Pioglitazone Attenuates Atherosclerosis in Diabetic Mice by Inhibition of Receptor for Advanced Glycation End-Product (RAGE) Signaling.

BACKGROUND Peroxisome proliferator-activated receptor-g (PPAR-g) exhibits anti-inflammatory and anti-diabetic properties, and is protective against cardiovascular diseases. This study aimed to determine the effects of a PPAR-g agonist pioglitazone on atherogenesis in an ApoE knockout mouse (ApoE-/-) diabetic mouse model and in a cultured vascular smooth muscle cells (VSMCs) model. MATERIAL AND METHODS Male ApoE-/- mice were rendered diabetic by 5 daily intraperitoneal injections of streptozotocin. Pioglitazone (20 mg/kg/d) or PPAR-γ inhibitor GW9662 (1 mg/kg/d) were administered for 12 weeks. At the end of treatment, mice were killed and the aortae were isolated. Oil Red O staining was used to evaluate atherosclerotic plaque area. H&E staining was used to evaluate the number of complicated plaques. Western blotting and immunohistochemistry were used to determine the expression of advanced glycation end-products (RAGE) and PPAR-γ. The effects of pioglitazone and GW9662 on RAGE and PPAR-g expression were examined in cultured primary mouse VSMCs in hyperglycemic conditions. RESULTS Administration of pioglitazone in diabetic ApoE-/- mice successfully reduced atherosclerotic plaque area and the number of complicated plaques. Moreover, pioglitazone inhibited RAGE and stimulated PPAR-γ protein expression in atherosclerotic plaques of diabetic ApoE-/- mice. In cultured VSMCs upon high-glucose challenge, pioglitazone downregulated RAGE mRNA and protein expression. Blockade of PPAR-γ activity by GW9662 remarkably attenuated the inhibitory actions of pioglitazone on atherogenesis, both in diabetic ApoE-/- mice and in cultured VSMCs, upon high-glucose challenge. CONCLUSIONS Pioglitazone has a therapeutic effect on atherosclerosis in diabetes, and inhibition of RAGE signaling plays a critical role in mediating the beneficial effects of pioglitazone.

Comparison between left bundle branch area pacing and right ventricular pacing: ventricular electromechanical synchrony and risk of atrial high-rate episodes.

Background: The electromechanical dyssynchrony associated with right ventricular pacing (RVP) has been found to have adverse impact on clinical outcomes. Several studies have shown that left bundle branch area pacing (LBBAP) has superior pacing parameters compared with RVP. We aimed to assess the difference in ventricular electromechanical synchrony and investigate the risk of atrial high-rate episodes (AHREs) in patients with LBBAP and RVP. Methods: We consecutively identified 40 patients with atrioventricular block and no prior atrial fibrillation. They were divided according to the ventricular pacing sites: the LBBAP group and the RVP group (including the right ventricular apical pacing (RVA) group and the right side ventricular septal pacing (RVS) group). Evaluation of ventricular electromechanical synchrony was implemented using electrocardiogram and two-dimensional speckle tracking echocardiography (2D-STE). AHRE was defined as event with an atrial frequency of ≥176 bpm lasting for ≥6 min recorded by pacemakers during follow-up. Results: The paced QRS duration of the LBBAP group was significantly shorter than that of the other two groups: LBBAP 113.56 ± 9.66 ms vs. RVA 164.73 ± 14.49 ms, p < 0.001; LBBAP 113.56 ± 9.66 ms vs. RVS 148.23 ± 17.3 ms, p < 0.001. The LBBAP group showed shorter maximum difference (TDmax), and standard deviation (SD) of the time to peak systolic strain among the 18 left ventricular segments, and time of septal-to-posterior wall motion delay (SPWMD) compared with the RVA group (TDmax, 87.56 ± 56.01 ms vs. 189.85 ± 91.88 ms, p = 0.001; SD, 25.40 ± 14.61 ms vs. 67.13 ± 27.40 ms, p < 0.001; SPWMD, 28.75 ± 21.89 ms vs. 99.09 ± 46.56 ms, p < 0.001) and the RVS group (TDmax, 87.56 ± 56.01 ms vs. 156.46 ± 55.54 ms, p = 0.003; SD, 25.40 ± 14.61 ms vs. 49.02 ± 17.85 ms, p = 0.001; SPWMD, 28.75 ± 21.89 ms vs. 91.54 ± 26.67 ms, p < 0.001). The interventricular mechanical delay (IVMD) was shorter in the LBBAP group compared with the RVA group (-5.38 ± 9.31 ms vs. 44.82 ± 16.42 ms, p < 0.001) and the RVS group (-5.38 ± 9.31 ms vs. 25.31 ± 21.36 ms, p < 0.001). Comparing the RVA group and the RVS group, the paced QRS duration and IVMD were significantly shorter in the RVS group (QRS duration, 164.73 ± 14.49 ms vs. 148.23 ± 17.3 ms, p = 0.02; IVMD, 44.82 ± 16.42 ms vs. 25.31 ± 21.36 ms, p = 0.022). During follow-up, 2/16 (12.5%) LBBAP patients, 4/11 (36.4%) RVA patients, and 8/13 (61.5%) RVS patients had recorded novel AHREs. LBBAP was proven to be independently associated with decreased risk of AHREs than RVP (log-rank p = 0.043). Conclusion: LBBAP generates narrower paced QRS and better intro-left ventricular and biventricular contraction synchronization compared with traditional RVP. LBBAP was associated with a decreased risk of AHREs compared with RVP.

Urotensin II promotes the production of LTC4 in rat aortic adventitial fibroblasts through NF-κB-5-LO pathway by p38 MAPK and ERK activations.

Adventitia is the outer part of the arterial wall where the inflammatory response often occurs. Urotensin II (UII) is a potent vasoconstrictive peptide that also promotes the inflammatory process in patients with cardiovascular disease. Leukotriene C4 (LTC4), a lipid mediator, was recently found to play a role in the inflammatory process in the artery. We hypothesized that the adventitia is one of the resources of LTC4 and that UII may promote LTC4 production through the 5-LO (5-lipoxygenase) pathway in adventitial fibroblasts. Rat adventitial fibroblasts were isolated and incubated in serum-free medium with either UII alone or in combination with inhibitors of p38 MAPK, ERK, and UII receptors. The expression of 5-LO was detected using real-time polymerase chain reaction and Western blot. The translocation and binding activity of nuclear factor (NF)-κB were measured using immunofluorescence and electrophoretic mobility shift assay, respectively. The production of LTC4 was measured by enzyme-linked immunosorbent assay. The results indicated that: (1) adventitial fibroblasts were a source of LTC4 production; (2) UII increased the expression of the 5-LO mRNA and the protein by NF-κB activation through p38 MAPK and ERK pathways; and (3) UII promoted the LTC4 release in fibroblasts through the 5-LO pathway by p38 MAPK and ERK activations. The 5-LO pathway mediates LTC4 production, which may be a new mechanism in the pathogenesis of the vascular adventitial inflammation caused by UII.

Urotensin II stimulates vascular endothelial growth factor secretion from adventitial fibroblasts in synergy with angiotensin II.

BACKGROUND: The adventitia plays an important role in and is considered to be the initiating site for vascular remodeling. Urotensin II (UII) and angiotensin II (Ang II) are the two most important vascular peptides involved in vascular remodeling in the adventitia. Nevertheless, little is known about their effect on the expression of vascular endothelial growth factor (VEGF). It was hypothesized that both UII and Ang II could induce VEGF expression in adventitial fibroblasts and VEGF may play a role in cell proliferation and collagen I synthesis induced by UII or Ang II. METHODS AND RESULTS: Growth-arrested adventitial fibroblasts were incubated in serum-free medium with UII and/or Ang II and inhibitors of the mitogen-activated protein kinase (MAPK) pathway or VEGF-neutralizing antibodies. The VEGF expression was evaluated using enzyme-linked immunosorbent assay (ELISA), while the proliferation and collagen I synthesis were detected using methyl thiazol tetrazolium (MTT) assay and ELISA. It was found that: (1) both UII and Ang II could stimulate VEGF expression in adventitial fibroblasts and they had a synergistic effect; (2) MAPK pathway inhibitors could inhibit VEGF secretion induced by UII and/or Ang II; and (3) VEGF-neutralizing antibodies could inhibit UII/Ang II-induced cell proliferation and collagen synthesis in adventitial fibroblasts. CONCLUSIONS: Induction of VEGF expression may be a new mechanism involved in vascular remodeling for UII and Ang II.

Contemporary impact of circadian symptom-onset patterns of acute ST-Segment elevation myocardial infarction on long-term outcomes after primary percutaneous coronary intervention.

BACKGROUND: Daytime variation with regard to onset time of ST-elevation myocardial infarction (STEMI) symptoms has been observed. Nevertheless, with the advanced medical therapy, it is not uncertainty if a similar circadian pattern of STEMI symptom onset occurs, as well as its possible impact on clinical outcomes. Few long-term data are available. We assess the impact of circadian symptom-onset patterns of STEMI on major adverse cardiovascular events (MACE) in more contemporary patients treated with primary percutaneous coronary intervention (PPCI). METHODS AND RESULTS: A total of 1099 consecutive STEMI patients undergoing PPCI ≤12h from symptom onset during 2013 to 2019 were classified into 4 groups by 6-h intervals according to time-of-day at symptom onset: night (0:00-5:59), morning (6:00-11:59), afternoon (12:00-17:59), and evening (18:00-23:59). Incidence of MACE including cardiovascular death and nonfatal MI during a median follow-up of 48 months was compared among the 4 groups. A morning peak of symptom onset of STEMI was detected during the period 06:00-11:59 (p < .001). Compared with other three 6-h intervals, the incidence of long-term MACE during night onset-time (18.8%, 10.1%, 10.7% and 12.4%, p = .020) was significant higher that was driven by more mortality (13.1%, 6.5%, 7.1%and 7.7%, p = .044). Night symptom-onset STEMI was independently associated with subsequent MACE (hazard ratio = 1.57, 95%CI: 1.09-2.27, p = .017) even after multivariable adjustment. CONCLUSIONS: Circadian variation of STEMI symptom-onset with morning predominance still exists in contemporary practice. Night symptom-onset STEMI was independently associated with increased risk of MACE in Chinese patients treated with PPCI.

Contemporary Implications of ECG to Activation Time on Long-term Outcomes in Patients With ST-Segment Elevation Myocardial Infarction Treated With Primary Percutaneous Coronary Intervention.

PURPOSE: The prognosis of door to balloon time (DBT) after primary percutaneous coronary intervention (PPCI) in ST-segment elevation myocardial infarction (STEMI) has been extensively studied. However, the clinical implications of the ECG to activation time (EAT), as a component of DBT, on long-term outcomes are less well established. This study evaluates the association of EAT with major adverse cardiovascular events (MACEs) in more contemporary patients undergoing PPCI and investigates factors that influence EAT. METHODS: A total of 1082 consecutive patients with STEMI who underwent PPCI from 2013 to 2019 were classified into 3 groups according to EAT: EAT ≤30 minutes, EAT of 30 to 60 minutes, and EAT >60 minutes. We analyzed the incidence of MACEs, including all-cause death, nonfatal recurrence of MI, or nonfatal stroke during a median follow-up of 37 months. FINDINGS: The median EAT was 58 minutes (interquartile range, 44-80 minutes), which explained 90% of the variability in DBT and had the strongest correlation with DBT (r = 0.95, P < 0.001). Achieving an EAT of ≤30 minutes resulted in a 94.1% chance of achieving a DBT ≤90 minutes. The incidence of MACEs had a concurrent increase with increased EAT (10.2% for EAT ≤30 minutes, 14.3% for EAT of 30-60 minutes, and 17.3% for EAT >60 minutes; P = 0.027) that was driven by more mortality (4.2% for EAT ≤30 minutes, 6.9% for EAT of 30-60 minutes, and 9.8% for EAT >60 minutes; P = 0.020). An EAT >30 minutes was independently associated with risk-adjusted long-term MACEs (hazard ratio = 1.99; 95% CI, 1.07-3.69; P = 0.030). Critically ill status in emergency department (P = 0.001) and time required for consent of revascularization (P < 0.001) were significantly associated with delayed EAT. IMPLICATIONS: Achieving an EAT ≤30 minutes was key to achieving the guideline-recommended target time of DBT in contemporary practice. As a strong driver of overall DBT, EAT >30 minutes was associated with worse clinical outcome in patients with STEMI undergoing PPCI. These data suggest that efforts to minimize EAT are needed to reduce long-term MACEs in contemporary population. © 2021 Elsevier HS Journals, Inc.

Assessment of ventricular mechanical synchronization after left bundle branch pacing using 2-D speckle tracking echocardiography.

BACKGROUND: The left bundle branch pacing (LBBP) makes the ventricular depolarization closer to the physiological state and shortens QRS duration. The purpose of this study is to explore the ventricular systolic mechanical synchronization after LBBP in comparison with traditional right ventricular pacing (RVP) using two-dimensional strain echocardiography (2D-STE). METHODS: Thirty-two patients who received LBBP (n = 16) or RVP (n = 16) from October 2018 to October 2019 and met the inclusion criteria were included in this retrospective study. Electrocardiogram (ECG) characteristics, pacing parameters, pacing sites, and safety events were assessed before and after implantation. Acquisition and analysis of ventricular systolic synchronization were implemented using 2D-STE. RESULTS: In RVP group, ECG showed left bundle branch block patterns. At LBBP, QRS morphology was in the form of right bundle branch block, and QRS durations were significantly shorter than that of the RVP QRS (109.38 ± 12.89 vs 149.38 \± 19.40 ms, P < .001). Both the maximum time differences (TD) and SDs of the 18-segments systolic time to peak systolic strain were significantly shorter under LBBP than under RVP (TD, 66.62 ± 37.2 vs 148.62 ± 43.67 ms, P < .01; SD, 21.80 ± 12.13 vs 52.70 ± 17.72 ms, P < .01), indicating that LBBP could provide better left ventricular mechanical synchronization. Left and right ventricular pre-ejection period difference was significantly longer in RVP group than in LBBP group (10.23 ± 3.07 vs 39.94 ± 14.81 ms, P < .05), indicating left and right ventricular contraction synchronization in LBBP group being better than in RVP group. CONCLUSION: LBBP is able to provide a physiologic ventricular activation pattern, which results in ventricular mechanical contraction synchronization.

Liraglutide inhibited AGEs induced coronary smooth muscle cell phenotypic transition through inhibiting the NF-κB signal pathway.

Vascular smooth muscle cell (VSMC) phenotype transition is involved in diabetes-associated cardiovascular diseases. The mechanism of VSMCs phenotypic transition in T2DM was still unclear. Rat coronary artery SMCs were pretreated with liraglutide alone, liraglutide and H89(a PKA inhibitor), neutralizing anti-RAGE antibody or the antioxidant pyrrolidine dithiocarbamate (PDTC; a nuclear factor-κB (NF-κB) inhibitor), followed by treatment with AGE. The morphological change of the SMCs was observed. We also observed the α-actin positive myofilaments and F-actin distribution in SMC through immunofluorescence microscopy. Smooth muscle myosin heavy chain 11(MYH11), α-smooth muscle actin (α-SMA) and myocardin protein expression were detected by Western blot. Collagen I productionS and NF-κB nuclear translocation were also investigated. AGEs induced a transition of SMC from contractile to synthetic phenotype, which was associated with decreased SMC differentiation markers such as α-SMA, MYH11 and myocardin by activating the NF-κB pathway. AGE also increased collagen I production and secretion by SMCs. Liraglutide inhibited AGEs induced SM phenotypic transition and down-regulation of α-SMA, MYH11 and myocardin. Liraglutide also inhibited AGEs induced NF-κB pathway activation and collagen I production. Pretreatment with liraglutide and H89 together did not exhibit this inhibitory effect as mentioned above. Blockade of RAGE in SMCs with neutralizing antibody inhibited AGEs induced phenotypic transition of SMC, and up-regulated α-SMA and MYH11 expression. Liraglutide inhibited AGE induced SMC phenotypic transition, increased SMC contractile markers expression, and decreased collagen production through down-regulation of myocardin, inhibition of NF-κB pathway, and activation of PKA signaling pathway.

Is Coronary Artery Ectasia a Thrombotic Disease?

Coronary artery ectasia (CAE) is a rare disease and a substantial portion of patients with CAE are first diagnosed with acute myocardial infarction (AMI). The question was raised if CAE was a kind of thrombotic disease. We assessed a consecutive series of 119 patients with CAE including 32 patients with AMI (CAE + AMI group) and 87 patients without AMI (CAE group). During the same period, 90 patients with coronary heart disease, 90 patients with normal coronary arteries (control), and 120 AMI patients without CAE (AMI group) were randomly selected and evaluated. Both current and previous AMI prevalence rates in the CAE population were higher than the AMI rate for the other patients undergoing coronary angiograms; the mean platelet volume and fibrinogen were increased in the CAE + AMI and CAE groups. For patients with CAE with AMI, most of the thrombotic lesions were in the ectasia site. After dividing the patients with CAE into with and without antiplatelet treatment groups before admission, the AMI rate was lower in the antiplatelet group. Platelets may participate in the thrombotic process in CAE. Antiplatelet treatment may decrease the AMI rate of patients with CAE.

AGEs impair Kv channel-mediated vasodilation of coronary arteries by activating the NF-κB signaling pathway in ZDF rats.

Excessive formation of advanced glycation end products (AGEs) impairs voltage-gated potassium (Kv) channels in rat coronary artery smooth muscle cells (CSMCs), resulting in weakened Kv-mediated coronary vasodilation. We hypothesized that induction of the nuclear factor-κB (NF-κB) signaling pathway by AGEs plays a significant role in the regulation of Kv channel-mediated vasodilation in Zucker diabetic fatty (ZDF) rats. Assays of mRNA transcripts, protein expression, and intracellular localization as well as patch-clamp experiments in cultured CSMCs revealed that AGEs significantly induced activation of the NF-κB signaling pathway, reduced Kv1.2/1.5 expression, and inhibited Kv currents. In addition, silencing of the receptor for AGEs (RAGE) or p65 with siRNA and treatment with alagrebrium (ALA) or pyrrolidine dithiocarbamate (PDTC) alleviated the AGE-induced impairment of Kv channels in CSMCs. Compared with Zucker lean (ZL) rats, the amount of AGEs, RAGE protein expression, and NF-κB activity in coronary arteries were higher in ZDF rats; whereas Kv1.2/1.5 expression was significantly lower in ZDF rats. Reduced Kv1.2/1.5 expression in coronary arteries and impaired Kv-mediated coronary relaxation tested by wire myography in ZDF rats were markedly improved by treatment with aminoguanidine (AG), ALA, or PDTC. These effects were accompanied by diminished NF-κB activity, inflammation, and oxidative stress. Taken together, these results indicate that an increased interaction between AGEs and RAGE in diabetic rats leads to impaired Kv channel-mediated coronary vasodilation. Moreover, activation of the NF-κB signaling pathway and a subsequent increase of inflammation and oxidative stress may play an important role in AGE-induced impairment of coronary vasodilation in diabetes.

Pioglitazone inhibits high glucose-induced expression of receptor for advanced glycation end products in coronary artery smooth muscle cells.

Receptor for advanced glycation end products (RAGE) is critical in inflammatory diseases, including diabetes and atherosclerosis. The mechanism underlying the effect of peroxisome proliferator­activated receptor γ (PPARγ) agonist pioglitazone (PIO) on RAGE expression in coronary artery smooth muscle cells (SMCs) stimulated by high glucose concentrations remains to be elucidated. In the present study, the effect and mechanism of action of PIO on RAGE expression in SMCs was investigated following treatment with high glucose concentrations. Rat coronary artery SMCs were pretreated with PIO alone, PIO and GW9662 (a PPARγ antagonist), diphenyleneiodonium (DPI; a nicotinamide adenine dinucleotide phosphate (NADPH) oxidase inhibitor) or the antioxidant pyrrolidine dithiocarbamate (PDTC; a nuclear factor­κB (NF­κB) inhibitor), followed by treatment with high glucose. RAGE mRNA and protein expression, reactive oxygen species (ROS) production and NF­κB nuclear translocation were investigated. Glucose induced RAGE expression in a dose­dependent manner, with maximal effect at a concentration of 25 mmol/l following treatment for 48 h. PIO, DPI and PDTC reduced high glucose­induced increases in RAGE protein and mRNA expression. PIO prominently downregulated RAGE expression and inhibited high glucose­induced increases in ROS production and NF­κB activation (P<0.05). Pretreatment with PIO and GW9662 did not exhibit this inhibitory effect. High glucose may stimulate RAGE expression in coronary artery SMCs through NADPH oxidase­mediated ROS generation and NF­κB activation. PIO downregulated RAGE expression and inhibited ROS production and NF­κB activation via PPARγ activation, which may prevent the inflammatory effect of AGE/RAGE system in diabetes.

Urotensin II-induced collagen synthesis in cultured smooth muscle cells from rat aortic media and a possible involvement of transforming growth factor-ß1/Smad2/3 signaling pathway.

BACKGROUND: Recent studies suggest that urotensin II (UII) and transforming growth factor-ß1 (TGF-ß1) both have critical roles in vascular remodeling. UII is a recently discovered vasoconstrictive peptide that is involved in the pathogenesis of atherosclerosis, restenosis and hypertension. TGF-ß1 is an important factor that has a pivotal role in vascular fibrosis. This study aimed to explore whether TGF-ß1 is involved in UII-induced collagen synthesis in rat aortic vascular smooth muscle cells (VSMCs) and examined the effects and mechanisms of UII on collagen synthesis and secretion in VSMCs. METHODS: VSMCs were prepared by the explant culture method. TGF-ß1 and collagen I secretions from the cells were determined by enzyme-linked immunosorbent assay (ELISA). The mRNA and protein expressions of TGF-ß1, collagen I, Smad2 and Smad3 were determined using Real-time RT-PCR and Western blotting. RESULTS: UII dose-dependently promoted TGF-ß1 protein expression and secretion from VSMCs, with maximal effect at 10(-8) mol/l at 24 h for protein expression and 10(-7) mol/l at 24 h for protein secretion (both P<0.01). Moreover, UII dose-dependently promoted Smad2 and Smad3 mRNA expression in VSMCs, with maximal effect at 10(-8) mol/l for 12 h (both P<0.01). The effects of UII were significantly inhibited by its receptor antagonists urantide (10(-6) mol/l) or SB-710411 (10(-6) mol/l), and by the mitogen-activated protein kinase (MAPK/ERK) inhibitor PD98059 (10(-6) mol/l). UII dose-dependently promoted collagen I mRNA expression and protein secretion in VSMCs, with maximal effect at 10(-8) mol/l at 12h for mRNA expression and 10(-6) mol/l at 24 h for protein secretion (both P<0.01). Collagen synthesis and secretion from VSMCs induced by UII were inhibited significantly by a TGF-ß1-specific neutralizing antibody, SB-431542 (an antagonist of the TGF-ß1 type II receptor) and PD98059 (all P<0.01). CONCLUSIONS: This study suggests that UII could induce collagen synthesis and secretion through upregulation of TGF-ß1 expression and secretion in VSMCs, and that TGF-ß1/Smad2/3 signaling might be one of the important pathways by which UII is involved in vascular fibrosis.

Plasma catecholamine release-inhibitory peptide catestatin in patients with essential hypertension.

BACKGROUND: Catestatin plays an important role in the adjustment of blood pressure and cardiac function. We investigated levels of plasma catestatin in essential hypertension and the relationship between catestatin and left ventricular hypertrophy. METHODS: Plasma was collected from 136 patients with essential hypertension and 61 healthy controls. Plasma catestatin was measured by enzyme-linked immunosorbent assay (ELISA). Plasma norepinephrine was measured by high-performance liquid chromatography. All patients underwent echocardiography, measurement of fasting blood glucose, body mass index (BMI) and lipid levels. RESULTS: Plasma levels of catestatin and norepinephrine were significantly higher in patients with essential hypertension than in normal controls (both P<0.01). The ratio of catestatin to norepinephrine was significantly lower in patients with essential hypertension than in normal controls (P<0.01). In patients with essential hypertension, plasma norepinephrine level was significantly higher in patients with than without left ventricular hypertrophy (P<0.01). Plasma catestatin level was lower, but not significantly, in patients with than without left ventricular hypertrophy. The ratio of catestatin to norepinephrine was significantly lower in patients with than without left ventricular hypertrophy (P<0.01). CONCLUSION: Plasma catestatin is elevated in patients with essential hypertension. The ratio of catestatin to norepinephrine was lower in patients with left ventricular hypertrophy. Catestatin might participate in the development of hypertension and left ventricular hypertrophy.