CAJ | 학술논문

目的探讨环磷酰胺( cyclophosphamide,CP)对术成熟睾丸血睾屏障紧密连接的损伤及其机制.方法建立未成熟SD大鼠CP化疗损伤模型,电镜技术观察睾丸支持细胞间紧密连接的改变,S-P免疫组化法及免疫印迹法检测细胞间紧密连接蛋白 Occludin及ZO-1的表达、免疫荧光检测细胞骨架F-actin的表达情况,初步探讨体内CP损伤血睾屏障的机制.结果 ①电镜检测:对照组可见支持细胞连接处光滑清楚,走行自然；而处理组连接处细胞膜分离,其间出现较大间隙,连接旁内质网也出现程度不等的扩张；②SP免疫组化检测:Occludin及ZO-1的表达与对照组比较未见明显异常(P＞0.05):③免疫荧光检测:在对照组中可见F-actin在生精上皮及支持细胞中呈强荧光表达,而环磷酰胺处理后F-actin的荧光染色明显减少；④免疫印迹检测:结果发现F-actin的表达较对照组明显下降(P＜0.05),而紧密连接蛋白Occludin及ZO-1的表达与对照组比较差异未见明显异常(P＞0.05).结论环磷酰胺可能通过下调F-actin的表达,干扰细胞紧密连接蛋白的功能,使血睾屏障紧密连接受到损伤.
목적 탐토배린선알( cyclophosphamide,CP)대술성숙고환혈고병장긴밀련접적손상급기궤제.방법 건립미성숙SD대서CP화료손상모형,전경기술관찰고환지지세포간긴밀련접적개변,S-P면역조화법급면역인적법검측세포간긴밀련접단백 Occludin급ZO-1적표체、면역형광검측세포골가F-actin적표체정황,초보탐토체내CP손상혈고병장적궤제.결과 ①전경검측:대조조가견지지세포련접처광활청초,주행자연；이처리조련접처세포막분리,기간출현교대간극,련접방내질망야출현정도불등적확장；②SP면역조화검측:Occludin급ZO-1적표체여대조조비교미견명현이상(P＞0.05):③면역형광검측:재대조조중가견F-actin재생정상피급지지세포중정강형광표체,이배린선알처리후F-actin적형광염색명현감소；④면역인적검측:결과발현F-actin적표체교대조조명현하강(P＜0.05),이긴밀련접단백Occludin급ZO-1적표체여대조조비교차이미견명현이상(P＞0.05).결론 배린선알가능통과하조F-actin적표체,간우세포긴밀련접단백적공능,사혈고병장긴밀련접수도손상.
Objective To investigate the impairment of cyclophospbamide on the tight-junction and blood-testis barrier in immature testis.Methods Cyclophosphamide was administered to immature SD rats to induce testicular injury.The control rats were administered with saline.Electron microscopy was employed to study the morphological changes of testicular tissue.The expressions of Occludin and ZO-1 were detected by immunohistochemical staining and western blotting.The expression of Factin,a cytoskeleton protein,was detected by immunofluorescence.Results In control testis,the tight junctions of Sertoli cell was smooth and clear.However,after cyclophosphamide treatment,abnormal wide gaps were found between junctions,and the endoplasmic reticulum closed to the junctions expanded.Immunohistochemical staining showed that there were no significant difference of the expressions of Occludin and ZO-1 between the control testis and cyclophosphamide treated testis ( P＞0.05).However,the fluorescence of F-actin in sertoli cells and spcrmatogenic cell was significantly decreased after cyclophosphamide treatment (P＜0.05).Western blotting showed the expression of Factin was significantly decreased in the eyclophosphamide treated testis (P＜0.05).No Significant difference of Occludin and ZO-1 was found between the control testis and cyclophosphamide treated testis (P＞0.05).Conclusions Cyclophosphamide decreases the expression of F-actin and disturbs the function of tight-junction protein to compromise the stability of blood-testis barrier in immature testis.