The Study Of Curative Effect And Pathogenesis Of PRC In Intervening Insulin Resistance Existed In An Experimental Rat Model Of NASH

Objectives:The experimental NASH rat model were established by using high-fat diet to feed SD rat and intervened with Polysaccharide of radix clematidis (PRC)by gavage. By detecting the content of fasting glucose,fasting insulin and retinol binding protein 4 in serum among the experimental NASH rat model group, PRC group and the polyene phosphatidyl choline group. Calculating and analyzing the difference of HOMA-IR and retinol binding protein 4 in each group. We aimed to investigate the curative effect that PRC intervene the insulin resistance and the correlation of insulin resistance and serum retinol binding protein 4 in the experimental NASH rat.Methods:1 The modeling method:14 were selected from 50 clean healthy male SD rats and assigned to the blank, group, then the remaining 36 to were used as a model group. Blank group and model group were fed 16 weeks, with a normal diet and high-fat diet(feed ingredients:82.5% normal diet + 10% lard +5% yolk powder +2% cholesterol+0.5% sodium cholate) respectively.1 rat in each group on 4th weekend,1 rat in each group on 8th weekend,1 rat in each group on 12th weekend,1 rat in each group on 16th weekend, were selected randomly, their body weight, wet liver weight and liver index were measured. Then their blood of inferior vena cava were used to detect ALT,AST, TC,TG,RBP4, fasting glucose and fasting insulin. HOMA-IR was calculated based on data of fasting glucose and fasting insulin. The central portion of the right lobe of the rats liver patho-wax were made and further stained by HE to observe the steatosis, inflammation, fibrosis of hepatic tissues.To observe the changes in the content of the above indicators in serum of each group by statistical methods. Finally, according to the results of the detection and observation of related indicators to determine whether the experimental rat model of NASH had been established successfully.2 The intervening scheme:After the experimental rat model of NASH established successfully. The remaining 30 rats of the original model group were divided into model group, the polyene phosphatidyl choline group, PRC group randomly, each group contained 10 rats. The remaining eight original blank group rats were used as the blank group. Four groups were respectively intervened with 0.9%NS, polyene phosphatidyl choline [195.4 mg/(kg.d)], PRC [50mg/(kg.d)], and 0.9%NS by gavage for 8 weeks, dose is 3ml/d. During this period, the general conditions of all groups were observed carefully. After eight weeks, four groups of rats bleed from the inferior vena cava to test related indicators. To observed the change of fasting glucose, fasting insulin and serum RBP4 in each group and correlation of HOMA-IR and serum retinol binding protein 4 by statistical methods.Results:1. Results of the modeling:1.1 General conditions of rats:The model group occurred the following phenomena that furs were poor, activity quantity were reduced, appetite were bad, Stool became soft and thin, water intake were increased gradually compared with the blank group. While the situation of blank group was that activity quantity, appetite, water intake, and stool were normal and their furs were smooth.1.2 Body weight, liver wet weight and liver index of rats:The body weight of model group and blank group were increased gradually, but the statistical difference between the two groups was not significant (P=0.351). The liver wet weight of model group and blank group were increased gradually, the statistical difference between the two groups was significant (P=0.011). Liver index of model group showed a gradual upward trend, While liver index of blank group showed no increase, statistical difference between the two groups had observably significance (P=0.000)1.3 Fasting glucose,fasting insulin and HOMA-IR of rats:Fasting glucose of model group showed a gradual upward trend, While fasting glucose of blank group showed no obviously increase, but the statistical difference between the two groups was not significant(P=0.305). Fasting insulin of model group showed a gradual upward trend, While fasting insulin of blank group showed no obviously increase, the statistical difference between the two groups was significant (P=0.020). HOMA-IR of model group showed a gradual upward trend, While HOMA-IR of blank group showed no obviously increase, the statistical difference between the two groups was significant (P=0.047).1.4 Serum retinol binding protein 4 of rats:Retinol binding protein 4 of model group showed a gradual upward trend, While retinol binding protein 4 of blank group showed no obviously increase, the statistical difference between the two groups was significant (P=0.049)1.5 Transaminases and lipid indexes of rats:AST、ALT、TG of model group showed a gradual upward trend, TC remained stable from the fourth weekend, While AST、 ALT、TG、TC of blank group showed no obviously increase. AST, ALT, TC and TG of model group were significantly higher than blank group. AST, ALT, TC, TG compared with each other between two groups, the statistical differences were significant (/P=0.000, P=0.002, P=0.000, P=0.023)1.6 Observation of the rat liver:Direct observation:The model group occur the following phenomena that surface gloss of liver was poor, color was yellow, volume was increased, edge was blunt slightly. While the situation of blank group was that surface gloss of liver was smooth, color was pink, volume was moderate, edge was sharp. Macroscopic observations:Hepatic histology of the model group showed a lesions law of normal-NAFL-NASH. At the end of 16 weeks, Hepatic steatosis degree of model group was Ⅳ (F4), hepatic inflammation grade was 2 (G2),hepatic fibrosis staging was 0(S0), NAS score was 8 scores. Statistical differences of hepatic steatosis degree, hepatic inflammation grade and NAS score between model group and blank group were significant (P=0.005, P=0.021, P=0.000)2. Intervention results:2.1 General conditions of rats:The phenomena that poor furs, reduced activity quantity and bad appetite of polyene phosphatidyl choline group and PRC group were improved after gavage intervention. It was better than model group. At the same time, the increase of weight of those four groups was slow relatively. The situation of blank group was that activity quantity, appetite, water intake, and stool were normal and their furs were smooth. The stool of the model group, the polyene phosphatidyl choline group and PRC group were still soft and thin, it was the same situation with the modeling period.2.2 HOMA-IR of rats:The statistical difference of HOMA-IR between blank group and model group was observably significant (P=0.000). The statistical difference of HOMA-IR between PRC group and model group was observably significant (P=0.000). The statistical difference of HOMA-IR between the polyene phosphatidyl choline group and model group was observably significant (P=0.000) HOMA-IR of the blank group, PRC group and the polyene phosphatidyl choline group compared with each one group, the statistical difference of it was no significance (P>0.05).2.3 Serum retinol binding protein 4 of rats:The statistical difference of RBP4 between blank group and model group was observably significant (P=0.006). The statistical difference of RBP4 between PRC group and model group was significant (P=0.006). The statistical difference of RBP4 between the polyene phosphatidyl choline group and model group was significant (P=0.006). RBP4 of the blank group, PRC group and the polyene phosphatidyl choline group compared with each one group, the statistical difference of it was no significance (P>0.05)2.4 The relationship of HOMA-IR and serum RBP4:HOMA-IR and RBP4 were positively correlated with each other (r=0.541, P=0.001)Conclusions:1. SD rats were fed with high-fat diet for 16 weeks can be established into the experimental rat model of NASH, but not associated with liver fibrosis.2. Insulin resistance exist in the experimental rat model of NASH.3. Serum RBP4 of the experimental rat model of NASH is positively related to insulin resistance of rat.4.The therapy of PRC and polyene phosphatidyl choline can both decrease the insulin resistance in the experimental rat model of NASH effectively and their curative effects are comparable.5. PRC has no obvious toxicity.