Indiana University-Purdue University Indianapolis (IUPUI) / Metabolic syndrome (MetS) affects 1/3 of all Americans and is the clustering of three or more of the following cardiometabolic risk factors: obesity, hypertension, dyslipidemia, glucose intolerance, and insulin resistance. MetS drastically increases the incidence of coronary artery disease (CAD), which is the leading cause of mortality globally. A cornerstone of CAD is arterial remodeling associated with coronary smooth muscle (CSM) cytodifferentiation from a contractile phenotype to proliferative and osteogenic phenotypes. This cytodifferentiation is tightly coupled to changes in intracellular Ca2+ handling that regulate several key cellular functions, including contraction, transcription, proliferation, and migration. Our group has recently elucidated the time course of Ca2+ dysregulation during MetS-induced CAD development. Ca2+ transport mechanisms, including voltage-gated calcium channels, sarcoplasmic reticulum (SR) Ca2+ store, and sarco-endoplasmic reticulum Ca2+ ATPase (SERCA), are enhanced in early, mild disease and diminished in late, severe disease in the Ossabaw miniature swine. Using this well-characterized large animal model, I tested the hypothesis that this Ca2+ dysregulation pattern occurs in multiple etiologies of CAD, including diabetes and aging. The fluorescent intracellular Ca2+ ([Ca2+]i) indicator fura-2 was utilized to measure [Ca2+]i handling in CSM from lean and diseased swine. I found that [Ca2+]i handling is enhanced in mild disease with minimal CSM phenotypic switching and diminished in severe disease with greater phenotypic switching, regardless of CAD etiology. We are confident of the translatability of this research, as the Ca2+ influx, SR Ca2+ store, and SERCA functional changes in CSM of humans with CAD are similar to those found in Ossabaw swine with MetS. Single-cell RNA sequencing revealed that CSM cells from an organ culture model of CAD exhibited many different phenotypes, indicating that phenotypic modulation is not a discreet event, but a continuum. Transcriptomic analysis revealed differential expression of many genes that are involved in the osteogenic signaling pathway and in cellular inflammatory responses across phenotypes. These genes may be another regulatory mechanism common to the different CAD etiologies. This study is the first to show that CSM Ca2+ dysregulation is common among different CAD etiologies in a clinically relevant animal model.

Indiana University-Purdue University Indianapolis (IUPUI) / Adenosine is widely thought to elicit coronary vasodilation and attenuate smooth muscle cell (SMC) proliferation, thereby providing cardioprotection. We cloned the porcine adenosine A1 receptor (A1R) subtype and found that it paradoxically stimulated proliferation of cultured coronary SMC by the extracellular signal-regulated protein kinases 1 and 2 (ERK1/2) signaling pathways, thus suggesting A1R dysregulation could play a role in coronary artery disease (CAD), restenosis, and regulation of coronary blood flow (CBF). We utilized the Ossabaw swine model of metabolic syndrome (MetS) to test the hypothesis that A1R activation contributes to development of CAD, in-stent stenosis, and CBF regulation. Swine were fed standard chow (Lean) or excess calorie atherogenic diet for over 20 weeks, which elicited MetS characteristics and coronary atherosclerosis compared to Lean. We observed increased A1R in native CAD in MetS, which was reversed by exercise training, and upregulation of A1R expression and A1R-ERK1/2 activation in an in vitro organ culture model of CAD. Intracoronary stent deployment followed by different durations of recovery showed A1R upregulation occurred before maximal in-stent stenosis in vi vivo. More importantly, selective A1R antagonism with 8-cyclopentyl-1, 3-dipropylxanthine (DPCPX)-eluting stents decreased coronary ERK1/2 activation and reduced in-stent stenosis comparable to Taxus® (paclitaxel-eluting stents). A1R antagonism potentiated vasodilatory effects of some vasodilators other than adenosine in porcine coronary microcirculation under basal conditions. Short-term exercise training around stenting prevented stent-induced microvascular dysfunction and attenuated native atheroma in the genetically lean Yucatan swine. Conclusions: A1R upregulation and activation contributes to coronary in-stent stenosis in vivo in MetS, plays a role in the development of coronary atherosclerosis in vitro, and might involve in CBF dysregulation in dyslipidemia and stenting. Exercise training decreased A1R expression in atherosclerosis, reduced native atheroma, and prevented stent-induced microvascular dysfunction. Selective pharmacological antagonism of A1R holds promise for treatment of CAD.