Fang Qi Qigongkangxian Kangxian (QF) anti-immune injury rat liver fibrosis (HF) experimental study molecular mechanisms
|School||Changchun University of Traditional Chinese Medicine|
|Course||Chinese medical science|
|Keywords||qigongkangxianfang ( QF ) rat Hepatic fibrosis induced by immunologic injury signal transduction of TGF-β1 /Smads molecular mechanism|
Objective: The hepatosplenic disorder, deficiencies of vital energy and stagnant blood are the main pathogenesis and mechanism of liver fibrosis. we adopted Qigongkangxian prescription(QF) to treat liver fibrosis and carried out basic study. QF were composed of pure herbal medicine,such as Radix Astragali, Rhizoma Atractylodis Macrocephalae, Radix Curcumae, Radix Notoginseng, Semen Persicae, Scolopendra, and appendiculate cremastra pseudobulb, which had function of nourishing qi to invigorate spleen, and regulating blood to soften hard mass. The experiment adopted methods of the enzyme-linked immunosorbent assay (ELISA) and immunohistochemistry(IHC) to detect the transmembrane signaling transduction of TGF-β1/Smads, and the changes of pathomorphology of liver tissue and ultramicrostructure of hepatocyte were observed with light and electron microscope, for exploring the molecular mechanism of QF on liver fibrosis in Rats.Methods:(1)Models of liver fibrosis induced by immunologic injury with intraperitoneal injection of porcine serum(with 0.5 ml each time,and two times every week)in rats.40 SD rats were randomly divided into the normal control group(n=10), the model group(n=10), the high–dosage QF treatment group(n=10), and the low-dosage QF treatment group (n=10). The normal control group were intraperitoneal injected with normal saline. The model group and QF treatment group were intraperitoneal injected with porcine serum, for eight weeks.The normal control group and model group were intragastric administration with normal saline,the rest groups were intragastric administration with corresponding of QF(with concentration of 1 g/ ml,the high-dosage QF treatment group with 2 ml/200 g,and the low-dosage QF treatment group with 1 ml/200 g), for four weeks.The serum of rats were collected before and after the experiment and after the end of model duplication to detect the hepatic function or other indexes. After the experiment, rats in each group were killed with decapitation, HE and Mallory staining for observing the changes of pathomorphology of liver tissue with light microscope; the changes of ultramicrostructure of hepatocyte were observed with electron microscope; the contents of LN、HA、TGF-β1 of serum were detected with method of ABC-ELISA; the protein expression of TGF-β1, Smad2, Smad3 and Smad7 of liver tissue were detected with method of immunohistochemistry. RT-PCR method that there was significant difference of TGF-β1 mRNA expressionResults:(1)The body weight of model group rats gained slowly, the weight and index number of spleen increased obviously. After treatment of QF, the body weight, weight and index number of spleen were improved, and the QF high-dosage group had notable effects.(2) The observation of light microscope and electron microscope: the normal structure of hepatic lobuli were injuried, proliferation of collagen fiber, fatty degeneration of hepatocyte, cellular nucleus deformed and collapsed, nucleoli disappeared, chromatin concentrated, mitochondria deformation, the rough endoplasmic reticulum arranged disorder, and the number of smooth endoplasmic reticulum increased. After treatment of QF, hepatic lobuli, hepatocyte and liver tissue were improved with certain extents. The intervention effects of large-dosage of QF were obvious.(3)Compared with the normal control group, the ALT levels and contents of TGF-β1, LN, and HA in serum increased notablely (P<0.05 or P<0.01), but A /G decreased(P<0.01).With the treatment of QF, the ALT levels and contents of TGF-β1, LN, and HA in serum lowered obviously, and A/G ascended(P<0.05).Which indicated QF had the regulation effects of hepatic function and liver fibrosis.(4)The results of immunohistochemistry indicated there were strong expression of TGF-β1 in cytoplasm, the expression of Smad 2 in cytoplasm transfered into cellular nucleus, and the expression of Smad7 was negative in the model group. The expression of TGF-β1, Smad2, and Smad3 decreased(P<0.01) in two QF treatment groups, and above all, the effect was excellence in the QF large-dosage group.(5)The results of RT-PCR indicated that there was significant difference of TGF-β1 mRNA expression between the blank control group and the model group, and the TGF-β1 mRNA expression in two treatment groups had decreased obviously compared with that of model group, and there was no difference between the high-dosage QF treatment group and the blank control group.Conclusion:1. QF could protect and repair the impaired hepatocyte, inhibit the formation of collagen fibers, and improve hepatic function, to treat liver fibrosis induced by immunologic injury, and has obvious relations of dose dependent.2. QF could regulate the transmembrane signal transduction of TGF-β1 /Smads for treating Hepatic fibrosis.