Katsuhiko Sumiyoshi

Single Administration of α-Glucosidase Inhibitors on Endothelial Function and Incretin Secretion in Diabetic Patients With Coronary Artery Disease - Juntendo University Trial: Effects of Miglitol on Endothelial Vascular Reactivity in Type 2 Diabetic Patients With Coronary Heart Disease (J-MACH) -

Background: Post-prandial hyperglycemia, hyperlipidemia, and endothelial dysfunction play an important role in the pathogenesis of atherosclerosis. Improvement in post-prandial hyperglycemia on α-glucosidase inhibitors (α-GIs) is associated with a risk reduction of cardiovascular diseases, but the post-prandial effects of α-GIs on endothelial function and incretin secretion in type 2 diabetic patients with coronary artery disease (CAD) remain unclear. Methods and Results: The post-prandial effects of a single administration of miglitol and voglibose on endothelial function and changing levels of glucose, insulin, lipids, glucagon-like peptide (GLP)-1, and gastric inhibitory polypeptide (GIP) were compared after a standard meal loading in 11 diabetic patients with CAD, using a placebo-controlled cross-over design. The changing levels of glucose, insulin and triglycerides at 60 min were significantly lower in the miglitol group than in the voglibose and placebo groups (all P<0.01). GLP-1 levels were significantly higher at 120 min (P<0.05) and GIP levels were significantly lower at 30 min and 60 min (P<0.05) in the miglitol group compared to other treatments. The reactive hyperemia duration at 120 min was significantly maintained in the miglitol group compared to the other groups. Conclusions: A single administration of miglitol significantly improved post-prandial glucose/lipid metabolism, incretin secretion, and endothelial dysfunction in diabetic patients with CAD, suggesting that miglitol may be a useful anti-atherogenic agent (UMIN000002264).  (Circ J 2010; 74: 1471 - 1478)

Serum levels of remnant lipoprotein cholesterol and oxidized low-density lipoprotein in patients with coronary artery disease

BACKGROUND Oxidized low-density lipoprotein (OxLDL) and remnant lipoprotein play a crucial role in the development of atherosclerosis. Recently, a novel method for measuring remnant cholesterol levels (remnant lipoproteins cholesterol homogenous assay: RemL-C) has been established. However, the correlation between OxLDL and remnant lipoprotein, including RemL-C, has not been fully investigated. METHODS We enrolled 25 consecutive patients with documented coronary artery disease (CAD) and 20 controls. Remnant-like particle cholesterol (RLP-C) and RemL-C were used to determine the levels of remnant lipoprotein cholesterol. Serum levels of malondialdehyde-modified LDL (MDA-LDL) and OxLDL using a monoclonal antibody DLH3 (OxPC) were used to measure the concentration of circulating OxLDL. RESULTS The CAD group had high levels of fasting glucose and glycosylated hemoglobin (HbA1c), and low levels of high-density lipoprotein cholesterol compared with the control group. Serum levels of total cholesterol or LDL cholesterol were not significantly different between the two groups. The levels of RemL-C (p = 0.035), MDA-LDL (p = 0.018), and MDA-LDL/LDL-C (p = 0.036) in the CAD group were significantly higher than those in the control group. The levels of RLP-C tended to be higher in the CAD group than those in the control group (p = 0.096). Positive correlations were demonstrated between remnant lipoprotein cholesterol and OxLDL (RLP-C and MDA-LDL/LDL-C, r = 0.45, p = 0.0024, RLP-C and OxPC, r = 0.51, p = 0.0005, RemL-C and MDA-LDL/LDL-C, r = 0.42, p = 0.0044, RemL-C and OxPC, r = 0.43, p = 0.0043). Similar trends were observed in non-diabetic subjects and in subjects without metabolic syndrome. Positive correlations were also observed between RLP-C and RemL-C (r = 0.94, p < 0.0001) and between MDA-LDL/LDL-C and OxPC (r = 0.40, p = 0.0074). CONCLUSIONS These results suggest that the association between high levels of remnant lipoprotein cholesterol and high OxLDL levels might be linked to atherogenesis in patients with CAD.

Dynamin-2 Regulates Oxidized Low-Density Lipoprotein–Induced Apoptosis of Vascular Smooth Muscle Cell

Background—On exposure to oxidized low-density lipoprotein (oxLDL), vascular cells generally undergo apoptosis, which is one of the major pathogenic factors of atherosclerosis. In this study, we examined the role of dynamin (a crucial GTPase protein in endocytosis) in oxLDL-induced apoptosis of vascular smooth muscle cells (VSMC). Methods and Results—After oxLDL stimulation, dynamin-2 colocalized with LOX-1 around the cell surface, as well as oxLDL in the cytoplasm, suggesting that dynamin-2 was involved in scavenger receptor–mediated oxLDL endocytosis. Downregulation of dynamin-2 induced by dynamin-2 dominant negative plasmid (K44A) resulted in a decrease of oxLDL uptake and thereby in a reduction of apoptosis. These data demonstrated that dynamin-2 was involved in oxLDL-induced apoptosis via the oxLDL endocytotic pathway. On the other hand, dynamin-2 wild-type plasmid transfection promoted oxLDL-induced apoptosis without increasing oxLDL uptake. Interestingly, the p53 inhibitor pifithrin-α (PFT) significantly reduced apoptosis promoted by wild-type dynamin-2 (78% reduction compared with the PFT[−] condition). These results indicated that dynamin-2 enhanced oxLDL-induced apoptosis of VSMC by participating in the p53 pathway, probably as a signal transducer. Moreover, we demonstrated that, in advanced plaques of apolipoprotein E−/− mice, dynamin-2 expression was often enhanced in apoptotic VSMC, suggesting that dynamin-2 might participate in apoptosis of VSMC even in vivo. Conclusions—Our data demonstrated that dynamin-2 at least partially regulated oxLDL-induced apoptosis of VSMC by participating in 2 independent pathways: the oxLDL endocytotic pathway and the p53 pathway. These findings suggest that dynamin-2 may serve as a new research or therapeutic target in vascular disease.

Involvement of cholesterol-enriched microdomains in class A scavenger receptor-mediated responses in human macrophages

OBJECTIVE Lipid rafts are cholesterol-enriched microdomains on cell membranes. We hypothesized that these microdomains could involve modified low-density lipoprotein (LDL) uptake. METHODS AND RESULTS Co-localizations of cholesterol-enriched microdomains and CD204 during the uptake of acetyl LDL (AcLDL) and oxidized LDL were observed using Alexa488-labeled polyethylene glycol cholesteryl ester, which is a sensitive probe used to analyze the dynamics of cholesterol-rich lipid microdomains in living cells. The lipid raft disruptors, methyl-β cyclodextrin and filipin, inhibited the uptake of AcLDL. CD204 siRNA treatments significantly reduced AcLDL uptake by 80%. We also demonstrated the presence of CD204 in the detergent-resistant membrane fraction (DRM) by immunoblotting analysis. The ratio of CD204/flotillin-1 in DRM was increased 11.5-fold by modified LDL administration. The PI3 kinase inhibitor LY294002, but not the Src kinase inhibitor PP1 or the Gαi/o inhibitor pertussis toxin, inhibited modified LDL uptake. The production of interleukin (IL)-8, but not CCL2, CXCL2, CXCL3, IL-6 or tumor necrosis factor-α was increased by AcLDL administration. The AcLDL-induced IL-8 production was inhibited by LY294002 and filipin. CONCLUSIONS These data firstly demonstrated that PI3 kinase-associated cholesterol-enriched microdomains are involved in CD204-mediated modified LDL uptake in human macrophages. Cholesterol-enriched microdomains may play a critical role in inflammatory processes.

High levels of saturated very long-chain fatty acid (hexacosanoic acid; C26:0) in whole blood are associated with metabolic syndrome in Japanese men

OBJECTIVE Hexacosanoic acid (C26:0) is a saturated very long-chain fatty acid and high levels of C26:0 in red blood cells are reported to be closely related with risk factors of atherosclerosis. However, the relationship between absolute levels of C26:0 in whole blood and metabolic syndrome (MS) has not been determined. MATERIALS AND METHOD We divided 218 consecutive apparently healthy male subjects into an MS group (n=78) and a non-MS group (n=140) according to the definition of the International Diabetes Federation. The levels of C26:0 in whole blood were measured by gas liquid chromatography-mass spectrometry. RESULTS The MS group had significantly higher levels of C26:0 than the non-MS group (2.42+/-0.31mug/ml vs. 2.25+/-0.29mug/ml, P=0001). There was a significant association between the levels of C26:0 and the number of factors of MS. The levels of C26:0 positively correlated with age, blood pressure, triglyceride and fasting plasma glucose. Multivariate analysis revealed that the level of C26:0 is still an independent variable for the presence of MS after adjustment for age and each criterion of MS. CONCLUSION The absolute levels of C26:0 in whole blood appear to be associated with MS independent of its component parts.

Presence of α-smooth muscle actin-positive endothelial cells in the luminal surface of adult aorta

Alpha-smooth muscle actin-positive endothelial cells have not been found in adult aortic endothelium except valve leaflets. Here, using en face immunostaining method, we identified alpha-smooth muscle actin-positive endothelial cells in the luminal surface of rat, mouse and human thoracic aortas. These cells express both endothelial markers and definite smooth muscle cell markers and were only occasionally observed in thoracic aorta of wild type mice and rats. Their density did not increase with aging. Given that alpha-smooth muscle actin-positive endothelial cells express low level of vascular endothelial-cadherin that is important for the maintenance of cell contact, these cells were frequently detected in the thoracic aorta of 5-week-old apolipoprotein-E deficient mice. In 20- to 24-week-old apolipoprotein-E deficient mice, marked accumulation of alpha-smooth muscle actin-positive endothelial cells was observed especially in the luminal surface of atheromatous plaques. Our findings indicate the existence of alpha-smooth muscle actin-positive endothelial cells in adult aortic endothelium and the possible association with progression of atherosclerosis.

Enhanced production of nitric oxide, reactive oxygen species, and pro-inflammatory cytokines in very long chain saturated fatty acid-accumulated macrophages

BackgroundDeterioration of peroxisomal β-oxidation activity causes an accumulation of very long chain saturated fatty acids (VLCSFA) in various organs. We have recently reported that the levels of VLCSFA in the plasma and/or membranes of blood cells were significantly higher in patients with metabolic syndrome and in patients with coronary artery disease than the controls. The aim of the present study is to investigate the effect of VLCSFA accumulation on inflammatory and oxidative responses in VLCSFA-accumulated macrophages derived from X-linked adrenoleukodystrophy (X-ALD) protein (ALDP)-deficient mice.ResultsElevated levels of VLCSFA were confirmed in macrophages from ALDP-deficient mice. The levels of nitric oxide (NO) production stimulated by lipopolysaccharide (LPS) and interferon-γ (IFN-γ), intracellular reactive oxygen species (ROS), and pro-inflammatory cytokines, including tumor necrosis factor-α (TNF-α), interluekin-6 (IL-6), and interleukin-12p70 (IL-12p70), were significantly higher in macrophages from ALDP-deficient mice than in those from wild-type mice. The inducible NO synthase (iNOS) mRNA expression also showed an increase in macrophages from ALDP-deficient mice.ConclusionThese results suggested that VLCSFA accumulation in macrophages may contribute to the pathogenesis of inflammatory diseases through the enhancement of inflammatory and oxidative responses.

Deletion of the Fc receptors γ chain preserves endothelial function affected by hypercholesterolaemia in mice fed on a high-fat diet

AIMS To clarify the role of Fc receptors (FcR) for immunoglobulin in endothelial dysfunction induced by hypercholesterolaemia, we evaluated the effect of deletion of the FcR gamma chain on endothelium-dependent relaxation and oxidative stress after 10 weeks on a high-fat diet in FcR gamma(-/-) mice compared with that in wild-type mice. METHODS AND RESULTS Plasma cholesterol levels of those on the high-fat diet were significantly increased compared with those on the normal chow diet in both groups of mice. Endothelium-dependent relaxation of the aortic ring with acetylcholine in wild-type mice was significantly reduced by the high-fat diet (ED(50): 0.22 vs. 0.43 nM, P < 0.002), whereas the relaxation in FcR gamma(-/-) mice was not inhibited (ED(50): 0.22 vs. 0.23 nM, NS). Furthermore, superoxide detection by dihydroethidium-derived fluorescence and immunohistochemical staining of p22phox expression were significantly increased in wild-type mice fed on the high-fat diet, while these oxidative stresses in FcR gamma(-/-) mice were not enhanced by the high-fat diet. Oil Red O-staining showed no significant lipid accumulation at the aortic sinus in both groups of mice. CONCLUSION This study demonstrates that the deletion of the FcR gamma chain preserves the endothelial function and attenuates oxidative stress affected by hypercholesterolaemia in FcR gamma(-/-) mice. These results indicate that FcR may play the pivotal role in endothelial dysfunction through oxidative stress induced by hypercholesterolaemia.

Effects of fenofibrate on lipid profiles, cholesterol ester transfer activity, and in-stent intimal hyperplasia in patients after elective coronary stenting

BackgroundThe association between modulation of detailed lipoprotein profiles and cholesterol ester transfer (CET) activity by peroxisome proliferator-activated receptor (PPAR)-a agonists in patients with coronary artery disease remains unclear. We assessed lipid profiles, plasma CET activity, and in-stent intimal hyperplasia after fenofibrate treatment in patients who underwent elective coronary stenting.MethodsForty-three consecutive patients who underwent elective coronary stenting were randomized to the fenofibrate group (300 mg/day for 25 weeks, n = 22) or the control group (n = 21). At baseline and follow up, CET activity and lipoprotein profiles were measured, and quantitative coronary angiography was performed.ResultsIn the fenofibrate group, the levels of large very low-density lipoprotein cholesterol, and small low-density lipoprotein (LDL) cholesterol decreased and those of small high-density lipoprotein (HDL) cholesterol increased. Besides, CET activity decreased independent of the effect of fenofibrate on total and LDL cholesterol. The reduction of CET activity significantly correlated with the increase in LDL particle size (r = 0.47, P = 0.03) and the decrease of triglycerides in large HDL subclasses (r = 0.48, P = 0.03). Although there were no significant differences in restenosis parameters between the two groups, low CET activity significantly correlated with the inhibition of neointimal hyperplasia (r = 0.56, P = 0.01).ConclusionsFenofibrate inhibited CET activity and thereby improved atherogenic lipoprotein profiles, and reduced intimal hyperplasia after coronary stenting.

Skeletonization with an ultrasonic scalpel is as safe as a non-skeletonized dissection in preserving the endothelial function of the human gastroepiploic artery

The right gastroepiploic artery (GEA) is frequently used as another in situ artery, other than the internal thoracic artery (ITA) in coronary artery bypass grafting (CABG). Skeletonizing the graft with an ultrasonic scalpel is now regarded as a useful technique; however, this technique may damage the endothelial function during harvesting the graft resulting in postoperative graft stenosis or occlusion. In the present study, GEA segments from nine patients were excised in both a skeletonized and non-skeletonized manner with an ultrasonic scalpel, and then were transported to the laboratory. The vessels were trimmed as rings, and were allotted to the group of skeletonized or non-skeletonized, accordingly. The force development in response to 1 mumol/l norepinephrine did not differ between the skeletonized and non-skeletonized groups. Endothelium-dependent relaxation induced by either acetylcholine or bradykinin was not impaired in the skeletonized group in comparison to the non-skeletonized group. No significant difference was observed in endothelium-independent relaxation elicited by sodium nitroprusside. Therefore, the skeletonization of the GEA with an ultrasonic scalpel was thus found to be as safe as a non-skeletonized dissection in preserving the vascular contractile ability or endothelium-dependent and -independent relaxation of the graft.