Kit ligand and c-Kit are expressed during early human ovarian follicular development and their interaction is required for the survival of follicles in long-term culture

Abstract

I B Carlsson and M P E Laitinen contributed equally to this paper AbstractThe receptor tyrosine c-Kit and its cognate ligand, c-Kit ligand (KL, stem cell factor, SCF), are involved in ovarian follicular development in several animal species. We studied the expression of KL and c-Kit using in situ hybridization and immunohistochemistry in donated human ovarian cortical tissue. The KL transcripts were expressed in granulosa cells of primary follicles, whereas the expression of c-Kit was confined to the oocyte and granulosa cells in primary and secondary follicles. We employed an ovarian organ culture using firstly serum-containing and then serum-free medium to study the effects of KL and an anti-c-Kit antibody, ACK2, on the development and survival of ovarian follicles in vitro. Culture of ovarian cortical slices for 7 days resulted in a 37% increase in the number of primary follicles and a 6% increase in secondary follicles. The proportion of viable follicles decreased in all cultures. The addition of KL (1, 10 and 100 ng/ml) into the culture media did not affect the developmental stages of the follicles or the proportion of atretic follicles. Inclusion of ACK2 (800 ng/ml) in the culture medium significantly increased the proportion of atretic follicles on days 7 (49 vs 28% in control cultures) and 14 (62 vs 38%) of culture. In conclusion, c-Kit and KL are expressed in human ovaries during follicular development. Blocking the c-Kit receptor induces follicular atresia. The KL/c-Kit signaling system is likely to control the survival of human ovarian follicles during early follicular development. Reproduction (2006) 131 641-649Introduction During a woman's reproductive life less than 1% of the follicles present in the human ovary at birth reach the stage of ovulation, while the rest undergo atresia and are lost. Some of the follicles present at birth start to grow during infancy and childhood, but most of them remain in the resting stage until they either degenerate or enter the growth phase during adult life (Adashi 1991, Gougeon 1996. It is likely that the mechanism behind the loss of germ cells is apoptotic cell death (Tilly 1996). Follicle-stimulating hormone (FSH) has been considered to be a key survival factor for follicles from the antral follicular stage onwards (McGee et al. 1998). However, the regulators of early follicular survival are still poorly characterized.The c-Kit ligand (KL, also called stem cell factor, SCF), a ligand for the c-Kit proto-oncogene receptor tyrosine kinase, is a pluripotent growth factor involved in the differentiation and growth of certain stem cell lineages including hematopoietic stem cells, neuroblasts, melanoblasts and primordial germ cells (PGCs) (Galli et al. 1994). The expression of KL has been detected in granulosa cells in mouse ovaries, while c-Kit receptor is confined to the oocytes and theca interna cells (Manova et al. 1990, Horie et al. 1991, Keshet et al. 1991, Motro et al. 1991, Motro & Bernstein 1993. In contrast to mice, c-Kit protein and mRNA have been detected...

KIT-KIT Ligand in the Growth of Porcine Oocytes in Primordial Follicles

“…It was unclear whether KL supported oocyte viability during in vitro treatment or in the subsequent xenografting in this experiment. Similar results have been reported by Carlsson et al [26], who employed organ culture for human ovarian tissues. They showed that KL does not affect follicular development, although the number of viable follicles is increased.…”

Dynamic Medium Containing Kit Ligand and Follicle-Stimulating Hormone Promotes Follicular Survival, Activation, and Growth during Long-Term in vitro Culture of Caprine Preantral Follicles

“…In mice, KL inhibited the apoptosis of oocytes from primordial follicles after in vitro culture [Jin et al, 2005] through an increase in the expression of antiapoptotic proteins Bcl-2 and Bcl-xL and a reduction in the expression of proapoptotic factor Bax. Other authors have also demonstrated that inhibition of the KL/c-kit interaction increases the proportion of atretic follicles at days 7 and 14 of culture in humans [Carlsson et al, 2006] and promotes oocyte death in mice Doneda et al, 2002;. In our study, KL addition until day 8 of culture was important for maintaining follicle and oocyte growth regardless of the substance tested from day 8 to day 16 of culture.…”

“…Similarly, in goat preantral follicles, different concentrations of KL (1, 10, 50, 100, or 200 ng/ ml) promoted follicular activation as early as after 1 day of in vitro culture [Celestino et al, 2010]. Nevertheless, in humans KL did not have an effect on preantral follicle development after 7 days of culture [Carlsson et al, 2006], indicating that the roles of KL differ between species. In the present study, after 8 days of culture the presence of KL in vitro as well as its replacement by FSH+KL or FSH alone maintained (KL/KL and KL/FSH+KL) or significantly increased (KL/FSH) the percentage of follicular activation until day 16.…”

Growth of Primordial Oocytes in Neonatal and Adult Mammals

“…This result suggests that KIT-KL might not be associated with the activation of porcine primordial oocytes, although they are necessary for the survival of the oocytes. Similar results have been reported by Carlsson et al [38], who employed organ culture for human ovarian tissues. They showed that KL did not affect follicular development, although the number of viable follicles was increased.…”