Low molecular weight heparin promotes transcription and release of placental growth factor from endothelial cells.

Circulating levels of placental growth factor (PlGF) are significantly reduced in women who develop preeclampsia. Low molecular weight heparin (LMWH) has been shown to acutely elevate circulating PlGF levels in pregnant women at increased risk of preeclampsia. The objective of the current investigation was to determine the mechanisms by which LMWH mediates the extracellular release of PlGF from endothelial cells. Cultured human aortic endothelial cells (HAECs) and human umbilical vein endothelial cells (HUVECs) were exposed to LMWH; PlGF transcription, translation, mobilization, and secretion were then assessed. LMWH significantly increased the release of PlGF from both HAECs and HUVECs. LMWH treatment promoted a significant increase of PlGF-1 mRNA expression in HAECs, accompanied by the intracellular transport and release of PlGF into the conditioned media. LMWH-mediated release of PlGF from HAECs was not directly mediated by extracellular mobilization, synthesis, or stability of PlGF mRNA/protein. LMWH exposure promotes the release of PlGF from endothelial cells through the upregulation of PlGF-1 mRNA expression. Stimulation of circulating PlGF levels by LMWH may be an important mechanism by which LMWH could reduce the risk of preeclampsia or minimize disease severity.NEW & NOTEWORTHY There are few therapeutic options available for the prevention of preeclampsia, a serious hypertensive disorder of pregnancy. Women who subsequently develop preeclampsia exhibit significantly reduced circulating levels of the proangiogenic placental growth factor protein. Low molecular weight heparin (LMWH) has previously been investigated as a preventative therapy against the development of preeclampsia; however, its mechanism of action is not known. The current study determined that LMWH promotes the transcription and release of placental growth factor protein from endothelial cells, providing a mechanistic basis by which LMWH could reduce the risk of preeclampsia or minimize disease severity.

The antithrombin binding regions of heparin mediate fetal growth and reduced placental damage in the RUPP model of preeclampsia†.

Preeclampsia is a serious hypertensive disorder of pregnancy, which is only cured with delivery of the placenta, thereby commonly necessitating preterm birth of the fetus. Low-molecular-weight heparin (LMWH) has demonstrated potential to reduce the incidence of preeclampsia in high-risk pregnant women, although the underlying mechanism by which LMWH protects against preeclampsia is unknown. Given the complex structure and biologic actions of heparin, we tested the hypothesis that heparin can mediate preeclampsia prevention via nonanticoagulant pathways. We compared the effects of a nonanticoagulant, glycol-split LMWH (gsHep)-rendered nonanticoagulant through disruption of the antithrombin binding regions-with the LMWH dalteparin in the rat reduced uterine perfusion pressure (RUPP) surgical model of preeclampsia. Although RUPP animals exhibit significantly elevated blood pressure and reduced plasma levels of placental growth factor (PGF) compared to sham, neither dalteparin nor gsHep treatment significantly impacted these parameters. However, the observed positive correlation between PGF levels and number of viable fetuses in RUPP-induced animals suggests that reduced PGF levels were predominately due to placental loss. Daily subcutaneous injections of low-dose dalteparin but not gsHep significantly restored fetal growth that was impaired by RUPP surgery. Placentas from RUPP animals exhibited an abnormal labyrinth structure, characterized by expanded sinusoidal blood spaces, relative to sham-operated animals. Morphometric analysis demonstrated that dalteparin but not gsHep treatment normalized development of the labyrinth in RUPP-exposed conceptuses. These data suggest that the antithrombin-binding regions of LMWH are required to confer its protective effects on fetal growth and placental development.

Effects of glycol-split low molecular weight heparin on placental, endothelial, and anti-inflammatory pathways relevant to preeclampsia.

Low molecular weight heparin (LMWH) is being investigated as a potential preventative therapy against preeclampsia. There is evidence suggesting that LMWH may prevent preeclampsia through anticoagulation-independent mechanisms. In this study, we compared the in vitro placental, endothelial, and anti-inflammatory effects of an LMWH (dalteparin) with a nonanticoagulant, glycol-split heparin derivative (gsHep). In contrast with dalteparin, gsHep did not interact with antithrombin III, possess significant anti-Factor Xa activity, or significantly prolong in vitro plasma clotting time. However, dalteparin and gsHep were otherwise mechanistically similar, both interacting with soluble fms-like tyrosine kinase-1 (sFlt1) and promoting release of the pro-angiogenic protein placental growth factor, but not the antiangiogenic sFlt1, from healthy placental villous explants. Placental explant media pretreated with dalteparin or gsHep significantly stimulated endothelial cell tube formation compared to untreated explants. Lastly, dalteparin and gsHep both significantly suppressed inflammation by inhibiting complement activation and leukocyte adhesion to endothelial cells that were activated using serum from preeclamptic women. Our data suggest that nonanticoagulant heparin derivatives may be utilized as a tool to distinguish the anticoagulation-independent mechanisms of LMWH, and provide insight into the role of anticoagulation in the prevention of preeclampsia.

Polyphosphate suppresses complement via the terminal pathway.

Polyphosphate, synthesized by all cells, is a linear polymer of inorganic phosphate. When released into the circulation, it exerts prothrombotic and proinflammatory activities by modulating steps in the coagulation cascade. We examined the role of polyphosphate in regulating the evolutionarily related proteolytic cascade complement. In erythrocyte lysis assays, polyphosphate comprising more than 1000 phosphate units suppressed total hemolytic activity with a concentration to reduce maximal lysis to 50% that was 10-fold lower than with monophosphate. In the ion- and enzyme-independent terminal pathway complement assay, polyphosphate suppressed complement in a concentration- and size-dependent manner. Phosphatase-treated polyphosphate lost its ability to suppress complement, confirming that polymer integrity is required. Sequential addition of polyphosphate to the terminal pathway assay showed that polyphosphate interferes with complement only when added before formation of the C5b-7 complex. Physicochemical analyses using native gels, gel filtration, and differential scanning fluorimetry revealed that polyphosphate binds to and destabilizes C5b,6, thereby reducing the capacity of the membrane attack complex to bind to and lyse the target cell. In summary, we have added another function to polyphosphate in blood, demonstrating that it dampens the innate immune response by suppressing complement. These findings further establish the complex relationship between coagulation and innate immunity.