| Objective: To investigate the effects of cholesterol on smooth muscle of gallbladder. In vivo the calcium signal of gallbladder was observed using isolated gallbladder smooth muscle strips. In vitro the morphological changes of mitochondria and the alterations of active form transform growth factor-β1( TGF-β1) expression was studied by using guinea pig model with high-level bile cholesterol.Methods:Force transducer was used to measure the contraction of isolated gallbladder smooth muscle strips ,which were induced by cholecystokinin(CCK)both in presence and absence of extracellular calcium in the chamber which was administrated with cholesterol.Sixty-four guinea pigs were randomly divided into four groups,every group contains 16 guinea pigs. The control group was fed standard chow and the high-cholesterol group was fed standard chow plus 2% cholesterol, after two weeks' that diet, the natural group was fed standard chow and the treated group was orally administrated with the traditional Chinese medicine.Gallbladder were exposed with a midline incision under anesthesia and gallbladder volume was measured and the bile was aspirated to measure its volume and concentration of bile cholesterol.Gallbladder was removed from the liver bed intactly. A small mass of gallbladder(about 1cm3) was obtained for electron-microscope, then the morphosis was observed and the mean volume(V) and the numerical density(Nv) of mitochondrial of smooth muscle cells were counted by an image analysis system.The rest gallbladder tissues were used to study the expression of active form TGF-β1. Results:(1) Cholesterolcan inhibit the contraction of the isolated smooth muscle strips(1.76±0.22VS2.20±0.20,P=0.002;1.16±0.34VS2.20±0.20,P=0.0001) and the effect is more dramatic than that of the nifedipine(1.16±0.34VS1.76±0.22,P=0.0001).(2) cholesterol can both block the extracellular calcium entry and prevent the intracellular calcium releasing(1.08±0.10VS1.25±0.15, P=0.007;0.72±0.06VS0.89±0.09, P=0.0001) but the nifedipine can only effect on the extracellular calcium entry(0.64±0.05VS0.89±0.09,P=0.0001) and has no impact on the intracellular calcium releasing(1.20±0.08VS1.25±0.15, P=0.330).(3) Compared with control group, after the two-week high-cholesterol feeding, the mean volume(V) of mitochondria was increased(0.08056±0.0069Vs0.0653±0.02868,P=0.0001) and the numerical density(Nv) of it was decreaed(0.2677±0.0138Vs0.3180±0.02868,P=0.0001). (4) Compared with control group, after the two-week high-cholesterol feeding, the expression of active form TGF-β1 increased(P=0.0001).(5) After two-week of herbs of shuganlidan liqihuoxue, compared the natural group , the concentration of bile cholesterol, the mean volume(V) and the numerical density(Nv) of mitochondria and the expression of active form TGF-β1 come back to normal.Conclusion:In vitro,the cholesterol can both block the extracellular calcium entry and prevent the intracellular calcium releasing and thus effect the excite-contraction coupling of gallbladder muscle.In vivo,the cholesterol can change the morphosis of the gallbladder smooth muscle cells'mitochondria and at the same time lead to overexpression of active form TGF-β1 to initial the process of gallbladder fibrosis. All these alterations can cause the gallbladder muscle contractile dysfuntion.Herbs shuganlidan liqihuoxue can help the gallbladder's fuction recover.The mechanism is correlated with reducing bile cholesterol concentration. |
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