Naoyuki Yao

Roles of calcineurin and calcium/calmodulin-dependent protein kinase II in pressure overload-induced cardiac hypertrophy

Calcineurin and calcium/calmodulin-dependent protein kinase (CaMK) II have been suggested to be the signaling molecules in cardiac hypertrophy. It was not known, however, whether these mechanisms are involved in cardiac hypertrophy induced by pressure overload without the influences of blood-derived humoral factors, such as angiotensin II. To elucidate the roles of calcineurin and CaMK II in this situation, we examined the effects of calcineurin and CaMK II inhibitors on pressure overload-induced expression of c-fos, an immediate-early gene, and protein synthesis using heart perfusion model. The hearts isolated from Sprague-Dawley rats were perfused according to the Langendorff technique, and then subjected to the acute pressure overload by raising the perfusion pressure. The activation of calcineurin was evaluated by its complex formation with calmodulin and by its R-II phosphopeptide dephosphorylation. CaMK II activation was evaluated by its autophosphorylation. Expression of c-fos mRNA and rates of protein synthesis were measured by northern blot analysis and by 14C-phenylalanine incorporation, respectively. Acute pressure overload significantly increased calcineurin activity, CaMK II activity, c-fos expression and protein synthesis. Cyclosporin A and FK506, the calcineurin inhibitors, significantly inhibited the increases in both c-fos expression and protein synthesis. KN62, a CaMK II inhibitor, also significantly prevented the increase in protein synthesis, whereas it failed to affect the expression of c-fos. These results suggest that both calcineurin and CaMK II pathways are critical in the pressure overload-induced acceleration of protein synthesis, and that transcription of c-fos gene is regulated by calcineurin pathway but not by CaMK II pathway.

Potential impact of renin-angiotensin system inhibitors and calcium channel blockers on plasma high-molecular-weight adiponectin levels in hemodialysis patients.

Although metabolic syndrome confers an increased risk of cardiovascular disease in the general population, little is known about the alteration of abdominal adiposity and its association with adipocytokines in hemodialysis patients. We investigated the plasma high-molecular-weight (HMW) adiponectin level and its relationship to visceral fat area (VFA) and various markers of atherosclerosis in hemodialysis patients. In a cross-sectional study, conventional cardiovascular risk factors, plasma total and HMW adiponectin, the number of components of the metabolic syndrome and, using computed tomography, the distribution of abdominal adiposity were assessed in 144 hemodialysis patients (90 men and 54 women; mean age, 60.7 years) and 30 age- and sex-matched patients with chronic kidney disease (CKD). Plasma HMW adiponectin levels in hemodialysis patients were significantly higher than those in patients with CKD, negatively associated with VFA and serum triglycerides and positively associated with plasma total adiponectin, as well as the HMW-to-total adiponectin ratio in men and women (all P<0.05) in a simple regression analysis. In a multiple regression analysis, VFA was a significant determinant of HMW adiponectin in hemodialysis patients. Furthermore, after adjustment for classical risk factors, HMW adiponectin levels were significantly higher in patients undergoing treatment with renin–angiotensin system inhibitors or calcium channel blockers compared with patients not undergoing such treatment. This study shows that plasma HMW adiponectin levels were negatively associated with VFA and positively associated with treatment with blockade of the renin–angiotensin system and of the calcium channel. Therefore, these drugs might be effective for improving adipocytokine-related metabolic abnormalities in hemodialysis patients.

Impact of metabolic disturbances and malnutrition-inflammation on 6-year mortality in Japanese patients undergoing hemodialysis.

Metabolic syndrome confers an increased risk of cardiovascular disease (CVD) in the general population. The relationship between adiponectins, and clinical outcomes in patients undergoing hemodialysis remains controversial. We investigated whether adiponectins, biomarkers of inflammation, nutrition status and clinical features predict the mortality of patients undergoing hemodialysis for 6 years. We measured baseline plasma total and high‐molecular‐weight (HMW) adiponectins, tumor necrosis factor (TNF)‐α, serum high sensitivity C‐reactive protein (hsCRP), and clinical characteristics including visceral fat area (VFA) and the Geriatric Nutritional Risk Index (GNRI) in 133 patients undergoing chronic hemodialysis. Forty‐one of the 133 patients died during follow‐up. The deceased patients were significantly older, had more prior CVD and diabetes, higher TNF‐α and hsCRP levels but lower GNRI. VFA, and total and HMW adiponectin did not significantly differ between the two groups. TNF‐α and hsCRP levels and GNRI score were significant for predicting all‐cause and cardiovascular mortality in receiver operating curve analyses. When stratified by a GNRI score of 96, Cox proportional hazards analyses identified TNF‐α as a significant predictor of all‐cause mortality (hazard ratio [HR] 1.23; P = 0.038) and hsCRP as a significant predictor of all‐cause and cardiovascular mortality (HR, 2.32, P = 0.003; HR 2.30, P = 0.012, respectively) after adjusting for age, sex, diabetes mellitus, and prior CVD, only in malnourished patients. These results demonstrate that malnutrition and the inflammatory markers TNF‐α and hsCRP, but not metabolic markers, including VFA and adiponectins have a significant impact on 6‐year all‐cause and cardiovascular mortality in Japanese patients undergoing hemodialysis.

The induction of human CL-P1 expression in hypoxia/reoxygenation culture condition and rat CL-P1 after ischemic/reperfusion treatment

BACKGROUND Oxidative stress-induced endothelial dysfunction and oxidized low-density lipoprotein (LDL) might play a key role in the pathogenesis of atherosclerosis. We recently identified a vascular endothelial scavenger receptor, collectin placenta 1 (CL-P1), which acts as a receptor for oxidized LDL as well as for microbes. METHODS We demonstrate how hypoxic and oxidative stress induced CL-P1 expression and compared their effects with the expression of lectin-like oxidized low-density lipoprotein receptor 1 (LOX-1), an endothelial scavenger receptor expressed by oxidative stress. RESULTS Hypoxia/reoxygenation induced CL-P1 mRNA and protein expression in human umbilical vein endothelial cells (HUVECs). The expression of LOX-1 mRNA in these cells peaked slightly at 24 h, while the expression of CL-P1 had an onset at 72 h and was sustained for 120 h after reoxygenation. Furthermore, the exposure of rat carotid artery endothelium to ischemia/reperfusion increased the maximal CL-P1 mRNA expression at 72 h and expression of its protein peaked at 7 days after this treatment. We demonstrate that CL-P1 up-regulation is induced in vitro and in vivo by oxidative stress. GENERAL SIGNIFICANCE The inducible expression of CL-P1 by oxidative stress might play a crucial role in endothelial dysfunction or chronic activation leading to the pathogenesis of atherosclerosis.

Management of calcium, phosphorus and bone metabolism in dialysis patients using sevelamer hydrochloride and vitamin D therapy.

Abstract:  Abnormalities of mineral metabolism, including those of calcium (Ca), phosphorus (P), and parathyroid hormone (PTH) in patients on maintenance hemodialysis induce severe bone involvement, which manifests as renal osteodystrophy. Recently, vascular calcification caused by abnormal mineral metabolism has been attracting attention because cardiovascular diseases (CVD) are a major cause of death in hemodialysis patients. Since 2000, the treatment standard for overt secondary hyperparathyroidism (SHPT) in our facilities has shifted from conventional or pulse therapy with oral vitamin D3 (VitD) to intravenous pulse therapy with maxacalcitriol or calcitriol. After selecting the criterion of overt SHPT as intact‐PTH > 500 pg/mL, the proportion of overt SHPT cases among all hemodialysis patients decreased from 12% at the start of intravenous pulse treatment to 6.4% after 4 years’ treatment. However, the number of patients who had an interruption to pulse treatment because of hypercalcemia and/or hyperphosphatemia was high and it became a bottleneck for the continuation of the therapy. The major cause of hypercalcemia is considered to be Ca load derived from oral calcium carbonate. In Japan, sevelamer hydrochrolide (SH), which does not contain Ca, has been available commercially since 2003 and potentially should enable a reduction in the incidence of overt SHPT during long‐term intravenous treatment when combined with careful adjustment of the dose of VitD and strict monitoring of Ca and P level concentrations. In this study, we found that the proportion of patients who satisfy the recommended serum concentrations of Ca and P reported by K/DOQI guideline was low irrespective of the serum concentration of intact‐PTH. The aortic calcification index was high in the patient group with lower intact‐PTH level concentration, probably because of reduced Ca and P buffering ability associated with reduced bone turnover. We consider that VitD treatment with SH might give better control of the intact‐PTH level concentration within the range recommended by the K/DOQI guideline.