Ping Xin

Repeated remote ischemic postconditioning protects against adverse left ventricular remodeling and improves survival in a rat model of myocardial infarction.

Rationale: Remote ischemic conditioning induced by repeated episodes of transient limb ischemia is a clinically applicable method for protecting the heart against injury at the time of reperfusion. Objective: To assess the effect of chronic, repeated, remote conditioning on infarct size and long-term remodeling after myocardial infarction. Methods and Results: Rats with ischemia/reperfusion injury received different protocols of remote limb conditioning. While a single early episode of remote ischemic conditioning during coronary occlusion (perconditioning) resulted in a decrease in infarct size on both day 4 and day 28, when it was repeated (postconditioning) intermittently (every 3 days) and intensively (every day), it was not associated with a further decrease in infarct size. However, the protection against adverse remodeling offered by a single episode of limb perconditioning was further enhanced by repeated remote postconditioning therapy in a dose-dependent manner. In separate experiments there was a dose-dependent improvement in survival at 84 days by Kaplan–Meier analysis. Conclusions: Whereas a single early episode of remote perconditioning reduces infarct size, repeated remote postconditioning further reduces adverse LV remodeling and improves survival in a dose-dependent fashion. These data may have clinical implications for the treatment of patients with evolving myocardial infarction.

Apelin-13 protects the heart against ischemia-reperfusion injury through inhibition of ER-dependent apoptotic pathways in a time-dependent fashion

Endoplasmic reticulum (ER) stress is activated during and contributes to ischemia-reperfusion (I/R) injury. Attenuation of ER stress-induced apoptosis protects the heart against I/R injury. Using apelin, a ligand used to activate the apelin APJ receptor, which is known to be cardioprotective, this study was designed to investigate 1) the time course of changes in I/R injury after ER stress; 2) whether apelin infusion protects the heart against I/R injury via modulation of ER stress-dependent apoptosis signaling pathways; and 3) how phosphatidylinositol 3-kinase (PI3K)/Akt, endothelial nitric oxide synthase (eNOS), AMP-activated protein kinase (AMPK), and ERK activation are involved in the protection offered by apelin treatment. The results showed that, using an in vivo rat I/R model induced by 30 min of ischemia followed by reperfusion, infarct size (IS) increased from 2 h of reperfusion (34.85 ± 2.14%) to 12 h of reperfusion (48.98 ± 3.35, P < 0.05), which was associated with an abrupt increase in ER stress-dependent apoptosis activation, as evidenced by increased CCAAT/enhancer-binding protein homologous protein (CHOP), caspase-12, and JNK activation (CHOP: 2.49-fold increase, caspase-12: 2.09-fold increase, and JNK: 3.38-fold increase, P < 0.05, respectively). Administration of apelin at 1 μg/kg not only completely abolished the activation of ER stress-induced apoptosis signaling pathways at 2 h of reperfusion but also significantly attenuated time-related changes at 24 h of reperfusion. Using pharmacological inhibition, we also demonstrated that PI3K/Akt, AMPK, and ERK activation were involved in the protection against I/R injury via inhibition of ER stress-dependent apoptosis activation. In contrast, although eNOS activation played a role in decreasing IS at 2 h of reperfusion, it failed to modify either IS or ER stress-induced apoptosis signaling pathways at 24 h after reperfusion.

Resveratrol protects cardiomyocytes from oxidative stress through SIRT1 and mitochondrial biogenesis signaling pathways

Reactive oxygen species (ROS) is generated by oxidative stress and plays an important role in various cardiac pathologies. The SIRT1 signaling pathway and mitochondrial biogenesis play essential roles in mediating the production of ROS. SIRT1 activated by resveratrol protects cardiomyocytes from oxidative stress, but the exact mechanisms by which SIRT1 prevents oxidative stress, and its relationship with mitochondrial biogenesis, remain unclear. In this study, it was observed that after stimulation with 50μMH2O2 for 6h, H9C2 cells produced excessive ROS and downregulated SIRT1. The mitochondrial protein NDUFA13 was also downregulated by ROS mediated by SIRT1. Resveratrol induced the expression of SIRT1 and mitochondrial genes NDUFA1, NDUFA2, NDUFA13 and Mn-SOD. However, the production of these genes was reversed by SIRT1 inhibitor nicotinamide. These results suggest that resveratrol inhibits ROS generation in cardiomyocytes via SIRT1 and mitochondrial biogenesis signaling pathways.

Fasudil Protects the Heart against Ischemia-Reperfusion Injury by Attenuating Endoplasmic Reticulum Stress and Modulating SERCA Activity: The Differential Role for PI3K/Akt and JAK2/STAT3 Signaling Pathways

Disordered calcium homeostasis can lead to endoplasmic reticulum (ER) stress. Our previous data showed that time course activation of ER stress contributes to time-related increase in ischemia-reperfusion (I/R) injury. However, it has not been tested whether PI3K/Akt and JAK2/STAT3 pathways play differential roles in reducing ER stress to protect the heart. In the present study, using fasudil which is a specific inhibitor of ROCK, we aimed to investigate whether improved SERCA expression and activity accounts for reduced ER stress by ROCK inhibition, specifically whether PI3K/Akt and JAK2/STAT3 pathways are differentially involved in modulating SERCA activity to reduce ER stress and hence I/R injury. The results showed that during the reperfusion period following 45 min of coronary ligation the infarct size (IS) increased from 3 h of reperfusion (45.4±5.57%) to 24 h reperfusion (64.21±5.43, P<0.05), which was associated with ER stress dependent apoptosis signaling activation including CHOP, Caspase-12 and JNK (P<0.05, respectively).The dynamic ER stress activation was also related to impaired SERCA activity at 24 h of reperfusion. Administration of fasudil at 10 mg/Kg significantly attenuated ROCK activation during reperfusion and resulted in an improved SERCA activity which was closely associated with decreases in temporal activation of ER stress and IS changes. Interestingly, while both PI3K/Akt and JAK2/STAT3 signaling pathways played equal role in the protection offered by ROCK inhibition at 3 h of reperfusion, the rescued SERCA expression and activity at 24 h of reperfusion by fasudil was mainly due to JAK2/STAT3 activation, in which PI3K/Akt signaling shared much less roles.

Favorable effects of resveratrol on sympathetic neural remodeling in rats following myocardial infarction.

Oxidative stress and inflammatory response induced by myocardial infarction play important roles in the development of sympathetic neural remodeling. The present study was designed to investigate whether resveratrol can improve sympathetic neural remodeling and hence cause less arrhythmias via its anti-oxidant and anti-inflammatory effects. Male Sprague Dawley rats were randomly assigned to either vehicle or resveratrol (1 mg/kg) treatment for 4 weeks post myocardial infarction. Another group of sham operated rats served as controls. Cardiac electrophysiology examination was performed to evaluate the severity of ventricular arrhythmias. Sympathetic neural remodeling characterized by heterogeneous nerve sprouting and sympathetic hyperinnervation was assessed by immunohistochemistry study. Western blotting and ELISA were used to evaluate inflammatory responses and oxidative stress was also quantified. Resveratrol treatment resulted in less episodes of inducible ventricular arrhythmias which was closely associated with attenuated sympathetic neural remodeling (P<0.001, respectively). Decreased nerve growth factor (NGF) expression was also observed in resveratrol treated rats in the peri-infarct area at 4 weeks after myocardial infarction (P<0.001). Interestingly, beneficial effects of resveratrol were also associated with less inflammatory responses and oxidative stress. Our data indicated that resveratrol can suppress sympathetic neural remodeling process after myocardial infarction via attenuated inflammatory responses and oxidative stress, which in turn leads to less inducibility of ventricular arrhythmias.

Therapeutic effects of continuous infusion of brain natriuretic peptides on postmyocardial infarction ventricular remodelling in rats.

BACKGROUND Previous studies have shown protective effects of brain natriuretic peptide (BNP) against the postmyocardial infarction (MI) remodelling process. The transcription factor NF-κB is known to play an important role after MI. AIMS To investigate if NF-κB is involved in the protective effects of BNP against adverse post-MI remodelling. METHODS Rats were randomly assigned to five groups: sham-operation; MI by coronary ligation; MI treated with chronic BNP infusion; MI treated with enalapril; MI treated with BNP+enalapril. Rats were closely monitored for survival rate analysis. Rats from each group were sacrificed on days 3, 7 and 28 postoperation. RESULTS The results showed that chronic continuous BNP infusion achieved similar effects to enalapril therapy, as evidenced by improved survival rate within the 28-day observation period compared with MI group rats; this effect was closely associated with preserved cardiac geometry and performance. The treatment combination did not offer extra benefits in terms of survival rate. Both BNP and enalapril therapy produced higher heart tissue concentrations of cyclic guanosine monophosphate and lower expression levels of inflammatory cytokines, including tumour necrosis factor-α, interleukin-1 and interleukin-6. These benefits were associated with lower phosphorylation levels of NF-κB subunits IκBα, p50 and p65. While enalapril significantly inhibited extracellular matrix remodelling via regulation of the protein expression ratio of matrix metalloproteinase/tissue inhibitor of metalloproteinase and the activity of matrix metalloproteinase, these variables were not affected by BNP, indicating that the two therapies involve different mechanisms. CONCLUSION Chronic BNP infusion can provide beneficial effects against adverse post-MI remodelling.

Combination of remote ischemic perconditioning and remote ischemic postconditioning fails to increase protection against myocardial ischemia/reperfusion injury, compared with either alone

Remote ischemic perconditioning (RIPerC) and remote ischemic postconditioning (RIPostC) have been previously demonstrated to protect the myocardium against ischemia/reperfusion (IR) injury. However, their combined effects remain to be fully elucidated. In order to investigate this, the present study used an in vivo rat model to assess whether synergistic effects are produced when RIPerC is combined with RIPostC. The rats were randomly assigned to the following groups: Sham, IR, RIPerC, RIPostC and RIPerC + RIPostC groups. The IR model was established by performing 40 min of left coronary artery occlusion, followed by 2 h of reperfusion. RIPerC and RIPostC were induced via four cycles of 5 min occlusion and 5 min reperfusion of the hindlimbs, either during or subsequent to myocardial ischemia. On measurement of infarct sizes, compared with the IR group (49.45±6.59%), the infarct sizes were significantly reduced in the RIPerC (34.36±5.87%) and RIPostC (36.04±6.16%) groups (P<0.05). However, no further reduction in infarct size was observed in the RIPerC + RIPostC group (31.43±5.43%; P>0.05), compared with the groups treated with either RIPerC or RIPostC alone. Activation of the reperfusion injury salvage kinase (RISK) Akt, extracellular signal-regulated kinase 1/2 and glycogen synthase kinase-3β, and survivor activating factor enhancement (SAFE) signal transducer and activator of transcription-3 pathways were enhanced in the RIPerC, RIPostC and the RIPerC + RIPostC groups, compared with the IR group, with no difference among the three groups. Therefore, whereas RIPerC and RIPostC were equally effective in providing protection against myocardial IR injury, the combination of RIPerC and RIPostC failed to provide further protection than treatment with either alone. The cardioprotective effects were found to be associated with increased activation of the RISK and SAFE pathways.

GRIM‑19‑mediated Stat3 activation is a determinant for resveratrol‑induced proliferation and cytotoxicity in cervical tumor‑derived cell lines.

Resveratrol is a natural phenol, produced from red grapes, berries and peanuts. Previous studies have suggested that resveratrol exerts anticancer effects. Activation of the signal transducer and activator of transcription 3 (Stat3) is important in cancer. However, the mechanisms by which resveratrol suppresses the Stat3 signaling pathway remain to be elucidated. The aim of the present study was to investigate the effects of resveratrol on GRIM‑19‑Stat3 signaling in HeLa cells, derived from a cervical tumor. HeLa cells were divided into experimental groups and treated with resveratrol. Western blotting was used to analyze the expression levels of p‑Stat3, Stat3, GRIM‑19 and β‑actin. Cell viability was determined using an MTT assay. The results showed that 100 µM resveratrol suppressed the proliferation and Stat3 phosphorylation in HeLa cells, and induced the expression of the gene associated with retinoid‑IFN‑induced mortality 19 (GRIM‑19) protein. Overexpression of GRIM‑19 suppressed the Stat3 signaling pathway in HeLa cells. The Stat3 signaling pathway was activated following the downregulation of GRIM‑19 expression using short interfering RNAs (siRNAs). Resveratrol suppressed cell proliferation, however, this effect was decreased through the use of siRNAs. The suppression of Stat3 phosphorylation by resveratrol decreased following treatment with siRNAs. To the best of our knowledge, the present study is among the first to identify GRIM‑19‑Stat3 signaling as a target of resveratrol, and further elucidates the mechanisms underlying the antitumor activity of resveratrol.

Myocardial infarction accelerates glomerular injury and microalbuminuria in diabetic rats via local hemodynamics and immunity.

BACKGROUND Clinically, approximately one-third of patients with chronic heart failure (CHF) exhibit some degree of renal dysfunction. This renal dysfunction is referred to as cardiorenal syndrome (CRS) and plays an important role in the poor prognosis of CHF. Mounting evidence suggests that diabetes is the most common underlying risk factor for CRS. However, the underlying pathophysiological mechanisms are poorly understood. METHODS We performed the following comparisons in two separate protocols: 1) surgically induced myocardial infarction rats (MI, n=10), sham operation rats (Ctr, n=10) and MI rats treated with Fasudil, a Rho-kinase inhibitor (MI+Fas, n=9); and 2) STZ-induced type 1 diabetic rats (DB, n=10), DB+MI rats (n=10) and DB+MI rats treated with Fasudil (DB+MI+Fas, n=9). Renal hemodynamics and vasoconstrictor reactivity were evaluated using the DMT myograph system. Renal immunity was evaluated by flow cytometry, electron microscopy, immunofluorescence, etc. RESULTS Twelve weeks after the operation, compared with DB or MI rats, DB+MI rats exhibited the following characteristics: 1) significantly increased glomerular enlargement, fibrosis, glomerulosclerosis, podocyte injury and microalbuminuria; 2) significantly increased vasoconstrictor reactivity of the renal interlobular arteries and renal venous pressure; 3) significantly increased infiltration of CD₃+ and CD₄+ T cells and decreased Treg/Th17 ratios; and 4) significantly increased glomerular deposition of IgG and C₄. In contrast, rats with MI only showed mildly accelerated glomerular remodeling and microalbuminuria, with little change in renal hemodynamics and immunity. Fasudil treatment significantly improved the renal lesions in DB+MI rats but not MI rats. CONCLUSIONS Post-MI cardiac dysfunction significantly accelerated glomerular remodeling, podocyte injury and microalbuminuria in STZ-induced diabetic rats. These changes were accompanied by altered local hemodynamics and immunity.

GW24-e0422 Myocardial Infarction Worsens the Podocyte Injury and Albuminuria in Potential Kidney Disease Rats probably via local hemodynamics-immunity way

Objectives Amounts of evidence demonstrated increased deterioration of renal function after chronic heart failure (CHF), particularly in patients who already had potential impairment such as diabetic nephropathy. Numberous studies suggested that the inflammation after abnormal renal hemodynamics played important roles in this interaction. Due to the inseparable relation between immunity and inflammation, we hypothesised that the aberrance of renal cellular, humoral immunity were also involved in this progression, and rho-kinase inhibitor (fasudil) which was thought to improve the renal vascular function would improve it. Methods There are 8 rat groups in this study: 1. Ctr (n = 12): Sprague-Dawley (SD) rats with sham myocardial infarction (MI) operation then treated with vehicle for 3 months; 2. MI (n = 12): SD rats with surgically induced MI then treated with vehicle for 3months; 3. DB (n = 11): STZ-induced type 1 diabetic rats (16ws) with sham MI operation then treated with vehicle for 3months; 4. DB + MI (n = 11): STZ-induced type 1 diabetic rats (16ws) with MI operation then treated with vehicle for 3months; 5. DB + MI + Fas (n = 11): STZ-induced type 1 diabetic rats (16ws) with MI operation then treated with fasudil for 3months; 6. UNX (n = 11): SD rats underwent unilateral nephrectomy (UNX, 2ws) then treated with vehicle for 3 months; 7. UNX + MI (n = 11): UNX rats (2ws) underwent MI operation then treated with vehicle for 3 months; 8. UNX + MI + Fas (n = 11): UNX rats (2ws) underwent MI operation then treated with fasudil for 3 months. Renal hemodynamics/vascular reactivity were evaluated with DMT multi wire myograph system. The parameters of cellular, humoral immunity were evaluated with flow cytometry analysis, electron microscopy, Immunofluorescence and Immunohistochemistry. In addition the marker of glomerular podocyte injury such as desmin, nephrin were observed with Immunohistochemistry. Results Compared with their sham-operation groups (DB, UNX), two operation groups (DB + MI, UNX + MI) showed: 1. significantly decreased renal reperfusion pressure (RPR) and increased vasoconstriction of renal interlobular arteries; 2. significantly increased infiltration of monocytes and the proportion of CD3+ cell, CD4+ T cells, producing-IFN-γ, IL-17, IL-4 CD4+ T cell and regulatory T cell, the deposition of immune complex (IgG) and complement C4 in glomeruli. 3. significantly aberrant albuminuria and the marker of glomerular podocyte injury such as desmin and nephrin. Two treated groups (DB + MI + Fas, UNX + MI + Fas) showed the improved parameters of hemodynamics/vascular reactivity, albuminuria and glomerular podocyte injury in varying degrees. At last the proportion of renal CD Conclusions In this study, post-MI CHF significantly deteriorated the renal function and remodelling in potential renal injury rats (DB or UNX). They were accompanied with the aberrated renal hemodynamics/vascular reactivity and renal immune system. Rho-kinase inhibitor fasudil prevented the aberration of them and the deterioration of renal function, providing new insight into the treatment of cardiorenal syndrome.