| Objective:Pumlonary interstitial fibrosis is a chronic in?ammatory interstitial lung disease,characterized by the accumulation of alveolar macrophages, neutrophils, and lymphocytes in the distal airspaces, and by parenchymal cell injury and fibrosis of the alveolae. The etiology and pathogenesis of the disease are still unknown.The survival rate of five year is less than 50%,and there are no effective treatments at present.Therefore,furthermore study about its pathogenesis and exploitation effective method have became the focus of present research.This study aims to observe the therapeutic effect of reduced glutathione (GSH) in bleomycin- induced pulmonary fibrosis of rat and compared with glucocorticoid. Explore its mechanisms via measurement the contents of GSH,hypdroxyproline(HYP) and transforming growth factorβ1(TGF-β1).Study the antioxidative effect of GSH of different dose in curing bleomycin-induced pulmonary fibrosis.Method : 90 healthy male Sprague-Dawley(SD) rats (provided by Center Experiment Animal of Hebei Medical University) weighting 220±10g were randomly divided into five groups:(1)normal control group (group N): 18 rats, They were injected about 1ml normal saline in trachea, and were injected normal saline intraperitoneally once a day at one day after fake establishing model, killed randomly at 7th day, 14th day and 28th day (n=6, at each time point).(2)Bleomycin group (group A): 18 rats. Pumlonary fibrosis was induced by intratracheal injection of Bleomycin A5 (5mg/kg). They were injected normal saline intraperitoneally once a day at one day after establishing model, killed randomly at 7th day, 14th day and 28th day. (3) Dexamethasone group(group B): 18 rats,they were injected Bleomycin A5 (5mg/kg) in trachea, and were treated with dexamethasone(0.5mg/kg) intraperitoneally once a day at one day after establishing model,and killed ditto.(4)GSH low dose group(group C): 18 rats,they were injected bleomycin A5 (5mg/kg) in trachea, and were treated with GSH(100mg/kg) intraperitoneally once a day at one day after establishing model,and killed ditto. (5)GSH high dose group(group D): 18 rats,they were injected bleomycin A5 (5mg/kg) in trachea, and were treated with GSH(400mg/kg) intraperitoneally once a day at one day after establishing model,and killed ditto.Before the rats were killed, abdominal aorta were pricked to make arterial blood gas analysis. Six rats in each group were sacrificed at days 7,14,28 after anaesthesia with abdominal aorta depletion method. The level of TGF-β1 was measured by immunohistochemical test,and positive area ratio of TGF-β1 was determined by sem quantitative picture analysis.The contents of GSH and HYP were detected in homogenate made of the inferior lobe of right lung. Estimate the therapeutic effect via hematoxylin-eosin(HE) staining,Masson staining and determination of concentration of HYP of lung tissue.Alveolitis and pulmonary fibrosis were divided by the method of Szapiel into 4 grades(1 point represented no alveolitis or pulmonary fibrosis; 4 points represented severe alveolitis or pulmonary fibrosis,more than a half of area were involved).Results:1 The results of pulmonary pathology: The level of alveolitis of group A was more serious than that of the normal group(P<0.05).The level of fibrosis of group A was more serious than that of the normal group(P<0.05)on the 14th day and 28 th day.These indicate the model of pulmonary fibrosis estabished successfully. The levels of alveolitis and fibrosis of group B and C were less serious than group A. The level of alveolitis of B7 and C7 was less serious than that of A7(P<0.05). The level of alveolitis of B14 and C14 was less serious than that of A14(P<0.05). The level of fibrosis of B28 and C28 was less serious than A28(P<0.05),but there were no significant differences between group B28 and C28 (P>0.05).The level of alveolitis and fibrosis of group D was not less serious than group B.There were no significant differences between them(P>0.05).2 The results of arterial blood gas analysis: On the 7th day,the rats of model group show hypoxemia,hypercapnia and respiratory acidosis.The changes took a turn for the better on the 14th day,but hypoxemia and hypercapnia were still existent.Compared with the normal control group,there were significant differences(P<0.05).On the 28th day,the results of arterial blood gas analysis had no significant differences with the control group(P>0.05).These indicate 7th day and 14th day are the best time points to evaluate the arterial blood gas.There were significant differences between the model group and the treatment group on the 7th day and 14th day (P<0.05). There were no significant differences between group B and C (P>0.05), but there were significant differences between group B and D (P<0.05).3 The change of the contents of GSH in lung tissue: on the 7th day,the content of GSH was significantly lower than that of the normal group (P<0.01); the content of GSH of group B and C was both higher than group A (P<0.05), and there were both significant differences between group A and B and between group A and C, but there were no significant differences between group B and C(P>0.05); the content of GSH of group D was higher than that of group A and B, and there were significant differences(P<0.05). On the 14th day and 28th day, he content of GSH of group A was still lower than that of the normal,but there was no significant differences(P>0.05).4 The change of the content of HYP in lung tissue: The content of HYP of the model group shown an increase tendency accompany with the time.On the 7th day, the content of HYP was significantly higher than that of the normal group(P<0.01), and reached a peak on the 28th day. The content of HYP of the rats in treatment groups also increased,but took on a low level. That of treatment groups was all lower than group A on 7th day,14th day,28th day (P<0.01). The contents of HYP of group B and C were both lower than group A (P<0.01), but there were no significant differences among the group B and C(P>0.01).The content of HYP of group D was significantly lower than group A (P<0.01).5 The expression of TGF-β1 in lung tissue: The expression of TGF-β1 of group B was significantly higher than that of normal group and treatment groups on 7th day,14th day,28th day(P<0.01). That of group B and C were both lower than group A (P<0.01),but there were no significant differences among the two groups on 7th day,14th day,28th day (P>0.01). But the level of TGF-β1 of group D was significantly lower than group B (P<0.01). There is a few of TGF-β1 expressed in the kytoplasm of tunica mucosa bronchiorum endothelial cell in the normal rats, and endothelial cell can express TGF-β1 as well.Conclusions:1 The GSH used in early stage can prevent bleomycin-induced pulmonary fibrosis advance,alleviate the interstitial collagen and raise the level of GSH in the lung.2 Giving the rats GSH 400mg/kg cannot lead to oxidative stress.3 This effct relates with its early antioxidant,and can come true by inhibiting the expression of transforming growth factorβ1. |
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