Michitsugu Nakamura

Vascular Inflammation Is Negatively Autoregulated by Interaction Between CCAAT/Enhancer-Binding Protein-δ and Peroxisome Proliferator-Activated Receptor-γ

Abstract— CCAAT/enhancer-binding proteins (C/EBPs) upregulate transcription of various inflammatory cytokines and acute phase proteins, such as interleukin (IL)-1&bgr;, IL-6, tumor necrosis factor-&agr;, and cyclooxygenase-2. Recent studies have demonstrated that peroxisome proliferator-activated receptor (PPAR)-&ggr; is present in atherosclerotic lesions, and negatively regulates expression of these genes. Interestingly, PPAR-&ggr; gene promoter has tandem repeats of C/EBP-binding motif, and C/EBP-&dgr; plays a pivotal role in transactivation of PPAR-&ggr; gene. It has been well known that the interaction between C/EBPs and PPAR-&ggr; plays a central role in maintaining adipocyte differentiation and glucometabolism; however, the relationship between PPAR-&ggr; and C/EBPs in the vessel wall remains unclear. In the present study, we showed that a high level of C/EBP-&dgr; expression induced by inflammation positively regulated transcription and protein expression of PPAR-&ggr; in vascular smooth muscle cells (VSMCs). On the other hand, PPAR-&ggr; ligands troglitazone, pioglitazone, and 15-deoxy-&Dgr;12,14-prostaglandin J2 inhibited IL-1&bgr;-induced IL-6 expression at a transcriptional revel in VSMCs. Functional promoter analysis revealed that PPAR-&ggr; ligands inhibited IL-1&bgr;-induced transactivation of IL-6 gene via suppression of not only nuclear factor-&kgr;B but also C/EBP-DNA binding. Moreover, PPAR-&ggr; ligands suppressed protein expression and transcription of C/EBP-&dgr; through dephosphorylation of signal transducer and activator of transcription 3. These findings strongly suggest that C/EBP-&dgr; is negatively autoregulated via transactivation of PPAR-&ggr;. This feedback mechanism probably downregulates transcription of inflammatory cytokines and acute phase proteins, and modulates inflammatory responses in the early process of atherosclerosis.

Troglitazone induces apoptosis via the p53 and Gadd45 pathway in vascular smooth muscle cells

Thiazolidinediones, activators of peroxisome proliferator-activated receptor (PPAR)gamma, have been reported to induce apoptosis in many types of cells. In the present study, we investigated the effects of thiazolidinediones, troglitazone, and pioglitazone on the cell growth of vascular smooth muscle cells, and identified a specific effect of troglitazone in addition to PPARgamma activation. Subconfluent rat culture vascular smooth muscle cells were treated with or without PPARgamma activators, troglitazone (1-30 microM), or pioglitazone (1-30 microM) for 72 h. After treatment, cell viability was significantly reduced by troglitazone in concentrations of 5-30 microM but not by pioglitazone. Vascular smooth muscle cells appeared to float and shrink 48 h after treatment with 20 microM of troglitazone. In situ DNA labeling showed that the nuclei of these cells were positively stained, and genomic DNA extracted from the cells showed nucleosomal laddering. Messenger RNA expression levels of c-myc, p21, bax, bcl-2, and bcl-x were not changed by the treatment with troglitazone. In contrast, along with the induction of vascular smooth muscle cell apoptosis, both the mRNA and protein expression levels of p53 and Gadd45 markedly increased in response to troglitazone. These results strongly suggest that troglitazone can induce vascular smooth muscle cell apoptosis and that this effect is caused primarily by activation of the p53 and Gadd45 pathway but not by PPARgamma activation.

Soluble Fas ligand and atherosclerosis in hypertensive patients.

Background The Fas–Fas ligand (FasL) system is involved in apoptosis in many types of cells. Recently, the expression of FasL on endothelial cells was reported. FasL is cleaved by a metalloproteinase and released in serum as soluble FasL (sFasL). Vasoactive substances, including metalloproteinase, are modulated by endothelial dysfunction. Advanced atherosclerosis and impaired endothelial function are seen in hypertensive patients. The inflammatory response has an important role in the development of atherosclerosis, whereas C-reactive protein (CRP) is associated with the presence and severity of atherosclerosis. Objective To measure the intima–media thickness of the common carotid artery and evaluate the relationship between atherosclerosis and serum sFasL concentrations in hypertensive patients. Patients and main outcome measures Forty-seven patients with hypertension participated in the study. The intima–media thickness of the common carotid artery was evaluated by ultrasound imaging. Serum concentrations of sFasL were measured by enzyme-linked immunosorbent assay. Results Intima–media thickness correlated positively with age (r = 0.362, P = 0.012) and sFasL concentrations (r =0.332, P = 0.022), and negatively with creatinine clearance (r = −0.399, P = 0.0055). A general linear model analysis with atherosclerotic risk factors and sFasL revealed that age, sFasL, high-density lipoprotein-cholesterol and systolic blood pressure were significantly associated with intima–media thickness. Furthermore, we demonstrated that serum sFasL is directly associated with CRP concentration (r = 0.316, P = 0.030). Conclusions These results indicated that serum sFasL concentration is associated with atherosclerosis and inflammatory disease, in patients with hypertension.

Nuclear Factor 1 Is a Negative Regulator of gadd153 Gene Expression in Vascular Smooth Muscle Cells

Growth arrest and DNA damage inducible gene 153 ( gadd153 ) is expressed at very low levels in growing cells but is markedly induced in response to cellular stresses, including glucose deprivation, exposure to genotoxic agents, and other growth-arresting situations. Forced expression of GADD153 can induce cell cycle arrest and/or apoptosis in many types of cells. Recently, we reported that GADD153 was induced in vascular smooth muscle cells (VSMCs) in neointimal lesions of balloon-injured carotid arteries. To investigate the underlying molecular mechanisms of gadd153 gene expression in VSMCs, we isolated and characterized a promoter region of the rat gadd153 gene. Sequence alignments of this region revealed 1 TATA-like sequence and several well-known cis elements. The 5′-deletion analysis for this region showed that a domain spanning −447 through −368 drastically reduced the promoter activity to almost equal levels of promoterless control. Because this domain contained a consensus sequence for the nuclear factor 1 family of proteins (NF1), DNA-binding studies were performed by use of 2 types of NF1 consensus probes. Both probes were specifically shifted by nuclear extracts from proliferating VSMCs and were supershifted by antiserum against CCAAT transcription factor/NF1. In addition, promoter activity of a mutant luciferase vector, which was generated by a point mutation at the NF1 binding motif of the gadd153 gene, was 14-fold higher than that of a wild-type one. These results suggest that gadd153 gene expression in VSMCs is negatively regulated by an NF1-binding motif, and NF1 may act as an antiapoptotic factor by continuously suppressing gadd153 gene expression in growing VSMCs.

Large Thrombus Originating From Left Atrial Diverticulum A New Concern for Catheter Ablation of Atrial Fibrillation

A 45-year-old man was admitted to our hospital for evaluation of a left atrial (LA) mass. The patient had a history of dilated hypertrophic cardiomyopathy, atrial fibrillation (AF), diabetes mellitus, and sustained ventricular tachycardia that was treated by implantation of a cardioverter-defibrillator. He had developed AF 1 year earlier and was followed up at the outpatient clinic. He had been treated with warfarin at a dose that resulted in an international normalized ratio of 2.0 to 3.0. The LA mass was detected incidentally on routine transthoracic echocardiography. He was referred to our hospital because the size of the LA mass was unchanged after intensive anticoagulation with intravenous heparin infusion. The mobile LA mass along the interatrial septum was detected by transthoracic echocardiography in the apical 4-chamber view (Figure 1 and Movie I in the online-only Data Supplement). Transesophageal echocardiography showed a huge mobile mass (2.6×1.4 cm) originating from the roof of the LA (Figure 2A and Movie II in the online-only …

Rest-redistribution thallium-201 myocardial scintigraphic study in cardiac amyloidosis

Background: Histopathological study in amyloid heart demonstrates that myocyte destructed by the extracellular deposition of amyloid protein together with viable myocyte is present. We hypothesized that rapid thallium washout may be found in amyloid heart as in regions which have a mixture of viable myocyte and scar tissue in patients with myocardial infarction. Thus, the purpose of this study was to evaluate the extent and severity of myocardial damage due to amyloid deposits using the washout rate of the tracer on rest-redistribution thallium-201 (201Tl) myocardial scans in cardiac amyloidosis patients. Methods: Rest-redistribution 201Tl myocardial scintigraphy was performed in 5 patients with biopsy-proved systemic amyloidosis with cardiac involvement (amyloidosis group). The initial and delayed images were obtained 15 min and 4 h, respectively, after intravenous injection of the tracer of 111 MBq. Washout rate of the tracer was calculated. Twelve patients with no apparent heart disease served as controls (control group). Results: Mean washout rate of the whole heart was higher in the amyloidosis group than in the control group (56 ± 9% vs 36 ± 6%, p < 0.001). Particularly, 4 of the 5 patients in the amyloidosis group presented a very high rate of thallium clearance which ranged from 57 to 61%, and died in less than a year. In the remaining 1 patient who had a normal washout rate of the tracer in the first study, it changed from 40 to 53% during the 5-year follow-up period. Conclusions: Washout rate in the setting of rest and delayed 201Tl images may represent the severity of amyloid depositions in the myocardium and may provide prognostic information.