| ObjectiveResearch on the role of Jian-Pi-Ruan-Gan-Fang( JPRGF) on liver basement membrane formation,Whether or not to interfere with it,it is to explore its mechanism of action, as Chinese Medicine to form a more effective therapies and formulas provide theoretical and experimental research foundation.MethodsFirst of 72 male Wistar rats were numbered, according to the random number randomly divided into six groups, namely normal group, model group, positive control(Fu zheng hua yu capsules) group, JPRGF high, medium and low dose group, a total of six groups, each with 12 animals. Replication model using intraperitoneal injection of 30% CCl4 and 70% olive oil,according to 2mL/1kg calculated in twice a week for 4 weeks. At the same time the normal group and model group were given normal saline from the first week of administration,High,medium and low dose group in IPRGF were given various doses of JPRGF gavage, and the positive control group received intragastric in Fuzhenghuayu. Observation following 4 weekend indicators: The use of biopsy(HE staining, Mallory staining) to observe the degree of hepatic fibrosis in rats;Using electron microscopy under sinusoidal endothelial basement membrane ultrastructure; using Q-PCR test ColⅣ, LN mRNA expression levels; It is using immunohistochemical ABC method which to detect the expression of LN protein levels.Results1. The general situation: Normal activity of rats is usual, shiny hair, normal food intake; It is to reduce rat model of group in activities, in apathetic, in dull hair, in diarrhea, in food intake;positive control group, JPRGF high, medium and low dose group are better than model group in food intake,in activity,in lustrous coat. Model group were higher dead than others.2.The morphology and quality in gross liver: Normal rat liver are dark red,the smooth,encapsulated,shiny,soft,sharp,edges;model group rat lives volume is increased,it is the lighter the color,it is tight envelope,there are fine particulate projections,dull,hard texture,blunt edge;The positive control group,JPRGF high,medium and low dose group in rat liver color than normal group.The Positive control group,JPRGF high,medium and low dose group,the quality of the liver modelgroup were significantly higher than the normal group,the normal group and the positive control group,middle dose group and low dose group rat liver mass compared with model group decreased;there was no significant difference in the quality of liver and high dose groups compared with the model group;But the high,medium and low dose group of the decrease of quality in liver.3. Light microscopy observations(HE dyeing 、 Mallory dyeing): normal group lobular architecture intact rats, the central vein is located in the central lobule, It is hepatic cord in the central vein of presenting radial, uniform size of liver cells, The liver cells were round, It shows a small amount of collagen fibersin the portal area lobular intervals; The model rats visible lobular structure was destroyed, It is divise out of thick collagen fiber strips from the central vein andperiportal cable, It is surrounded with steatosis and edema generation, hepatic cord disorder, a large number of the collagen fibers deposition, diffuse distribution;The positive control group, The JPRGF high, medium and low dose groups in rat liver tissue show the liver tissue damage reduction, It is to reduce the deposition of collagen fibers, collagen fiber area is relatively smaller,reducing the amount of fat vacuoles, the degree of liver fibrosis were alleviated.4.Electron microscope observations: The normal rat liver sinusoidal endothelial cells were flat, distal cytoplasmic into a sheet with a plurality of discrete apertures, It is no subendothelial basement membrane in the space of Disse from a lot of liver and microvilli a small amount of collagen fibers;It is reducing the number of rat liver sinusoidal endothelial fenestrae model group or even disappear within the Disse space collagen fibers visible, even under partial can be seen continuous endothelial basement membrane;The positive control group, JPRGF high, medium and low dose group rat liver sinusoidal endothelial fenestrae diameter becomes smaller, reducing the number of small range It is compared with the model group, it did not see the subendothelial in basement membrane, it have a small amount of liver microvilli.5.The ColⅣ, LN mRNA expression levels: a comparison with the normal group, model group ColⅣ, LN mRNA expression levels increased, the difference is statistically significant(P<0.05); Compared with the model group, middle dose group, positive control group ColⅣmRNA expression levels was reduced, the difference is statistically significant(P<0.05); But between high, medium and low dose groups ColⅣ mRNA expression levels is not a certain drug dependency. LN mRNA expression level of positive control group, high, medium and low dose group were lower, the difference is statistically significant(P<0.05);And between high, medium and low dose group of the decrease of LN mRNA expression levels between certain drug dependency.6.The LN protein expression levels: Compared with the model group, the normal group LN protein were significantly increased, the difference is statistically significant(P<0.05); compared with the model group, positive control group, high, medium and low dose group LN protein content decreased significantly, the difference is statistically significant(P<0.05).And between high, medium and low dose group of the decrease of LN protein expression levels between certain drug dependency.Conclusion JPRGF basement membrane of rat liver fibrosis formation has the effect, The mechanism and JPRGF is related to the inhibition of the expression of Col IV and LN in liver tissue. |
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