Khurana M L, Pandey K N
Department of Biochemistry and Molecular Biology, Medical College of Georgia, School of Medicine, Augusta 30912-2100.
Endocrinology. 1993 Nov;133(5):2141-9. doi: 10.1210/endo.133.5.8404664.
We have investigated the mechanism by which different natriuretic peptides stimulate steroidogenesis in purified mouse Leydig cells. In addition to atrial natriuretic factor (ANF), we show that brain natriuretic peptide (BNP) and C-type natriuretic peptide (CNP) also stimulate testosterone production in these cells. Testosterone production was increased dramatically to 14-fold with ANF (EC50 = 0.3 nM) and 15-fold with BNP (EC50 = 0.2 nM); however, the CNP-stimulated level of testosterone production was only 2.5-fold compared with controls. ANF and BNP enhanced the stimulatory effect of LH on testosterone production. The C-ANF(4-23) (a truncated form of ANF) had no effect on testosterone production in these cells. ANF, BNP, and CNP stimulated the production of intermediate precursors of testosterone biosynthesis, which included progesterone, 17 alpha-hydroxy progesterone, androstenedione, pregnenolone, 17 alpha-hydroxy pregnenolone, and dehydroepiandrosterone sulfate. The conversion of pregnenolone and progesterone to testosterone was also significantly enhanced after treatment of Leydig cells with these peptides. All three natriuretic peptides (ANF, BNP, and CNP) stimulated the activity of particulate guanylate cyclase by 8.4-, 8.5-, and 4.8-fold and the accumulation of intracellular cGMP by 52-, 58-, and 19-fold, respectively. The cGMP inhibitor LY83583 attenuated both the generation of cGMP as well as testosterone in response to these natriuretic peptides, suggesting the involvement of cGMP as a second messenger. Leydig cells were found to contain high affinity and low capacity binding sites for ANF [dissociation constant (Kd), 2.0 x 10(-10) M; maximum binding capacity (Bmax). 20 fmol/1 x 10(5) cells], BNP (Kd, 2.2 x 10(-10) M; Bmax, 19 fmol/1 x 10(5) cells), and CNP (Kd, 3.1 x 10(-10) M; Bmax, 8.6 fmol/1 x 10(5) cells). The results presented here document that a family of different natriuretic peptides stimulates Leydig cell steroidogenesis in receptor-mediated fashion, beginning at the cholesterol side-chain cleavage enzyme. The data also show that these peptide hormones induce testosterone production in mouse Leydig cells by involving both delta 4- and delta 5-pathways of steroidogenesis.
我们研究了不同利钠肽刺激纯化的小鼠睾丸间质细胞类固醇生成的机制。除了心房利钠因子（ANF）外，我们还发现脑利钠肽（BNP）和C型利钠肽（CNP）也能刺激这些细胞产生睾酮。ANF（EC50 = 0.3 nM）可使睾酮生成显著增加至14倍，BNP（EC50 = 0.2 nM）可使其增加至15倍；然而，与对照组相比，CNP刺激的睾酮生成水平仅为2.5倍。ANF和BNP增强了促黄体生成素（LH）对睾酮生成的刺激作用。C-ANF(4 - 23)（ANF的截短形式）对这些细胞的睾酮生成没有影响。ANF、BNP和CNP刺激了睾酮生物合成中间前体的生成，这些前体包括孕酮、17α-羟孕酮、雄烯二酮、孕烯醇酮、17α-羟孕烯醇酮和硫酸脱氢表雄酮。用这些肽处理睾丸间质细胞后，孕烯醇酮和孕酮向睾酮的转化也显著增强。所有三种利钠肽（ANF、BNP和CNP）分别使颗粒型鸟苷酸环化酶的活性提高了8.4倍、8.5倍和4.8倍，使细胞内cGMP的积累分别增加了52倍、58倍和19倍。cGMP抑制剂LY83583减弱了这些利钠肽刺激引起的cGMP生成以及睾酮生成，提示cGMP作为第二信使参与其中。发现睾丸间质细胞含有ANF的高亲和力和低容量结合位点[解离常数（Kd），2.0×10(-10) M；最大结合容量（Bmax），20 fmol/1×10(5)个细胞]、BNP（Kd，2.2×10(-10) M；Bmax，19 fmol/1×10(5)个细胞）和CNP（Kd，3.1×10(-10) M；Bmax，8.6 fmol/1×10(5)个细胞）。此处呈现的结果表明，一类不同的利钠肽以受体介导的方式刺激睾丸间质细胞类固醇生成，起始于胆固醇侧链裂解酶。数据还表明，这些肽类激素通过参与类固醇生成的δ4-和δ5-途径诱导小鼠睾丸间质细胞产生睾酮。
