CAJ | 학술논문

目的观察金黄色葡萄球菌外毒素及中件粒细胞对视网膜色素卜皮(RPE)细胞的毒性作用.方法分离人外周血L中性粒细胞,和人RPE细胞株D407(RPE D407)共培养,并加入金黄色葡萄球菌ATCC29213外毒素建立细菌性眼内炎体外模型.通过观察RPE细胞培养液中乳酸脱氢酶(LDH)水平、流式细胞仪分析RPE细胞存活状况改变、以及Hoechst 33342/Propidium Iodide(P1)染色观察细胞核的荧光染色等方法,验证金黄色葡萄球菌外毒素及中性粒细胞对RPE细胞的损伤作用.结果金黄色葡萄球菌外毒素单独作用RPE细胞时,细胞外LDH水平及RPE细胞的坏死细胞比率随着外毒素剂量增加而升高,但仪250、500 μl组差异有统计学意义.当加入中性粒细胞共同培养后,100μl外毒素作用下,仪6 h,RPE细胞外LDH水平即高于无外毒素组,差异具有统计学意义;在6 h时,流式细胞仪分析显示,RPE细胞发生了明显的细胞凋亡,并且随着中性粒细胞数量增多而加剧,差异具有统计学意义.结论金黄色葡萄球菌外毒素对RPE细胞具有毒性作用,其机制主要以细胞坏死的方式进行.中性粒细胞超过一定数量后对RPE细胞具有损伤作用.外毒素和中性粒细胞共同参与时,对RPE细胞的毒性作用大丁两者单独对RPE细胞的损害.
목적 관찰금황색포도구균외독소급중건립세포대시망막색소복피(RPE)세포적독성작용.방법 분리인외주혈L중성립세포,화인RPE세포주D407(RPE D407)공배양,병가입금황색포도구균ATCC29213외독소건립세균성안내염체외모형.통과관찰RPE세포배양액중유산탈경매(LDH)수평、류식세포의분석RPE세포존활상황개변、이급Hoechst 33342/Propidium Iodide(P1)염색관찰세포핵적형광염색등방법,험증금황색포도구균외독소급중성립세포대RPE세포적손상작용.결과 금황색포도구균외독소단독작용RPE세포시,세포외LDH수평급RPE세포적배사세포비솔수착외독소제량증가이승고,단의250、500 μl조차이유통계학의의.당가입중성립세포공동배양후,100μl외독소작용하,의6 h,RPE세포외LDH수평즉고우무외독소조,차이구유통계학의의;재6 h시,류식세포의분석현시,RPE세포발생료명현적세포조망,병차수착중성립세포수량증다이가극,차이구유통계학의의.결론 금황색포도구균외독소대RPE세포구유독성작용,기궤제주요이세포배사적방식진행.중성립세포초과일정수량후대RPE세포구유손상작용.외독소화중성립세포공동삼여시,대RPE세포적독성작용대정량자단독대RPE세포적손해.
Objective To evaluate the toxic effects of staphylococcus aurcus exotoxins andneutrophils on retinal pigment epithelium(RPE)cells(RPEC).Methods An in-vitro model ofbacteroidal endophthalmitis was established by co-culturing of human RPE cell line D407 and humanperipheral blood neutrophils in the present of staphylococcus aureus exotoxins ATCC29213.The level oflactate dehydrogcnase hydroxide(LDH)in the cuture supernant was measured,and the viability of RPEwas evlauated by flow cytometry and Hoechst 33342/propidium iodide(PI)staining.Results WhenRPE cells were cultured with the exotoxin ATCC29213,the LDH level and necrotic RPE cells werepositive proportional to the dosage of exotoxin,but only 250 μl or 500 μl of ATCC29213 had a statisticalsignificant effect.When RPE cells were co-cultured with neutrophils in the present of ATCC29213 for 6hours,100 μl of ATCC29213 already had a statistical significant effect on LDH level and necrotic RPEC,and the effect was proportional to the amount of neutrophils in the culture.Conclusion Bothstaphylococcus aureus exotoxins and neutrophils can damage the RPEC by inducing necrosis,and theirfunction had synergetic effect.