The relevance of toxicant-induced apoptosis to reproductive impairment in fish and mammals

Scope and method of study. A variety of toxicants have been reported to stimulate apoptosis, alter expression of heat shock protein 70 (HSP70), and affect sex steroid production in the vertebrate ovary. The objective of this study was to determine effects of selected toxicants and hormones on ovarian cell apoptosis, HSP70 expression and steroidogenesis in cultured pre-ovulatory ovarian follicles of two teleosts with contrasting reproductive strategies; fathead minnows (Pimephales promelas) (i.e. an asynchronous spawner) and channel catfish (Ictalurus punctatus ) a synchronous spawner. Isolated follicles were exposed in vitro for 48 and 24 hours to putative apoptogens (dimethylbenz[a]anthracene (DMBA), cadmium chloride (CdCl2), as well as survival factors (estradiol, epidermal growth factor (EGF), phorbol myracetate (PMA), or a combination of forskolin plus isobutyl-methylxanthine (IBMX). Untreated (control) follicles were cultured in serum-free media for 48 or 24 hours. We measured the extent of apoptotic DNA fragmentation using 3′-end labeling of isolated ovarian follicular (granulosa and theca) cell DNA. Western blotting and densitometry were used to quantify expression of HSP70 in follicular cells, and sex steroid hormone (testosterone and 17β-estradiol) concentrations in culture media were measured using enzyme-linked immunosorbent assays (ELISA).; Finding and conclusions. Fathead minnow trials revealed that in comparison to control follicles cultured for 48 hours in serum-free media, 17β-estradiol, EGF and PMA treatments suppressed apoptosis and CdCl2 caused a concentration-dependent increase in follicular cell apoptosis. In both fathead and catfish models, although all follicle survival factors increased HSP70 overexpression compared to control follicles, this result was only significant in the EGF treatment. Of all apoptogens tested in fathead minnow model, only testosterone significantly induced HSP70 expression in follicular cells.