BIOLOGY OF REPRODUCTION 51, 700-705 (1994) Effects of Interferon-i and Interferon-a on Proliferation of Bovine Endometrial Cells'

The ruminant conceptus secretes a unique interferon, interferon-, that regulates endometrial prostaglandin secretion during early pregnancy. Because one of the pleiotropic effects of interferons is to inhibit cellular proliferation, a series of experiments was conducted to determine whether or not the bovine endometrium is sensitive to the antiproliferative effect of interferon-r and the related interferon, interferon-a. Endometrial epithelial and stromal cells were prepared from the endometrium of cows from Days 11-17 after estrus and incubated with recombinant bovine interferon-T (rbIFNr; 1-1000 ng/ml), recombinant bovine interferon-al (rbIFNca; 1-1000 ng/ml), recombinant human interferon-a2b (rhIFNa; 100 ng/ml), or ovine interferon-,r (olFNT; 100 ng/ml). Proliferation was determined by monitoring uptake of [ H]thymidine into DNA. Generally, interferons did not inhibit proliferation of endometrial epithelial cells. Exceptions were for 1000 ng/ml rblFNT, which inhibited proliferation by 23%; 100 ng/ml rbIFNa, which inhibited proliferation by 28% in one of two experiments only; and 100 ng/ml oFNT, which inhibited proliferation by 17%96. Proliferation of endometrial stromal cells was not inhibited by any concentration of any interferon in two separate experiments. Therefore, unlike other bovine cells tested previously (lymphocytes and oviductal cells), bovine endometrial cells were not consistently inhibited by IFNr or IFNa. Such reduced responsiveness of endometrial cells to the antiproliferative effects of type I interferons could allow for growth of the endometrium during the period of pregnancy when the conceptus produces IFNT. INTRODUCTION Interferon-T is a secretory product of the ruminant trophoblast that in bovine species is produced as early as the blastocyst stage [1] and produced maximally between Days 15 and 25 [2]. This protein, also called trophoblast protein1, plays a key role in the maintenance of CL function during early pregnancy by blocking the release of prostaglandin F2 from endometrial epithelial cells [3-5]. Interferon-T is a type I interferon that has about 50% amino acid sequence homology with interferon-a [6-7] but is now recognized as a separate class of interferon because of significant differences in regulatory regions of the gene [8-9]. Like other interferons, however, interferon-T is a pleiotropic molecule that can alter synthesis of endometrial proteins [10], induce an antiviral state in target cells [11-13], and enhance activity of 2',5'-oligoadenylate synthetase in endometrial stromal and epithelial cells [14]. Interferons are also well known antiproliferative molecules [15-16], and interferon-T has been shown to inhibit proliferation of lymphocytes [17-20], oviductal epithelial cells [21], WISH cells, and MDBK cells [22]. Given the importance of the endometrium for the developing conceptus, inhibition of endometrial cell prolifAccepted May 31, 1994. Received September 29, 1993. 'Research supported by USDA Grant 89-37240-4583. Dr. Tiemann was supported by Deutsche Forschungsgemeinschaft and Dr. Kamwanja received support through a Fulbright Scholarship. This is Journal Series No. R-03223 of the Florida Agricultural Experiment Station. 2 Correspondence: PJ. Hansen, P.O. Box 110920, University of Florida, Gainesville, FL 32611-0920. FAX: (904) 392-5595. 3 Current address: Department of Biological Sciences, East Texas State University, Commerce, Texas 75429. eration by interferon-T might potentially be deleterious to conceptus development. Previous results, however, indicate that cell lines differ in their sensitivity to antiproliferative effects of interferon-ot [23-25]. Therefore, it is possible that bovine endometrium is more resistant to the inhibitory effects of interferon-r and interferon-oa than other cells. This was tested by evaluating effects of these interferons on proliferation of endometrial epithelial and stromal cells. Cells were collected from Days 11-17 after estrus because these times represent the period when embryonic synthesis of interferon-T takes place. Endometrium was separated into epithelial and stromal cells because these two cell populations have clear functional differences with respect to prostaglandin secretion and regulation by interferon-T [5]. MATERIALS AND METHODS Materials Recombinant bovine interferon-T (rbIFNT) was donated by RM. Roberts (University of Missouri, Columbia, MO), and recombinant bovine interferon-al (rbIFNa) by CIBA-GEIGY (Basle, Switzerland). The antiviral activities of rbIFNT and rbIFNa were 2.4 x 106 U/mg and 5.7 x 106 units/mg, respectively. Natural ovine interferon-T (oIFNT) was donated by F.W. Bazer (Texas A&M University, College Station, TX). Human recombinant interferon-a2b (rhIFNa) was obtained from Hoffman-LaRoche (Nutley, NJ). Batches of bovine interferons used in the present studies were shown to be effective in inhibiting bovine peripheral blood lymphocytes (results not shown) and bovine oviductal cells [21]. Ultra-