Rosiglitazone Alleviated Bleomycin-induced Rats’ Lung Fibrosis By Caspase-3 And ERK Signal Pathway

Objective: Idiopathic pulmonary fibrosis(IPF) is a kind of progressive and fatal diffused interstitial lung disease,and the morbidity and mortality increased steadily at present. The characteristics of IPF are the proliferation of fibroblasts and the deposition of extracellular matrix components, but the etiology and pathogenesis are not fully unknown and lack of effective treatment. In our study, we preliminary explore whether rosiglitazone(RSG) inhibit the proliferation and transformation of fibroblasts, and reduce synthesis and deposition of collagen to alleviate pulmonary fibrosis by activating PPAR-γ and regulating apoptosis, and explore whether extracellular signal-regulated kinase(ERK) signal pathway participate in the anti-fibrosis process of RSG.Methods:54 healthy male Wistar rats weighted 125±5g,were divided into three groups with random table:Control group, BLM group(BLM) and BLM+RSG group(BLM+RSG). Each group contains 18 rats. The rats with 10% chloral hydrate(3 ml/kg) abdominal cavity anesthesia, supine fixed on the rats table, the neck skins were disinfect conventionally, the middle of the neck skin were cut, the tissue were separated layer by layer, tracheas were exposed. The rats in group BLM and group BLM+RSG received intratracheal injection of bleomycin(BLM)(5mg/kg) to establish the model of pulmonary fibrosis, while equal volume normal saline were injected into trachea in Control group. The rats were upringted and rotated immediately after injection in order to distribute the liquid in the lungs. 24 hours after modeling, rats in group BLM+RSG were given intragastic administration with RSG 3mg/kg.d, while equal volume normal saline were injected in Control group and group BLM, 6 rats were taken randomly in each group after BLM administration on the 7th day 、 14 th day and 28 th day. The rats were exsanguination by bilateral carotidartery after abdominal cavity anesthesia with 10% chloral hydrate(3ml/Kg), the sternum was cut alonging the xiphoid, thoracic cavity was opened, and tracheas and lung tissue were separated. Middle lobe of right lung was used for pathological examination by Hematoxylin and eosin(HE)staining and Masson staining, and the alveolitis and pulmonary fibrosis were divided by the method of Szapiel. Superior lobe of right lung was used to measure the mRNA expressions of α-smooth muscle actin, Collagen typeⅠ, Collagen type Ⅲ, PPAR-γ and caspase-3 by real time RT-PCR. Right lower lung was used to evaluate the relative protein expression of P-ERK by Western blot. Statistical analysis of values was performed with SPSS13.0 software, P<0.05 indicated that there was statistical significance.Results1 The results of pulmonary pathology: on the 7th day、14th day、28th day,the Control group showed the normal alveolar structures. In group BLM, there were many inflammatory cells and obvious alveolitis. The alveolitis were alleviated on the 14 th day and the 28 th day, but the level of alveolitis of group BLM was more serious than that in Control group, there were significant difference at each time points(P<0.05). On the 14 th day and 28 th day, the number of pulmonary fibroblasts was increased and focal or a sheet of blue collagen were observed, the level of fibrosis of group BLM was more serious than that in Control group(P<0.05). Thus the model of BLM-induced pulmonary fibrosis in rats was established successfully. After the treatment of RSG, the level of alveolitis and fibrosis of group BLM+RSG was ameliorated than that in group BLM, there were significant difference at the oberserved time points(P<0.05).2 The results of real time RT-PCR: On day 7, 14 and 28, the expression of α-SMA、Collagen type Ⅰ、Collagen type Ⅲ in group BLM was increased obviously than that in the Control group, there were significant difference(P<0.05). After the administration of RSG, the expression of α-SMA、Collagen typeⅠ、Collage type Ⅲ was reduced than that in the group BLM, there were significant difference at the oberserved time points(P<0.05). After the therapy of RSG, the expression of PPARγ mRNA was higer than that in the group BLM, there were significant difference on each time point(P<0.05). On day 7, 14 and 28, the expression of caspase-3 in group BLM+RSG was upregulated than group BLM, there were significant difference at the oberserved time points(P<0.05)3 The results of Western blot: On day 7, 14 and 28, the protein expression of P-ERK in group BLM was upregulated than Control group, there were significant difference on each time point(P<0.05). However, the P-ERK expression was downregulated in group BLM+RSG than that in the group BLM after the treatment of RSG, there were significant difference on each time point(P<0.05).Conclusions:1 RSG remarkably ameliorated BLM-induced pulmonary fibrosis in rats by activating PPAR-γ.2 RSG upregulated the expression of caspase-3 and promoted the apoptosis of fibroblasts to delay the process of pulmonary fibrosis.3 The interaction between PPAR-γand ERK signal pathway maybe taking part in the therapeutic effect of RSG on BLM-induced pulmonary fibrosis in rats.