Characterization of heparin-binding endothelial mitogen(s) produced by the ovine endometrium during early pregnancy.
To characterize mitogenic factors produced by ovine endometrium during early pregnancy, endometrial explant-conditioned media (ECM) were obtained from ewes on day 12, 18, 24, or 30 after mating. These ECM contained mitogenic activity for both endothelial and 3T3 cells across all days. The endothelial mitogenic activity was greatest on day 24, whereas mitogenic activity for 3T3 cells did not differ across days. By ultrafiltration, ion exchange, and heparin-affinity chromatography, the endothelial mitogenic activity was found to have a molecular mass greater than 100 kDa, to be anionic, and to be heparin binding, respectively. Three peaks of endothelial mitogenic activity were recovered from heparin-affinity chromatography. The major peak, H3, was mitogenic for endothelial but not for 3T3 cells. H3 was further purified, and the single peak of heparin-binding activity, designated H3b, represented a 681-fold purification of endothelial mitogenic activity from endometrial ECM. H3 and H3b were heat labile and trypsin sensitive, and their biological activity was heparin enhanced. The majority of the endothelial mitogenic activity was immunoneutralized by antibodies against acidic and basic FGF. Nevertheless, we were unable to detect bFGF in H3 or H3b by Western immunoblot analysis. Thus, in this study we have extended our previous observations and demonstrated that (i) during early pregnancy the ovine endometrium produces mitogenic activity for both endothelial and 3T3 cells, (ii) the endothelial mitogenic activity is greatest on day 24 after mating. which corresponds with the onset of endometrial vascular growth, and (iii) the major endothelial mitogen has a high affinity for heparin, and although it is immunologically related to FGF, it differs from known FGF in its apparent molecular size and biological activities.
Department of Animal and Range Sciences, North Dakota State University, Fargo 58105, USA.
J
J
Zheng
D A
DA
Redmer
S D
SD
Killilea
L P
LP
Reynolds
D016475:3T3 Cells; D000818:Animals; D002846:Chromatography, Affinity; D004717:Endometrium / Q000098:blood supply / Q000166:cytology / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D005260:Female; D016222:Fibroblast Growth Factor 2 / Q000032:analysis; D005865:Gestational Age; D006493:Heparin / Q000378:metabolism*; D051379:Mice; D008934:Mitogens / Q000096:biosynthesis*; D011247:Pregnancy; D011270:Pregnancy, Animal / Q000378:metabolism*; D012756:Sheep
D008934:Mitogens; D016222:Fibroblast Growth Factor 2; D006493:Heparin