摘要
目的观察脂多糖(lipopolysac charide,LPS)作用下大鼠肺泡Ⅱ型上皮(typeⅡalveol arepith elial,AT-Ⅱ)细胞过氧化物酶增殖体激活受体γ(PPARγ)的表达变化情况。方法通过酶消化分离和免疫黏附法纯化原代培养AT-Ⅱ细胞,并进行碱性磷酸酶、肺表面活性物质相关蛋白A(SP-A)和电镜鉴定。采用RT-PCR法检测细胞经LPS作用后PPARγ、TNF-α、SP-AmRNA的改变,Westernblot法检测LPS作用后细胞PPARγ表达,ELISA法检测TNF-α蛋白表达。结果经碱性磷酸酶、SP-A和电镜鉴定,原代培养AT-Ⅱ细胞成功;在致炎因子LPS作用下,PPARγmRNA和蛋白表达均下降,这一变化伴随炎性因子TNF-α的升高。结论在LPS作用下,AT-Ⅱ细胞内PPARγ表达降低,提示PPARγ参与炎症反应。
Objective To determine the effect of LPS stimulation on the expression of PPARγ in rat type Ⅱ alveolar epithelial （AT-Ⅱ） cells. Methods Rat lung tissues were digested by trypsase and collagenase. AT-Ⅱ cells were isolated and purified by immuno-adherence, and then primarily cultured AT-Ⅱ cells were identified by alkaline phosphatase （AKP）, pulmonary surfactant-associated protein A （SP-A） staining and electron microscopy. PPARγ, TNF-α and SP-A mRNA levels in AT-Ⅱcells were assayed by RT-PCR after being activated by lipopolysaccharide （LPS） stimulation, while the PPARγ and TNF-α protein levels in culture supernatants were measured by Western blotting and ELISA respectively. Results Primary culture of rat AT-Ⅱ cells were successfully established as confirmed by the expression of AKP and SP-A and unltrastructural features. PPARγ expression in AT-Ⅱ cells were decreased when stimulated with LPS, which were accompanied by increased TNFα expression. Conclusion Pro-inflammatory substance LPS can markedly inhibit the expression of PPARγ in AT-Ⅱcells, which could be involved in the loss of control of inflammation.
出处
《第三军医大学学报》
CAS
CSCD
北大核心
2009年第20期1960-1963,共4页
Journal of Third Military Medical University
基金
国家自然科学基金(30570808)
全军医学科研“十一五”计划科技攻关项目(06G083)~~
