Investigation Of The Effect And Mechanism Of Rapamycin Against Paraquat-induced Pulmonary Fibrosis In Mice

Background:Ingestion of paraquat (PQ), a widely used herbicide, can cause severe toxicity in humans leading to a poor survival rate and prognosis. One of the main causes of death by PQ is PQ-induced pulmonary fibrosis, for which there are no effective therapies. Rapamycin (RAPA) is a potent inhibitor of the mammalian target of rapamycin (mTOR), which binds intracellularly to the FK506(tacrolimus)-binding protein to form a complex which subsequently binds to mTOR to inhibit its ability to signal adequately to its downstream effectors, and eventually inhibit the growth and proliferation of cells. RAPA has been widely used in clinical, including preventing graft rejection in solid organ transplantation because of its immunosuppressive effects. Nowadays, RAPA is also attractive in clinical practice due to its potent antifibrotic property.Aims:To evaluate the effects of RAPA on inhibiting PQ-induced pulmonary fibrosis in mice, and to explore its possible mechanisms.Methods:Male C57BL/6J mice were exposed to either saline (control group) or PQ (10mg/kg body weight, intraperitoneally)(test group). The test group was divided into four sub-groups:a PQ group (PQ-exposed, non-treated), a PQ+RAPA group (PQ-exposed, treated with RAPA at1mg/kg intragastrically), a PQ+MP group (PQ-exposed, treated with methylprednisolone (MP) at30mg/kg intraperitoneally) and a PQ+MP+RAPA group (PQ-exposed, treated with MP at30mg/kg intraperitoneally and with RAPA at1mg/kg intragastrically). The survival rate and body weight of all the mice were recorded every day. Three mice in each group were sacrificed at14days and the rest at28days after intoxication. Lung tissues were excised and stained with hematoxylin-eosin (H&E) and Masson’s Trichrome stain for histopathological analysis. The hydroxyproline (HYP) content in lung tissues was detected using an ELISA kit. The expression of transforming growth factor-β1(TGF-β1), a-smooth muscle actin (a-SMA) and E-cadherin in lung tissues was detected by immunohistochemical staining and western blotting.Results:A mice model of PQ-induced pulmonary fibrosis was established. Histological examination of lung tissues showed that RAPA treatment moderated the pathological changes of pulmonary fibrosis, including alveolar collapse and interstitial collagen deposition. HYP content in lung tissues increased soon after PQ intoxication but had decreased significantly by the28th day after RAPA treatment. Immunohistochemical staining and western blotting showed that RAPA treatment significantly down-regulated the enhanced levels of TGF-β1and a-SMA in lung tissues caused by PQ exposure, but could not recover the expression of E-cadherin. Meanwhile, RAPA treatment alone could not significantly ameliorate the lower survival rate and weight loss of treated mice. MP treatment enhanced the survival rate, but had no significant effects on attenuating PQ-induced pulmonary fibrosis or reducing the expression of TGF-β1and α-SMA.Conclusions:This study demonstrates that RAPA treatment effectively suppresses PQ-induced alveolar collapse and collagen deposition in lung tissues through reducing the expression of TGF-β1and α-SMA. Thus, RAPA has potential value in the treatment of PQ-induced pulmonary fibrosis.