kg.d(soluble extract of pu-erh tea, PTE); 0.6g/kg.d(soluble extract of green tea,GTE); 0.8g/kg.d(soluble extract of black tea,BTE); 0.8g/kg.d(soluble extract of oolong tea,OTE) respectively for PTG; GTG; BTG; OTG for 4weeks, rats in NCG and NMG were given normal diet and high-fat diet espectively till the end of the 8th weeks. The body weight, liver weight, blood lipid parameter, fatty acid related protein, tight junction proteins mRNA were detected by RT-PCR after the rest rats were sacrificed at 8 weeks.Results:(1 Establishment of NAFLD model in rats:Data show that at the fourth weekend, compared with the normal control group, the rats body weight and wet liver weight were significantly elevated in the NAFLD model group(p<0.01; p<0.01,respectively); serum TG, LDL-C level were also increased(p<0.01; p<0.01, respectively). While the HDL-C level was remarkably reduced (p<0.01); compared with the NMG, the rats body weight and wet liver weight were notabely reduced(p<0.01; p<0.01,respectively), serum TG, LDL-C levels were markedly decreased(p<0.01;p<0.01,respectively), and serum HDL-C strikingly increased (p<0.01)in the PPG. Hepatic pathology showed a significant hepatic steatosis in the NMG. Which suggest us that rats could be established as a model of NAFLD by fed a HFD for 4 weeks.(2)Effects of soluble extract of tea(SET)on rat body weight and liver weight:Data from the 8th weekend display that, Compared with NMG, rats body weight and wet liver weight were decreased significantly(P<0.01; P<0.01)in PTG; the change of body weight was not significant (p>0.05; p>0.05,respectively)in the GTG and in BTG, while the wet liver weight of rats decreased significantly(P<0.01;P<0.01, respectively)in these two groups.Regarding as the OTG,the change of rats' body weight and the wet liver weight were not significant(p>0.05; p>0.05,respectively).The difference of the body weight and liver weight were not significant among the four groups(p>0.05).(3)Effects of SET on the serum TG?LDL-C?HDL-C dectection:Compared with NMG, serum LDL-C decreased strikingly(p<0.01; p<0.01) in the PTG and GTG, but the change of serum TG and HDL-C was not remarkable(p>0.05; p>0.05; p>0.05; p>0.05, respectively); serum LDL-C decreased notablely (p<0.01),serum HDL-C increased markedly (p<0.01), and serum TG decreased but not overtly(p>0.05)in the BTG;while for OTG, the serum TG and LDL-C decreased markedly (p<0.01; p<0.01, respectively), and serum HDL-C elevated significantly(p<0.01).The difference of the blood lipid parameters were not significant among the four groups(p>0.05)(4) Effects of SET on the gene expression of intestinal long-chain fatty acids absorption portiens CD36, FATP4:Compared with NCG, the expression of intestinal CD36 increased markedly(p<0.01)in NMG, but the expression difference of intestinal FATP4 was not significant(p>0.05). Compared with NMG, in all four treatment groups, the expression of intestinal CD36 decreased markedly (p<0.01; p<0.01; p<0.01; p<0.01, respectively), but FATP4 was not significantly changed (p>0.05; p>0.05; p>0.05; p>0.05, respectively). The expression levels of CD36 were lower in both PTG and OTG campared with the GTG(p<0.01; p<0.01).(5)Effects of SET on the gene expression of intestinal tight junction proteins Occludin and ZO-1:Compared with NCG, the expression difference of intestinal Occludin and ZO-1 were not significant(p>0.05; p>0.05). Compared with NMG, in the PTG, GTG, BTG and OTG, the expression of intestinal Occludin and ZO-1 all increased markedly(p<0.01; p<0.01, respectively). The expression level of Occludin was higher in the BTG than the other three groups(p<0.01; p<0.01; p<0.01, respectively).(6)Effects of SET on the gene expression of intestinal TNF-a and iNOS:Compared with NCG, the expression of intestinal TNF-a increased markedly(p<0.01) in NMG, but the expression difference of intestinal iNOS was not significant(p>0.05). Compared with NMG, in all four treating groups, the expression of intestinal TNF-a all reduced markedly(p<0.01; p<0.01; p<0.01; p<0.01, respectively);and the expression difference of iNOS were also notabely(p<0.01; p<0.01) in the BTG and the OTG, while iNOS expression did not reduced significantly(p>0.05; p>0.05) in the PTG and the GTG.The expression levels of TNF-a and iNOS were both lower in PTG, BTG and OTG than those in GTG(p<0.01).Conclusions:(1)We successfully replicated the rat NAFLD model by fed a high-fat diet, which had the following characteristics:?rats body weight and wet liver weight increased significantly; ?rats serum TG and LDL-C increased markedly, and serum HDL-C reduced overtly; ?Liver gross observation showed a larger volume and the color turned yellow in the model group; ?Hepatic pathology showed a significant hepatic steatosis in the NMG.This model is applicable for NAFLD research.(2) In all four SET groups, the rats body weight, liver weight and serum TG, LDL-C and HDL-C levels were improved compared to the NMG to varying degrees, which suggests that SET intervention can improve NAFLD rats' obesity and dyslipidemia level.(3) SET could significantly decrease NAFLD rats' intestinal CD36 gene expression.These results may indicate that SET may reduce the uptake of long-chain fatty acids by intestine via inhibiting the expression of fatty acids absorbing proteins on enterocyte membrane.(4) SET could upregulate expression of tight junction proteins Occludin and ZO-1, prevent rats' intestinal tight junction, and prevent macromolecular substances into the liver to some extent. the mechanisms involved in reducing the expression of intestinal TNF-a and iNOS. |
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