In Vitro Culture of Hamster Ovarian Primary Interstitial Cells: Effect of Serum¹

^a Department of Obstetrics and Gynecology, Leland J. and Dorothy H. Olson Center for Women's Health, Division of Reproductive Endocrinology, and ^b Department of Physiology and Biophysics, University of Nebraska Medical Center, Omaha, Nebraska 68198–4515

The function of ovarian interstitial cells has been largely addressed using rat theca-interstitial cell culture. However, this preparation is primarily enriched with theca and secondary interstitial cells, which make it difficult to address selectively the function of the primary interstitial cells. We have developed an in vitro culture of hamster ovarian primary interstitial cells. Cells were isolated from postnatal hamster ovaries by collagenase digestion and purified over a Percoll gradient. The preparation contained 90% viable, pure interstitial cells, which anchored to the plastic and glass culture surface in the presence of fetal bovine serum. Cell proliferation was noted in the presence of serum dosages higher than 0.2%; however, reduction of serum concentration to 0.1% or complete serum starvation did not affect cell viability but almost completely abolished cell proliferation as determined by [³H&; incorporation, labeling index, and DNA content of the culture. All cells exhibited active 3ß-hydroxysteroid dehydrogenase and P450 side chain cleavage immunoreactivity, which corresponded to basal progesterone and androstenedione accumulation. Replacement of serum to starving cells resulted in the induction of the "S" phase and "M" phase specific cyclins, and resumption of cell proliferation. Our results indicate that hamster primary interstitial cells can be cultured in vitro as a monolayer, and the anchorage and proliferation of these cells depend on serum supplement; however, a viable monolayer can be maintained for several days without serum. This model will be useful for addressing the mechanisms of differentiation of ovarian interstitial cells.

¹ This work was supported by grants HD28165 from the National Institute of Child Health and Human development, NIH, and the Olson Foundation to S.K.R.

² Correspondence: Shyamal K. Roy, Bennett Hall 5005, Departments of OB/GYN and Physiology/Biophysics, 984515 University of Nebraska, Medical Center, Omaha, NE 68198–4515. FAX: 402 559 6164.

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