The influence of blood cells and PDGF on porcine theca cell function in vitro

The role of red and white blood cells in the regulation of porcine theca cell function is poorly understood. Interactions between these cell types and a potential mediator of any interaction, PDGF, were investigated using a serum-free culture system. Theca cells were collected from 6-9mm antral follicles and plated at 50x10(3) viable cells/well. In the first experiment, macrophages were removed and theca cells+/-macrophages were cultured with a range of PDGF doses (0.1, 1, and 10ng/ml)+/-IGF-1. In the second experiment, red blood cells were removed with lysing buffer. In both experiments the effect of treatment on steroidogenesis and viable cell number was examined. Macrophage removal decreased oestradiol production but increased androstenedione output irrespective of the presence of IGF-1 (oestradiol+/-IGF-1, P<0.001; androstenedione P=0.02 without IGF-1, P<0.001 with IGF-1). PDGF increased oestradiol synthesis by whole and macrophage-free theca cell preparations but only in the presence of IGF-1 (P<0.001). In contrast, androstenedione production was unaffected by PDGF dose in the presence of IGF-1 (P=0.67). Without IGF-1, 10ng/ml PDGF tended to decrease androstenedione levels (P=0.06). Macrophage removal increased viable cell number at 144h (P<0.001+/-IGF-1) as did PDGF (P<0.001+/-IGF-1). In the absence of IGF-1, there was a PDGF x cell type interaction (P=0.02). Macrophage-free cultures with 10ng/ml PDGF had twice as many viable cells as whole preparations with no PDGF. In the second experiment, red blood cell removal did not affect steroidogenesis or the number of viable cells present at 144h when cells were cultured with IGF-1. The data show that theca cell/macrophages interactions do occur, and influence both steroidogenesis and viable cell number during culture. The macrophage product(s) enhanced oestradiol synthesis but reduced androstenedione production and the number of viable cells. As all these interactions were not mimicked by PDGF, PDGF cannot be the only factor mediating the theca/macrophage interaction. When cultured under optimised conditions the presence of red blood cells was not detrimental to theca cell steroidogenesis or the number of viable cells.