CAJ | 학술논문

目的：通过探索适宜的培养条件，建立小鼠睾丸器官培养模型。方法利用旋转通气培养法在体外培养睾丸，优化培养条件，并与Transwell小室培养法进行形态学比较。HE染色观察睾丸组织结构的变化，BrdU染色法观察睾丸在体外的细胞增殖，放射免疫法观察体外培养睾丸的睾酮分泌能力，免疫组化法观察睾丸中功能蛋白的表达。结果通过培养结局的对比，旋转通气培养法培养的睾丸组织形态更完整。睾丸大小以0.3~0.8 mm3为宜。各组精原细胞增殖指数呈上升趋势，Sertoli细胞增殖指数呈下降趋势：3 d组精原细胞增殖指数的增加与1 d组相比有统计学差异（P<0.05），5 d组Sertoli细胞增殖指数的减小与1 d组相比有统计学差异（P<0.05）。睾酮分泌量随培养天数的增加而减少且差异均有统计学意义（P<0.05）。培养3 d后，Leydig细胞胞浆内可见3β-羟基类固醇脱氢酶（3β-HSD）、细胞色素P45017α羟化酶（P450c17）、胆固醇侧链裂解酶（P450Scc）蛋白的表达， Sertoli细胞胞浆内可见波形蛋白（Vimentin）表达。结论利用旋转通气培养法成功建立睾丸器官体外培养的模型。
목적：통과탐색괄의적배양조건，건립소서고환기관배양모형。방법이용선전통기배양법재체외배양고환，우화배양조건，병여Transwell소실배양법진행형태학비교。HE염색관찰고환조직결구적변화，BrdU염색법관찰고환재체외적세포증식，방사면역법관찰체외배양고환적고동분비능력，면역조화법관찰고환중공능단백적표체。결과통과배양결국적대비，선전통기배양법배양적고환조직형태경완정。고환대소이0.3~0.8 mm3위의。각조정원세포증식지수정상승추세，Sertoli세포증식지수정하강추세：3 d조정원세포증식지수적증가여1 d조상비유통계학차이（P<0.05），5 d조Sertoli세포증식지수적감소여1 d조상비유통계학차이（P<0.05）。고동분비량수배양천수적증가이감소차차이균유통계학의의（P<0.05）。배양3 d후，Leydig세포포장내가견3β-간기류고순탈경매（3β-HSD）、세포색소P45017α간화매（P450c17）、담고순측련렬해매（P450Scc）단백적표체， Sertoli세포포장내가견파형단백（Vimentin）표체。결론이용선전통기배양법성공건립고환기관체외배양적모형。
Objective To establish an in vitro model of cultured mouse testis using rotary aerobic culture. Methods Rotary aerobic incubation with optimized culture conditions was used for in vitro culture of mouse testis, and the morphology of the cultured testicular tissues was compared with that cultured in Transwell chambers. The changes in the testicular tissue structure were examined using HE staining, and the cell proliferation was assessed with BrdU staining. Testosterone concentrations in the culture medium were tested with radioimmunoassay and the expression of the functionally related proteins in the testis was detected using immunohistochemistry. Results The testicular tissue cultured by optimized rotary aerobic culture presented with more intact histological structure with the size of the testis ranged from 0.3 to 0.8 mm3. In the two culture systems, the prolifeation index of the spermatogonia increased and that of Sertoli cells decreased with time, and such changes in spermatogonia and Sertoli cell proliferation indices became statistically significant at 3 days (P<0.05) and 5 days (P<0.05) of culture, respectively, as compared with those at 1 day. The concentration of testoerone in the culture media decreased significantly with incubation time (P<0.05). At 3 days of culture, the protein expression of 3β-hydroxysteroid dehydrogenase, cytochrome P450 17α-hydroxylase and cholesterol side-chain cleavage enzyme was detected in Leydig cell cytoplasm and vimentin expression in Sertoli cell cytoplasm. Conclusion An in vitro model of cultured mouse testis has been successfully established using rotary aerobic incubation.