The endothelial glycocalyx is a specialized extracellular matrix that covers the apical side of vascular endothelial cells, projecting into the lumen of blood vessels. The composition of the glycocalyx has been studied in great detail and it is known to be composed of a mixture of proteoglycans, glycosaminoglycans, and glycoproteins. Although this structure was once believed to be a passive physical barrier, it is now recognized as a multifunctional and dynamic structure that participates in many vascular processes; including but not limited to vascular permeability, inflammation, thrombosis, mechanotransduction, and cytokine signaling. Because of its participation in many physiological and pathophysiological states, comprehensive knowledge of the glycocalyx will aid future vascular biologists in their research. With that in mind, this review will discuss the biochemical structure of the glycocalyx and its function in many vascular physiological processes. We will also briefly review a more recent discover in glycocalyx biology, the placental glycocalyx.
The human placenta is of vital importance for proper nutrient and waste exchange, immune regulation, and overall fetal health and growth. Specifically, the extracellular matrix (ECM) of placental syncytiotrophoblasts, which extends outward from the placental chorionic villi into maternal blood, acts on a molecular level to regulate and maintain this barrier. Importantly, placental barrier dysfunction has been linked to diseases of pregnancy such as preeclampsia and intrauterine growth restriction. To help facilitate our understanding of the interface, and develop therapeutics to repair or prevent dysfunction of the placental barrier, in vitro models of the placental ECM would be of great value. In this study we aimed to characterize the ECM of an in vitro model of the placental barrier using syncytialized BeWo choriocarcinoma cells. Syncytialization caused a marked change in syndecans, integral proteoglycans of the ECM, which matched observations of in vivo placental ECM. Syndecan-1 expression increased greatly and predominated the other variants. Barrier function of the ECM, as measured by electrical impedance, increased significantly during and after syncytialization, while the ability of THP-1 monocytes to adhere to syncytialized BeWos was greatly reduced compared to non-syncytialized controls. Furthermore, ECIS measurements indicated that ECM degradation with MMP-9, but not heparanase, decreased barrier function. This decrease in ECIS-measured barrier function was not associated with any changes in THP-1 adherence to syncytialized BeWos treated with heparanase or MMP9. Thus, syncytialization of BeWos provides a physiologically accurate placental ECM with a barrier function matching that seen in vivo.
During pregnancy, the placenta produces significant amounts of pro and anti‐angiogenic factors, notably VEGF, PlGF, and sFlt‐1. The exact cell population(s) most responsible for this production is not definitively known. In the event of placental insufficiency, placental ischemia can cause alterations in the production of these factors, favoring the anti‐angiogenic sFlt‐1 protein. This mechanism is believed to be a major driver of preeclampsia. Within the placenta, the maternal/fetal interface is defined by a layer of specialized syncytiotrophoblasts (ST) which line the maternal side of the underlying fetal chorionic vessels. These large multinucleated cells define the selectivity of nutrient/waste exchange from the maternal to the fetal circulation. In studying trophoblast function, immortalized trophoblast‐derived cell lines are commonly utilized, such as the BeWo choriocarcinoma cell line. These cells however, do not spontaneously syncytialize, and therefore do not likely reflect in vivo ST function. We hypothesized that inducing syncytialization of BeWo cells would alter their production of angiogenic factors in vitro. Syncytialization of BeWo cells was induced with forskolin treatment for 48 hours. At 48 hours, cellular morphology shifted from dense cobblestone patterning into majority large, multinucleated cells. Syncytialization caused ~5 fold increase in both VEGF and PlGF mRNA (p<0.0001). There were significant (p<0.0001) increases in the levels of sFlt‐1 splice variants 2 (~5 fold), 3 (~50 fold), and 4 (~3fold). As a result, there were significant (p,<0.005) ~2 fold increases in secreted VEGF and PlGF protein, while sFlt‐1 went from not detectable to 53 pg/ml as measured by ELISA. Perhaps most interestingly, when cultured in oxygen concentrations mimicking healthy and ischemic placentas (8% and 1% respectively), there was differential regulation of these factors at the transcriptional level. There was no change in VEGF, while sFlt V2 and V3 were significantly upregulated (2‐fold each, p<0.0005), while PlGF was significantly down regulated by 40% (p<0.0001). Taken together, these data suggest that syncytialization significantly alters expression of both pro and anti‐angiogenic factors in vitro. Future studies should take syncytialization state into account when extrapolating from cell culture to in vivo placental function.
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