Liangrong Zheng

Serum levels of calcitonin gene-related peptide and substance P are decreased in patients with diabetes mellitus and coronary artery disease.

OBJECTIVE: This study evaluated serum levels of the neuropeptides calcitonin gene-related peptide (CGRP) and substance P (SP) in coronary artery disease (CAD) patients with and without a history of diabetes mellitus (DM). METHODS: Patients undergoing coronary angiography for suspected myocardial ischaemia were divided into four groups depending on their clinical status: control group (no CAD or DM; n = 44), DM group (DM without CAD; n = 46), CAD group (stable CAD without DM; n = 44) and DM + CAD group (stable CAD with DM; n = 50). Serum levels of CGRP and SP were determined using radioimmunoassays. RESULTS: CGRP and SP levels in the DM and CAD groups were significantly lower than in the control group. The lowest levels of CGRP and SP were observed in the DM + CAD group. There were no significant differences in CGRP and SP levels between the DM group and the CAD group. CONCLUSION: CGRP and SP may have a role in the pathogenesis of CAD in patients with diabetes.

Low dose cyclophosphamide rescues myocardial function from ischemia-reperfusion in rats.

OBJECTIVES The effect of low dose of cyclophosphamide (CP) protecting cardiac function from ischemia-reperfusion injury was studied on rats. The premise is that CP inhibits immune and inflammatory process, thereby limits I/R injury and improves myocardial function. METHODS Open chest rats were submitted to 30 min of ischemia followed by 3h, 12h or 24h of reperfusion. Rats were divided into sham group, I/R group and CP group, and each group included 3 time-point subgroups (3h, 12h and 24h; n=8 for each subgroup). A total of 750 mg/m(2) cyclophosphamide was intraperitoneally administrated in CP group and saline was given to I/R group. A polyethylene tube was inserted into the left ventricular cavity to detect left ventricular systolic pressure (LVSP) and +/-dp/dt(max). At the end of the experiment, hearts were harvested for histopathological assessment and infarct size determination. RESULTS Compared with I/R group, rats treated with low dose CP showed a significant recovery in myocardial function with improved LVSP (88+/-11 vs 69+/-11 mmHg of 3h; 92+/-11 vs 64+/-14 mmHg of 12h; 90+/-11 vs 64+/-14 mmHg of 24h; p<0.01 respectively). The +/-dp/dt(max) also had the similar trends. The myocardial infarct size was reduced in CP group compared to that in I/R group; the infiltration of polymorph nuclear leukocytes (PMNs) in myocardium was decreased in CP group. The histopathological damage score was also attenuated. CONCLUSIONS These findings suggest that low dose CP rescues cardiac function from ischemia-reperfusion injury by alleviating histopathological damage in rat myocardium.

Captopril administered at night restores the diurnal blood pessure rhythm in adequately controlled, nondipping hypertensives

The aim of our study was to evaluate whether captopril administered at night, can shift the circadian blood pressure (BP) from a nondipper to a dipper pattern in adequately controlled hypertensive patients, who continued their antihypertensive therapy. In a prospective, randomized, double blind, placebo-controlled designed study, we enrolled 121 treated, adequately controlled nondipping hypertensive patients. All patients were randomly assigned to 12.5 mg captopril or placebo treatment administered at night. In case of nondippers, the dosage of captopril or placebo was doubled after two weeks of treatment, while for dippers antihypertensive regimens were not changed. After another two weeks, all patients underwent ambulatory BP monitoring. Our results show that at the end of the active treatment period, the prevalence of a dipping diurnal BP pattern in the captopril group (70%) was significantly higher than that in the placebo group (9.8%, P < 0.001). Nighttime BP, night/day BP ratio, nighttime BP load and 24-h systolic BP were significantly lower after 4 weeks nighttime captopril treatment compared to baseline. In conclusion, the present study demonstrates for the first time that captopril administered at night can restore the diurnal BP rhythm and decrease the elevated night/day BP ratio in appropriately controlled, nondipper hypertensive patients. These results were mainly due to the decrease of nighttime BP.

TRPV1 activation exacerbates hypoxia/reoxygenation-induced apoptosis in H9C2 cells via calcium overload and mitochondrial dysfunction.

Transient potential receptor vanilloid 1 (TRPV1) channels, which are expressed on sensory neurons, elicit cardioprotective effects during ischemia reperfusion injury by stimulating the release of neuropeptides, namely calcitonin gene-related peptide (CGRP) and substance P (SP). Recent studies show that TRPV1 channels are also expressed on cardiomyocytes and can exacerbate air pollutant-induced apoptosis. However, whether these channels present on cardiomyocytes directly modulate cell death and survival pathways during hypoxia/reoxygenation (H/R) injury remains unclear. In the present study, we investigated the role of TRPV1 in H/R induced apoptosis of H9C2 cardiomyocytes. We demonstrated that TRPV1 was indeed expressed in H9C2 cells, and activated by H/R injury. Although neuropeptide release caused by TRPV1 activation on sensory neurons elicits a cardioprotective effect, we found that capsaicin (CAP; a TRPV1 agonist) treatment of H9C2 cells paradoxically enhanced the level of apoptosis by increasing intracellular calcium and mitochondrial superoxide levels, attenuating mitochondrial membrane potential, and inhibiting mitochondrial biogenesis (measured by the expression of ATP synthase β). In contrast, treatment of cells with capsazepine (CPZ; a TRPV1 antagonist) or TRPV1 siRNA attenuated H/R induced-apoptosis. Furthermore, CAP and CPZ treatment revealed a similar effect on cell viability and mitochondrial superoxide production in primary cardiomyocytes. Finally, using both CGRP8–37 (a CGRP receptor antagonist) and RP67580 (a SP receptor antagonist) to exclude the confounding effects of neuropeptides, we confirmed aforementioned detrimental effects as TRPV1−/− mouse hearts exhibited improved cardiac function during ischemia/reperfusion. In summary, direct activation of TRPV1 in myocytes exacerbates H/R-induced apoptosis, likely through calcium overload and associated mitochondrial dysfunction. Our study provides a novel understanding of the role of myocyte TRPV1 channels in ischemia/reperfusion injury that sharply contrasts with its known extracardiac neuronal effects.

Nerve growth factor rescues diabetic mice heart after ischemia/reperfusion injury via up-regulation of the TRPV1 receptor.

AIMS Nerve growth factor (NGF), a member of the neurotrophin family, plays an essential role in diabetic neuropathy and ischemic heart disease. In the present study, we explored the potential role of NGF and the involvement of TRPV1 receptor in isolated diabetic mouse hearts following ischemia/reperfusion (I/R) injury. METHODS Adenovirus-mediated NGF gene delivery was performed on diabetic and sham hearts 8weeks after streptozotocin treatment. The sciatic nerve conduction velocity was recorded using a biological signal acquisition system. Forty-eight hours after heart surgery, mice were subjected to I/R injury using a Langendorff system. Several cardiac parameters and the expression of associated molecules were analyzed during the experiment. RESULTS The sciatic nerve conduction velocity was reduced in diabetic mice compared with that in control mice. Decreased expression of NGF, TRPV1, and the downstream neurotransmitters CGRP and SP was observed in the diabetic hearts. Adenovirus-mediated NGF overexpression reversed the reduction in TRPV1 and downstream neuropeptides, resulting in improved cardiac recovery post-I/R injury in diabetic hearts. The protective effect of NGF was abolished by CGRP8-37 (a selective CGRP antagonist), while it was preserved by low-dose capsaicin. CONCLUSIONS The NGF-induced up-regulation of TRPV1 via the increased synthesis and release of endogenous CGRP leads to improved cardiac performance in I/R-injured diabetic heart.

Up-regulation of calcitonin gene-related peptide protects streptozotocin-induced diabetic hearts from ischemia/reperfusion injury

BACKGROUND Diabetic hearts are vulnerable to ischemia/reperfusion (I/R) injury. Pretreatment with exogenous calcitonin gene-related peptide (CGRP) exerts a cardioprotective effect against myocardial I/R injury. Our previous study found that the CGRP level was decreased in diabetic hearts. This study aimed to investigate whether up-regulation of CGRP could reduce I/R injury in diabetic hearts. METHODS AND RESULTS Adenovirus encoding the CGRP gene (Ad-CGRP) was injected intramyocardially in mice with or without streptozotocin (STZ) treatment. Three days after injection, the hearts were subjected to in vivo and in vitro I/R. Myocardial infarct size, cardiac function, lactate dehydrogenase (LDH) level in plasma and effluents, and cell mitochondrial function were measured. After ischemia (30 min) and reperfusion (24h) in vivo, diabetes mellitus (DM) mice had greater myocardial infarct size than their nondiabetic counterparts, and released more LDH in plasma. However, CGRP gene transfer reduced myocardial infarct size and plasma LDH level in both non-DM and DM hearts. After 30 min global ischemia and 40 min reperfusion in vitro, the DM hearts demonstrated increased left ventricular end-diastolic pressure (LVEDP) and effluent LDH level, and decreased left ventricular developed pressure (LVDP), coronary flow (CF), as well as cell mitochondrial function, when compared with the non-DM hearts. Again, CGRP gene transfer could protect against I/R injury in both non-DM and DM hearts. CONCLUSIONS Adenovirus-mediated up-regulation of CGRP gene expression protects diabetic hearts against I/R injury.

Lovastatin Inhibits Gap Junctional Communication in Cultured Aortic Smooth Muscle Cells

Background: Gap junctions, which serve as intercellular channels that allow the passage of ions and other small molecules between neighboring cells, play an important role in vital functions, including the regulation of cell growth, differentiation, and development. Statins, the 3-hydroxy-3-methylglutaryl-coenzymeA (HMG-CoA) reductase inhibitors, have been shown to inhibit the migration and proliferation of smooth muscle cells (SMCs) leading to an antiproliferative effect. Recent studies have shown that statins can reduce gap junction protein connexin43 (Cx43) expression both in vivo and in vitro. However, little work has been done on the effects of statins on gap junctional intercellular communication (GJIC). We hypothesized in this study that lovastatin inhibits vascular smooth muscle cells (VSMCs) migration through the inhibition of the GJIC. Methods: Rat aortic SMCs (RASMCs) were exposed to lovastatin. Vascular smooth muscle cells migration was then assessed with a Transwell migration assay. Gap junctional intercellular communication was determined by using fluorescence recovery after photobleaching (FRAP) analysis, which was performed with a laser-scanning confocal microscope. Results: The migration of the cultured RASMCs were detected by Transwell system. Cell migration was dose-dependently inhibited with lovastatin. Compared with that in the control (110 ± 26), the number of migrated SMCs was significantly reduced to 72 ± 24 (P < .05), 62 ± 18 (P < .01), and 58 ± 19 (P < .01) at the concentration of 0.4, 2, and 10 umol/L, per field. The rate of fluorescence recovery (R) at 5 minutes after photobleaching was adopted as the functional index of GJIC. The R- value of cells exposed to lovastatin 10 umol/L for 48 hours was 24.38% ± 4.84%, whereas the cells in the control group had an R- value of 36.11% ± 10.53%, demonstrating that the GJIC of RASMCs was significantly inhibited by lovastatin (P < .01). Smaller concentrations of lovastatin 0.08 umol/L did not change gap junction coupling (P > .05). Conclusions: These results suggest that lovastatin inhibits migration in a dose-dependent manner by attenuating JIC. Suppression of gap junction function could add another explanation of statin-induced antiproliferative effect.

Paeoniflorin protects diabetic mice against myocardial ischemic injury via the transient receptor potential vanilloid 1/calcitonin gene-related peptide pathway

BackgroundDiabetes mellitus has multiple complications including neuropathy and increases cardiovascular events. Paeoniflorin (PF), a monoterpene glycoside, plays an essential role in neuroprotection and ischemic heart disease. In this study, we aimed to investigate the hypothesis that PF protects mice with diabetes mellitus against myocardial ischemic injury, and determine its associated mechanisms.ResultsMyocardial infarction (MI) was generated in the streptozotocin-mediated diabetic mice, which were pretreated with either vehicle or PF, respectively. Myocardial infarct size, myocardial enzyme, cardiac function, circulating calcitonin gene-related peptide (CGRP) concentration, histological analysis and the expression of associated molecules were determined and compared among different experimental groups. Compared to diabetic hearts pretreated with vehicle, hearts pretreated with PF exhibited less tissue damage and better CGRP concentration in serum when subjected to myocardial ischemia. Transient receptor potential vanilloid 1(TRPV1) gene knockout attenuated PF-mediated cardioprotection. Moreover, a specific Ca2+/calmodulin-dependent protein kinase (CaMK) inhibitor, KN-93, increased tissue damage and decreased CGRP activity in serum. Meanwhile, pretreated with PF increased the phosphorylation of cAMP response element binding protein (CREB).ConclusionsTaken together, these findings demonstrate that PF protects diabetic mice against MI at least partially via the TRPV1/CaMK/CREB/CGRP signaling pathway.

Cross-talk between macrophages and atrial myocytes in atrial fibrillation

Increased macrophage accumulation occurs in the atria of patients with atrial fibrillation (AF). However, the phenotype and functions of the macrophages in AF remain unclear. We investigated the macrophage-atrial myocyte interaction in AF patients and found that the increased macrophages were mainly pro-inflammatory macrophages (iNOS+, Arg1−). Tachypacing of HL-1 atrial myocytes also led to pro-inflammatory macrophage polarization. In addition, lipopolysaccharide (LPS)-stimulated pro-inflammatory macrophages-induced atrial electrical remodeling, evidenced by increased AF incidence and decreased atrial effective refractory period and L-type calcium currents (ICa-L) in both canine and mouse AF models. Depletion of macrophages relieved LPS-induced atrial electrical remodeling, confirming the role of pro-inflammatory macrophages in the pathogenesis of AF. We also found that the effect of LPS-stimulated macrophages on atrial myocytes was mediated by secretion of interleukin 1 beta (IL-1β), which inhibited atrial myocyte quaking protein (QKI) expression. IL-1β knockout in macrophages restored the LPS-stimulated macrophage-induced inhibition of QKI and CACNA1C (α1C subunit of L-type calcium channel) in atrial myocytes. Meanwhile, QKI overexpression in atrial myocytes restored the LPS-stimulated macrophage-induced electrical remodeling through enhanced binding of QKI to CACNA1C mRNA, which upregulated the expression of CACNA1C as well as ICa-L. In contrast, QKI knockout inhibited CACNA1C expression. Finally, using transcription factor activation profiling plate array and chromatin immunoprecipitation, we revealed that special AT-rich sequence binding protein 1 activated QKI transcription. Taken together, our study uncovered the functional interaction between macrophages and atrial myocytes in AF. AF induced pro-inflammatory macrophage polarization while pro-inflammatory macrophages exacerbated atrial electrical remodeling by secreting IL-1β, further inhibiting QKI expression in atrial myocytes, which contributed to ICa-L downregulation. Our study demonstrates a novel molecular mechanism underlying the pathogenesis and progression of AF and suggests that QKI is a potential therapeutic target.

Effects of diltiazem on platelet activation and cytosolic calcium during percutaneous transluminal coronary angioplasty

Aims: To evaluate effects of diltiazem on platelet hyper-reactivity in situations associated with endothelial injury and their possible relationship to cytosolic calcium concentration. Methods: Blood samples were collected at seven time points from 35 patients undergoing percutaneous transluminal coronary angioplasty (PTCA) who received combined diltiazem and aspirin/ticlopidine therapy or aspirin/ticlopidine therapy alone. Platelet expression of glycoprotein IIb/IIIa and P-selectin, production of thromboxane B2, and cytosolic calcium concentration were measured, respectively, by whole blood flow cytometry, radioimmunoassay, and fluorospectrophotometry. The effects of diltiazem of different concentrations on expression of glycoprotein IIb/IIIa and P-selectin were also studied in vitro in blood samples from patients with chronic stable angina. Results: Of the two treatments, aspirin/ticlopidine therapy did not prevent an acute increase of expression of glycoprotein IIb/IIIa and P-selectin and plasma thromboxane B2 five minutes and 10 minutes after first inflation and 10 minutes after PTCA, whereas combined diltiazem and aspirin/ticlopidine therapy had a significant inhibitory effect. In the group receiving aspirin/ticlopidine therapy, there was a short term increase of platelet [Ca2+]i immediately after PTCA which was significantly reduced by diltiazem treatment. Expression of glycoprotein IIb/IIIa and P-selectin was significantly inhibited in vitro by diltiazem in the concentration of 200 ng/ml or higher, but not 50 ng/ml. Conclusions: Combined diltiazem and aspirin/ticlopidine therapy significantly inhibited platelet activation that continued in the presence of conventional aspirin/ticlopidine treatment. Antiplatelet effects of diltiazem were probably a consequence of reduction of platelet [Ca2+]i and may only be achieved in higher than therapeutic concentrations.