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Journal of Clinical Endocrinology & Metabolism Vol. 73, No. 5 1134-1140
doi:10.1210/jcem-73-5-1134
Copyright © 1991 by the Endocrine Society.
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3β-Hydroxysteroid Dehydrogenase Activity in Glandular and Extraglandular Human Fetal Tissues*

LEON MILEWICH, CYNTHIA E. SHAW, KATHLEEN M. DOODY, WILLIAM E. RAINEY, J. IAN MASON and BRUCE R. CARR

Department of Obstetrics and Gynecology, University of Texas Southwestern Medical Center Dallas, Texas 75235
Biochemistry, University of Texas Southwestern Medical Center Dallas, Texas 75235

Address all correspondence and requests for reprints to: Dr. L. Milewich, Department of Obstetrics and Gynecology, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, Texas 75235.

The expression of 3β-hydroxysteroid dehydrogenase (3βHSD) in steroidogenic tissues is an absolute requirement for mammalian reproduction, fetal growth, and life maintenance. We sought to identify extraglandular tissue sites in the human fetus where 3βHSD is expressed. To this effect, we conducted in vitro studies by use of homogenates prepared from second trimester fetal tissues. To facilitate the determination of 3βHSD activity, an abbreviated technique was developed that consisted in the use of [3{alpha}-3H]dehydroepiandrosterone ([3{alpha}-3H] DHEA) as the substrate and NAD+ as the cofactor. With these reagents, the enzymatic reaction leads to the production of both nonradiolabeled androstenedione and NAD3H in equimolar amounts, and the radioactivity associated with NAD3H is used for quantification of 3βHSD activity. The kinetic isotope effect introduced by substitution of tritium for hydrogen at the C-3{alpha} position of DHEA, determined with six different tissues, was 2.5 ± 0.7 (mean ± SD). The specific activities of the enzyme in peripheral tissues and ovary were relatively low, in the range of 0.03 nmol/mg protein h for stomach (n = 2) to 0.18 ± 0.14 nmol/mg protein h for liver (mean ± SD; n = 13), while in fetal testis and placenta the specific activities were relatively high, viz. 3.4 ± 0.7 nmol/mg protein h (mean ± SD; n = 4) and 2.8 ± 1.8 nmol/mg protein h (mean ± SD; n = 13), respectively. The findings of this study serve to demonstrate that 3βHSD is distributed widely among tissues of the human fetus. Although the enzymatic activity was easily demonstrated in peripheral tissues by the use of radiolabeled DHEA as the substrate, 3βHSD protein was not readily detected by Western analysis.

* This work was supported in part by NIH Grants HD-11149 and T32-HD-07190.

Received January 7, 1991.




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