The Ubiquitin-Proteasome System and Inflammatory Activity in Diabetic Atherosclerotic Plaques: Effects of Rosiglitazone Treatment

Diabetes, Mar 2006

The role of ubiquitin-proteasome system in the accelerated atherosclerotic progression of diabetic patients is unclear. We evaluated ubiquitin-proteasome activity in carotid plaques of asymptomatic diabetic and nondiabetic patients, as well as the effect of rosiglitazone, a peroxisome proliferator–activated receptor (PPAR)-γ activator, in diabetic plaques. Plaques were obtained from 46 type 2 diabetic and 30 nondiabetic patients undergoing carotid endarterectomy. Diabetic patients received 8 mg rosiglitazone (n = 23) or placebo (n = 23) for 4 months before scheduled endarterectomy. Plaques were analyzed for macrophages (CD68), T-cells (CD3), inflammatory cells (HLA-DR), ubiquitin, proteasome 20S activity, nuclear factor (NF)-κB, inhibitor of κB (IκB)-β, tumor necrosis factor (TNF)-α, nitrotyrosine, matrix metalloproteinase (MMP)-9, and collagen content (immunohistochemistry and enzyme-linked immunosorbent assay). Compared with nondiabetic plaques, diabetic plaques had more macrophages, T-cells, and HLA-DR+ cells (P < 0.001); more ubiquitin, proteasome 20S activity (TNF-α), and NF-κB (P < 0.001); and more markers of oxidative stress (nitrotyrosine and O2− production) and MMP-9 (P < 0.01), along with a lesser collagen content and IκB-β levels (P < 0.001). Compared with placebo-treated plaques, rosiglitazone-treated diabetic plaques presented less inflammatory cells (P < 0.01); less ubiquitin, proteasome 20S, TNF-α, and NF-κB (P < 0.01); less nitrotyrosine and superoxide anion production (P < 0.01); and greater collagen content (P < 0.01), indicating a more stable plaque phenotype. Similar findings were obtained in circulating monocytes obtained from the two groups of diabetic patients and cultured in the presence or absence of rosiglitazone (7.0 μmol/l). Ubiquitin-proteasome over-activity is associated with enhanced inflammatory reaction and NF-κB expression in diabetic plaques. The inhibition of ubiquitin-proteasome activity in atherosclerotic lesions of diabetic patients by rosiglitazone is associated with morphological and compositional characteristics of a potential stable plaque phenotype, possibly by downregulating NF-κB-mediated inflammatory pathways.

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The Ubiquitin-Proteasome System and Inflammatory Activity in Diabetic Atherosclerotic Plaques: Effects of Rosiglitazone Treatment

Raffaele Marfella 1 2 Michele D'Amico 0 1 Katherine Esposito 1 2 Alfonso Baldi 6 Clara Di Filippo 0 1 Mario Siniscalchi 2 Ferndinando Carlo Sasso 2 Michele Portoghese 5 Francesca Cirillo 4 Federico Cacciapuoti 2 Ornella Carbonara 2 Basilio Crescenzi 4 Feliciano Baldi 6 Antonio Ceriello 3 7 Giovanni Francesco Nicoletti 7 Francesco D'Andrea 7 Mario Verza 2 Ludovico Coppola 2 Francesco Rossi 0 1 Dario Giugliano 1 2 0 Department of Experimental Medicine, Second University Naples , Naples, Italy ; the 1 Cardiovascular Research Center, Second University Naples , Naples, Italy ; the 2 Department of Geriatrics and Metabolic Diseases , Second Univer- sity Naples, Naples, Italy ; the 3 Department of Pathology and Medi- cine, Experimental and Clinical, University of Udine , Udine, Italy ; and the 4 Cardiovascular Unit, Hospital V. Monaldi , Naples, Italy ; the 5 Cardiovas- cular Surgery Unit, Sassari Hospital , Sassari, Italy ; the 6 Department of Biochemistry, Section of Pathology, Second University Naples , Naples, Italy ; the 7 Department of Surgery, Second University of Naples , Naples, Italy . Emilio Scaglione 141, 80145 Napoli, Italy The role of ubiquitin-proteasome system in the accelerated atherosclerotic progression of diabetic patients is unclear. We evaluated ubiquitin-proteasome activity in carotid plaques of asymptomatic diabetic and nondiabetic patients, as well as the effect of rosiglitazone, a peroxisome proliferator-activated receptor (PPAR)- activator, in diabetic plaques. Plaques were obtained from 46 type 2 diabetic and 30 nondiabetic patients undergoing carotid endarterectomy. Diabetic patients received 8 mg rosiglitazone (n 23) or placebo (n 23) for 4 months before scheduled endarterectomy. Plaques were analyzed for macrophages (CD68), T-cells (CD3), inflammatory cells (HLADR), ubiquitin, proteasome 20S activity, nuclear factor (NF)-B, inhibitor of B (I B)-, tumor necrosis factor (TNF)-, nitrotyrosine, matrix metalloproteinase (MMP)9, and collagen content (immunohistochemistry and enzyme-linked immunosorbent assay). Compared with nondiabetic plaques, diabetic plaques had more macrophages, T-cells, and HLA-DR cells (P < 0.001); more ubiquitin, proteasome 20S activity (TNF-), and NF-B ( P < 0.001); and more markers of oxidative stress (nitrotyrosine and O2 production) and MMP-9 (P < 0.01), along with a lesser collagen content and IB- levels (P < 0.001). Compared with placebo-treated plaques, rosiglitazone-treated diabetic plaques presented less inflammatory cells (P < 0.01); less ubiquitin, proteasome 20S, TNF-, and NF-B ( P < 0.01); less nitrotyrosine and superoxide anion production (P < 0.01); and greater collagen content (P < 0.01), indicating a more stable plaque phenotype. Similar findings were obtained in circulating monocytes obtained from the - two groups of diabetic patients and cultured in the presence or absence of rosiglitazone (7.0 mol/l). Ubiquitinproteasome over-activity is associated with enhanced inflammatory reaction and NF-B expression in diabetic plaques. The inhibition of ubiquitin-proteasome activity in atherosclerotic lesions of diabetic patients by rosiglitazone is associated with morphological and compositional characteristics of a potential stable plaque phenotype, possibly by downregulating NF-B-mediated inflammatory pathways. Diabetes 55:622 632, 2006 Ccause of death in patients with type 2 diabetes ardiovascular disease represents the leading (1). Diabetes leads to increased vulnerability for plaque disruption and mediates increased incidence and severity of clinical events (2). Inflammation, particularly in diabetes, plays a central role in the cascade of events that result in plaque erosion and fissuring (2). There is emerging evidence that the ubiquitin-proteasome system, the major pathway for nonlysosomal intracellular protein degradation in eucaryotic cells, induces inflammation in both initial stage and progression of atherosclerosis (3). The ubiquitin-mediated proteolytic pathway involves the conjugation of multiple moieties of ubiquitin, a 76amino acid polypeptide, to cellular proteins in a multienzymatic process, targeting these proteins to degradation (4). This ligation of ubiquitin by a series of ubiquitinconjugating enzymes produce polyubiquitin chains, which serve as targeting signals for degradation of the protein by the proteasome. The multicatalytic proteasome consists of a central catalytic core, the 20S proteasome, and two regulatory 19S complexes (5). Moreover, the ubiquitinproteasome pathway is required for activation of nuclear factor kappa B (NF-B), a central transcription factor that regulates inflammatory genes, by degradation of its inhibitory IB proteins (6). Although it has been demonstrated that diabetes may upregulate ubiquitin-proteasome pathway in rat muscle (7), still no evidence exists about the potential role of ubiquitin-proteasome system in the evolution of atherosclerotic plaques of diabetic patients. We hypothesized that by inc (...truncated)


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Raffaele Marfella, Michele D’Amico, Katherine Esposito, Alfonso Baldi, Clara Di Filippo, Mario Siniscalchi, Ferndinando Carlo Sasso, Michele Portoghese, Francesca Cirillo, Federico Cacciapuoti, Ornella Carbonara, Basilio Crescenzi, Feliciano Baldi, Antonio Ceriello, Giovanni Francesco Nicoletti, Francesco D’Andrea, Mario Verza, Ludovico Coppola, Francesco Rossi, Dario Giugliano. The Ubiquitin-Proteasome System and Inflammatory Activity in Diabetic Atherosclerotic Plaques: Effects of Rosiglitazone Treatment, Diabetes, 2006, pp. 622-632, 55/3, DOI: 10.2337/diabetes.55.03.06.db05-0832