| Idiopathic pulmonary fibrosis(IPF) is a chronic,diffused and progressive interstitial lung disease of unknown etiology with its pathologic feature of usual interstitial pneumonia(UIP).Corticosteroids or corticosteroids associated with immunodepressants are currently used for the treatment of IPF,but no evident curative effect was shown.Arsenic trioxide(ATO) has been efficaciously used as a medical agent for acute progranulocyte leukemia(APL) with significant curative effect and tiny side effect,and its application in other diseases has also been reported,for instance,it has been observed that asthma in animals could be relieved by ATO for its induction of acidophil apoptosis.Based on the above findings,it is inferred that whether ATO could be used to treat IPF by inducing apoptosis of some kinds of cells in lung tissue(inflammatory cells or fibroblasts).Furthermore,being a classical agent for IPF,corticosteroids are still worth studying.Therefore,this article is to observe the curative effect of ATO and its association with dexamethasone to treat IPF,and to investigate its mechanism from the point of inducing cell apoptosis.Part 1Objective To observe the therapeutic and side effect of different dosages of dexamethasone(DXM) on bleomycin-induced lung fibrosis in rats and to investigate the effective and safe dosage of DXM on lung fibrosis models for the preparation of formal experiment.Methods 144 healthy male Sprague-Dawley(SD) rats were randomly divided into four groups including control group(n=18),DXM side effect group(n=54),model group(n=18) and DXM treated group(n=54).DXM side effect group and DXM treated group were then divided into three subgroups respectively(18 rats each subgroup) according to different dosages of DXM including 1mg/kg,2mg/kg and 3mg/kg subgroup.Finally,each group was divided into three groups randomly(6 ratseach group), which were sacrificed on 14d,28d and 56d.Survival analysis was done and HE staining was performed on the left lung tissue sections to observe the extent of alveolitis and fibrosis.Hydroxyproline(HYP) assays of the right lung tissue of rats in control group, model group and DXM treated group were also made to estimate the collagen content.The liver and kidney tissue sections were got HE stained to observe the structure and inflammation.Results(1) The general status of rats in control group was much better than that of the other three experimental groups,and the rats in the three experimental groups were much lighter than that of the control group,especially for that of the DXM side effect groups and treated groups.(2) The death rate by 56d of 3mg/kg DXM side effect group and treated group was significantly higher as compared to that of homologous control group, model group,1mg/kg DXM side effect group and treated group(P＜0.05).(3) The alveolitis and fibrosis induced by bleomycin could be relieved by DXM in some extent,while the infection rate was getting higher with the increase of its dosage and treated durations,and no more improvement was shown on lung fibrosis.(4) HYP analysis showed that most treated groups had a lower content of collagen than homologous model groups and there is no significant difference of HYP among the treated groups which were treated by three different dosages of DXM respectively.(5) The livers in DXM side effect groups and treated groups got fatty degeneration generally,the renal tubules in the above groups suffered infection.Conclusion 1mg/kg DXM is an effective and safe choice for the formal experiment to ameliorate the formation of bleomycin-induced lung fibrosis in rats,which wouldn't result in serious infection and high death rate.Part 2Objective To observe the therapeutic effect of arsenic trioxide against bleomycin-induced pulmonary fibrosis in rats and to investigate its mechanism on cell apoptosis.Methods 240 Sprague-Dawley(SD) rats were randomly divided into five groups including control group,model group,arsenic trioxide treated group,dexamethasone treated group and arsenic trioxide plus dexamethasone treated group(48 rats for each group).Each group was separated into two subgroups according to different time-points (14 days and 28 days) at which those rats started receiving treatment after pulmonary fibrosis models were made.These two subgroups were eventually divided into three groups for different treatment durations(14days,28days and 56days).In different stages, the rats were sacrificed respectively,HE staining was performed on the lung tissue sections to observe the extent of alveolitis and fibrosis,the hydroxyproline assays of the lung tissues of rats were also made to estimate the collagen content,the ultrastructure changes were analyzed by transmissional electromicroscope,immunohistochemistry was used to detect the expression of those apoptosis-related genes such as P53,C-myc,Bax and Bcl-2 and the survival rate was also evaluated.Results(1) Pathological scores:the pathological scores in the control groups were significantly lower than that of any other groups(P＜0.05).The pathological scores in all of the treated groups exclusive of the dexamethasone treated 28d and 56d groups after fibrosis models were made for 28 days,were lower than that of the model groups,but only arsenic trioxide treated groups,arsenic trioxide plus dexamethasone treated groups and dexamethasone treated 14d group(all these treated groups got treatment 14 days after fibrosis models were made) showed statistical difference compared to the model groups.(2) Hydroxyproline analysis:hydroxyproline analysis showed that control groups had a much lower content of collagen than any other groups,and content of Hydroxyproline in treated groups was lower than that of model groups except in dexamethasone treated 28d,56d groups and in dexamethasone plus arsenic trioxide treated 28d,56d groups after fibrosis models were made for 28 days,but only part of the treated groups that get treatment 14 days after fibrosis models were made had a significantly lower content than that of model groups(P＜0.05).(3) Transmissional electromicroscope observations:the collagen deposition was much less in the arsenic trioxide treated group than that of the model group and the number of apoptotic inflammatory cells was reverse under transmissional electromicroscope.(4) Expression of apoptosis-related genes:the expression of P53 and C-myc in all treated groups was much higher than that of model groups(P＜0.05).The expression of Bax in most treated groups was significantly higher than that of model groups(P＜0.05),while the expression of Bcl-2 in arsenic trioxide treated groups was the reverse.Most of the dexamethasone treated groups had a significantly higher expression of Bcl-2 than that of model groups(P＜0.05).Conclusion Suitable dosage of arsenic trioxide had an effect of reducing collagen deposition on pulmonary fibrosis induced by Bleomycin in rats and earlier treatment was much better,its infection rate was significantly lower than dexamethasone treated group.The mechanism of it was probably related to the effect of inducing apoptosis in certain kinds of cells,especially the inflammatory cells.
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