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探讨骨桥蛋白(osteopontin,OPN)对博莱霉素(bleomycin,BLM)诱导的大鼠肺纤维化肺组织基质金属蛋白酶-9(MMP-9)、金属基质蛋白酶抑制剂-1(TIMP-1)和Ⅰ、Ⅲ型胶原mRNA表达的影响。取72只健康雌性Wistar大鼠随机分成生理盐水对照组(NS组)、博莱霉素模型组(BLM组)和骨桥蛋白抗体干预组(OPN-Ab组)各24只,气管内灌注BLM复制肺纤维化模型(BLM组、OPN-Ab干预组),NS组气管内灌注生理盐水,OPN-Ab组于0d、2d、4d、6d尾静脉注射骨桥蛋白抗体(1:32),BLM组和NS组尾静脉注射生理盐水,各组分别于7d、14d、28d处死动物8只。收集肺组织作切片行HE、MASSON染色测定肺泡炎症和纤维化改变,免疫组织化学PAP法测定肺组织中OPN、MMP-9、TIMP-1的表达;RT-PCR测定肺组织Ⅰ/Ⅲ型胶原mRNA的表达。结果与BLM组比较,在第7天和第14天OPN-Ab组肺泡炎明显减轻(P<0.01);而肺纤维化程度在各时间点均明显减轻(P<0.05);与NS组比较,BLM组、OPN-Ab组肺组织中OPN、MMP-9、TIMP-1表达水平均显著升高(P<0.01)。BLM组第7天OPN、MMP-9、TIMP-1在肺组织中表达水平显著升高,随后下降,第28天时仍高于NS组(P<0.01)。与BLM组比较,OPN-Ab组各时间点OPN、MMP-9、TIMP-1表达水平降低(P<0.01)。OPN-Ab组第7、14、28天肺组织Ⅰ、Ⅲ型胶原mRNA表达均较同时间点BLM组明显降低(P<0.05)。OPN可能是肺纤维化形成机制中的重要因素,骨桥蛋白抗体可能通过抑制细胞因子MMP-9、TIMP-1表达,减少肺组织中胶原的生成,最终抑制肺纤维化。
To investigate the effects of osteopontin (OPN) on lung fibrosis induced by bleomycin (BLM) in rats with pulmonary fibrosis and the expression of matrix metalloproteinase-9 (MMP-9), matrix metalloproteinase-1 ) And type Ⅰ, Ⅲ collagen mRNA expression. Totally 72 healthy female Wistar rats were randomly divided into saline control group (NS group), bleomycin model group (BLM group) and osteopontin antibody intervention group (24), and intratracheal instillation of BLM The lung fibrosis model (BLM group, OPN-Ab intervention group) was injected into the tail vein of rats in NS group. The osteopontin antibody (1:32) was injected into the caudal vein on the 0d, 2d, 4d and 6d after intratracheal instillation of saline and OPN- The rats in NS group and NS group were injected with saline, and the animals in each group were sacrificed at 7d, 14d and 28d respectively. Pulmonary tissue was collected for HE staining and MASSON staining for alveolar inflammation and fibrosis. Immunohistochemical PAP method was used to detect the expression of OPN, MMP-9 and TIMP-1 in lung tissue. RT-PCR was used to detect the expression of type Ⅰ / Ⅲ collagen mRNA expression. Results Compared with the BLM group, the alveolitis of OPN-Ab group was significantly reduced on the 7th and 14th days (P <0.01), while the degree of pulmonary fibrosis was significantly reduced at all time points (P <0.05). Compared with the NS group The expression of OPN, MMP-9 and TIMP-1 in BLM group and OPN-Ab group were significantly increased (P <0.01). On day 7, the expression of OPN, MMP-9 and TIMP-1 in BLM group was significantly increased in lung tissue, and then decreased, still higher than that in NS group on the 28th day (P <0.01). Compared with BLM group, the expression of OPN, MMP-9 and TIMP-1 at each time point in OPN-Ab group decreased (P <0.01). The mRNA expression of type I and type III collagen in lung tissue of OPN-Ab group was significantly lower than that of BLM group on the 7th, 14th and 28th day (P <0.05). OPN may be an important factor in the formation of pulmonary fibrosis. Osteopontin antibody may inhibit pulmonary fibrosis by inhibiting the expression of cytokines MMP-9 and TIMP-1, reducing the production of collagen in lung tissue.