1 sFlt1 production in human endometrial stromal cells is reversibly regulated during hormone stimulation and withdrawal

Abstract

Introduction Preeclampsia is a major complication of pregnancy in which the placenta is known to have shallow implantation into the decidua. Studies have implicated sFlt1, a soluble vascular endothelial growth factor (VEGF) receptor protein, in the pathogenesis of preeclampsia through its ability to bind with VEGF and thereby neutralize its angiogenic functions. The presence of sFlt1 and its action in the endometrium is yet to be determined. We hypothesize that endometrial cells at the maternal–fetal interface may play a role in sFlt1 regulation during pregnancy. In this study, we are seeking to understand the dynamic regulation of sFlt1 production in human endometrial stromal cells (HESC) during reversible decidualization as a result of hormone stimulation and withdrawal. Aim To investigate the regulation of sFlt1 production in primary HESC during in an in vitro model of reversible decidualization that mimics the biphasic differentiation changes that occur during the ovulatory cycle. Methods Primary HESC were isolated and cultured according to published reports. Decidualization of HESC was induced either by adding a cocktail of estrogen, progesterone and cAMP (EPC) or cAMP alone for 4, 8 and 12 days. sFlt levels were assessed by Western blot or ELISA in the culture supernatant. sFlt1 mRNA expression was quantified by real-time RT-PCR. After decidualization, a subset of HESC were exposed to cocktail-free (no EPC) or cAMP free medium for 4, 8, and 12 days to study reversal of decidualization on sFlt1 expression. Results sFlt1 was detected in the culture supernatant of HESC. sFlt1 production decreased in a time-dependent manner during decidualization induced by the EPC cocktail and was totally abrogated after 12 days of treatment. This effect coincided with an increased production of VEGF and prolactin, and cellular morphologic changes characteristic of HESC decidualization. Upon hormonal and cAMP withdrawal, sFlt1 production began to increase in a time-dependent manner in concert with a reversal of decidualization as indicated by morphologic and biochemical criteria (e.g. prolactin and VEGF production). Although estrogen and progesterone are critical players during decidualization, our results indicated that cAMP is the dominant factor which regulates sFlt production. Conclusion HESC synthesize and secrete measurable quantities of sFlt1 that is downregulated during decidualization as a result of increased intracellular cAMP levels. With hormonal and cAMP withdrawal, sFlt1 production was regained, reflecting the plasticity of HESC. These findings may have important implications in diseases such as preeclampsia that involve abnormal decidualization and angiogenesis at the maternal-fetal interface. Studies are underway to understand the precise mechanisms of sFlt1 reduction by cAMP in the endometrium.