Potentiation response of cultured human uterine leiomyoma cells to various growth factors by endothelin-l:: role of protein kinase C

Objective: Factors responsible for the abnormal proliferation of myometrial cells that accompanies leiomyoma formation are unknown, although steroid hormones and peptide growth factors have been implicated. We hypothesized that endothelin-1 (ET-1) is a physiological regulator of tumor growth. Design: In this study, we investigated the role of ET-1 on growth of human leiomyoma cells and its synergistic effect with growth factors, as well as the signaling pathway involved in this interaction. Methods: Leiomyoma cell proliferation was assayed by [H-3]thymidine incorporation and cell number. Protein kinase C (PKC) isoforms were analyzed by Western blot using specific antibodies. Results: ET-1 on its own was unable to stimulate DNA synthesis but potentiated the leiomyoma cell growth effects of basic fibroblast growth factor (bFGF), epidermal growth factor (EGF), IGF-I and IGF-II, The failure of a protein tyrosine kinase (PTK) inhibitor, tyrphostin 51, to affect the potentiating effect of ET-1, supports the hypothesis of non-involvement of PTK in this process. The inhibition of PKC by calphostin C or its down-regulation by phorbol 12,13-dibutyrate (PDB) eliminated the potentiating effect of ET-1, but did not block cell proliferation induced by the growth factors alone. Five PKC isoforms (alpha, beta1, epsilon, delta and xi) were detected in leiomyoma cells, but only phorbol ester-sensitive PKC isoforms (PK alpha, epsilon and delta) contribute to the potentiating effect of leiomyoma cell growth by ET-1. Conclusions: We have demonstrated that ET-1 potentiates leiomyoma cell proliferation to growth factors through a PKC-dependent pathway. These findings suggest a possible involvement of ET-1 in the pathogenesis of leiomyomas.