11β-HSD2 downregulation leads to upregulation of ADAM17 expression
Increased sFlt1 release is associated with either increased sFlt-1 transcripts or increased shedding of sFlt1 from membrane Flt1. ADAM10 and ADAM17 are the sheddases for shedding of sFlt1form Flt1 (Zhao et al., 2010). Cortisol (10-6M) treatment significantly increased ADAM17 but not ADAM10 protein expression in placental explants in the presence of CBX (supplemental Fig.S9). Further, we found that cortisol (10-6M) treatment significantly enhanced ADAM17 expression in primary syncytiotrophoblasts transfected with 11β-HSD2 siRNA (Fig.7D). We then confirmed that sFlt1 release was mediated by ADAM17 by showing that sFlt1 secretion was significantly decreased in syncytiotrophoblasts with ADAM17 knockdown compared with the cells transfected with scramble siRNA (Fig.7E).
In the classic model of steroid hormone action, upon binding a ligand steroid receptors interact with the DNA at specific hormone response elements to modulate gene transcription. Bioinformatic analysis shows that there are seven putative glucocorticoid response elements (GREs) in the upstream (2kb) ofADAM17gene. We therefore investigated whether GCs have an impact on ADAM17gene transcriptional activity and the role of these elements in GC regulation of ADAM17 transcription in placental cells. As shown in Fig.7F, DEX (10-6M) significantly increased transcriptional activity of ADAM 17 reporter. When these elements were mutated, DEX did not affect transcriptional activity of the mutant GRE ADAM17 reporter in primary syncytiotrophoblasts.
In the animal models, we found that systemic administration of CBX (1.2 and 2.4 mg/kg) led to a significant increase in ADAM17 expression in the placenta compared with vehicle treatment (Fig.8A). Specific inhibition of placental 11β-HSD2 by administration of CBX-CSA could also significantly promote ADAM17 expression in placentas compared with vehicle control (Fig.8B). Administration of CBX-NPs and CBX-SCR had no effect on ADAM17 expression in placenta.
We then examined ADAM17 expression in human placental samples recruited from a Shanghai hospital. It was shown that ADAM17 levels were significantly increased in PE placentas compared with normal ones (Fig.8C). In addition, we also determined 11β-HSD2 protein expression in placenta and confirmed that placental 11β-HSD2 level was significantly decreased in PE patients.