| Objective: To investigate the therapeutic effect of Qingbidajinfang on arthritis and pulmonary fibrosis and its anti-fibrosis mechanism.Methods: Thirty-six seven-week-old healthy male SPF Wistar rats were randomly divided into control group,model group,Qingbidajinfang high-dose group,Qingbidajinfang medium-dose group,Qingbidajinfang low-dose group and positive drug group,with 6 rats in each group.Except for the control group,the other 30 rats were injected with bleomycin in tracheal and bovine type ⅱ collagen in tail to establish collagen-induced arthritis combined with pulmonary fibrosis in rats model.After modeling,the rats were treated with corresponding drugs for two weeks,during which the general conditions of the rats were observed,arthritis index(AI)score and changes in paw volume were recorded.The rats were sacrificed two weeks later.Blood from the abdominal aorta and lung tissue was collected.ELISA was used to detect the levels of TGF-β1 and Fstl-1 in serum.The degree of inflammation in lung tissues was observed by HE staining.The degree of fibrosis in lung tissues was observed by Masson staining.The expressions of TGF-β1 and Fstl-1 in lung tissues was detected by immunohistochemical method.The m RNA expressions of TGF-β1,Smad3,Fstl-1,E-cadherin and α-SMA in lung tissues were detected by q-PCR.The protein contents of TGF-β1,Smad3,Fstl-1,E-cadherin and α-SMA in lung tissues were detected by Western-blot.Results:1.General condition,AI score and paw volume of rats: During the experiment,the rats in control group were in good condition,with steady weight increase and normal diet and water condition.The rats in model group had poor mental state,slow weight growth,hind foot swelling,even joint deformity,shortness of breath,reduced diet and water.The condition of rats in each administration group was improved compared with that in model group.Two weeks after administration,AI score in Qingbidajinfang high,medium dose groups and positive groups was decreased than model group(P<0.01 or P<0.05);The volume of paw in Qingbidajinfang high,medium,low dose groups and positive group was significantly decreased than model group(P<0.01).2.Contents of TGF-β1 and Fstl-1 in serum: The contents of Fstl-1 and TGF-β1 in serum of Qingbidajinfang high,medium and low dose groups and positive group were decreased than those of model group(P<0.01 or P<0.05).3.HE staining results: The alveolar structure of rats in control group was intact;Alveolar wall thickened and alveolar septum disappeared in model group.The alveolar walls of rats in Qingbidajinfang high,medium and low dose groups were thickened with the decrease of the dose,but the alveolar structures were more intact than those in the model group.The alveolar wall of rats in positive group was slightly thickened and the structure was intact.4.Masson staining results: The alveolar structure of the control group was normal,and only a few light blue collagen fibers were distributed.The alveolar structure of model group was severely damaged,and large area of blue collagen fiber hyperplasia appeared.Compared with the model group,the lung tissue of Qingbidajinfang high,medium and low dose groups and the positive group were improved,and the statistical results showed that compared with model group,the degree of pulmonary fibrosis in Qingbidajinfang high dose group and positive group was significantly decreased(P<0.01).5.Immunohistochemical results of Fstl-1 and TGF-β1: In the control group,the alveolar structure was clear and intact,and a small amount of positive expression of TGF-β1 and Fstl-1 was occasionally observed.Compared with the model group,the alveolar structure of Qingbidajinfang high,medium and low dose groups and the positive group were more intact,the lesion range was reduced,and the expression of TGF-β1 and Fstl-1 was significantly lower than that of the model group(P<0.01).6.Expression results of TGF-β1,Fstl-1,Smad3,α-SMA and E-cadherin m RNA in lung tissues: Compared with the control group,the m RNA contents of TGF-β1,Smad3,Fstl-1 andα-SMA in lung tissues of model group were significantly increased(P<0.01),while the m RNA content of E-cadherin was significantly decreased(P<0.01).Compared with model group,the m RNA contents of TGF-β1,Smad3,Fstl-1 and α-SMA in Qingbidajinfang high,medium and low dose groups and positive groups were decreased(P<0.01 or P<0.05).The E-cadherin m RNA content in lung tissues of Qingbidajinfang high,medium and low dose groups and positive group was increased(P<0.01).7.Relative expression results of TGF-β1,Fstl-1,Smad3,E-cadherin and α-SMA proteins in lung tissues: Compared with the control group,the relative expression levels of TGF-β1,Smad3,Fstl-1 and α-SMA proteins in the lung tissues of model group were significantly increased(P<0.01),while the relative expression levels of E-cadherin protein was significantly decreased(P<0.01).Compared with model group,the relative expression levels of TGF-β1,Smad3,Fstl-1 and α-SMA in lung tissues of Qingbidajinfang high,medium and low dose groups and positive groups were decreased(P<0.01 or P<0.05).The relative expression of E-cadherin protein in lung tissues of Qingbidajinfang high,medium and low dose groups and positive group was increased(P<0.01 or P<0.05).Conclusion: Qingbidajinfang alleviates the symptoms of arthritis and pulmonary fibrosis in rats,and its anti-pulmonary fibrosis effect may be exerted by inhibiting TGF-β1 signaling pathway. |
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