Yukai Liu

LincRNA-p21 Regulates Neointima Formation, Vascular Smooth Muscle Cell Proliferation, Apoptosis and Atherosclerosis by Enhancing p53 Activity

Background— Long noncoding RNAs (lncRNAs) have recently been implicated in many biological processes and diseases. Atherosclerosis is a major risk factor for cardiovascular disease. However, the functional role of lncRNAs in atherosclerosis is largely unknown. Methods and Results— We identified lincRNA-p21 as a key regulator of cell proliferation and apoptosis during atherosclerosis. The expression of lincRNA-p21 was dramatically downregulated in atherosclerotic plaques of ApoE−/− mice, an animal model for atherosclerosis. Through loss- and gain-of-function approaches, we showed that lincRNA-p21 represses cell proliferation and induces apoptosis in vascular smooth muscle cells and mouse mononuclear macrophage cells in vitro. Moreover, we found that inhibition of lincRNA-p21 results in neointimal hyperplasia in vivo in a carotid artery injury model. Genome-wide analysis revealed that lincRNA-p21 inhibition dysregulated many p53 targets. Furthermore, lincRNA-p21, a transcriptional target of p53, feeds back to enhance p53 transcriptional activity, at least in part, via binding to mouse double minute 2 (MDM2), an E3 ubiquitin-protein ligase. The association of lincRNA-p21 and MDM2 releases MDM2 repression of p53, enabling p53 to interact with p300 and to bind to the promoters/enhancers of its target genes. Finally, we show that lincRNA-p21 expression is decreased in patients with coronary artery disease. Conclusions— Our studies identify lincRNA-p21 as a novel regulator of cell proliferation and apoptosis and suggest that this lncRNA could serve as a therapeutic target to treat atherosclerosis and related cardiovascular disorders.

Plasma long non-coding RNA, CoroMarker, a novel biomarker for diagnosis of coronary artery disease.

Long non-coding RNAs (lncRNAs) have been reported to be involved in the pathogenesis of cardiovascular disease (CVD), but whether circulating lncRNAs can serve as a coronary artery disease (CAD), biomarker is not known. The present study screened lncRNAs by microarray analysis in the plasma from CAD patients and control individuals and found that 265 lncRNAs were differentially expressed. To find specific lncRNAs as possible CAD biomarker candidates, we used the following criteria for 174 up-regulated lncRNAs: signal intensity ≥8, fold change >2.5 and P<0.005. According to these criteria, five intergenic lncRNAs were identified. After validation by quantitative PCR (qPCR), one lncRNA was excluded from the candidate list. The remaining four lncRNAs were independently validated in another population of 20 CAD patients and 20 control individuals. Receiver operating characteristic (ROC) curve analysis showed that lncRNA AC100865.1 (referred to as CoroMarker) was the best of these lncRNAs. CoroMarker levels were also stable in plasma. The predictive value of CoroMarker was further assessed in a larger cohort with 221 CAD patients and 187 control individuals. Using a diagnostic model with Fishers criteria, taking the risk factors into account, the optimal sensitivity of CoroMarker for CAD increased from 68.29% to 78.05%, whereas the specificity decreased slightly from 91.89% to 86.49%. CoroMarker was stable in plasma because it was mainly in the extracellular vesicles (EVs), probably from monocytes. We conclude that CoroMarker is a stable, sensitive and specific biomarker for CAD.

Protective effects of aliskiren on ischemia-reperfusion-induced renal injury in rats.

The protective effect of aliskiren on ischemia-reperfusion (I/R) injury in the heart and brain has been reported. Whether or not this protective effect extends into the alleviation of renal I/R injury is not known. Therefore, we investigated the protective effect of aliskiren in the kidney in this study. Sprague-Dawley rats were randomly divided into four groups: sham control group; sham control with aliskiren pretreatment; I/R group and I/R with aliskiren pretreatment. Aliskiren (3mg/kg) or vehicle was administrated intravenously via vena cava. Blood samples and the left kidneys were then collected to check for renal function, angiotensin II (Ang II), apoptosis and oxidative stress levels. Compared with the sham rats, serum creatinine (SCR) and blood urea nitrogen (BUN) were significantly increased in the I/R rats, accompanied by histopathological damage to the kidney, which included tubular cell swelling, desquamation, and cast formation. There were also more apoptotic cells and leukocyte infiltration in the I/R rats than in the sham rats. Pretreatment with aliskiren ameliorated I/R induced renal injury, i.e. reduced SCR and BUN levels, ameliorated renal histopathological changes, and decreased the apoptosis of cells and leukocyte infiltration in kidney. I/R injury also decreased superoxide dismutase (SOD) and glutathione (GSH-reduced form) levels, which were blocked with the aliskiren pretreatment. Aliskiren pretreatment exerts a protective effect on ischemia/reperfusion injury in the kidney, via amelioration of oxidative stress, and reduction in leukocyte infiltration and cellular apoptosis.

Irisin Lowers Blood Pressure by Improvement of Endothelial Dysfunction via AMPK‐Akt‐eNOS‐NO Pathway in the Spontaneously Hypertensive Rat

Background Exercise is a major nonpharmacological treatment for hypertension, but its underlying mechanisms are still not completely elucidated. Irisin, a polypeptide containing 112 amino acids, which is secreted mainly by skeletal muscle cells during exercise, exerts a protective role in metabolic diseases, such as diabetes mellitus and obesity. Because of the close relationship between irisin and metabolic diseases, we hypothesized that irisin may play a role in the regulation of blood pressure. Methods and Results Blood pressures of male Wistar‐Kyoto (WKY) rats and spontaneously hypertensive rats (SHRs) were monitored through the carotid artery. Our study found that acute intravenous injection of irisin reduced blood pressure in SHRs, but not WKY rats. Irisin, by itself, had no direct vasorelaxing effect in phenylephrine‐preconstricted mesenteric arteries from SHRs. However, irisin augmented the acetylcholine‐induced vasorelaxation in mesenteric arteries from SHRs that could be reversed by Nω‐nitro‐l‐arginine‐methyl ester (L‐NAME; 100 μmol/L), indicating a role of nitric oxide (NO) in this action. Indeed, irisin increased NO production and phosphorylation of endothelial nirtic oxide synthase (eNOS) in endothelial cells. 5′‐AMP‐activated protein kinase (AMPK) was involved in the vasorelaxing effect of irisin because compound C (20 μmol/L), an AMPK inhibitor, blocked the irisin‐mediated increase in phosphorylation of eNOS and protein kinase B (Akt) in endothelial cells and vasodilation in mesenteric arteries. Conclusions We conclude that acute administration of irisin lowers blood pressure of SHRs by amelioration of endothelial dysfunction of the mesenteric artery through the AMPK‐Akt‐eNOS‐NO signaling pathway.

MG53 permeates through blood-brain barrier to protect ischemic brain injury

Ischemic injury to neurons represents the underlying cause of stroke to the brain. Our previous studies identified MG53 as an essential component of the cell membrane repair machinery. Here we show that the recombinant human (rh)MG53 protein facilitates repair of ischemia-reperfusion (IR) injury to the brain. MG53 rapidly moves to acute injury sites on neuronal cells to form a membrane repair patch. IR-induced brain injury increases permeability of the blood-brain-barrier, providing access of MG53 from blood circulation to target the injured brain tissues. Exogenous rhMG53 protein can protect cultured neurons against hypoxia/reoxygenation-induced damages. Transgenic mice with increased levels of MG53 in the bloodstream are resistant to IR-induced brain injury. Intravenous administration of rhMG53, either prior to or after ischemia, can effectively alleviate brain injuries in rats. rhMG53-mediated neuroprotection involves suppression of apoptotic neuronal cell death, as well as activation of the pro-survival RISK signaling pathway. Our data indicate a physiological function for MG53 in the brain and suggest that targeting membrane repair or RISK signaling may be an effective means to treat ischemic brain injury.

Stimulation of Dopamine D3 Receptor Attenuates Renal Ischemia-Reperfusion Injury via Increased Linkage With Gα12.

Background Renal ischemia-reperfusion (I/R) injury causes renal tubular necrosis, apoptosis, and inflammation leading to acute renal dysfunction. Recent studies have revealed that deletion of G&agr;12 mitigates the renal damage due to I/R injury. Our previous study showed that activation of dopamine D3 receptor (D3R) increased its linkage with G&agr;12, and hampered G&agr;12-mediated stimulation of renal sodium transport. In the present study, we used an in-vivo rat model and an in vitro study of the renal epithelial cell line (NRK52E) to investigate whether or not an increased linkage between D3R and G&agr;12 contributes to the protective effect of D3R on renal I/R injury. Methods For in vivo studies, I/R injury was induced in a rat renal unilateral clamping model. For in vitro studies, hypoxia/reoxygenation and cold storage/rewarming injuries were performed in NRK52E cells. PD128907, a D3R agonist, or vehicle, was administered 15 minutes before clamping (or hypoxia) in both the in vivo or in vitro studies. Results In the rat renal unilateral clamping model, pretreatment with PD128907 (0.2 mg/kg, intravenous) protected against renal I/R injury and increased survival rate during a long-term follow-up after 7 days. A decrease in the generation of reactive oxygen species, apoptosis, and inflammation may be involved in the D3R-mediated protection because pretreatment with PD128907 increased renal glutathione and superoxide dismutase levels and decreased malondialdehyde levels in the I/R group. The increase in cytokines (TNF-&agr;, IL-1&bgr;, and IL-10) and myeloperoxidase in I/R injured kidney was also prevented with a simultaneous decrease in the apoptosis of the epithelial cells and expression of apoptosis biomarkers in kidney harvested 1 day after I/R injury. The increase in the coimmunoprecipitation between D3R and G&agr;12 with D3R stimulation paralleled the observed renal protection from I/R injury. Moreover, in vitro studies showed that transient overexpression of G&agr;12 in the NRK52E cells attenuated the protective effect of PD128907 on hypoxia/reoxygenation injury. The protective effect of PD128907 might be of significance to renal transplantation because cold storage/rewarming induced injury increased lactate dehydrogenase release and decreased cell viability in NRK52E cells. Conversely, in the presence of PD128907, the increased lactate dehydrogenase release and decreased cell viability were reversed. Conclusions These results suggest that activation of D3R, by decreasing G&agr;12-induced renal damage, may exert a protective effect from I/R injury.

Aliskiren and Amlodipine in the Management of Essential Hypertension: Meta-Analysis of Randomized Controlled Trials

Background Aliskiren is a novel renin-angiotensin aldosterone system (RAAS) inhibitor, the combination therapy of aliskiren and amlodipine for blood pressure control have been reported recently. The primary objective of this analysis is to review recently reported randomized controlled trials (RCTs) to compare antihypertensive effects and adverse events between mono (amlodipine or aliskiren alone) and combination therapy of both medicines. Methods Databases for the search included Pubmed, Embase and the Cochrane Central Register of Controlled Trials. Revman v5.0 statistical program was used to analyze the data. Weighted mean differences (WMD) with a 95% confidence interval (CI) were used for the calculation of continuous data, and relative risk (RR) with a 95% CI was used for dichotomous data. Results We analyzed the data from 7 RCTs for a total of 6074 participants in this meta-analysis. We found that the aliskiren/amlodipine combination therapy had a stronger effect in lowering blood pressure as compared with the monotherapy using aliskiren (SBP: WMD = −10.42, 95% CI −13.03∼−7.82, P<0.00001; DBP: WMD = −6.60, 95% CI −7.22∼−5.97, P<0.00001) or amlodipine (SBP: WMD = −4.85, 95% CI −6.88∼−2.81, P<0.00001; DBP: WMD = −2.91, 95% CI −3.85∼−1.97, P<0.00001). No differences were found in terms of adverse events between combination therapy and monotherapy, except for the rates of peripheral edema and hypokalaemia which were significantly lower in the combination therapy than in the amlodipine monotherapy (RR = 0.78, 0.66∼0.92, P = 0.004; RR = 0.51, 0.27∼0.97, P = 0.04). Similar antihypertensive effects were found in both obese (body mass index > = 30 kg/m2) hypertensive and non-obese (body mass index <30 kg/m2) hypertensive patients. Moreover, there was no difference with the blood pressure lowering or adverse effects with regards to the combination therapy in both subgroups. Conclusion We found that aliskiren/amlodipine combination therapy provided a more effective blood pressure reduction than monotherapy with either drug without increase in the occurrence of adverse events.

Protective effects of tirofiban on ischemia/reperfusion-induced renal injury in vivo and in vitro.

Tirofiban, a glycoprotein IIb/IIIa receptor inhibitor, is widely used in the management of patients with unstable angina or myocardial infarction, and shows protective effects on ischemia/reperfusion (I/R) injured heart. Whether or not it has protective effect on I/R injured kidney is not known. The present in vivo and in vitro study found that serum creatinine (SCR) and blood urea nitrogen (BUN) were significantly increased in I/R rats, accompanied by histopathological damage of the kidney. Apoptotic cells, leukocyte infiltration and ROS production were increased in I/R rats. Pretreatment by intravenous injection of tirofiban (200μg/kg) reduced SCR and BUN levels, ameliorated renal histopathological changes, and decreased ROS production, cell apoptosis and leukocyte infiltration in I/R injured kidney. Our further study showed that the protection of tirofiban might be associated with the restoration of eNOS/iNOS balance, since inhibition of NO production blocked the tirofiban-mediated renal protection on I/R injury. The present in vivo and in vitro study indicated that tirofiban pretreatment exerts a protective effect on I/R injury in kidney through regulation of eNOS/iNOS balance.

Dopamine d1-like receptors suppress proliferation of vascular smooth muscle cell induced by insulin-like growth factor-1

Abstract Objective: Proliferation of vascular smooth muscle cells (VSMCs) participates in the pathogenesis and development of cardiovascular diseases, including essential hypertension and atherosclerosis. Our previous study found that stimulation of D1-like dopamine receptors inhibited insulin-induced proliferation of VSMCs. Insulin-like growth factor-1 (IGF-1) and insulin share similar structure and biological effect. However, whether or not there is any effect of D1-like receptors on IGF-1-induced proliferation of VSMCs is not known. Therefore, we investigated the inhibitory effect of D1-like dopamine receptors on the IGF-1-induced VSMCs proliferation in this study. Method: VSMC proliferation was determined by [3H]-thymidine incorporation, the uptake of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay and cell number. Phosphorylated/non-phosphorylated IGF-1 receptor, Akt, mTOR and p70S6K expressions were determined by immunoblotting. The oligodeoxynucleotides were transfected to A10 cells to identify the effect of D1 and D5 receptors, respectively. Results: IGF-1 increased the proliferation of VSMCs, while in the presence of fenoldopam, IGF-1-mediated stimulatory effect was reduced. Use of either antisense for D1 or D5 receptor partially inhibited the fenoldopam-induced antiproliferation effect of VSMCs. Use of both D1 and D5 receptor antisenses completely blocked the inhibitory effect of fenoldopam. In the presence of PI3k and mTOR inhibitors, the IGF-1-mediated proliferation of VSMCs was blocked. Moreover, IGF-1 increased the phosphorylation of PI3k and mTOR. The inhibitory effect of fenoldopam on VSMC proliferation might be due to the inhibition of IGF-1 receptor expression and IGF-1 phosphorylation, because in the presence of fenoldopam, the stimulatory effect of IGF-1 on phosphorylation of IGF-1 receptor, PI3k and mTOR is reduced, the IGF-1 receptor expression was reduced in A10 cells. Conclusion: Activation of the D1-like receptors suppressed the proliferative effect of IGF-1 in A10 cells via the inhibition of the IGF-1R/Akt/mTOR/p70S6K pathway and downregulated the expression of IGF-1 receptor.

Irisin protects mitochondria function during pulmonary ischemia/reperfusion injury

Irisin protects against ischemia/reperfusion injury to the lung by improving mitochondrial function. Protecting the lungs with irisin Limb remote ischemic preconditioning, or subjecting a limb to temporary periods of brief ischemia, is a method that can be used to protect the lungs from ischemia/reperfusion damage due to an expected stressor such as surgery. Chen et al. investigated the mechanism by which this occurs and determined that remote ischemic preconditioning releases irisin, a myokine that targets mitochondria and blocks some of the deleterious effects of oxidative stress. The authors showed that this can occur naturally, as when infants with neonatal respiratory distress syndrome show transfer of endogenous irisin from the circulation into the lungs, or it can be induced therapeutically, as the authors demonstrated by treating mouse models with exogenous irisin, which protected their lungs from ischemia/reperfusion injury. Limb remote ischemic preconditioning (RIPC) is an effective means of protection against ischemia/reperfusion (IR)–induced injury to multiple organs. Many studies are focused on identifying endocrine mechanisms that underlie the cross-talk between muscle and RIPC-mediated organ protection. We report that RIPC releases irisin, a myokine derived from the extracellular portion of fibronectin domain–containing 5 protein (FNDC5) in skeletal muscle, to protect against injury to the lung. Human patients with neonatal respiratory distress syndrome show reduced concentrations of irisin in the serum and increased irisin concentrations in the bronchoalveolar lavage fluid, suggesting transfer of irisin from circulation to the lung under physiologic stress. In mice, application of brief periods of ischemia preconditioning stimulates release of irisin into circulation and transfer of irisin to the lung subjected to IR injury. Irisin, via lipid raft–mediated endocytosis, enters alveolar cells and targets mitochondria. Interaction between irisin and mitochondrial uncoupling protein 2 (UCP2) allows for prevention of IR-induced oxidative stress and preservation of mitochondrial function. Animal model studies show that intravenous administration of exogenous irisin protects against IR-induced injury to the lung via improvement of mitochondrial function, whereas in UCP2-deficient mice or in the presence of a UCP2 inhibitor, the protective effect of irisin is compromised. These results demonstrate that irisin is a myokine that facilitates RIPC-mediated lung protection. Targeting the action of irisin in mitochondria presents a potential therapeutic intervention for pulmonary IR injury.