Role of endothelin signaling in osteochondrogenic differentiation and matrix calcification of vascular smooth muscle cells
Miller, Stephanie Lauren
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Type II diabetes mellitus (T2DM) and high serum levels of endothelin-1 (ET1) are associated with accelerated ectopic calcification of blood vessels. Most often, this calcification occurs in the medial layer, a layer populated with vascular smooth muscle cells (SMCs). Additionally, T2DM-related hyperglycemia has been implicated in increased incidence and size of atherosclerotic plaques. We aimed to determine whether ET1 signaling through its type A receptor (ETA) is critical for matrix calcification of SMCs in culture conditions that mimic hyperglycemia. Furthermore, we aimed to determine whether this effect on calcification is associated with ET1-ETA interaction alone or its interplay with receptor for advanced glycation end-products (RAGE) signaling that has been implicated in diabetes-associated nephropathy and vascular calcification. Although glucose in hyperglycemic concentrations did not initiate calcification, high glucose in procalcific medium enhanced SMC calcification up to three-fold after seven days. Additionally, we found that adding ET1 to hyperphosphatemic-hyperglycemic cultures enhanced glucose accelerated SMC calcification by 21%, but ET1 had no effect on calcification under high-phosphate conditions alone. Delivery of ETA antagonists BQ-123 and atrasentan to high phosphate, high glucose cultures, resulted in a 71% inhibition of SMC calcification. In addition to a substantial inhibition of calcification, SMCs treated with 1 μM atrasentan had higher mRNA expression levels of myocardin (the SMC master transcription factor co-activator) and lower expression levels of Runx2 compared to SMCs treated with procalcific high glucose media alone, suggesting the preservation of SMC phenotype in their original fate decision. Moreover, blocking RAGE signaling with a neutralizing antibody reduced glucose-accelerated calcification of SMCs, and blocked ET1-enhanced calcification. These results show that ETA antagonism is an efficacious inhibitor of matrix calcification under diabetic conditions and imply interplay between the RAGE and ETA pathways.
- Bioengineering