Advanced age and reduced hyaluronan synthesis impair glycocalyx integrity resulting in endothelial dysfunction
Authors: Lim Jisok , Kim John , Gogulamudi Venkateswara , Adeyemo Adelola , Hall Shelby , Bramwell Colton , Lesniewski Lisa 
Published: May 2024

The glycocalyx (GC) is a densely structured, gel-like matrix that coats the inner lining of the vascular endothelium. The GC plays a pivotal role in regulating the integrity of the vascular endothelium, primarily through mechano-sensing and -transducing shear stress, while also preserving endothelial barrier function. With advancing age, the GC deteriorates concomitantly with its essential constituent, hyaluronan (HA), which may lead to endothelial dysfunction. Therefore, we hypothesized that advanced age and reduced HA synthesis alone or together will impair GC integrity resulting in endothelial dysfunction. We have generated a tamoxifen-inducible, endothelial cell-specific, knockout mouse model of a hyaluronan producing enzyme, hyaluronan synthase 2 (HAS2), in young (4-6 mo) and old (22-23 mo, n=9-12/group). HAS2 mRNA expression in carotid endothelial cell lysates were reduced by ~47% in endothelial cell HAS2 knockout (ecHAS2KO) vs wildtype (ecHAS2WT, p=0.004) after tamoxifen treatment. The perfused boundary region (PBR), indicative of GC barrier function, was evaluated in vivo in the mesenteric microcirculation. The PBR was significantly impaired with aging (young vs old ecHAS2WT, p=0.047) and genotype (young ecHAS2WT vs KO, p=0.012) but there was no additive effect of aging and genotype (old ecHAS2WT vs KO, p=0.989). Subsequently, GC thickness was determined by measuring the depth of GC collapsed by leukocyte passage through the mesenteric segments. GC thickness was diminished by ~53% with aging (young vs old ecHAS2WT, p<0.0001) and ~59% with genotype (young ecHAS2WT vs KO, p<0.0001) but there was no synergistic effect of aging and genotype (old ecHAS2WT vs KO, p=0.893). Mechano-sensing and -transducing properties of the GC were assessed using ex vivo flow-induced vasodilation in isolated carotid arteries with and without the intra-luminal infusion of HA degrading enzyme, hyaluronidase (HYAL). Flow-induced vasodilation was diminished with aging (young vs old ecHAS2WT, p<0.0001) but this age-related decline was dampened with HYAL (p=0.212). The vasodilatory response to flow was reduced with genotype (young ecHAS2WT vs KO, p<0.0001) but this effect was weakened with HYAL (p=0.953). There was no augmentative effect of aging and genotype with and without HYAL (all p≥0.883). Acetylcholine induced endothelium-dependent dilation (EDD), a measure of endothelial function, was examined ex vivo in isolated mesenteric arteries. The maximal EDD was diminished by ~23% with aging (young vs old ecHAS2WT, p=0.04) but not genotype (young ecHAS2WT vs KO, p=0.962). There was an additive reduction in maximal EDD by ~46% with aging and genotype (old ecHAS2KO vs WT, p=0.021). Endothelium-independent dilation in response to sodium nitroprusside, an exogenous nitric oxide donor, was not different across groups with either aging, genotype, or both (ALL p>0.05). In conclusion, aging and reduced HA synthesis independently impaired GC integrity. As a result, mechano-sensing and -transducing property of GC was disturbed and further diminished by HYAL activity. In addition, endothelial function was impaired with aging and further exacerbated after a reduction in HA synthesis. Funded in part by awards from National Institutes of Health Awards R01 AG060395, R01 AG077751, R01 AG076748, T32 HL007576, T32 HL139451, Veteran's Affairs Merit Review Award I01 BX004492 and Nora Eccles Treadwell Foundation. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.