Premature Luteinization and Cumulus Cell Defects in Ovarian-SpecificSmad4Knockout Mice

Abstract

SMAD4 is a central component of the TGFbeta superfamily signaling pathway. Within the ovary, TGFbeta-related proteins play crucial roles in controlling granulosa cell growth, differentiation, and steroidogenesis. To study the in vivo roles of SMAD4 during follicle development, we generated an ovarian conditional knockout of Smad4 using the cre/loxP recombination system. Smad4 ovarian-specific knockout mice are subfertile with decreasing fertility over time and multiple defects in folliculogenesis. Regulation of steroidogenesis is disrupted in the Smad4 conditional knockout, leading to increased levels of serum progesterone. In addition, severe cumulus cell defects are present both in vivo and when assayed in vitro. These findings demonstrate that disrupting signaling through SMAD4 in the ovarian granulosa cells leads to premature luteinization of granulosa cells and eventually premature ovarian failure, thereby demonstrating key in vivo roles of TGFbeta superfamily signaling in the timing of granulosa cell differentiation.

Interference RNA-based silencing of endogenous SMAD4 in porcine granulosa cells resulted in decreased FSH-mediated granulosa cells proliferation and steroidogenesis

“…Our findings showed that FSHR is significantly down-regulated in SMAD4-siRNA-treated cells, which is in agreement with a previous study in which FSHR was slightly downregulated in GCs from pregnant mare's serum gonadotrophin (PMSG)-stimulated SMAD4 cKO ovaries (Pangas et al 2006). Knockdown of SMAD4 also diminished the ability of FSH to upregulate its own receptor.…”

“…Recently, SMAD1/5/8 cKO mice have been generated by Pangas' group, and these mice developed metastatic tumors with increased signaling of SMAD2/3 in GCs . Moreover, ablation of all TGFB signaling in the ovarian GCs by conditionally inactivating SMAD4 resulted in premature luteinization of GCs, leading to the predisposition of premature ovarian failure (Pangas et al 2006). The R-SMADs form heteromeric complexes with SMAD4, which translocates to the nucleus and then modulates transcription of TGFB target genes.…”

Relative expression of genes encoding SMAD signal transduction factors in human granulosa cells is correlated with oocyte quality

“…5). This may be consistent with the cumulus cell defects, premature luteinization of granulosa cells and premature ovarian failure seen in Smad4 ovarianspecific knockout mice [27]. Moreover, these findings illustrate that oocyte number alone may not be predictive of oocyte quality and subsequent IVF outcomes.…”

“…As the SMAD signal transduction proteins are known to mediate signaling by members of the TGF-β superfamily, which is involved in the control of normal folliculogenesis and ovulation [7,8], and they associate with members of the Forkhead family of transcription factors, the FOXO class [34], thought to play important roles in oocyte maturation, ovulation, and possibly luteinization [34][35][36][37][38][39], with expression in human granulosa cells [40], it is possible that SMAD expression levels may also affect ovarian folliculogenesis and fertility. Female Smad3 −/− mice exhibit impaired folliculogenesis and reduced fertility [28,30] and Smad4 ovarian-specific knockout mice exhibit multiple defects in folliculogenesis and decreased fertility over time [27]. Female mice in which dominant negative Smad2 is expressed in granulosa cells exhibit impaired folliculogenesis and the females are subfertile [26].…”

“…In addition, ovarian-and granulosaspecific Smad knockout and transgenic mouse models exhibit a variety of defects in folliculogenesis, coupled with reduced fertility [26][27][28][29][30]. In humans, transcripts for SMAD2, 3 and 4 have been identified in granulosa cells collected at oocyte retrieval [31], which suggested that these and other SMADs might also be involved in human ovarian function and fertility.…”