Regulation of pregnancy-associated plasma protein A2 (PAPPA2) in a human placental trophoblast cell line (BeWo)

BACKGROUND Pregnancy-associated plasma protein A2 (PAPPA2) is an insulin-like growth factor-binding protein (IGFBP) protease expressed at high levels in the placenta and upregulated in pregnancies complicated by preeclampsia and HELLP (Hemolytic anemia, Elevated Liver enzymes, and Low Platelet count) syndrome. However, it is unclear whether elevated PAPPA2 expression causes abnormal placental development, or whether upregulation compensates for placental pathology. In the present study, we investigate whether PAPPA2 expression is affected by hypoxia, oxidative stress, syncytialization factors or substances known to affect the expression of PAPPA2's paralogue, PAPPA. METHODS BeWo cells, a model of placental trophoblasts, were treated with one of the following: hypoxia (2% O2), oxidative stress (20 microM hydrogen peroxide), forskolin (10 microM and 100 microM), TGF-beta (10 and 50 ng/mL), TNF-alpha (100 ng/mL), IL-1beta (100 ng/mL) or PGE2 (1 microM). We used quantitative RT-PCR (qRT-PCR) to quantify the mRNA levels of PAPPA2, as well as those of PAPPA and ADAM12 since these proteases have similar substrates and are also highly expressed in the placenta. Where we observed significant effects on PAPPA2 mRNA levels, we tested for effects at the protein level using an in-cell Western assay. RESULTS Hypoxia, but not oxidative stress, caused a 47-fold increase in PAPPA2 mRNA expression, while TNF-alpha resulted in a 6-fold increase, and both of these effects were confirmed at the protein level. PGE2 resulted in a 14-fold upregulation of PAPPA2 mRNA but this was not reflected at the protein level. Forskolin, TGF-beta and IL-1beta had no significant effect on PAPPA2 mRNA expression. We observed no effects of any treatment on PAPPA or ADAM12 expression. CONCLUSION Our study demonstrates that factors previously known to be highly expressed in preeclamptic placentae (PGE2 and TNF-alpha), contribute to the upregulation of PAPPA2. Hypoxia, known to occur in preeclamptic placentae, also increased PAPPA2 expression. These results are consistent with the hypothesis that PAPPA2 is upregulated as a consequence of placental pathology, rather than elevated PAPPA2 levels being a cause of preeclampsia.