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Background: Serum and plasma are used for measurements of microRNAs (miRNAs) as biomarkers of various diseases. However, no consistent findings have been obtained regarding differences in serum and plasma levels of miRNAs. The purpose of this study was to clarify differences in serum and plasma levels of total miRNAs and their time-course changes after blood collection. Methods: Venous blood was collected from healthy men, and samples were prepared at the time points of 0, 15, 30, 60 and 180 min after blood collection for plasma and after clot formation for serum. Levels of total miRNAs were analyzed by the hybridization method using the 3D-Gene miRNA Oligo chip. Results: About one third of 2632 miRNAs tested showed levels high enough for comparison of serum and plasma levels and for investigation of their time-course changes. Levels of 299 miRNAs at time 0 were significantly different in serum and plasma. Levels of representative platelet-derived miRNAs including miR-185-5p, -22-3p and -320b were significantly higher in plasma than in serum, while levels of representative erythrocyte-derived miRNAs including miR-451a, -486-5p and -92a-3p were not significantly different in serum and plasma. Plasma levels of 173 miRNAs and 6 miRNAs showed significant decreasing and increasing tendencies, respectively, while there were no miRNAs in serum that showed significant time-course changes. Conclusion: The results suggest that careful attention should be paid when comparing serum and plasma levels of miRNAs and that plasma samples should be prepared early after blood collection.

Ethanol and resveratrol have been shown to inhibit platelet aggregation. The aim of this study was to determine whether resveratrol has an additional effect on ethanol-induced inhibition of platelet aggregation. Ca2+ entry and subsequent aggregation of human platelets were measured by the fluorescence method and light transmittance method, respectively. Thromboxane B2 concentrations in media containing platelets were measured by using the enzyme-linked immunosorbent assay. Platelet aggregation induced by thrombin (0.025 U/ml) was significantly inhibited by preincubation of platelets with ethanol (0.5%). Preincubation with resveratrol (3.125 µM), which did not affect thrombin-induced platelet aggregation, significantly augmented the inhibitory effect of ethanol on platelet aggregation. Similar synergic effects of ethanol and resveratrol were found on aggregatory responses to collagen (2 µg/ml) and arachidonic acid (0.25 mM). On the other hand, the thrombin-induced increase in intracellular Ca2+ concentration ([Ca2+]i) was not affected by ethanol alone, resveratrol alone or both ethanol and resveratrol together. In nominally Ca2+-free medium, arachidonic acid (0.75 mM) caused a potent platelet aggregation, which was not affected by the presence of ethanol alone, resveratrol alone, or both of them together. Thromboxane B2 formation induced by thrombin was significantly inhibited by ethanol (0.5%) alone and resveratrol (3.125 µM) alone, and these inhibitory effects were significantly augmented in the presence of both ethanol and resveratrol together. Resveratrol shows an additive effect on ethanol-induced inhibition of platelet aggregation. This effect by resveratrol is partly explained by its inhibitory action on thromboxane A2 production in platelets. In addition, both ethanol and resveratrol attenuate platelet aggregation through acting on the Ca2+-dependent intra-platelet pathway after an increase in [Ca2+]i induced by thrombin.

Background and aims: We have recently proposed urinary pteridine level as a useful biomarker of oxidative stress in a general population. However, the significance of urinary pteridines in patients with diabetes is unknown. Methods: The relationships of the level of urinary pteridine derivatives with d-dimer, ankle-brachial pressure index (ABI), and known cardiovascular risk factors were investigated in patients with type 2 diabetes. Results: Urinary pteridine level showed significant positive correlations with urinary15-isoprostane F2t, female gender, history of smoking and d-dimer and significant inverse correlations with history of alcohol drinking, body mass index (BMI) and ABI. ABI was significantly lower and d-dimer was significantly higher in the highest tertile group of pteridines than in the lowest tertile group. The odds ratios of the highest vs. lowest tertiles for low ABI and high d-dimer were significantly higher than the reference level. The above relationships of urinary pteridines with ABI and d-dimer were not altered when age, gender, BMI, hemoglobin A1c and history of alcohol drinking were used as explanatory variables in multivariable analyses. History of smoking confounded the relation of pteridines with ABI but not that with d-dimer. However, in logistic regression analysis, the association between pteridines and ABI remained significant with adjustment for history of smoking. Conclusion: Urinary pteridine level was associated with d-dimer and ABI, which reflect blood coagulability and arterial flow to the lower extremities, respectively, and is thus thought to be a useful discriminator of thromboatherosclerotic risk in patients with diabetes.

Leukocyte count in peripheral blood is an acute-phase reactant and is associated with the risk of atherosclerotic diseases. Blood lipid profile, a major risk factor of cardiovascular disease, is known to be associated with leukocyte count, but it remains to be determined how this association is affected by other factors including lifestyle and age. The subjects were 11261 Japanese middle-aged men (30~65 years old) who had received health checkup examinations. The relationships of leukocyte count with lipid-related indices (ratio of LDL cholesterol to HDL cholesterol [LDL-C/HDL-C], ratio of triglycerides to HDL cholesterol [TG/HDL-C] and cardiometabolic index [CMI]) were investigated. Leukocyte count, LDL-C/HDL-C, TG/HDL-C and CMI were significantly higher in smokers than in nonsmokers, while leukocyte count and LDL-C/HDL-C were significantly lower in regular drinkers than in nondrinkers. Both in overall subjects and subjects without habits of smoking and drinking, LDL-C/HDL-C, TG/HDL-C and CMI were significantly higher in the 2nd and 3rd tertiles for leukocyte count than in the 1st tertile and tended to be higher with an increase of the tertile. Odds ratios for high TG/HDL-C and high CMI of the 3rd vs. 1st tertiles for leukocyte count tended to be lower with an increase of age, and odds ratios for high TG/HDL-C and high CMI of the interaction term, consisting of age (60~65 vs. 30~39 years) and tertile (3rd vs. 1st tertiles for leukocyte count), were significantly lower than the reference level. In conclusion, leukocyte count is associated with lipid-related indices, and the associations are independent of smoking and alcohol drinking and tend to be weaker with an increase of age in Japanese middle-aged men.

Platelets are a major source of microRNAs (miRNAs) in blood. Relationships between circulating platelet-derived miRNAs were investigated to elucidate their significance as biomarkers. Total miRNAs in serum were analyzed using the 3D-Gene miRNA Oligo chip. Among 22 miRNAs that are included in platelets and play functional roles, sufficient miRNA levels for comparison were detected for 11 miRNAs (let-7b-5p, miR-16–5p, miR-17–5p, miR-24–3p, miR-107, miR-126–3p, miR-150–3p, miR-191–5p, miR-197–3p, miR-223–3p, and miR-326). Among 55 pairs prepared by these miRNAs, relatively strong correlations (Spearman’s correlation coefficient >0.8) were shown between miRNAs of 7 pairs including let-7b-5p and miR-16-5p, let-7b-5p and miR-17-5p, let-7b-5p and miR-107, miR-16–5p and miR-17-5p, miR-16-5p and miR-107, miR-17-5p and miR-107, and miR-107 and miR-126-3p. In principal component analysis, the first principal component consisted of let-7b-5p, miR-16-5p, miR-17-5p, miR-107, miR-126–3p, and miR-191-5p. These six miRNAs may be useful biomarkers that reflect platelet condition and function.

Background: Hyperuricemia is a risk factor of cardiovascular disease. It remains to be elucidated how blood urate level is associated with hyperglycemia in women. Methods: The participants were 4612 middle-aged Japanese female workers. They were divided into four quartile groups by serum urate level, and cardiovascular risk factors were compared in the quartile groups. Results: With an increase of the quartile for urate, the means of waist-to-height ratio, systolic and diastolic blood pressure, log-transformed triglycerides, low-density lipoprotein (LDL) cholesterol, and cardiometabolic index (CMI) tended to be higher and high-density lipoprotein (HDL) cholesterol tended to be lower. Hemoglobin A1c was significantly higher in the 4th quartile for urate than in the 1st quartile, but this difference was not found when body mass index (BMI) was adjusted. The odds ratios versus the 1st quartile for high waist-to-height ratio, hypertension, hypertriglyceridemia, hypo-HDL cholesterolemia, hyper-LDL cholesterolemia, high CMI, and diabetes tended to be higher with an increase of the quartile. The odds ratios of the 4th versus 1st quartiles for these abnormalities except for high waist-to-height ratio and diabetes were significantly higher than the reference level even with adjustment for BMI. Hemoglobin A1c showed a weak but significant positive correlation with urate in analysis with adjustment for BMI. Conclusion: Blood urate was positively associated with adiposity, blood pressure, triglycerides, LDL cholesterol, and glycemic status and was inversely associated with HDL cholesterol in middle-aged women. The associations of urate with blood pressure, blood lipids, and glycemic status remained independent of adiposity, although being confounded by adiposity.

Background: The prevalence of obesity is higher in Western countries than in East Asian countries. It remains unknown whether microRNAs (miRNAs) are involved in the pathogenesis of the ethnic difference in obesity. The purpose of this study was to determine whether expression levels of circulating obesity-associated miRNAs are different in Europeans and Asians. Methods: The subjects were middle-aged healthy male Austrians (n = 20, mean age of 49.9 years) and Japanese (n = 20, mean age of 48.7 years). Total miRNAs in serum from each subject were analyzed using the 3D-Gene miRNA Oligo chip. miRNAs that showed significant differences between the Austrian and Japanese groups were uploaded into Ingenuity Pathway Analysis (IPA). Results: Among 16 miRNAs that were revealed to be associated with obesity in previous studies and showed expression levels that were high enough for a reasonable comparison, serum levels of 3 miRNAs displayed significant differences between the Austrian and Japanese groups: miR-125b-1-3p was significantly lower with a fold change of −2.94 and miR-20a-5p and miR-486–5p were significantly higher with fold changes of 1.73 and 2.38, respectively, in Austrians than in Japanese. In IPA including all 392 miRNAs that showed significant differences between Austrians and Japanese, three canonical pathways including leptin signaling in obesity, adipogenesis pathway and white adipose tissue browning pathway were identified as enriched pathways. Conclusions: miRNAs are thought to be involved in the ethnic difference in the prevalence of obesity, which may in part be caused by different expression levels of miR-125b-1-3p, miR-20a-5p and miR-486–5p.

Abstract Background Urinary liver-type fatty acid-binding protein (L-FABP) is a well-known marker of proximal tubular impairment. We evaluated the relationship between cardiovascular disease (CVD) risk factors and levels of L-FABP in a cross-sectional community-based study. Participants with normoalbuminuria and normal estimated glomerular filtration rate (eGFR), that is, non-chronic kidney disease (non-CKD), were enrolled in this study. To the best of our knowledge, this is the first study to focus on the association between CVD risk factors and a proximal tubular marker in the Japanese general population with normoalbuminuria and normal eGFR. Methods The present study is part of the Sasayama study. The participants included 1000 community residents (447 men and 553 women) aged 40–64 years without a history of CVD or renal dysfunction. Out of these participants 375 men and 477 women, defined as non-CKD, were included for further analysis. In each sex, the highest quintile group was considered to have high-normal L-FABP levels. A multiple logistic regression model was used to evaluate the relationship between risk factors for CVD and high-normal L-FABP levels in the non-CKD participants. We performed a similar analysis using the high-normal urinary albumin to creatinine ratio (UACR) as a dependent variable instead of L-FABP. Results Among the non-CKD participants, in the highest quintile group (Q5, top 20%), L-FABP was ≥2.17 μg/gCre in men and ≥ 2.83 μg/gCre in women. In women, the multivariate odds ratio was 3.62 (1.45–9.00) for high-normal L-FABP in the presence of diabetes mellitus (DM) compared with that in the group without DM. However, the relationship between DM and the UACR level was not significant. In men, DM was significantly associated with high-normal UACR. However, the relationship with L-FABP levels was not significant. Conclusions The presence of DM was more strongly related to high-normal L-FABP levels than to high-normal UACR in women even at the stage of normoalbuminuria and normal eGFR. Our results were also consistent with the findings of a previous study where women were more prone to nonalbuminuric renal impairment compared to men, although further studies are required to confirm the results.

Background: Lipid-related indices are useful for early detection of the risk of cardiovascular disease. The relationships of smoking and alcohol drinking with lipid-related indices in women with diabetes remain to be clarified. Methods: In female participants with hyperglycemia, four lipid-related indices, ratio of low-density lipoprotein cholesterol to high-density lipoprotein cholesterol (LDL-C/HDL-C ratio), atherogenic index of plasma (AIP), lipid accumulation product (LAP), and cardiometabolic index (CMI), were compared in smokers and nonsmokers and in occasional drinkers, regular drinkers, and nondrinkers. Analysis of covariance and logistic regression analysis were used for comparison with adjustment for age, hemoglobin A1c, history of regular exercise, and history of alcohol drinking or smoking. Results: Mean levels of LDL-C/HDL-C ratio, AIP, and CMI were significantly higher in smokers than in nonsmokers, and the odds ratios in smokers versus nonsmokers for high LDL-C/HDL-C ratio, high AIP, and high CMI were significantly higher than the reference level. These differences in mean levels and odds ratios were not found in analysis of LAP. Mean levels of LDL-C/HDL-C ratio, LAP, AIP, and CMI were significantly lower in regular drinkers than in nondrinkers. The odds ratios versus nondrinkers for high LDL-C/HDL-C ratio, high AIP, high LAP, and high CMI in regular drinkers were significantly lower than the reference level. The odds ratios versus nondrinkers for high LDL-C/HDL-C ratio, high LAP, and high CMI in occasional drinkers were also significantly lower than the reference level. Conclusions: In women with hyperglycemia, smoking was positively associated with LDL-C/HDL-C ratio, AIP, and CMI, and habitual alcohol drinking was inversely associated with LDL-C/HDL-C ratio, AIP, LAP, and CMI. Thus, LDL-C/HDL-C ratio, AIP, and CMI are thought to be affected by both smoking and alcohol drinking, which accelerates and suppresses atherosclerotic progression, respectively.

Abstract Background Resveratrol has been shown to inhibit platelet aggregation. However, the mechanism for this action of resveratrol remains to be clarified. The purpose of this study was to elucidate the Ca2+-related mechanism for the inhibitory action of resveratrol on platelet aggregation. Methods Ca2+ entry and subsequent aggregation of human platelets induced by different stimulants including thrombin, thapsigargin, and 1-oleoyl-2-acetylglycerol (OAG) were measured by the fluorescence method and light transmittance method, respectively. Each stimulant was added to a nominally Ca2+-free medium containing platelets, and then CaCl2 was added to the medium to induce Ca2+ influx into platelets. Results Thapsigargin-induced Ca2+ entry into platelets and subsequent platelet aggregation were significantly inhibited in the presence of resveratrol at 6.25 μM or higher concentrations, while OAG-induced Ca2+ entry and subsequent platelet aggregation were not affected by resveratrol at concentrations up to 50 μM. In the nominally Ca2+-free medium, thrombin induced a small transient increase in intracellular Ca2+ concentrations, which was attenuated in the presence of resveratrol at 12.5 μM or higher concentrations. Thrombin-induced Ca2+ entry into platelets and subsequent platelet aggregation were significantly inhibited in the presence of resveratrol at 12.5 μM or higher concentrations. Conclusions The results suggest that resveratrol inhibits thrombin-induced platelet aggregation through decreasing Ca2+ release from its stores and inhibiting store-operated Ca2+ influx into platelets.

Abstract Background Results of previous studies regarding the relationship between blood urate level and diabetes are conflicting. The aim of this study was to clarify the relations of urate with glycemic status and metabolic syndrome in a general population and in patients with diabetes. Methods The participants were 12,528 men receiving health-checkup examinations (29–70 years old) and they were divided into four quartile groups for serum urate levels. Relationships of urate with metabolic syndrome and its components were investigated after adjustment for age and lifestyle factors such as smoking, alcohol drinking and regular exercise. Results In the overall participants and the participants with diabetes (n = 802), hemoglobin A1c levels were significantly lower in the 2nd, 3rd and 4th quartiles for urate than in the 1st quartile and the levels tended to be lower with an increase in the quartile. In the overall participants, the adjusted odds ratios for diabetes vs. the 1st quartile of urate were significantly lower in the 2nd quartile (0.71 [0.59–0.87])), 3rd quartile (0.51 [0.41–0.63]) and 4th quartile [0.35 (0.28–0.44)) than the reference level and the odds ratio tended to be lower with an increase in the quartile. A high urate level was positively associated with visceral obesity, hypertension, dyslipidemia (high triglycerides and/or low HDL cholesterol) and metabolic syndrome, and these associations were less prominent in the diabetes group than in the overall participants. Conclusions Blood urate level is inversely associated with glycemic status in both a general population and patients with diabetes. Positive associations of urate with metabolic syndrome and its components except for diabetes are confounded in the presence of diabetes.

Presented here are the supplemental data of the research article “Urinary pteridines as a discriminator of atherosclerotic risk in patients with diabetes” [1]. These data provide the first information on variables that affect urinary levels of pteridines (oxidized-form pteridine derivatives) in patients with diabetes mellitus. In linear regression analysis, gender (women vs. men), current history of smoking and urinary albumin showed significant positive correlations with pteridines, while there were significant inverse correlations of pteridines with a history of alcohol drinking and body mass index. The above associations were confirmed by using analysis of covariance and logistic regression analysis. Among the eight variables (age, gender, medication therapy for diabetes, smoking, alcohol drinking, body mass index, hemoglobin A1c and urinary albumin) tested, smoking showed the strongest association with urinary pteridines.

Abstract Background The ratio of triglycerides to HDL cholesterol (TG/HDL-C ratio) is known as a good predictor for cardiovascular disease. The purpose of this study was to compare discrimination for cardiovascular risk by different cut-off values of the TG/HDL-C ratio. Methods Receiver operating characteristic (ROC) analysis was performed for the relationship between TG/HDL-C ratio and accumulation of cardio-metabolic risk factors including visceral obesity, hypertension and diabetes. Logistic regression analysis was performed for the relationships of TG/HDL-C ratio with cardio-metabolic risk factors using the cut-off values obtained by ROC analysis and conventional cut-off values (men, 3.75; women, 3.00). Results In ROC analysis, the optimal cut-off values for TG/HDL-C ratio were 2.967 in men and 2.237 in women, which were much smaller than the conventional cut-of values. Odds ratios for multiple cardio-metabolic risk factors of subjects with vs. subjects without a high TG/HDL-C ratio in men and women were 5.75 (4.43–7.46) and 18.76 (10.32–34.13), respectively, by using the new cut-off values and they were 5.03 (3.96–6.39) and 16.11 (9.20–28.20), respectively, by using the conventional cut-off values. The odds ratios for visceral obesity, hypertension and diabetes were comparable when using these two different cut-off values. Conclusion Cut-off values should be ideally calculated by ROC analysis. However, the discrimination power of cut-off values for the TG/HDL-C ratio calculated by ROC analysis for cardio-metabolic risk was similar to those by using the conventional cut-off values. Further studies using cardiovascular events as outcomes in the analysis may be needed to determine more suitable cut-off values of the TG/HDL-C ratio.

Abstract Background Smoking is a major risk factor for dyslipidemia. However, it remains to be clarified whether light smoking in Asian women affects lipid profiles and lipid-related indices. The aim of this study was to determine the relationships between lipid-related indices and smoking in Japanese women. Alcohol drinking influences blood lipid levels and is a potent confounder for the relationship between smoking and blood lipids. Thus, analysis for the relationships between smoking and blood lipid-related indices was also performed after stratification of drinking status. Methods The participants were 18,793 Japanese women aged 35–70 years. A cross-sectional study was performed using a local population-based database. The relationships of smoking with each index were investigated by using analysis of covariance and logistic regression analysis with adjustment for age and other lifestyle factors such as alcohol drinking and regular exercise. Results In multivariate logistic regression analysis, odds ratios of smokers vs. nonsmokers for high ratio of LDL cholesterol to HDL cholesterol (LDL-C/HDL-C), high ratio of triglycerides to HDL cholesterol (TG/HDL-C), high lipid accumulation product (LAP) and high cardio metabolic index (CMI) were significantly higher than the reference level of 1.00 in overall participants (2.17 [1.78–2.66], 1.70 [1.47–1.97], 1.17 [1.08–1.27] and 1.41 [1.30–1.53], respectively), nondrinking participants (2.29 [1.80–2.91], 1.68 [1.39–2.02], 1.21 [1.08–1.36] and 1.46 [1.30–1.63], respectively), and drinking participants (1.96 [1.35–2.85], 1.76 [1.39–2.21], 1.13 [1.01–1.27] and 1.38 [1.22–1.55], respectively). In overall participants, nondrinking participants, and drinking participants, LDL-C/HDL-C, TG/HDL-C, LAP and CMI were significantly higher in smokers than in nonsmokers. In nondrinking participants, triglycerides and LDL cholesterol were significantly higher in smokers than in nonsmokers, while the ratio of waist circumference to height and HDL cholesterol were significantly lower in smokers than in nonsmokers. Conclusion In women, all of the four lipid-related indices tested were higher in smokers than in nonsmokers, and these associations were independent of alcohol drinking. The high levels of the lipid-related indices in smokers result from the detrimental effects of smoking on levels of blood lipids such as triglycerides, HDL cholesterol and LDL cholesterol.

Alcohol inhibits platelet function, and platelet count is often reduced in individuals with alcohol use disorder. However, the relation of habitual alcohol drinking with platelet count in a general population remains to be determined. The participants were 6508 men (30 ~ 69 years old) who had received annual health checkup examinations, and most of them (98.6%) were nondrinkers or drinkers with an average ethanol intake of less than 66 g per day. Relationships of platelet count with alcohol intake were investigated by using analysis of covariance and multivariate linear and logistic regression analyses. Platelet count was significantly correlated with age, smoking, γ-glutamyl transpeptidase, body mass index and leukocyte count, which were thus used as explanatory variables in the multivariate analyses. Mean platelet counts in light (

Summary: Background: Cardio-ankle vascular index (CAVI) and ankle-brachial systolic pressure index (ABI) are used as indicators of atherosclerotic progression, which is more prominent in the elderly than in the young. The relationships of age and gender with these indicators were investigated in patients with diabetes mellitus. Methods: Subjects were outpatients with type 2 diabetes (113 men and 53 women). CAVI and ABI were simultaneously measured at rest. ABI was also measured after leg exercise. Patients with ABI higher than 1.3 were excluded from the subjects. Results: CAVI and ABI were significantly higher in men than in women, while % decrease in ABI after exercise was not significantly different in men and women. Both in men and women, CAVI was not significantly correlated with ABI and % decrease in ABI. Both in men and women, there were significant correlations between age and CAVI (men, r = 0.497 [p

Abstract Background Although ethanol is known to inhibit platelet aggregation, the effects of another variant of alcohol, methanol, have not been reported. The purpose of this study was to determine whether methanol and its metabolite, formic acid, affect Ca2+ entry into and subsequent aggregation of platelets in vitro. Methods Ca2+ entry into and aggregation of human platelets were measured by spectrofluorometry using Fura-2/AM as an indicator and the light transmission method, respectively. Results Thrombin-induced platelet aggregation was significantly augmented by methanol at pharmacological concentrations (0.5–2%) in a concentration-dependent manner. Methanol at 2% significantly attenuated thapsigargin-induced platelet aggregation, which was not significantly affected by lower concentrations (0.5 and 1%) of methanol. Methanol (0.5–2%) did not significantly affect platelet aggregation induced by 1-oleoyl-2-acetyl-sn-glycerol (OAG), or Ca2+ entry into platelets induced by thrombin, thapsigargin, or OAG. Platelet aggregation induced by thrombin, thapsigargin, or OAG was significantly inhibited by formic acid at toxic concentrations (0.01% or higher). Ca2+ entry into platelets induced by thrombin or thapsigargin was also significantly inhibited by formic acid at 0.01% or higher, while that induced by OAG was not affected by formic acid at 0.005 and 0.01% and augmented by that at 0.02%. Conclusions Methanol at pharmacological doses has diverse effects on platelet aggregation, depending on the aggregation stimuli, without affecting Ca2+ entry into platelets. Formic acid at toxic concentrations has an inhibitory action on platelets aggregation, which was partly explained by the reduction of Ca2+ entry into platelets.

A better understanding of pathogenic mechanisms is required in order to treat diseases. However, the mechanisms of diabetes mellitus and diabetic complications are extremely complex. Immune reactions are involved in the pathogenesis of diabetes and its complications, while diabetes influences immune reactions. Furthermore, both diabetes and immune reactions are influenced by genetic and environmental factors. To address these issues, animal models are useful tools. So far, various animal models of diabetes have been developed in rats, which have advantages over mice models in terms of the larger volume of tissue samples and the variety of type 2 diabetes models. In this review, we introduce rat models of diabetes and summarize the immune reactions in diabetic rat models. Finally, we speculate on the relationship between immune reactions and diabetic episodes. For example, diabetes-prone Biobreeding rats, type 1 diabetes model rats, exhibit increased autoreactive cellular and inflammatory immune reactions, while Goto-Kakizaki rats, type 2 diabetes model rats, exhibit increased Th2 reactions and attenuation of phagocytic activity. Investigation of immunological abnormalities in various diabetic rat models is useful for elucidating complicated mechanisms in the pathophysiology of diabetes. Studying immunological alterations, such as predominance of Th1/17 or Th2 cells, humoral immunity, and innate immune reactions, may improve understanding the structure of amplification circuits for diabetes in future studies.

Localization of cyclooxygenase-2 (COX-2) in rat aortae was investigated. In endothelium-intact aortic strips stimulated with interleukin-1 β (IL-1 β ), COX-2 was detected in adventitial fibroblasts and macrophages but not in tunica media. IL-1 β−induced COX-2 expression was also detected in medial smooth muscle cells of endothelium-denuded aortae and cultured rat aortic smooth muscle cells. IL-1β−induced COX-2 expression in all of the above cells was abolished in the presence of U0126, an inhibitor of extracellular signal-regulated kinases (ERK). Thus, IL-1β induces COX-2 through the ERK-mediated pathway in adventitial fibroblasts and macrophages of healthy aortae; and in addition, medial smooth muscle cells are also responsible for COX-2 expression in remodeled aortae. Keywords:: cyclooxygenase-2, remodeling, vascular smooth muscle

The aim of this study was to determine whether the pathway of inducible NO synthase (iNOS) in blood vessels is changed by type 2 diabetes. Lipopolysaccharide (LPS)-induced nitric oxide (NO) production and expression of iNOS and effects of LPS on phenylephrine-induced contractile force were compared in aortae isolated from Goto-Kakizaki (G-K) diabetes rats and aortae isolated from control Wistar rats. Both LPS-stimulated nitrite generation and iNOS expression levels were significantly higher in aortae from G-K rats than in those from control rats. Phenylephrine-induced contractile force in the presence of LPS was significantly lower in aortae from G-K rats than in those from control rats, while contractile force in the absence of LPS was comparable in the diabetic and control groups. On the other hand, incubation of aortae in high glucose-containing medium did not affect the LPS-stimulated nitrite accumulation and iNOS expression and the phenylephrine-induced contractile force, regardless of the presence of LPS. These results suggest that LPS-induced NO production through the iNOS pathway is increased and subsequent attenuation of contractile force by excess NO is enhanced in arteries of rats with type 2 diabetes. Keywords:: artery, glucose, lipopolysaccharide, nitric oxide synthase, type 2 diabetes

The purpose of this study was to explore the peptides that are related to acute reduction of blood pressure after alcohol drinking. Venous blood was collected from male healthy volunteers before and after drinking white wine (3 ml/kg weight) containing 13% of ethanol. Peptidome analysis for serum samples was performed using a new target plate, BLOTCHIP®. Alcohol caused significant decreases in systolic and diastolic blood pressure levels at 45 min. The peptidome analysis showed that the levels of three peptides of m/z 1467, 2380 and 2662 changed significantly after drinking. The m/z 1467 and 2662 peptides were identified to be fragments of fibrinogen alpha chain, and the m/z 2380 peptide was identified to be a fragment of complement C4. The intensities of the m/z 2380 and m/z 1467 peptides before drinking were associated with % decreases in systolic and diastolic blood pressure levels at 45 min after drinking compared with the levels before drinking, while there were no significant correlations between the intensity of the m/z 2662 peptide and % decreases in systolic and diastolic blood pressure levels after drinking. The m/z 1467 and 2380 peptides are suggested to be markers for acute reduction of blood pressure after drinking alcohol.

The purpose of this study was to investigate the effects of daunorubicin on lipopolysaccharide (LPS)-stimulated inducible-type nitric oxide synthase (iNOS) expression in macrophages. LPS-stimulated iNOS expression and NO production were significantly inhibited in alveolar macrophages from rats administrated daunorubicin (4 mg/kg body weight per day) for 5 consecutive days. Incubation of macrophages with daunorubicin at 1 µM but not at 0.1 and 0.5 µM significantly inhibited LPS-stimulated NO production and iNOS induction. Activation of extracellular regulated kinase (ERK) by LPS was markedly attenuated in both macrophages isolated from in vivo daunorubicin-treated rats and those incubated in vitro with daunorubicin at 1 µM. ERK activation, iNOS induction, and NO production following LPS stimulation were all markedly inhibited in the presence of U0126, an ERK inhibitor. The viability of macrophages was decreased by incubation with daunorubicin at 0.5 and 1 µM, while treatment of rats with daunorubicin did not affect viability of macrophages isolated from the rats. These results suggest that in vivo treatment of rats with daunorubicin attenuates LPS-induced iNOS expression of macrophages through inhibition of ERK activation, while inhibition of iNOS induction by in vitro incubation with daunorubicin may be mainly due to its cytotoxicity. Keywords:: daunorubicin, anthracycline antibiotic, nitric oxide, extracellular regulated kinase, macrophage

Urotensin II induced sustained contraction with an EC50 value of 2.29 ± 0.12 nM in rat aorta. Urotensin II (100 nM) transiently increased cytosolic Ca2+ level ([Ca2+]i), followed by a small sustained phase superimposed with rhythmic oscillatory change. In the presence of verapamil and La3+, the [Ca2+]i oscillation was completely inhibited, although a small transient increase in [Ca2+]i remained. The urotensin II-induced contraction was also partially inhibited by verapamil and La3+. Combined application of verapamil, La3+, and thapsigargin completely inhibited the increase in [Ca2+]i with only partial inhibition of the contraction elicited by urotensin II. Urotensin II increased myosin light chain (MLC) phosphorylation to a level greater than that induced by 72.7 mM KCl (high K+). Pretreatment with Go6983 (PKC inhibitor), U0126 (MEK inhibitor), or SB203580 (p38MARK inhibitor) partially inhibited the urotensin II-induced contraction with no effects on the high K+-induced contractions. Wortmannin (MLC kinase inhibitor) only partially inhibited urotensin II-induced contraction, although it completely inhibited the high K+-induced contraction. These results suggest that urotensin II-induced contraction is mediated by the Ca2+/calmodulin/MLC kinase system and modulated by the Ca2+ sensitization mechanisms to increase MLC phosphorylation. In addition, activations of PKC, p38MAPK, and ERK1/2 modulate the contractility mediated by urotensin II in rat aorta. Keywords:: urotensin II, rat aorta, Ca2+, myosin phosphorylation, Ca2+ sensitization