Heggland S J, Stalvey J R
Department of Biological Sciences, Kent State University, OH 44242, USA.
Steroids. 1996 May;61(5):309-16. doi: 10.1016/0039-128x(95)00235-i.
We previously reported that 3 beta-hydroxysteroid dehydrogenase-isomerase (3 beta HSD) activity is higher in Leydig cells from C57BL/6J mice than those from C3H/HeJ mice. This study examines whether the differences in 3 beta HSD activity in Leydig cells from the two strains of mice are due to the expression of different 3 beta HSD isoforms and if a specific isoform corresponds with the amount of 3 beta HSD activity under various culture conditions. Leydig cells were plated in Waymouth's +15% horse serum (HS) medium. Some cultures were terminated 24 h later after plating (day 1) and assayed for 3 beta HSD activity or immunoreactivity. The remaining cultures were maintained in HS or changed to serum-free medium. We demonstrate for the first time that two 3 beta HSD immunoreactive isoforms are expressed in freshly isolated Leydig cells and those cultured for 1 day. The same two 3 beta HSD isoforms are detectable in both strains. Thus, the previously reported strain-related differences in 3 beta HSD activity are not due to the expression of different isoforms. When cultured for 8 days, the higher molecular weight 3 beta HSD immunoreactive band is no longer detectable in Leydig cells from either strain. When maintained in HS, 3 beta HSD activity in C57BL/6J Leydig cells decreases significantly by day 8, while 3 beta HSD activity in C3H/HeJ Leydig cells does not change through day 8. When Leydig cells were cultured in serum-free medium, 3 beta HSD activity is maintained in cultured Leydig cells from C57BL/6J and significantly increases in C3H/HeJ 3 beta HSD by day 8. These data suggest that HS has a strain-specific inhibitory effect on 3 beta HSD activity, causing a significant decrease in C57BL/6J 3 beta HSD activity and preventing an increase in C3H/HeJ. Densitometric analysis reveals a correspondence between changes in 3 beta HSD activity and the lower molecular weight isoform but not the higher molecular weight isoform. Treatment with cAMP induces the immunoreactive mass of the lower molecular isoform but not the higher molecular isoform of 3 beta HSD. Currently, it is unclear what the function of the higher molecular weight 3 beta HSD isoform is in mouse Leydig cells.
我们之前报道过，C57BL/6J小鼠睾丸间质细胞中的3β-羟类固醇脱氢酶异构酶（3βHSD）活性高于C3H/HeJ小鼠的睾丸间质细胞。本研究旨在探讨这两种品系小鼠睾丸间质细胞中3βHSD活性的差异是否归因于不同3βHSD同工型的表达，以及在各种培养条件下是否有一种特定的同工型与3βHSD活性水平相对应。将睾丸间质细胞接种于含15%马血清（HS）的Waymouth培养基中。部分培养物在接种后24小时（第1天）终止培养，检测其3βHSD活性或免疫反应性。其余培养物继续在含HS的培养基中培养或更换为无血清培养基。我们首次证明，在新鲜分离的睾丸间质细胞和培养1天的细胞中表达了两种3βHSD免疫反应性同工型。在两个品系中均可检测到相同的两种3βHSD同工型。因此，先前报道的品系相关的3βHSD活性差异并非由于不同同工型的表达。培养8天后，两个品系的睾丸间质细胞中均无法再检测到分子量较高的3βHSD免疫反应性条带。在含HS的培养基中培养时，C57BL/6J小鼠睾丸间质细胞中的3βHSD活性在第8天时显著降低，而C3H/HeJ小鼠睾丸间质细胞中的3βHSD活性在第8天前未发生变化。当睾丸间质细胞在无血清培养基中培养时，C57BL/6J小鼠培养的睾丸间质细胞中的3βHSD活性得以维持，而C3H/HeJ小鼠的3βHSD活性在第8天时显著增加。这些数据表明，HS对3βHSD活性具有品系特异性抑制作用，导致C57BL/6J小鼠的3βHSD活性显著降低，并阻止C3H/HeJ小鼠的3βHSD活性增加。光密度分析显示，3βHSD活性变化与分子量较低的同工型相对应，而与分子量较高的同工型不对应。用cAMP处理可诱导3βHSD分子量较低的同工型的免疫反应性，但不能诱导分子量较高的同工型。目前尚不清楚分子量较高的3βHSD同工型在小鼠睾丸间质细胞中的功能。
