CAJ | 학술논문

目的探讨邻苯二甲酸二乙基己酯( di-2-ethylhexyl phthalate, DEHP)体外对大鼠睾丸支持细胞紧密连接的作用以及相关基因表达的影响。方法体外分离和原代培养大鼠睾丸支持细胞,建立支持细胞原代双室培养模型,以只加入二甲基亚砜(DMSO)的细胞孔作为对照,以10、50、100 nmol/L浓度的DEHP(DEHP低、中、高剂量组)染毒24 h。应用跨上皮电阻测定法检测单层细胞两侧跨上皮细胞电阻值( TER),荧光定量聚合酶链反应( PCR)法检测支持细胞闭锁蛋白(Occludin)、闭锁小带蛋白1(Zonula occluden-1, ZO-1)、紧密连接蛋白11(Claudin 11)基因mRNA表达,应用Weastern blot检测支持细胞Occludin、ZO-1、Claudin 11蛋白的表达。结果与对照组比较,不同浓度的DEHP各组细胞TER值均降低。同时大鼠睾丸支持细胞Occludin、ZO-1、Claudin 11基因mRNA表达随DEHP浓度增加而降低(P<0.05),Occludin、ZO-1、Claudin 11蛋白表达也随DEHP浓度增加而降低(P<0.05)。结论 DEHP可通过下调Occludin、ZO-1、Claudin 11基因表达而影响大鼠睾丸支持细胞的紧密连接,进而影响支持细胞的功能。
목적탐토린분이갑산이을기기지( di-2-ethylhexyl phthalate, DEHP)체외대대서고환지지세포긴밀련접적작용이급상관기인표체적영향。방법체외분리화원대배양대서고환지지세포,건립지지세포원대쌍실배양모형,이지가입이갑기아풍(DMSO)적세포공작위대조,이10、50、100 nmol/L농도적DEHP(DEHP저、중、고제량조)염독24 h。응용과상피전조측정법검측단층세포량측과상피세포전조치( TER),형광정량취합매련반응( PCR)법검측지지세포폐쇄단백(Occludin)、폐쇄소대단백1(Zonula occluden-1, ZO-1)、긴밀련접단백11(Claudin 11)기인mRNA표체,응용Weastern blot검측지지세포Occludin、ZO-1、Claudin 11단백적표체。결과여대조조비교,불동농도적DEHP각조세포TER치균강저。동시대서고환지지세포Occludin、ZO-1、Claudin 11기인mRNA표체수DEHP농도증가이강저(P<0.05),Occludin、ZO-1、Claudin 11단백표체야수DEHP농도증가이강저(P<0.05)。결론 DEHP가통과하조Occludin、ZO-1、Claudin 11기인표체이영향대서고환지지세포적긴밀련접,진이영향지지세포적공능。
Objective To explore the effect of di-2-ethylhexyl phthalate ( DEHP) on tight junction and related gene ex-pression of sertoli cells in vitro in rats. Methods The rats' sertoli cells were separated in vitro and primarily cultured, and dual-chamber models for sertoli primary culture were established, and then sertoli cells were treated with different concentrations of DEHP (10, 50 and 100 nmol/L) for 24 h. The transepithelia electrical resistance (TER) assay was used to detect TER value;fluorescent quantitation polymerase chain reaction (FQ-PCR) was used to detect mRNA of Occludin, Zonula occluden-1 (ZO-1) and Claudin 11 gene expressions, and the Western blot method was used to detect the protein expressions of Occludin, ZO-1 and Claudin 11. Results Compared with those in control group, the TER values decreased in different concentrations DEHP groups. Meanwhile, the Occludin, ZO-1 and Claudin 11 gene of mRNA expressions decreased with increasing DEHP concentra-tion (P<0. 05), and the Occludin, ZO-1 and Claudin 11 protein expressions also decreased with increasing DEHP concentra-tion (P<0. 05). Conclusion DEHP can downregulate Occludin, ZO-1 and Claudin 11 gene expressions to affect tight junction of sertoli cells in rats, which may affect sertoli cell function.