CHARACTERIZATION OF BOVINE OVARIAN SURFACE EPITHELIUM AND STROMAL CELLS - IDENTIFICATION OF SECRETED PROTEINS

The majority of ovarian cancers are derived from a single layer of epi thelial cells that covers the surface of the ovary termed the ovarian surface epithelium (OSE). Ovarian surface stromal cells underlie the O SE and have a morphology distinct from that of the interstitial stroma l cells between follicles. Because of the similarities between bovine and human ovarian physiology, and to allow an adequate supply of tissu e and cells, bovine OSE and stromal cell cultures Were established to investigate the cell biology of these cell types. Morphological analys is of bovine ovarian sections revealed that several layers of ovarian surface stromal cells can be identified and are structurally distinct from interstitial stromal cells. Both OSE and stromal cells can be iso lated and cultured for weeks in the absence or presence of serum. The cell populations were found, through use of a keratin immunocytochemic al stain for OSE, to be highly purified. To investigate the functional properties of the two cell types, radiolabeled secreted proteins were collected and electrophoretically analyzed. The radiolabeled secreted protein profiles of OSE and stromal cells mere found to be distinct w ith a discrete number of secreted proteins. Major OSE secretory produc ts were obtained from serum-free concentrated conditioned medium, elec trophoretically separated, blotted, and sequenced. TWO OSE secretory p roducts of 28 kDa and 40 kDa were sequenced and found to match a seque nce in the computerized database. The 28-kDa OSE protein was identifie d as a tissue inhibitor of metalloproteinase, TIMP. The 40-kDa OSE pro tein was identified as the insulin-like growth factor (IGF) binding pr otein-2 (IGFBP2). The TIMP may play a role in regulating the local pro teolytic activity of the OSE, and IGFBP2 in regulating IGF-I actions o n OSE. These proteins provide biochemical markers for OSE that can be used to further investigate the regulation of OSE function. The growth properties of cultured bovine OSE were also investigated. OSE prolife ration was stimulated by epidermal growth factor (EGF) and was not inf luenced by keratinocyte or hepatocyte growth factors. Transforming gro wth factor-beta was found to inhibit the growth stimulatory actions of EGF on OSE. Concentrated serum-free stromal cell-conditioned medium w as also found to influence OSE growth. Therefore, the ovarian surface stromal cells appear to produce factors that can regulate the growth o f the OSE. Combined results indicate that a culture system has been es tablished to investigate the biology of OSE and ovarian surface stroma l cells. Two OSE secretory products were identified as TIMP and IGFBP2 . Preliminary data are presented that the ovarian surface stromal cell s can regulate OSE growth. This stromal-epithelial cell interaction ma y have an important role in the normal cell biology of the OSE and be involved in the transformation of this cell to form an ovarian cancer.