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A Sirotkin, Research Institute of Animal Production, Nitra, 949 92, Slovakia
D Ovcharenko, Division of Pharmacology and Toxicology, College of Pharmacy, University of Texas at Austin, Austin, TX 78712, Austin, United States
M Mlyncek, Polyclinics and Hospital of Nitra, Nitra, Slovakia

Correspondence: Av Sirotkin, Email: sirotkin{at}scpv.sk

Abstract

The goal of this study was to identify protein kinases (PKs) that control the secretory activity of human ovarian cells. Cultured ovarian granulosa cells were transfected with 264 siRNA constructs that selectively block the expression of 88 known PKs. The efficiency of transfection and of silencing marker molecules (GAPDH and CDC2/p34 PK) were validated by fluorescence microscopy, real-time reverse transcription-PCR, and immunocytochemistry. Release of steroid hormones (progesterone, P4) and insulin-like growth factor I (IGF-I) were determined by radioimmunoassay (RIA). siRNA suppressed the expression of marker molecules by up to 84%. P4 release was suppressed after inhibiting 34 individual PKs and was stimulated after inhibiting 12 PKs. Blocking nine individual PKs inhibited IGF-I release, while the inactivation of 17 others stimulated IGF-I release. Together, these results demonstrate that the release of both steroid and peptide hormones by human ovarian cells is controlled by a large number of PKs, and that siRNA constructs may be useful tools for further defining the role of PKs in controlling ovarian secretory function.