CTGF is a heparin-binding multi-domain growth factor, stimulated by TGF- beta, and implicated as a paracrine regulator of mitosis, angiogenesis, cellular taxis and remodelling of the extracellular matrix in diverse epithelial cell layers. To examine the roles of CTGF in the ovarian paracrine system, we studied rat granulosa cell CTGF gene expression both in vitro and in vivo. Untreated granulosa cell monolayers from DES-treated immature rats expressed abundant approx. 2.5 kb CTGF mRNA, determined by Northern analysis, but had low levels of aromatase enzyme activity (an index of granulosa cell differentiation). FSH (0.1-10 ng/ml) inhibited (50% at 10 ng/ml) CTGF mRNA expression but enhanced aromatase activity after 48 h. Studies of in vivo ovarian CTGF expression in FSH-beta knockout mice by Northern blot and in situ hybridization analyses demonstrated high levels of CTGF expression in the granulosa cells of preantral follicles blocked from further development owing to the absence of FSH. FSH action was mimicked in vitro by 8br-AMP (1.0 mM) and augmented by the additional presence of androgen (1 microM 5alpha-DHT), consistent with mediation by intracellular cAMP. Conversely, TGF-beta1 (0.1-10 ng/ml) increased CTGF mRNA levels up to 12-fold (at 10 ng/ml), without affecting aromatase activity. Co-treatment with FSH (0.1-10 ng/ml) suppressed the stimulatory action of TGF-beta1 on CTGF mRNA, but substantially enhanced aromatase activity beyond levels induced by FSH alone. Importantly, other ovarian expressed TGF-beta superfamily members (GDF-9 and activin, 10 ng/ml) stimulated granulosa cell CTGF mRNA in a similar fashion to TGF-beta1 (10 ng/ml), and this was also inhibited by FSH (10 ng/ml). Our results show that CTGF gene expression in granulosa cells is inversely related to the degree of cellular differentiation, and inhibited by FSH via cAMP-mediated intracellular signalling. TGF-beta, GDF-9 and activin up-regulate CTGF mRNA in vitro, suggesting a paracrine/autocrine involvement of these growth/differentiation factors in regulating CTGF in the ovary.
08 - 11 Apr 2002
British Endocrine Societies