Inhibition of murine transformed Leydig cell proliferation by leukotrienes in serum-free culture.

ABSTRACT

We have previously shown that estrogen-dependent growth enhancement of murine transformed Leydig cells (B-1 F) is mediated through inhibition of arachidonic acid metabolite formation. In the present study, the growth-inhibitory ability of leukotrienes (LTs) on B-1 F cells in serum-free culture was directly addressed. All peptidyl LTs (LTC4, LTD4, and LTE4) inhibited B-1 F cell growth in a dose-dependent manner and exhibited maximum inhibition of DNA synthesis (60-80%) compared with that of untreated cells in a range of 10(-9) to 10(-8) M. To examine the mechanism of this LT-dependent inhibition, binding studies of LTD4 toward plasma membrane were conducted. Specific binding sites for LTD4 were identified. Scatchard analyses indicated the presence of a single class of high-affinity sites (Kd = 0.9 +/- 0.2 nM; maximum binding sites, 61 +/- 18 fmol/mg protein). This binding of LTD4 to the high-affinity site was markedly inhibited by ICI 198615, a specific inhibitor for LTD4. These results would suggest that inhibitory effects of LTs, at least LTD4, are elicited as a receptor-mediated event. In addition, this LT-dependent growth inhibition could not be blocked by simultaneous exposure of cells to estrogen, whereas estrogen partially protected arachidonic acid-dependent growth inhibition. Furthermore, treatment of cells with estrogen resulted in marked suppression of 5-lipoxygenase activity. Collectively, the present data clearly show that LTs play an important role as intermediates in an autocrine loop for B-1 F cells to exhibit estrogen-dependent growth.