Expression and activity of 17β-hydoxysteroid dehydrogenase type 3 in the human apocrine sweat gland

人类顶泌汗腺是一种独特的雄激素靶组织，在青春期发育，男性和女性都有。尽管女性的血清睾酮和双氢睾酮水平比男性低得多，但这些活性雄激素的核内浓度在两性之间没有差异。17 β-羟基类固醇脱氢酶（17 β-HSD）3型在睾丸和腹部脂肪组织中表达艾德，并具有将雄烯二酮转化为睾酮的有效活性。本研究对人顶泌腺中是否存在3型同工酶进行了研究。从腋臭患者腋窝皮肤新鲜分离的大汗腺保存在-70 ℃。将顶泌腺匀浆与200 nM ^3H-雄烯二酮和1 mM NADPH孵育20 min。在中性pH或碱性条件下，混合物中加入MK 386（一种1型5 α-还原酶抑制剂），以防止酶快速消耗底物。以下内容 ...更多信息 获得了结果。(1)辅因子的影响：在NADPH存在下，酶反应比NADPH生成系统或NADH更快。(2)pH的影响：在pH5.5和pH9.0时，酶活力有两个高峰。(3)酶活性的亚细胞定位：在pH 5.0和9.0的细胞质组分中观察到最高活性，尽管它低于匀浆的活性。其他组分，如粗核，线粒体和微粒体组分表现出较小的活动。结果不同于3型同工酶定位在微粒体部分的本地化。然而，它们与Hodgins等人先前的报告一致，Hodgins等人也发现了细胞质中存在主要活性。(4)RT-PCR检测3型同工酶的mRNA表达：在35个循环时，在顶泌腺的同源物和来自腺体周围结缔组织的培养的成纤维细胞（分离的顶泌腺含有少量结缔组织）中检测到3型同工酶的mRNA表达。因此，尽管人类顶泌腺表达17 β-HSD 3型的mRNA，并在pH 5.0时像3型同工酶一样主动将雄烯二酮转化为睾酮，但亚细胞定位与该同工酶不同。需要进一步的研究来检查我们实验中的亚细胞分离是否成功。此外，还必须研究是否存在一类新的同工酶。少

The human apocrine sweat gland is a unique androgen target tissue which develops in adolescence both in males and females. In spite of much lower serum levels of testosterone and dihydrotestosterone in females than males, the intranuclear concentrations of these active androgens do not differ between the two sexes. 17β-Hydroxysteroid dehydrogenase (17β-HSD) type 3 is express ed in the testes and abdominal adipose tissues and has potent activity in converting androstenedione to testosterone. In the present study, we studied whether the type 3 isozyme exhists in the human apocrine gland. Freshly isolated apocrine glands from axillary skin of patients with osmidrosis were stored at -70 C. Homogenates of apocrine glands were incubated with 200 nM of ^3H-androstenedione and 1 mM of NADPH for 20 min. At neutral pH or alkaline conditions, MK 386, a type 1 5α-reductase inhibitor, was included in the mixture in order to prevent rapid consumption of the substrate by the enzyme. The following res … More ults were obtained. (1) Effect of cofactors : the enzyme reaction was faster in the presence of NADPH than NADPH-generating system or NADH. (2) Effect of pH : two peaks of the enzyme activities were obtained at pH 5.5 an pH 9.0. (3) Subcellular localization of the enzyme activity : the highest activity was observed in the cytosol fraction at both pH 5.0 and 9.0, although it was lower than the activity of homogenates. Other fractions such as crude nuclear, mitochondrial and microsomal fractions exhibited lesser activities. The results differ from the localization of the type 3 isozyme which localizes in the microsomal fraction. However, they are consistent with the previous report by Hodgins et al. who also found the presence of the major activity in the cytosol. (4) mRNA expression of the type 3 isozyme by RT-PCR : type 3 expression was detected at 35 cycles in the homogemate of the apocrine gland and cultured fibroblasts derived from the connective tissue surrounding the gland (the isolated apocrine gland contained a small amount of connecive tissues). Thus, although human apocrine glands express mRNA of 17β- HSD type 3 and actively converts androstenedione to testosteron at pH 5.0 like the type 3 isozyme, subcellular localization is different from that of this isozyme. Further studies are required to check whether the subcellular fractionation in our experiment was successful. In addition, the possibility of the presence of a new class of the isozyme must be investigated. Less