PGE2对气道平滑肌细胞分泌和凋亡影响的研究

目的前列腺素E2（prostaglandin E2,PGE2）通过调控气道平滑肌细胞（airway smooth muscle cells,ASMCs）的迁移、分泌、增殖功能,对哮喘气道重塑有关键性影响。研究PGE2对ASMCs凋亡及其分泌白介素4（interleukin 4,IL-4）、白介素10（interleukin 10,IL-10）和γ-干扰素（interferon-γ,IFN-γ）的影响,对阐明PGE2通过调控ASMCs分泌Th1/Th2细胞因子对气道炎症发挥作用机制有重要意义。方法用大鼠气道平滑肌细胞株与终浓度为10-9～10-6的PGE2共培养24h,分6组,共4个浓度梯度。用AnnexinV-EGFP/PI双染细胞凋亡检测试剂盒在流式细胞仪测定凋亡率,同时用IL-4、IL-10和IFN-γELISA试剂盒在酶标仪检测其浓度。每4个复孔为1组,实验重复2次,数据为8孔的平均值。结果应用不同浓度的PGE2后ASMCs各组细胞凋亡率与对照组相比无明显变化。但IL分泌与PGE2有显著的量效关系,IL-4和IL-10随PGE2浓度下降而分泌量显著下降,但IFN-γ随PGE2浓度下降而分泌量呈升高趋势,两者呈分离现象（空白对照、阴性对照、10-9PGE2、10-8PGE2、10-7PGE2、10-6PGE2组,IL-4分别为6.77±1.58、9.24±2.45、38.17±11.33、29.27±11.12、26.05±9.49、21.11±7.21;IL-10为4.8±1.06、6.35±2.11、71.89±12.03、43.68±11.25、28.89±8.75、24.31±6.67;IFN-γ为2.17±1.97、3.25±1.48、10.63±4.75、13.4±5.57、15.47±6.16、19.54±6.35）。结论 PGE2能调控ASMCs分泌细胞因子,随着浓度梯度增加能促进Th1型细胞因子IFN-γ分泌,同时抑制Th2型细胞因子IL-4、IL-10分泌,但对ASMCs凋亡无显著影响。认为在正常状态下PGE2的分泌与维持气道内Th1/Th2的平衡有关。

Effect of PGE2 on secretion function and apoptosis of airway smooth muscle cells

Objective Airway remodeling is closely related to migration, proliferation, secretion function of airway smooth muscle cells (ASMCs). Prostaglandin E2 ( PGE2) have a regulatory role in these functions. To culture with different concentration gradient of PGE2 and ASMCs in vitro and to study ASMCs secretion functions of interleukin 4 (IL-4) and interleukin 10 (IL-10) and Interferon-γ(IFN- γ),and apoptosis of ASMCs,this is favorable to understand ASMCs immune regulation mechanism in asthma, and the regulation role of PGE2.Methods The rat airway smooth muscle cells and the final concentration of 10-9~10-6PGE2 were cultured for 24 hours, divided into 6 groups, and 4 concentration gradient. Flow cytometric were used determination of apoptosis with Annexin V-EGFP/PI double staining apoptosis detection kit, United States Bio-Rad 680 enzyme mark instrument were used detection of its concentration with IL-10, IL-4, IFN-γ ELISA kit. 4 complex holes into 1 groups, the experiment was repeated 2 times, average data for 8 holes.Results There were no significant changes in ASMCs cell apoptosis rate after application of the different concentrations of PGE2 compared with the control group. But interleukin secretion and PGE2 has significant dose-effect relationship.When PGE2 concentration were decreased, the secretion of IL-4 and IL-10 decreased significantly, but the IFN- γ secretion was increased.( Control, negative,10-9PGE2,10-8PGE2,10-7PGE2,10-6PGE2 group, IL-4 was respectively 6.77±1.58,9.24±2.45,38.17±11.33,29.27±11.12,26.05±9.49,21.11±7.21;IL-10 was 4.8±1.06,6.35±2.11,71.89±12.03,43.68±11.25,28.89±8.75,24.31±6.67;2.17±1.97,IFN-γ 3.25±1.48,10.63±4.75,13.4 ±5.57,15.47±6.16,19.54±6.35).Conclusions PGE2 can regulate the secretion of ASMCs cytokines, increased with concentration gradient can promote Th1 cytokines IFN-γ increased secretion, while inhibition of type Th2 cytokine secretion of IL-10, IL-4, but no significant effect on the apoptosis of ASMCs. We think the key role of the normal state of PGE2 secretion were related to maintaining Th1/ Th2 balance in the airway.