Production of nitric oxide by Sertoli cell and its influence upon the cells

从幼年大鼠分离的生精上皮细胞(Sertoli cell，SC)在双室系统和15 mm圆形玻片上进行培养，以验证一氧化氮(NO)对SC蛋白分泌和SC单层形成的通透性屏障(紧密连接)功能的影响。研究结果如下：1.研究结果。结果1.SC在培养过程中自发产生的NO量相对较少。在条件培养液中加入内毒素(1μg/ml)对条件培养液中的NO浓度无明显影响。NO供体(NOC18)释放的外源NO对SC单层的电阻(ER)没有影响。SC分泌的总转铁蛋白(Tf)在对照组和NOC18处理组上皮片间差异无统计学意义。然而，对于NO-DO…，SC分泌TF的极性明显较低更多也不包含文化。在外源添加NOC18(50和100μM)的培养中，2天采集期的TF分泌比率(顶端/基底端=1.86±0.22)分别下降到1.26±0.04和1.47±0.08。在未经处理的SC中，occludin和ZO-1的免疫组织化学染色显示，在细胞-细胞连接复合体的细胞周围有连续的标记，并且这些标记与TJ相关的F-肌动蛋白细丝共定位。然而，与NOC18(200μM和400μM)孵育后，48H.7内阻滞素染色完全解体，ZO-1染色部分破坏。此外，在NO供体处理的SC中，occludin、ZO-1和Cortactin的表达呈剂量依赖性地下降。这些结果表明，在急、慢性病理生理条件下，睾丸内过量产生的NO扰乱了SC的TJ相关蛋白，并可能扰乱血-睾丸屏障的形成，进而调节正常的精子发生。较少

Rat seminiferous epithelial cells (Sertoli cells : SC) isolated from immature rats were cultured either in the bicameral chamber system or on 15-mm round glass cover slide to validate the effect of nitric oxide (NO) on the protein secretion from the SC and the function of permeability barrier (tight junction) formed by the SC monolayer. The results were as follows.1. The amount of NO produced spontaneously by SC in culture was relatively minute.2. The NO concentration in the conditioned media was not significantly increased by the addition of LPS (1 μg/ml) to the basal reservoir of the culture system.3. The electrical resistance (ER) of SC monolayer was not influenced by the addition of the exogenous NO that released from a NO donor (NOC18).4. The amount of the total transferrin (TF) secreted by SC was not significantly different statistically among the control and the NOC18 treated epithelial sheets. However, the polarity of the TF secretion by SC was significantly lower for the NO-do … More nor contained cultures. The secretion ratio (apical/basal=1.86 ± 0.22) of TF during 2-day collection period was significantly declined to 1.26 ± 0.04 and 1.47 ± 0.08 in the culture with the exogenously added NOC18 (50 and 100 μM), respectively.5. Immunostaining for occludin and ZO-1 in untreated-SC showed continuous labeling around the cell periphery in the region of the cell-cell junctional complex, and these colocalized with TJ-associated F-actin filaments.6. However, incubation with NOC 18 (200 μM and 400 μM) caused complete disorganization of occludin staining and partial disruption of ZO-1 staining within 48 h.7. Furthermore, descending expressions of occludin, ZO-1 and cortactin were observed in a dose-dependent manner of NOC 18 in NO donor treated SC.These results suggest that NO produced excessively within testis during acute or chronic pathophysiological conditions disrupts TJ-associated proteins of SC and that NO may perturb blood-testis barrier formation and in turn the regulation of normal spermatogenesis. Less