All ETDs from UAB

Advisory Committee Chair

Rakesh P Patel

Advisory Committee Members

Scott Ballinger

Jarrod Barnes

Susan Bellis

Silvio Litovsky

Jan Novak

Document Type


Date of Award


Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine


Monocyte extravasation through the endothelial layer is a hallmark of atherosclerotic plaque development and is mediated by heavily glycosylated surface adhesion molecules, such as intercellular adhesion molecule-1 (ICAM-1). Human monocytes have been classified into three distinct groups: classical (anti-inflammatory; CD14+/CD16-), nonclassical (patrolling; CD14+/CD16++), and intermediate (pro-inflammatory; CD14++/CD16+). The CD16+ nonclassical / intermediate monocytes have been implicated in atherosclerosis progression and their levels positively associate with adverse cardiac events. However, there is a relative lack of understanding as to whether there are distinct mechanisms that regulate CD16+ vs. CD16- monocyte adhesion to the inflamed endothelium. Our previous data identified a high-mannose (HM) glycoform of ICAM-1 present during inflammation, but the function of different endothelial N-glycoforms is not yet known. Here, we test the hypothesis that distinct ICAM-1 N-glycoforms mediate recruitment of different monocyte subsets and specifically that endothelial HM-ICAM-1 promotes adhesion of CD16+ monocytes. We measured monocyte rolling and adhesion to i) TNFalpha-activated HUVEC, in which N-glycosylation was modulated by inhibiting alpha- mannosidase activity or by lectin-based blocking or ii) Cos-1 cells expressing HM- or complex ICAM-1. Expression of HM-ICAM-1 selectively enhanced CD16+ monocyte adhesion under flow with no effect on CD16- monocytes noted. This effect was dependent upon both HM epitopes and ICAM-1; adhesion was abrogated by blocking either. iv Further, using the proximity ligation assay (PLA) we show that HM-ICAM-1 is present in human and mouse atherosclerosis, increasing with disease severity and positively correlating with CD68 macrophage staining. Finally, we addressed the mechanism behind the formation of HM-ICAM-1 during inflammation by studying endothelial alpha-mannosidases, a class of enzymes responsible for early N-glycan processing. We show that TNFalpha decreases class I alpha-mannosidase activity in a time-dependent manner, resulting in formation of HM-ICAM-1 on the cell surface. We also show that this decrease in activity is independent of NF-kappaB, suggesting a parallel mechanism of N-glycan regulation distinct from up-regulation of adhesion molecule protein expression. Together, these data highlight a high-mannose ICAM-1 present in human and mouse atherosclerotic lesions as a key mediator of CD16+ monocyte recruitment to the endothelium, and identify alpha-mannosidases as a potential therapeutic target in atherosclerosis.



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