In some patients with type 2 diabetes mellitus (DM) without hypertension, cardiac hypertrophy and attenuated cardiac function are observed, and this insult is termed diabetic cardiomyopathy. To date, ...microRNA (miRNAs or miR) functions in diabetic cardiomyopathy remain to be elucidated.
To clarify the functions of miRNAs involved in diabetic cardiomyopathy caused by type 2 DM.
C57BL/6 mice were fed a high-fat diet (HFD) for 20 weeks, which induced obesity and type 2 DM. miRNA microarray analyses and real-time polymerase chain reaction revealed that miR-451 levels were significantly increased in the type 2 DM mouse hearts. Because excess supply of saturated fatty acids is a cause of diabetic cardiomyopathy, we stimulated neonatal rat cardiac myocytes with palmitic acid and confirmed that miR-451 expression was increased in a dose- and time-dependent manner. Loss of miR-451 function ameliorated palmitate-induced lipotoxicity in neonatal rat cardiac myocytes. Calcium-binding protein 39 (Cab39) is a scaffold protein of liver kinase B1 (LKB1), an upstream kinase of AMP-activated protein kinase (AMPK). Cab39 was a direct target of miR-451 in neonatal rat cardiac myocytes and Cab39 overexpression rescued the lipotoxicity. To clarify miR-451 functions in vivo, we generated cardiomyocyte-specific miR-451 knockout mice. HFD-induced cardiac hypertrophy and contractile reserves were ameliorated in cardiomyocyte-specific miR-451 knockout mice compared with control mice. Protein levels of Cab39 and phosphorylated AMPK were increased and phosphorylated mammalian target of rapamycin (mTOR) was reduced in cardiomyocyte-specific miR-451 knockout mouse hearts compared with control mouse hearts.
Our results demonstrate that miR-451 is involved in diabetic cardiomyopathy through suppression of the LKB1/AMPK pathway.
Nardilysin (NRDC) has been shown to be involved in post-translational histone modifications, in addition to enhancement in ectodomain shedding of membrane-anchored protein, which play significant ...roles in various pathophysiology, including glucose homeostasis, inflammatory diseases and cancer. The present study sought to determine roles of NRDC in the liver on lipid and lipoprotein metabolism. We established liver-specific NRDC deficient mice by use of NRD1 floxed mice and albumin promoter-Cre recombinase (Cre) transgenic mice, and found that their serum low-density lipoprotein (LDL) cholesterol levels were significantly lower than those in control littermate mice. In the liver, LDL receptor (LDLR) mRNA expression was significantly upregulated, while inducible degrader of LDLR (IDOL) and microsomal triglyceride transfer protein (MTP) mRNA expression was significantly downregulated, in liver-specific NRDC deficient mice. Hepatic cell-surface LDLR expression levels were significantly elevated and serum pro-protein convertase subtilisin–kexin type 9 (PCSK9) levels were significantly reduced in mice with hepatic NRDC deficiency. In cultured hepatocytes, NRDC deficiency significantly reduced secreted PCSK9 and increased cell-surface LDLR expression. On the other hand, NRDC overexpression in cultured hepatocytes significantly increased secreted PCSK9 and lowered cell-surface LDLR expression. Thus, NRDC in murine hepatocytes appears to play key roles in cholesterol homeostasis, although the precise molecular mechanisms remain to be determined.
Sterol regulatory element-binding protein 2 (SREBP-2) transcription factor has been identified as a key protein in cholesterol metabolism through the transactivation of the LDL receptor and ...cholesterol biosynthesis genes. Here, we generated mice lacking microRNA (miR)-33, encoded by an intron of the Srebp2, and showed that miR-33 repressed the expression of ATP-binding cassette transporter A 1 (ABCA 1) protein, a key regulator of HDL synthesis by mediating cholesterol efflux from cells to apolipoprotein A (apoA)-I. In fact, peritoneal macrophages derived from miR-33—deficient mice showed a marked increase in ABCA 1 levels and higher apoA-I—dependent cholesterol efflux than those from WT mice. ABCA 1 protein levels in liver were also higher in miR-33—deficient mice than in WT mice. Moreover, miR-33—deficient mice had significantly higher serum HDL cholesterol levels than WT mice. These data establish a critical role for miR-33 in the regulation of ABCA 1 expression and HDL biogenesis in vivo.
Heart failure and atherosclerosis share the underlying mechanisms of chronic inflammation followed by fibrosis. A highly conserved microRNA (miR), miR-33, is considered as a potential therapeutic ...target for atherosclerosis because it regulates lipid metabolism and inflammation. However, the role of miR-33 in heart failure remains to be elucidated.
To clarify the role of miR-33 involved in heart failure.
We first investigated the expression levels of miR-33a/b in human cardiac tissue samples with dilated cardiomyopathy. Increased expression of miR-33a was associated with improving hemodynamic parameters. To clarify the role of miR-33 in remodeling hearts, we investigated the responses to pressure overload by transverse aortic constriction in miR-33-deficient (knockout KO) mice. When mice were subjected to transverse aortic constriction, miR-33 expression levels were significantly upregulated in wild-type left ventricles. There was no difference in hypertrophic responses between wild-type and miR-33KO hearts, whereas cardiac fibrosis was ameliorated in miR-33KO hearts compared with wild-type hearts. Despite the ameliorated cardiac fibrosis, miR-33KO mice showed impaired systolic function after transverse aortic constriction. We also found that cardiac fibroblasts were mainly responsible for miR-33 expression in the heart. Deficiency of miR-33 impaired cardiac fibroblast proliferation, which was considered to be caused by altered lipid raft cholesterol content. Moreover, cardiac fibroblast-specific miR-33-deficient mice also showed decreased cardiac fibrosis induced by transverse aortic constriction as systemic miR-33KO mice.
Our results demonstrate that miR-33 is involved in cardiac remodeling, and it preserves lipid raft cholesterol content in fibroblasts and maintains adaptive fibrotic responses in the remodeling heart.
Aims: Although intensive statin therapy reduced cardiovascular risks, cardiovascular events have not been completely prevented. Probucol is a potent antioxidant and reduces tendon xanthomas in ...familial hypercholesterolemia patients despite reduction of high-density lipoprotein (HDL)-cholesterol (HDL-C). We investigated whether probucol can reduce cardiovascular events on top of conventional lipid-lowering therapy in patients with coronary heart disease (CHD). Methods: PROSPECTIVE is a multicenter, randomized, prospective study that recruited 876 Japanese patients with CHD and dyslipidemia with a low-density lipoprotein (LDL)-cholesterol (LDL-C) level of ≥ 140 mg/dL without medication or those treated with lipid-lowering drugs. Lipid-lowering agents were administered during the study period in the control group (n=438), and probucol 500 mg/day was added to lipid-lowering therapy in the probucol group (n=438). Patients were randomly assigned to two treatment groups by adjusting the LDL-C level and presence of diabetes and hypertension and followed up for more than 3 years. The primary end point was a composite of cerebrovascular and cardiovascular events (cardiovascular disease death including sudden death, nonfatal myocardial infarction, nonfatal stroke, hospitalization for unstable angina, hospitalization for heart failure, or coronary revascularization). The secondary end point was carotid intima–media thickness in a subset of patients. Results: The incidence of the primary end point showed a trend to be lower in the probucol group compared with that in the control group despite reduced HDL-C without serious adverse events. Anti-atherogenic effects of probucol may be attributed to its potent antioxidative function and enhancement of reverse cholesterol transport. Conclusion: Since there was no statistical significance between the probucol and control groups despite a marked reduction of HDL-C, further studies on the clinical outcomes of probucol on top of conventional therapy may be necessary in the future (UMIN000003307).
Background: In the short- to mid-term, cardiomyocytes generated from human-induced pluripotent stem cells (hiPSC-CMs) have been reported to be less mature than those of adult hearts. However, the ...maturation process in a long-term culture remains unknown. Methods and Results: A hiPSC clone generated from a healthy control was differentiated into CMs through embryoid body (EB) formation. The ultrastructural characteristics and gene expressions of spontaneously contracting EBs were analyzed through 1-year of culture after cardiac differentiation was initiated. The 14-day-old EBs contained a low number of myofibrils, which lacked alignment, and immature high-density Z-bands lacking A-, H-, I-, and M-bands. Through the long-term culture up to 180 days, the myofibrils became more tightly packed and formed parallel arrays accompanied by the appearance of mature Z-, A-, H-, and I-bands, but not M-bands. Notably, M-bands were finally detected in 360-day-old EBs. The expression levels of the M-band-specific genes in hiPSC-CMs remained lower in comparison with those in the adult heart. Immunocytochemistry indicated increasing number of MLC2v-positive/MLC2a-negative cells with decreasing number of MLC2v/MLC2a double-positive cells, indicating maturing of ventricular-type CMs. Conclusions: The structural maturation process of hiPSC-CMs through 1-year of culture revealed ultrastructural sarcomeric changes accompanied by delayed formation of M-bands. Our study provides new insight into the maturation process of hiPSC-CMs. (Circ J 2013; 77: 1307–1314)
In chronic kidney disease, fibroblast dysfunction causes renal fibrosis and renal anemia. Renal fibrosis is mediated by the accumulation of myofibroblasts, whereas renal anemia is mediated by the ...reduced production of fibroblast-derived erythropoietin, a hormone that stimulates erythropoiesis. Despite their importance in chronic kidney disease, the origin and regulatory mechanism of fibroblasts remain unclear. Here, we have demonstrated that the majority of erythropoietin-producing fibroblasts in the healthy kidney originate from myelin protein zero-Cre (P0-Cre) lineage-labeled extrarenal cells, which enter the embryonic kidney at E13.5. In the diseased kidney, P0-Cre lineage-labeled fibroblasts, but not fibroblasts derived from injured tubular epithelial cells through epithelial-mesenchymal transition, transdifferentiated into myofibroblasts and predominantly contributed to fibrosis, with concomitant loss of erythropoietin production. We further demonstrated that attenuated erythropoietin production in transdifferentiated myofibroblasts was restored by the administration of neuroprotective agents, such as dexamethasone and neurotrophins. Moreover, the in vivo administration of tamoxifen, a selective estrogen receptor modulator, restored attenuated erythropoietin production as well as fibrosis in a mouse model of kidney fibrosis. These findings reveal the pathophysiological roles of P0-Cre lineage-labeled fibroblasts in the kidney and clarify the link between renal fibrosis and renal anemia.
Summary Background Epidemiological and clinical evidence suggests that an increased intake of long-chain n-3 fatty acids protects against mortality from coronary artery disease. We aimed to test the ...hypothesis that long-term use of eicosapentaenoic acid (EPA) is effective for prevention of major coronary events in hypercholesterolaemic patients in Japan who consume a large amount of fish. Methods 18 645 patients with a total cholesterol of 6·5 mmol/L or greater were recruited from local physicians throughout Japan between 1996 and 1999. Patients were randomly assigned to receive either 1800 mg of EPA daily with statin (EPA group; n=9326) or statin only (controls; n=9319) with a 5-year follow-up. The primary endpoint was any major coronary event, including sudden cardiac death, fatal and non-fatal myocardial infarction, and other non-fatal events including unstable angina pectoris, angioplasty, stenting, or coronary artery bypass grafting. Analysis was by intention-to-treat. The study was registered at ClinicalTrials.gov , number NCT00231738. Findings At mean follow-up of 4·6 years, we detected the primary endpoint in 262 (2·8%) patients in the EPA group and 324 (3·5%) in controls—a 19% relative reduction in major coronary events (p=0·011). Post-treatment LDL cholesterol concentrations decreased 25%, from 4·7 mmol/L in both groups. Serum LDL cholesterol was not a significant factor in a reduction of risk for major coronary events. Unstable angina and non-fatal coronary events were also significantly reduced in the EPA group. Sudden cardiac death and coronary death did not differ between groups. In patients with a history of coronary artery disease who were given EPA treatment, major coronary events were reduced by 19% (secondary prevention subgroup: 158 8·7% in the EPA group vs 197 10·7% in the control group; p=0·048). In patients with no history of coronary artery disease, EPA treatment reduced major coronary events by 18%, but this finding was not significant (104 1·4% in the EPA group vs 127 1·7% in the control group; p=0·132). Interpretation EPA is a promising treatment for prevention of major coronary events, and especially non-fatal coronary events, in Japanese hypercholesterolaemic patients.
MicroRNAs (miRs) are small non-protein-coding RNAs that bind to specific mRNAs and inhibit translation or promote mRNA degradation. Recent reports have indicated that miR-33, which is located within ...the intron of sterol regulatory element-binding protein (SREBP) 2, controls cholesterol homoeostasis and may be a potential therapeutic target for the treatment of atherosclerosis. Here we show that deletion of miR-33 results in marked worsening of high-fat diet-induced obesity and liver steatosis. Using miR-33(-/-)Srebf1(+/-) mice, we demonstrate that SREBP-1 is a target of miR-33 and that the mechanisms leading to obesity and liver steatosis in miR-33(-/-) mice involve enhanced expression of SREBP-1. These results elucidate a novel interaction between SREBP-1 and SREBP-2 mediated by miR-33 in vivo.
Aim: The Japan EPA Lipid Intervention Study (JELIS) was the first prospective randomized clinical trial to demonstrate prevention of coronary events by pure eicosapentaenoic acid (EPA). The aim of ...this study was to examine the relationships between various plasma fatty acid concentrations and the risk of coronary events in JELIS participants. Methods: In 15,534 participants, we calculated the hazard ratio for major coronary events (sudden cardiac death, fatal or nonfatal myocardial infarction, unstable angina pectoris, and angioplasty/stenting or coronary artery bypass grafting) relative to the on-treatment average level of plasma fatty acids with the Cox proportional hazard model. Results: As a result of EPA intervention, the plasma EPA concentration increased, but the docosahexaenoic acid (DHA) concentration did not. The other fatty acids measured decreased slightly. The higher plasma level of EPA (hazard ratio=0.83, p=0.049, in all participants and hazard ratio=0.71, p=0.018, in the EPA intervention group), but not of DHA, was inversely associated with the risk of major coronary events. The associations between other fatty acids and the risk of major coronary events were not significant. In all JELIS participants, the risk of major coronary events was significantly decreased (20%) in the group with high (150 µg/mL or more) on-treatment plasma EPA concentration compared with that in the low (less than 87 µg/mL) group. Conclusion: The risk of coronary artery disease is influenced by variations in plasma fatty acid composition. Among n-3 polyunsaturated fatty acids, EPA and DHA exhibited differences in the correlation with the risk of major coronary events.
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