Sen Aritro, Wright Marietta, Inskeep E Keith, Flores Jorge A
Department of Biology, Eberly College of Arts and Sciences, West Virginia University, Morgantown, WV 26506-6057, USA.
Domest Anim Endocrinol. 2006 Oct;31(3):284-99. doi: 10.1016/j.domaniend.2005.11.006. Epub 2005 Dec 19.
Expression of PKC alpha, beta I, beta II, epsilon and micro has been demonstrated in the whole bovine CL with PKC epsilon being differentially expressed as a function of development. In experiment 1 we have investigated the amount of mRNA encoding PKC epsilon at different stages of luteal development (days 1, 4, 10 and 17). In experiment 2, the cellular source of luteal PKC isozymes was determined. Enriched steroidogenic (SC) and endothelial (EC) cells from day-10 CL were used to examine this question by Western blot analysis and immuno-histochemistry. In experiment 3, Western blot analysis was used to examine the ability of ET-1 to activate luteal PKC isozymes in day-10 CL. In experiment 4, the role of luteal PKC isozymes in the ET-1 mediated inhibition of P(4) accumulation in steroidogenic cell cultures from day-4 and day-10 CL was examined. Abundance of PKC epsilon mRNA gradually increased from day-1 to -10 with no further increase on day-17. In experiment 2, PKC epsilon was exclusively detected in SC (LLC and SLC). In contrast, PKC alpha, beta I and beta II were detected in both SC and EC, with EC expressing higher amounts of PKC isozymes. In day-10 CL, ET-1 induced cellular redistribution of PKC alpha, beta I, epsilon but not beta II. Inhibitors specific for conventional PKC isozymes as well as PKC epsilon were able to negate the inhibitory effects of ET-1 on P4 accumulation in the day 10 CL. In the day-4 CL, the inhibitory effect of ET-1 might be mediated via conventional PKC. Thus, an exclusive presence of PKC epsilon in luteal steroidogenic cells, its higher expression along with the ability of ET-1 to stimulate its activation in day-10 CL strongly suggests that this PKC isoform may play an important regulatory role in decreasing P(4) during luteal regression. Inhibition of P(4) by ET-1 in the early CL may be mediated via conventional PKC isozymes.
已在整个牛黄体中证实蛋白激酶C（PKC）α、βI、βII、ε和μ的表达，其中PKCε的表达随发育过程而有差异。在实验1中，我们研究了黄体发育不同阶段（第1、4、10和17天）编码PKCε的mRNA量。在实验2中，确定了黄体PKC同工酶的细胞来源。使用来自第10天黄体的富集的类固醇生成细胞（SC）和内皮细胞（EC），通过蛋白质印迹分析和免疫组织化学来研究这个问题。在实验3中，使用蛋白质印迹分析来检测内皮素-1（ET-1）激活第10天黄体中PKC同工酶的能力。在实验4中，研究了黄体PKC同工酶在ET-1介导的抑制第4天和第10天黄体类固醇生成细胞培养物中孕酮（P4）积累中的作用。PKCεmRNA的丰度从第1天到第10天逐渐增加，在第17天没有进一步增加。在实验2中，仅在SC（大黄体细胞和小黄体细胞）中检测到PKCε。相比之下，在SC和EC中均检测到PKCα、βI和βII，且EC中PKC同工酶的表达量更高。在第10天的黄体中，ET-1诱导PKCα、βI、ε发生细胞内重新分布，但未诱导PKCβII重新分布。针对传统PKC同工酶以及PKCε的特异性抑制剂能够消除ET-1对第10天黄体中P4积累的抑制作用。在第4天的黄体中，ET-1的抑制作用可能是通过传统PKC介导的。因此，PKCε在黄体类固醇生成细胞中的独特存在、其较高的表达以及ET-1在第10天黄体中刺激其激活的能力，强烈表明这种PKC同工型可能在黄体退化过程中降低P4方面发挥重要的调节作用。ET-1在早期黄体中对P4的抑制作用可能是通过传统PKC同工酶介导的。
