脂多糖LPS对糖尿病大鼠肺泡巨噬细胞RBD-2表达的影响

摘要

目的 以肺泡巨噬细胞为研究对象,现察脂多糖(lipopolysaccharide,LPS)刺激下RBD-2(大鼠β防御素-2,Rat β-defensins-2)在正常大鼠及糖尿病大鼠肺泡巨噬细胞表达的变化.方法 以健康雄性SD大鼠制备糖尿病模型.32只大鼠随机分为四组:正常对照组（A组）、耱尿病组（B组）、LPS组（C组）和糖尿病+LPS组（D组）,每组均为8只.分离培养大鼠肺泡巨噬细胞,通过RT-PCR以及Western blotting分别检测RBD-2的RNA及蛋白质表达水平,同时通过Real Time PCR检测大鼠肺泡巨噬细胞的TLR-2以及TLR-4的mRNA的表达.结果 大鼠耱尿病模型构建成功.RT-PCR以及Western blotting结果显示,与正常组相比,糖尿病组、正常组+LPS组、糖尿病十脂多糖组的RBD-2 mRNA以及蛋白质表达水平依次增加,差异显著(P＜0.05).Real Time PCR结果显示,与正常组相比,糖尿病组、正常组+ LPS组、糖尿病十脂多糖组的TLR-4 mRNA的表达水平依次增加,差异显著(P＜0.05),而TLR-2差异则不明显.结论 LPS刺激后糖尿病大鼠肺泡巨噬细胞RBD-2表达增加较糖尿病组更为显著,说明RBD-2变化对于增强糖尿病机体的非特异性免疫能力具有明显的帮助,同时糖尿病大鼠肺泡巨噬细胞RBD-2表达较正常组增高,说明处于耱尿病时期的大鼠机体处于炎症状态,并且这一通路的表达受体主要是TLR-4受体.%Objective To observe the expression of rat β-defensin 2 (RBD-2) in alveolar macrophages of diabetic rats after lipopolysaccharide (LPS) stimulation. Methods Healthy SD rats were used to construct the diabetic models. Then they were randomly divided into four groups. Group A: the control group; Group B; the diabetic group; Group C; the LPS stimulated group; Group D; the diabetic group with LPS infection. Rat alveolar macrophages were captured and cultured. The RT-PCR and Western blotting method were utilized to detect the mRNA and protein level of RBD-2. The Real Time PCR method were used to determine the mRNA level of TLR-2 and TLR-4. Results The results of RT-PCR and Western blotting indicated that the expression level of RBD-2 mRNA and protein in group B, group C and group D followed by increased significantly (P<0. 05). Subsequently, the results of Real Time PCR indicated that the expression level of TLR-4 mRNA in group B, group C and group D followed by increased significantly (P<0. 05) but that of the TLR-2 was different. Conclusion The expression of RBD-2 is obvious after LPS stimulation, which suggests that RBD-2 help to enhance the nonspecific immunity in diabetic rat. Subsequently, the expression of RBD-2 of diabetic rats also indicates that diabetic bodies are in the pro-inflammatory state and the main receptor was TLR-4.