Dataset Information

The glycocalyx core protein Glypican 1 protects vessel wall endothelial cells from stiffness-mediated dysfunction and disease.

ABSTRACT:

Aims

Arterial stiffness is an underlying risk factor and a hallmark of cardiovascular diseases. The endothelial cell (EC) glycocalyx is a glycan rich surface layer that plays a key role in protecting against EC dysfunction and vascular disease. However, the mechanisms by which arterial stiffness promotes EC dysfunction and vascular disease are not fully understood, and whether the mechanism involves the protective endothelial glycocalyx is yet to be determined. We hypothesized that endothelial glycocalyx protects the endothelial cells lining the vascular wall from dysfunction and disease in response to arterial stiffness.

Methods and results

Cells cultured on polyacrylamide (PA) gels of substrate stiffness 10 kPa (mimicking the subendothelial stiffness of aged, unhealthy arteries) showed a significant inhibition of glycocalyx expression compared to cells cultured on softer PA gels (2.5 kPa, mimicking the subendothelial stiffness of young, healthy arteries). Specifically, gene and protein analyses revealed that a glycocalyx core protein Glypican 1 was inhibited in cells cultured on stiff PA gels. These cells had enhanced endothelial cell dysfunction as determined by enhanced cell inflammation (enhanced inflammatory gene expression, monocyte adhesion, and inhibited nitric oxide expression), proliferation, and EndMT. Removal of Glypican 1 using gene-specific silencing with siRNA or gene overexpression using a plasmid revealed that Glypican 1 is required to protect against stiffness-mediated endothelial cell dysfunction. Consistent with this, using a model of age-mediated stiffness, older mice exhibited a reduced expression of Glypican 1 and enhanced endothelial cell dysfunction compared to young mice. Glypican 1 gene deletion in knockout mice (GPC1-/-) exacerbated endothelial dysfunction in young mice, which normally had high endothelial expression, but not in old mice that normally expressed low levels. Endothelial cell dysfunction was exacerbated in young, but not aged, Glypican 1 knockout mice (GPC1-/-).

Conclusion

Arterial stiffness promotes EC dysfunction and vascular disease at least partly through the suppression of the glycocalyx protein Glypican 1. Glypican 1 contributes to the protection against endothelial cell dysfunction and vascular disease in endothelial cells.

SUBMITTER: Mahmoud M

PROVIDER: S-EPMC8152694 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:The endothelial glycocalyx is a dynamic signaling surface layer that is involved in the maintenance of cellular homeostasis. The glycocalyx has a very diverse composition, with glycoproteins, proteoglycans, and glycosaminoglycans interacting with each other to form a mesh-like structure. Due to its highly interactive nature, little is known about the relative contribution of each glycocalyx constituent to its overall function. Investigating the individual roles of the glycocalyx components to cellular functions and system physiology is challenging, as the genetic manipulation of animals that target specific glycocalyx components may result in the development of a modified glycocalyx. Thus, it is crucial that genetically modified animal models for glycocalyx components are characterized and validated before the development of mechanistic studies. Among the glycocalyx components, glypican 1, which acts through eNOS-dependent mechanisms, has recently emerged as a player in cardiovascular diseases. Whether glypican 1 regulates eNOS in physiological conditions is unclear. Herein, we assessed how the deletion of glypican 1 affects the development of the pulmonary endothelial glycocalyx and the impact on eNOS activity and endothelial function. Male and female 5-9-week-old wild-type and glypican 1 knockout mice were used. Transmission electron microscopy, immunofluorescence, and immunoblotting assessed the glycocalyx structure and composition. eNOS activation and content were assessed by immunoblotting; nitric oxide production was assessed by the Griess reaction. The pulmonary phenotype was evaluated by histological signs of lung injury, in vivo measurement of lung mechanics, and pulmonary ventilation. Glypican 1 knockout mice showed a modified glycocalyx with increased glycocalyx thickness and heparan sulfate content and decreased expression of syndecan 4. These alterations were associated with decreased phosphorylation of eNOS at S1177. The production of nitric oxides was not affected by the deletion of glypican 1, and the endothelial barrier was preserved in glypican 1 knockout mice. Pulmonary compliance was decreased, and pulmonary ventilation was unaltered in glypican 1 knockout mice. Collectively, these data indicate that the deletion of glypican 1 may result in the modification of the glycocalyx without affecting basal lung endothelial function, validating this mouse model as a tool for mechanistic studies that investigate the role of glypican 1 in lung endothelial function.

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