Background-Several clinical studies have demonstrated that levels of adiponectin are significantly reduced in patients with type 2 diabetes and that adiponectin levels are inversely related to the risk of myocardial ischemia. The present study was designed to determine the mechanism by which adiponectin exerts its protective effects against myocardial ischemia/reperfusion. Methods and Results-AdiponectinϪ/Ϫ or wild-type mice were subjected to 30 minutes of myocardial ischemia followed by 3 hours or 24 hours (infarct size and cardiac function) of reperfusion. Myocardial infarct size and apoptosis, production of peroxynitrite, nitric oxide (NO) and superoxide, and inducible NO synthase (iNOS) and gp91 phox protein expression were compared. Myocardial apoptosis and infarct size were markedly enhanced in adiponectin Ϫ/Ϫ mice (PϽ0.01). Formation of NO, superoxide, and their cytotoxic reaction product, peroxynitrite, were all significantly higher in cardiac tissue obtained from adiponectin Ϫ/Ϫ than from wild-type mice (PϽ0.01). Moreover, myocardial ischemia/ reperfusion-induced iNOS and gp91 phox protein expression was further enhanced, but endothelial NOS phosphorylation was reduced in cardiac tissue from adiponectin Ϫ/Ϫ mice. Administration of the globular domain of adiponectin 10 minutes before reperfusion reduced myocardial ischemia/reperfusion-induced iNOS/gp91 phox protein expression, decreased NO/superoxide production, blocked peroxynitrite formation, and reversed proapoptotic and infarctenlargement effects observed in adiponectin Ϫ/Ϫ mice. Conclusion-The present study demonstrates that adiponectin is a natural molecule that protects hearts from ischemia/ reperfusion injury by inhibition of iNOS and nicotinamide adenine dinucleotide phosphate-oxidase protein expression and resultant oxidative/nitrative stress.

SummaryThe key regulator of salicylic acid (SA)-mediated resistance, NPR1, is functionally conserved in diverse plant species, including rice ( Oryza sativa L.). Investigation in depth is needed to provide an understanding of NPR1 -mediated resistance and a practical strategy for the improvement of disease resistance in the model crop rice. The rice genome contains five NPR1 -like genes. In our study, three rice homologous genes, OsNPR1 / NH1 , OsNPR2 / NH2 and OsNPR3 , were found to be induced by rice bacterial blight Xanthomonas oryzae pv.oryzae and rice blast Magnaporthe grisea , and the defence molecules benzothiadiazole, methyl jasmonate and ethylene. We confirmed that OsNPR1 is the rice orthologue by complementing the Arabidopsis npr1 mutant. Over-expression of OsNPR1 conferred disease resistance to bacterial blight, but also enhanced herbivore susceptibility in transgenic plants. The OsNPR1-green fluorescent protein (GFP) fusion protein was localized in the cytoplasm and moved into the nucleus after redox change. Mutations in its conserved cysteine residues led to the constitutive localization of OsNPR1(2CA)-GFP in the nucleus and also abolished herbivore hypersensitivity in transgenic rice. Different subcellular localizations of OsNPR1 antagonistically regulated SA-and jasmonic acid (JA)-responsive genes, but not SA and JA levels, indicating that OsNPR1 might mediate antagonistic cross-talk between the SA-and JA-dependent pathways in rice. This study demonstrates that rice has evolved an SA-mediated systemic acquired resistance similar to that in Arabidopsis, and also provides a practical approach for the improvement of disease resistance without the penalty of decreased herbivore resistance in rice.

Objective Reduced plasma adiponectin (APN) in diabetic patients is associated with endothelial dysfunction. However, APN knockout animals manifest modest systemic dysfunction unless metabolically challenged. The protein family CTRPs (C1q/TNF-related proteins) has recently been identified as APN paralogs and some CTRP members share APN’s metabolic regulatory function. However, the vasoactive properties of CTRPs remain completely unknown. Methods and Results The vasoactivity of currently identified murine CTRP members was assessed in aortic vascular rings and underlying molecular mechanisms was elucidated in HUVECs. Of eight CTRPs, CTRPs 3, 5, and 9 caused significant vasorelaxation. The vasoactive potency of CTRP9 exceeded that of APN (3-fold), and is endothelium-dependent and nitric oxide (NO) mediated. Mechanistically, CTRP9 increased AMPK/Akt/eNOS phosphorylation and increased NO production. AMPK knockdown completely blocked CTRP9-induced Akt/eNOS phosphorylation and NO production. Akt knockdown had no significant effect upon CTRP9-induced AMPK phosphorylation, but blocked eNOS phosphorylation and NO production. Adiponectin receptor 1 (AdipoR1), but not receptor 2, knockdown blocked CTRP9-induced AMPK/Akt/eNOS phosphorylation and NO production. Finally, pre-incubating vascular rings with an AMPK-inhibitor abolished CTRP9-induced vasorelaxive effects. Conclusion We have provided the first evidence that CTRP9 is a novel vasorelaxive adipocytokine which may exert vasculoprotective effects via the AdipoR1/AMPK/eNOS dependent/NO mediated signaling pathway.

Background Obesity/diabetes adversely affects post-ischemic heart remodeling via incompletely understood underlying mechanisms. C1q/TNF-related protein-3 (CTRP3) is a newly identified adipokine exerting beneficial metabolic regulation, similar to adiponectin. The current study determined whether CTRP3 may regulate post-ischemic cardiac remodeling and cardiac dysfunction, and, if so, sought to elucidate the involved underlying mechanisms. Methods and Results Male adult mice were subjected to myocardial infarction (MI) via left anterior descending (LAD) coronary artery occlusion. Both the effect of MI upon endogenous CTRP3 expression/production and the effect of exogenous CTRP3 (adenovirus or recombinant CTRP3) replenishment upon MI injury were investigated. MI significantly inhibited adipocyte CTRP3 expression and reduced plasma CTRP3 level, reaching nadir 3 days post-MI. CTRP3 replenishment improved survival rate (P<0.05), restored cardiac function, attenuated cardiomyocyte apoptosis, increased revascularization, and dramatically reduced interstitial fibrosis (P values all <0.01). CTRP3 replenishment had no significant effect upon cardiac AMP-activated protein kinase (AMPK) phosphorylation, but significantly increased Akt phosphorylation and expression of hypoxia inducing factor-1α (HIF-1α) and vascular endothelial growth factor (VEGF). Surprisingly, treatment of human umbilical vascular endothelial cells (HUVECs) with CTRP3 did not directly affect NO production or tube formation. However, pre-conditioned medium from CTRP3-treated cardiomyocytes significantly enhanced HUVEC tube formation, an effect blocked by either pre-treatment of cardiomyocytes with a PI3K inhibitor, or pre-treatment of HUVECs with a VEGF inhibitor. Finally, pre-conditioned medium from CTRP3-knockdown 3T3 cells significantly enhanced hypoxia-induced cardiomyocyte injury. Conclusions CTRP3 is a novel anti-apoptotic, pro-angiogenic, and cardioprotective adipokine, whose expression is significantly inhibited following MI.

Background CTRP9 is a newly identified adiponectin paralog with established metabolic-regulatory properties. However, the role of CTRP9 in post-myocardial infarction (post-MI) remodeling remains completely unknown. This study determined whether C1q/TNF-related protein-9 (CTRP9) may regulate cardiac remodeling following acute myocardial infarction (AMI), and elucidated the underlying mechanisms. Methods and Results Male adult mice were subject to AMI by left anterior descending coronary artery (LAD) ligation or sham surgery, and treated with saline (vehicle) or globular CTRP9 via peritoneal implant osmotic-pumps for 6 weeks. H9C2 cardiac cell lines were utilized in vitro for determining underlying mechanisms. Adipocyte CTRP9 expression and plasma CTRP9 levels were both significantly reduced after AMI. Compared to vehicle, CTRP9 treatment improved animal survival rate (P<0.05), restored cardiac function (P<0.05), attenuated adverse remodeling (P<0.01), and ameliorated cardiomyocyte apoptosis and fibrosis following AMI (P<0.01). Among multiple anti-remodeling molecules determined, AMP-activated protein kinase (AMPK), protein kinase-A (PKA), and Akt were significantly activated in CTRP9-treated heart. Surprisingly, CTRP9 remains cardioprotective in cardiac-specific AMPK-DN mice. Additional in vitro experiments demonstrated that administration of either PKA inhibitor or PKA-specific siRNA virtually abolished CTRP9’s anti-apoptotic effect (P<0.05), whereas inhibition of Akt is less effective in blocking CTRP9 cardioprotection. Finally, CTRP9 phosphorylates BAD at its multiple anti-apoptotic sites, an effect blocked by PKA inhibitor. Conclusions We demonstrate that adipokine CTRP9 attenuates adverse cardiac remodeling following AMI, largely via a PKA-dependent pathway.

The porous architectural characteristics of biomaterials play an important role in scaffold revascularization. However, no consensus exists regarding optimal interconnection sizes for vascularization and its scaffold bioperformance with different interconnection sizes. Therefore, a series of disk-type beta-tricalcium phosphates with the same pore sizes and variable interconnections were produced to evaluate how the interconnection size influenced biomaterial vascularization in vitro and in vivo. We incubated human umbilical vein endothelial cells on scaffolds with interconnections of various sizes. Results showed that scaffolds with a 150 μm interconnection size ameliorated endothelial cell function evidenced by promoting cell adhesion and migration, increasing cell proliferation and enhancing expression of platelet-endothelial cell adhesion molecules and vascular endothelial growth factor. In vivo study was performed on rabbit implanted with scaffolds into the bone defect on femoral condyles. Implantation with scaffolds with 150 μm interconnection size significantly improved neovascularization as shown by micro-CT as compared to scaffolds with 100 and 120 μm interconnection sizes. Moreover, the aforementioned positive effects were abolished by blocking PI3K/Akt/eNOS pathway with LY-294002. Our study explicitly demonstrates that the scaffold with 150 μm interconnection size improves neovascularization via the PI3K/Akt pathway and provides a target for biomaterial inner structure modification to attain improved clinical performance in implant vascularization.

Recently identified as adiponectin (APN) paralogs, C1q/TNF-related proteins (CTRPs) share similar metabolic regulatory functions as APN. The current study determined cardiac expression of CTRPs, their potential cardioprotective function, and investigated whether and how diabetes may regulate cardiac CTRP expression. Several CTRPs are expressed in the heart at levels significantly greater than APN. Most notably, cardiac expression of CTRP9, the closest paralog of APN, exceeds APN by >100-fold. Cardiac CTRP9 expression was significantly reduced in high-fat diet-induced diabetic mice. In H9c2 cells, tumor necrosis factor-alpha (TNF-α) strongly inhibited CTRP9 expression (>60%), and significantly reduced peroxisome proliferator activated receptor-gamma (PPARγ), a known transcription factor promoting adiponectin expression. The inhibitory effect of TNF-α on PPARγ and CTRP9 was reversed by Tiron or rosiglitazone. CTRP9 knockdown significantly enhanced, whereas CTRP9 overexpression significantly attenuated simulated ischemia/reperfusion injury in H9c2 cells. In vivo CTRP9 administration to diabetic mice significantly attenuated NADPH oxidase expression and superoxide generation, reduced infarct size, and improved cardiac function. To the best of our knowledge, this is the first study providing evidence that downregulation of CTRP9, an abundantly expressed and novel cell survival molecule in the heart, by TNF-α-initiated oxidative PPARγ suppression contributes to exacerbated diabetic cardiac injury. Preservation of CTRP9 expression or augmentation of CTRP9-initiated signaling mechanisms may be the potential avenues for ameliorating ischemic diabetic cardiac injury.

Owing to their sessile nature, plants have evolved sophisticated genetic and epigenetic regulatory systems to respond quickly and reversibly to daily and seasonal temperature changes. However, our knowledge of how plants sense and respond to warming ambient temperatures is rather limited. Here we show that an increase in growth temperature from 22°C to 30°C effectively inhibited transgene-induced posttranscriptional gene silencing (PTGS) in Arabidopsis. Interestingly, warmth-induced PTGS release exhibited transgenerational epigenetic inheritance. We discovered that the warmth-induced PTGS release occurred during a critical step that leads to the formation of double-stranded RNA (dsRNA) for producing small interfering RNAs (siRNAs). Deep sequencing of small RNAs and RNA blot analysis indicated that the 22-30°C increase resulted in a significant reduction in the abundance of many trans-acting siRNAs that require dsRNA for biogenesis. We discovered that the temperature increase reduced the protein abundance of SUPPRESSOR OF GENE SILENCING 3, as a consequence, attenuating the formation of stable dsRNAs required for siRNA biogenesis. Importantly, SUPPRESSOR OF GENE SILENCING 3 overexpression released the warmth-triggered inhibition of siRNA biogenesis and reduced the transgenerational epigenetic memory. Thus, our study reveals a previously undescribed association between warming temperatures, an epigenetic system, and siRNA biogenesis.