Abstract Introduction Atherosclerosis is a chronic inflammatory disease characterized by endothelial dysfunction, immune cell activation, and plaque destabilization, leading to acute cardiovascular events. Junctional cadherin 5-associated protein (JCAD/KIAA1462) is an endothelial junctional protein identified through genome-wide association studies as a susceptibility factor for coronary artery disease and arterial thrombosis. However, its role in vascular inflammation, neutrophil activation, and residual cardiovascular risk remains incompletely understood. Objectives The aim of this PhD project was to investigate the role of JCAD as a mediator of inflammation in atherosclerosis through translational clinical and experimental approaches. Specifically, the study aimed to: evaluate the association between circulating JCAD levels, plaque vulnerability, and cardiovascular outcomes in patients undergoing carotid endarterectomy; investigate the relationship between JCAD and neutrophil activation and degranulation mechanisms; assess the prognostic role of JCAD in patients with acute coronary syndrome and residual cardiovascular risk. Methods A translational approach combining clinical and experimental studies was employed. In the first part, circulating JCAD levels were measured in 175 patients undergoing carotid endarterectomy for high-grade carotid stenosis. Correlations with systemic inflammatory markers, intraplaque histological features, and major adverse cardiovascular events (MACE) during 24-month follow-up were assessed. In the second part, in vitro experiments were performed on human neutrophils isolated from healthy donors. Confocal microscopy, qPCR, ELISA assays, and pharmacological inhibition studies were used to investigate JCAD localization, release, and involvement in neutrophil degranulation pathways following stimulation with PMA and fMLP. In the third part, the prognostic significance of JCAD was evaluated in patients with acute coronary syndrome enrolled in the multicentre SPUM-ACS cohort, focusing on residual lipid, inflammatory, and combined inflammatory-lipid risk. Results Elevated circulating JCAD levels were associated with increased plaque vulnerability and systemic inflammation. JCAD positively correlated with inflammatory mediators involved in neutrophil activation and degranulation, including myeloperoxidase (MPO), neutrophil elastase (NE), matrix metalloproteinases (MMP-8 and MMP-9), interleukin-6 (IL-6), resistin, and CCL4. At the plaque level, higher JCAD concentrations were associated with increased M1 macrophages, mast cells, lymphocytes, tissue factor, MMP-9, and C-reactive protein, while inversely correlating with type III collagen and M2 macrophages. Circulating JCAD independently predicted MACE, acute coronary syndrome, and ischemic stroke during follow-up. Experimental studies demonstrated intracellular localization of JCAD within neutrophils, with partial co-localization with MMP-9-containing tertiary granules. Activated neutrophils released JCAD together with classical degranulation products, and this release was modulated through PI3K/Akt, ERK1/2, and p38 MAPK pathways. In the SPUM-ACS cohort, JCAD emerged as an independent predictor of recurrent cardiovascular events in patients with residual lipid risk, residual inflammatory risk, and combined residual inflammatory-lipid risk, independently of LDL cholesterol and hs-CRP levels. Conclusions This thesis identifies JCAD as a novel biomarker and potential mediator of vascular inflammation, neutrophil activation, plaque vulnerability, and residual cardiovascular risk. The findings support a pathophysiological role for JCAD in atherosclerosis and acute cardiovascular disease and suggest its potential utility as both a prognostic biomarker and a future therapeutic target in cardiovascular medicine.
JCAD come potenziale mediatore dell'infiammazione nell'aterosclerosi
MORIERO, MARGHERITA
2026-05-25
Abstract
Abstract Introduction Atherosclerosis is a chronic inflammatory disease characterized by endothelial dysfunction, immune cell activation, and plaque destabilization, leading to acute cardiovascular events. Junctional cadherin 5-associated protein (JCAD/KIAA1462) is an endothelial junctional protein identified through genome-wide association studies as a susceptibility factor for coronary artery disease and arterial thrombosis. However, its role in vascular inflammation, neutrophil activation, and residual cardiovascular risk remains incompletely understood. Objectives The aim of this PhD project was to investigate the role of JCAD as a mediator of inflammation in atherosclerosis through translational clinical and experimental approaches. Specifically, the study aimed to: evaluate the association between circulating JCAD levels, plaque vulnerability, and cardiovascular outcomes in patients undergoing carotid endarterectomy; investigate the relationship between JCAD and neutrophil activation and degranulation mechanisms; assess the prognostic role of JCAD in patients with acute coronary syndrome and residual cardiovascular risk. Methods A translational approach combining clinical and experimental studies was employed. In the first part, circulating JCAD levels were measured in 175 patients undergoing carotid endarterectomy for high-grade carotid stenosis. Correlations with systemic inflammatory markers, intraplaque histological features, and major adverse cardiovascular events (MACE) during 24-month follow-up were assessed. In the second part, in vitro experiments were performed on human neutrophils isolated from healthy donors. Confocal microscopy, qPCR, ELISA assays, and pharmacological inhibition studies were used to investigate JCAD localization, release, and involvement in neutrophil degranulation pathways following stimulation with PMA and fMLP. In the third part, the prognostic significance of JCAD was evaluated in patients with acute coronary syndrome enrolled in the multicentre SPUM-ACS cohort, focusing on residual lipid, inflammatory, and combined inflammatory-lipid risk. Results Elevated circulating JCAD levels were associated with increased plaque vulnerability and systemic inflammation. JCAD positively correlated with inflammatory mediators involved in neutrophil activation and degranulation, including myeloperoxidase (MPO), neutrophil elastase (NE), matrix metalloproteinases (MMP-8 and MMP-9), interleukin-6 (IL-6), resistin, and CCL4. At the plaque level, higher JCAD concentrations were associated with increased M1 macrophages, mast cells, lymphocytes, tissue factor, MMP-9, and C-reactive protein, while inversely correlating with type III collagen and M2 macrophages. Circulating JCAD independently predicted MACE, acute coronary syndrome, and ischemic stroke during follow-up. Experimental studies demonstrated intracellular localization of JCAD within neutrophils, with partial co-localization with MMP-9-containing tertiary granules. Activated neutrophils released JCAD together with classical degranulation products, and this release was modulated through PI3K/Akt, ERK1/2, and p38 MAPK pathways. In the SPUM-ACS cohort, JCAD emerged as an independent predictor of recurrent cardiovascular events in patients with residual lipid risk, residual inflammatory risk, and combined residual inflammatory-lipid risk, independently of LDL cholesterol and hs-CRP levels. Conclusions This thesis identifies JCAD as a novel biomarker and potential mediator of vascular inflammation, neutrophil activation, plaque vulnerability, and residual cardiovascular risk. The findings support a pathophysiological role for JCAD in atherosclerosis and acute cardiovascular disease and suggest its potential utility as both a prognostic biomarker and a future therapeutic target in cardiovascular medicine.
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