M1 Macrophages Contribute to Endothelial Dysfunction and Oxidative Stress in Pre-eclampsia

Hypertensive disorders during pregnancy impact approximately 10% of pregnancies worldwide, with pre-eclampsia (PE) being a particularly significant burden on expecting mothers. PE is responsible for up to 14% of maternal deaths and contributes to 10-25% of perinatal deaths annually. This severe condition typically develops after the 20th week of pregnancy and is characterized by high blood pressure in conjunction with proteinuria or other end-organ dysfunction. It is associated with fetal growth restriction, maternal endothelial dysfunction, and chronic inflammation. In a healthy pregnancy, macrophages make up approximately 20-30% of the decidual leukocytes, being the second most abundant immune cell type in the placental bed. Macrophage populations undergo phenotype shifts throughout pregnancy and must maintain a delicate balance between pro-inflammatory M1 macrophages and anti-inflammatory M2 macrophages. This balance is disrupted in PE, where an excess of pro-inflammatory M1 macrophages is observed. While this imbalance in macrophage populations is well-documented, the precise consequences and role of altered macrophage phenotypes in PE pathophysiology remain unclear. In this study, we set out to determine the effect of PE-stimulated macrophages on endothelial cell activation and function. After obtaining informed consent from patients delivering at the University of Mississippi Medical Center, placental macrophages were isolated from normal pregnant (NP) and PE human placentas using magnetic beads. Isolated macrophages were then co-cultured overnight with Human Umbilical Vein Endothelial Cells (HUVECs). E-selectin, ICAM, and VCAM protein expression by HUVECs was assessed via Western blot, pre-proendothelin (PPET) and NLRP3 gene expression in HUVECs was assessed via RT-PCR, mitochondrial oxidative stress was measured via MitoSox Red using flow cytometry, and patient serum was used to measure cytokines and angiogenic factors on IsoSpark codeplex chips. Western blot analysis of endothelial activation markers revealed no significant differences in ICAM and E-selectin expression of HUVECs co-cultured with NP and PE macrophages (n=8/group; p=0.0953; p=0.7771, respectively). However, VCAM was significantly decreased in HUVECs co-cultured with PE placental macrophages vs NP placental macrophages (p=0.0411). PPET expression was 3.5 times higher, and NLRP3 expression was 2.7 times higher in HUVECs co-cultured with PE macrophages compared to those co-cultured with NP macrophages (n=8/group; p=0.3401; p=0.2176, respectively). HUVECs co-cultured with PE placental macrophages (n=10/group) exhibited significantly increased mitochondrial superoxide production compared to HUVECs co-cultured with NP macrophages (p=0.0273). Circulating levels of the pro-inflammatory cytokine IL-15 were significantly higher in PE patient plasma vs NP patient plasma (p=0.0026; n=8/group), while circulating IL-10 and IL-4 were significantly lower PE patient plasma vs NP (p=0.047 and p=0.0029, respectively; n=8/group). PE patients also showed significantly lower levels of circulating vascular endothelial growth factor (VEGF) than NP patients (p=0.0335; n=8/group). Additional analyses will include the determination of cytokine secretion profile of NP and PE macrophages, HUVEC migration and proliferation assays, and measurement of additional cytokines in patient serum and placental homogenates. The results thus far demonstrate that PE macrophages induce mitochondrial oxidative stress in endothelial cells and may play a role in endothelial dysfunction associated with PE pathophysiology. This Research was supported by funding from the National Institutes of Health Grants R01HL151407 This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.