Feixian Formula Extract Intervene Pulmonary Fibrosis In Rats VEGF / VEGFR1, VEGFR2, PAI-1, VCAM-1 MRNA And Pulmonary Microvascular Endothelial Cells In Vitro Expression Studies

Idiopathic pulmonary fibrosis (IPF) is a complex disease, there is currently a lack of treatment or safe and effective drugs, the incidence of patients with annual growth of11%.Since diagnosis, the median survival time is less than3years. In Traditional Chinese Medicine,idiopathic pulmonary fibrosis belongs to "Feiwei","Feibi",with lung and kidney deficiency, phlegm and blood stasis as the basic pathogenesis. Fei Xian Fang (FXF) is an effective prescription in treating pulmonary fibrosis, which can tonify lung and kidney,dissipate phlegm, promote blood circulation.Previous clinical studies have demonstrated that it has the advantages of improving symptoms and physiological indexes, animal experiments confirmed that it has effect of anti-inflammatory, antioxidant, anti matrix heavy plastic etc.Recent studies on the pathological mechanism of IPF has transfevered from "inflammation center" to "fibrosis repair",and angiogenesis, coagulation/fibrinolytic abnormalities has played a very important part in it. So this research has been divided into two parts:the whole animal experiment which studied FXF extract interventing bleomycin induced pulmonary fibrosis in rats model,detecting vascular endothelial adhesion factor VCAM-1, vascular endothelial growth factor VEGF and its receptor VEGFR1, VEGFR2, plasminogen activator inhibitor PAI-1mRNA expression level through RT-PCR. In vitro, pulmonary microvascular endothelial cell from Model rats of pulmonary fibrosis are primary cultured,and test their ability of adhesion, migration and angiogenesis. To further explore the mechanism FXF treating pulmonary fibrosis.Animal experiment:156rats were randomly divided into7groups:12rats in the sham operation group, model group24, FXF large dose group24, FXF middle dose group24, FXF small dose group24, control group of Western medicine were divided into two groups:Losartan group24,and prednisone group24. Saline perfusion in false operation group,the rest rats were infused BLM to induce model disease through tracheal intubation,3mg/kg.Intragastric administration started from the next day after inducing model disease.With intragastric volume:FXF big, middle, small dose group were2.4g/kg,1.2g/kg,0.6g/kg.Losartan group, prednisone group5mg/kg10mg/kg. The rats were deal with on14and28days after inducing model disease respectively,the rats were randomly divided into two group with equal number.The left lung were tested through histopathology examination with HE staining and MASSON staining, Semi quantitative detection had been introduced to figure out the degree of the alveolitis and pulmonary fibrosis. Fresh from the right lung for RT-PCR detection of VEGF/VEGFR1, VEGFR2, PAI-1, VCAM-1gene expression level. Results:the middle dose group could effectively improve the alveolitis and pulmonary fibrosis, inhibition of VEGF/VEGFR1, VEGFR2, PAI-1, VCAM-1gene expression level, there were significant differences compared with the model group; P<0.05; Prednisone can effectively improve the degree of alveolitis prednisone on14days, there were significant differences compared with the model group, P<0.05; losartan could inhibit the expression of PAI-1, VCAM-1gene on14days, with statistical significance compared with the model group;P<0.05.Conclusion:1.FXF can inhibit the expression of VCAM-1gene in acute inflammation; inhibit protein and collagen accumulation by inhibition of plasminogen activator inhibitor PAI-1gene expression; inhibit angiogenesis by inhibition of VEGF/VEGFR1,VEGFR2gene expression.The FXF middle dose group (1.2g/kg) has a more obvious effect than those in the model group with statistical significance(P<0.05).2.prednisone can alleviate the degree of alveolitis, losartan inhibited VCAM-1and PAI-1gene expression witch result in inhibition of inflammation、collagen and fibrin in the early period (14days)(there are significant differences compared with the model group, P<0.05). But FXF middle dose group has a more lasting effect.3.FXF has shown a certain advantage in both improving the mental state in rats and detect indicators.Cell culture experiments in vitro:isolation and identification of rat pulmonary microvascular endothelial cells, respectively with FXF big, median and small dosage group, prednisone, losartan stimulated cells.WST-1assay adhesion of endothelial cells with leukocytes, endothelial cell migration rate through detection of Transwell capacity, Matrigel adhesive detecting angiogenesis ability. Results:the Losartan and Fei Xian Fang formula of high dose group could promote the adhesion of leukocyte to pulmonary microvascular endothelial cells. The ability to inhibit migration and angiogenesis increase with dose of FXF.FXF high-dose completely inhibited the pulmonary microvascular formation. Prednisone promote the angiogenesis and migration of microvascular endothelial cells, losartan and FXF middle dosage group can effectively inhibit the migration and angiogenesis of lung microvascular endothelial cell formation. Conclusively,FXF can inhibit inflammation、angiogenesis and reduce fibrinolytic inhibition,thus slowing down the progress of pulmonary fibrosis.