摘要
目的观察睾酮对体外培养牛眼小梁细胞(bovine trabecular meshwork cells,BTMC)中的组织型谷氨酰胺转移酶(tis-sue transglutaminase,tTG)在mRNA水平和蛋白水平表达的影响,初步探讨睾酮与原发性开角型青光眼(primary open-angle glau-coma,POAG)发病的关系。方法取新生牛BTMC原代培养和传代培养,通过细胞形态学及免疫组化染色法进行鉴定后,取3～5代BTMC用于实验。用不同浓度的睾酮作用24h。采用RT-PCR与Western-blotting方法检测不同浓度睾酮作用前后BT-MCtTG表达水平的变化。结果分别加入浓度为18×10-3mmol.L-1、36×10-3mmol.L-1、54×10-3mmol.L-1、72×10-3mmol.L-1睾酮的培养液作用后,RT-PCR产物电泳结果显示正常对照组(不含睾酮)、睾酮组均在433bp处出现特异性的tTG基因条带,但睾酮组表达明显高于正常对照组;各组均在212bp处出现β-actin的扩增产物的基因条带,且表达无明显差别。激光灰度值扫描进行半定量分析后,不同浓度睾酮组的tTG相对积分光密度值分别为1.461±0.021、2.111±0.016、2.536±0.032、3.176±0.026,其中后三个浓度组与正常对照组的相对积分光密度值1.487±0.013比较,差异均有统计学意义(均为P<0.05),但18×10-3mmol.L-1睾酮组与正常对照组间差异无统计学意义。Western blotting检测结果显示tTG的蛋白表达水平也随睾酮浓度变化而变化。结论雄激素睾酮可显著增加BTMC中tTG的表达。tTG的增加可能与POAG的发病有关。提示雄激素可能通过tTG作用而与POAG发病有一定关系。
Objective To observe the effects of testosterone on the expression of tissue transglutaminase（tTG） at the mRNA and protein levels in cultured bovine tra- becular meshwork cells （ BTMC ）, and evaluate the relationship between testosterone and the pathogenesis of primary open-angle glaucoma（POAG）. Methods The newborn bovine BTMC was primary cultured and passaged, after the identification by mor- phological and immunohistochemical staining, the third to fifth generation BTMC were chosen for the experiment. Different concentrations of testosterone were applied in the chosen BTMC for 24 hours,then RT-PCR and Western-blotting were used to detect the tTG expression at different levels in these BTMC. Results After adding different con- centrations（ 18 × 10-3 mmol · L-1 ,36 ×10-3 mmol · L-1 ,54 × 10-3 mmol · L-1 ,72 ×10-3 mmol · L - 1 ） of testosterone culture media, the electrophoresis results of RT-PCR prod- ucts revealed that a specific tTG gene band was appeared in the 433 bp of both control group and testosterone group, while the gene expression in androgen group was signifi- cantly higher than that of control group. Meanwhile, each group showed 212 bp gene bands as β-actin amplification products, and there was no obvious difference between these expressions. After semi-quantitative analysis by laser scanning of gray value, the relative integrated values in four concentration groups were 1. 461 ± 0. 021,2. 111 ± 0. 016,2.536 ± 0. 032,3. 176 ± 0. 026, respectively, and the control group was 1. 487 ± 0. 013 ,there was no statistical difference between control group and 18 ± 10 -3 mmol · L-1 testosterone group, while there were statistical differences between control group and other testosterone groups （ all P 〈 0.05 ）. Conclusion Testosterone can significantly increase the expression of tTG in BTMC, and the increase of tTG may be associated with the onset of POAG, which indicates androgens may have some associations with the onset of POAG through the function of tTG.
出处
《眼科新进展》
CAS
北大核心
2012年第11期1027-1029,共3页
Recent Advances in Ophthalmology
