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

人顶浆汗腺中17β-羟基类固醇脱氢酶3型的表达和活性

• 批准号: 11670838
• 负责人: TAKAYASU Susumu
• 金额: $ 0.77万
• 依托单位: OITA MEDICAL UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11670838/
• 关键词: apocrine gland; 17β-hydroxysteroid dehydrogenase; type 3 isozyme

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

人类大汗腺是一种独特的雄激素靶向组织，在青春期发育，无论是男性还是女性。尽管女性的血清睾酮和双氢睾酮水平比男性低得多，但这些活跃的雄激素在细胞核内的浓度在两性之间没有差异。17-羟基类固醇脱氢酶(17-β-β)3型在睾丸和腹部脂肪组织中表达，具有将雄烯二酮转化为睾酮的活性。在本研究中，我们研究了人类大汗腺中是否存在3型同工酶。将腋臭患者腋窝皮肤新鲜分离的顶分泌腺保存于-70℃，顶分泌腺匀浆用200 nm~(-3)-雄烯二酮和1 mM NADPH孵育20min。在中性或碱性条件下，在混合物中加入15型α还原酶抑制剂MK386，以防止酶迅速消耗底物。以下是Res…获得了更多的结果。(1)辅因子的影响：在NADPH存在下，酶的反应速度比NADPH生成系统或NADH更快。(2)pH的影响：在pH为5.5和pH为9.0时出现两个酶活性高峰。(3)酶活性的亚细胞定位：在pH值为5.0和9.0时，胞质组分的酶活性最高，但低于匀浆的活性。其他组分，如粗核、线粒体和微粒体组分表现出较低的活性。这一结果与3型同工酶的定位不同，3型同工酶定位于微粒体组分。然而，它们与Hodgins等人之前的报告是一致的。世卫组织还在胞浆中发现了主要活性物质。(4)RT-PCR检测3型同工酶的mRNA表达：35个循环的大汗腺组织和培养的大汗腺周围结缔组织成纤维细胞(分离的大汗腺含有少量的连接组织)均有3型同工酶的表达。因此，尽管人类顶分泌腺表达17HSD3型β，并在pH值为5.0时类似于3型同工酶将雄烯二酮转化为睾酮，但亚细胞定位与该同工酶不同。我们还需要进一步的研究来检验我们实验中的亚细胞分离是否成功。此外，必须研究存在一类新的同工酶的可能性。较少