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Endocrinology Vol. 138, No. 3 1289-1298
Copyright © 1997 by The Endocrine Society

Articles

Regulation of Rat Testis Gonocyte Proliferation by Platelet-Derived Growth Factor and Estradiol: Identification of Signaling Mechanisms Involved1

Hua Li, Vassilios Papadopoulos, Branislav Vidic, Martin Dym and Martine Culty

Department of Cell Biology, Georgetown University Medical Center, Washington, D.C. 20007

Address all correspondence and requests for reprints to: Dr. Martine Culty, Department of Cell Biology, Georgetown University Medical Center, 3900 Reservoir Road NW, Washington D.C. 20007.

To determine what factors regulate gonocyte proliferation in newborn rats, we first examined the expression of several signal transduction molecules by immunocytochemistry in 3-day-old rat testis sections. We found that gonocytes specifically expressed the {iota} and {zeta} isoforms of protein kinase (PK) C (PKC) and the phosphatidylinositol 3-kinase (PI 3-K). Because both the {zeta}PKC and PI 3-K have been shown to play a role in platelet-derived growth factor (PDGF)-induced cell proliferation, we examined the effects of PDGF on gonocytes. For this, we developed a method to obtain highly purified and viable gonocytes in culture. After enzymatic digestion, differential adhesion, and two successive gradient fractionations, the gonocyte suspension obtained was over 90% pure, as assessed by light microscopy. The viability of cultured gonocytes exceeded 90% after 48 h in the presence of 2.5% FBS used as a survival factor. Immunodetection studies showed that isolated gonocytes expressed {zeta}PKC, PI 3-K, and the PDGF receptor. Treatment with 10 ng/ml PDGF induced a 4-fold increase of bromodeoxyuridine incorporation into gonocytes (from 5% proliferative gonocytes under basal conditions to 20% in the presence of PDGF). Because neonatal Sertoli cells secrete high levels of the growth promoting steroid, 17ß-estradiol, we also tested its effect and found that it induced gonocyte proliferation at a level comparable with that of PDGF and that this effect was blocked by the estrogen receptor antagonist, ICI 164384. The combination of PDGF and estradiol, however, was not additive, suggesting that their effects were mediated by common molecular target(s). These results demonstrate that PDGF and estradiol activate gonocyte proliferation in vitro, suggesting that they may act as the physiological regulators of gonocyte development in vivo.




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