Differential Regulation of sFlt‐1 in response to VEGF Family‐Based Therapeutics In Endothelial Cells
Authors: Chapman Heather , Eddy Adrian C , Williams London , Moore Kyle , George Eric Matthew 
Published: June 2021

Preeclampsia (PE) is a hypertensive disorder of pregnancy which affects ~5–7% of all pregnancies in the United States. Hallmarked by new‐onset hypertension and organ damage, it is a leading cause of premature birth and maternal/fetal perinatal morbidity. While the cause of PE remains elusive, placental dysfunction due to chronic ischemia is believed to be a central regulator of the disorder. One of the factors known to be released from the ischemic placenta is the soluble form of the VEGF receptor Flt‐1 (sFlt‐1), which is released into the maternal circulation and is believed to be a major contributor to the maternal endothelial dysfunction seen in PE. It has been hypothesized that restoring angiogenic balance by way of administration of VEGF family members could be beneficial in PE. To that end, we have recently described several novel VEGF family‐based chimeric proteins with enhanced bioavailability and improved pharmacokinetic profiles. Recently, we have demonstrated that in vivo administration of a VEGFA‐based chimeric therapeutic protein was able to relieve the symptoms of PE in a rodent model, but resulted in adverse effects at dose‐limiting toxicity; in particular a marked increase in the circulating levels of sFlt‐1. Here we hypothesize that chimeric therapeutics based on the VEGF family members Placental Growth Factor (PlGF) and VEGFB will cause reduced production of sFlt‐1 from placental and vascular cells when compared to their VEGFA counterpart. To that end, we utilized cultured rodent placental explants, cultured BeWo cells derived from placental syncytiotrophoblasts, and primary Human Umbilical Vein Endothelial Cells (HUVEC) cells to determine the relative effects of the VEGF chimeras on sFlt‐1 production. Incubation of HUVECs with chimeric VEGFA caused a significant increase in released sFlt‐1 (6593±199 vs 14547±364 pg/ml, p<0.0001), while there was no induction with either chimeric PlGF (6830±88pg/ml) or VEGFB (7217±71pg/ml). To further elucidate the effects of chimeric VEGFA on HUVEC sFlt‐1 production, we examined three splice variants of sFlt‐1 as well as the full length Flt‐1 by qRT‐PCR. In response to VEGF, the β‐Actin normalized expression increased in sFlt v2 (100±34% vs 401±36%, p<0.0001), a trend to increase full length Flt‐1 message (100±34% vs 177±20%, p=−0.11) and sFlt‐1 v3 (123±8%, p=0.16) with no effect on sFlt v4 (100±34% vs 68±8%, p=0.36). This VEGF induced sFlt‐1 release was significantly attenuated with either a dual VEGFR1/VEGFR2 antagonist or a VEGR2 specific antagonist, but not a neuropilin‐1 selective antagonist, suggesting the receptor responsible for the release of sFlt‐1 in response to VEGF is the VEGFR2 receptor. In conclusion we conclude that the significant in vivo increase in sFlt‐1 caused by chronic chimeric VEGF administration is due to increased endothelial production of sFlt‐1. This is associated with increase production of the sFlt‐1 v2 splice variant, and is likely due to activation of the VEGFR2 receptor.Support or Funding InformationThis work was funded in part by NIH grants P01HL51971, P20GM104357, T32HL105324, R00HL11677, R25 HL121042‐01, and R01HL 137791.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.