Wan Joo Shim

Gene expression profiling of oxidative stress on atrial fibrillation in humans

Atrial Fibrillation (AF) is thought be caused by oxidative stress. Oxidative stress at the cellular level results from many factors, including exposure to alcohol, medications, cold, toxins or radiation. In this study we investigated gene transcriptional profiles on the human myocardial tissues from AF and oxidative stress conditions. Right atrial appendages were obtained from AF patients (n = 26) undergoing the Maze procedure, and from control patients (n = 26) who were in normal sinus rhythm and undergoing coronary artery bypass graft operation. To examine the effects of oxidative stress on AF, we used radioactive complementary DNA (cDNA) microarrays to evaluate changes in the expression of 1,152 known genes. This technology, which monitors thousands of genes simultaneously, gives us a better picture of the interactions between AF and oxidative stress. Total RNAs prepared from the retrieved tissues were used to synthesize (33)P-labeled cDNAs by reverse transcription and hybridized to cDNA microarrays. Gene expression profiles showed that 30 genes were upregulated and 25 were downregulated in AF patients compared with control patients. Moreover, comparison rank analysis revealed that the expression of five genes related to reactive oxygen species (ROS)-including flavin containing monooxygenase 1, monoamine oxidase B, ubiquitin specific protease 8, tyrosinase-related protein 1, and tyrosine 3-monooxygenase-increased by more than 2.0 of the Z-ratio, and two genes related to anti-oxidants including glutathione peroxidase 1, and heme oxygenase 2-decreased to the Z-ratio levels of <= -2.0. Apparently, a balanced regulation of pro- and anti-oxidation can be shifted toward pro-oxidation and can result in serious damage similar to that of human AF. Western blotting analysis confirmed the upregulation of tyrosinase-related protein 1 and tyrosine 3-monooxygenase and the downregulation of heme oxygenase 2. These results suggested that the gene expression pattern of myocardial tissues in AF patients can be associated with oxidative stress, resulting in a significant increase in ROS. Thus, the cDNA microarray technique was useful for investigating transcription profiles in AF. It showed that the intracellular mechanism of oxidative stress plays a pivotal role in the pathologic progression of AF and offers novel insight into potential treatment with antioxidants.

Action potential duration restitution kinetics in human atrial fibrillation

OBJECTIVES We undertook this study to determine whether human atrial fibrillation (AF) relates to steeply sloped action potential duration restitution (APDR) kinetics and whether the spatial nonuniformity of APDR promotes persistence of AF. BACKGROUND A steeply sloped APDR curve is known to be an important determinant of the induction of more complex action potential duration (APD) dynamics and fibrillation. METHODS Patients with chronic atrial fibrillation (CAF) (n = 18), paroxysmal atrial fibrillation (PAF) (n = 14) and normal control subjects (n = 9) were studied. The monophasic action potential duration at 90% repolarization (APD(90)) and the effective refractory period (ERP) were measured at six sites in the right atrium. After AF was electrically converted, APDR was assessed by delivering a single extrastimulus after a train of stimuli at a cycle length of 600 ms (S(1)S(2)) at six different sites of the right atrium, as well as rapid pacing at cycle lengths that induced APD alternans. RESULTS The APD(90) and ERP in patients with CAF were shorter than those in patients with PAF and control subjects (p < 0.05); however, the dispersions of APD(90) and ERP in each group were similar. The maximal slopes of APDR by S(1)S(2) and rapid pacing in patients with CAF (1.2 +/- 0.4 and 1.7 +/- 0.2) and PAF (1.1 +/- 0.4 and 1.3 +/- 0.4) were higher than those in control subjects (0.5 +/- 0.3 and 0.8 +/- 0.2, respectively; p < 0.01). The maximal slope obtained by S(1)S(2) did not differ from that obtained by rapid pacing in any group. The inter-regional difference of the maximal slope in patients with CAF (1.6 +/- 0.4, p < 0.05) was greater than that in patients with PAF (1.2 +/- 0.3, p = NS vs. control) and control subjects (0.4 +/- 0.2). CONCLUSIONS Atrial fibrillation was related to steeply sloped (>1) APDR kinetics. The spatial dispersion of APDR in patients with chronic AF was greater than that of patients with paroxysmal AF and control subjects, indicating that the heterogeneity of APDR of the atrium plays an important role in the persistence of AF.

5-azacytidine induces cardiac differentiation of P19 embryonic stem cells.

The P19 embryonal carcinoma cell line is a useful model cells for studies on cardiac differentiation. However, its low efficacy of differentiation hampers its usefulness. We investigated the effect of 5-azacytidine (5-aza) on P19 cells to differentiate into a high-efficacy cardiomyocytes. Embryoid-body-like structures were formed after 6 days with 1 µM of 5-aza in a P19 cell monolayer culture, beating cell clusters first observed on day 12, and, the production of beating cell clusters increased by 80.1% (29 of 36-wells) after 18 days. In comparison, the spontaneous beating cells was 33.3% (12 of 36-wells) for the untreated control cells. In response to 1 µM of 5-aza, P19 cells expressed bone morphogenetic protein-2 (BMP-2), BMP-4, Bmpr1a and Smad1 at day 6 or 9, and also cardiac markers such as GATA-4, Nkx2.5, cardiac troponin I, and desmin were up-regulated in a time-dependent manner after induction of BMP signaling molecules. Immunocytochemistry revealed the expression of smooth muscle a-actin, sarcomeric a-actinin, cardiac myosin heavy chain, cardiac troponin T and desmin, respectively. The proportion of sarcomeric a-actinin positive cells accounted for 6.48% on day 15 after 5-aza exposure as measured by flow cytometry. This study has demonstrated that 5-aza induces differentiation of P19 cells into cardiomyocytes in a confluent monolayer culture in the absence of prior embryoid formation and dimethyl sulfoxide exposure, depending in part on alteration of BMP signaling molecules. These results suggest that 5-aza treatment could be used as a new method for cardiac differentiation in P19 cells.

Fibroblast growth factor-2 and -4 promote the proliferation of bone marrow mesenchymal stem cells by the activation of the PI3K-Akt and ERK1/2 signaling pathways.

Bone marrow mesenchymal stem cells (BMMSCs) have the capacity for self-renewal, and differentiation into a variety of cell types. They thus represent an attractive source of material for cell therapy. However, little is known about the mechanisms underlying the proliferation of BMMSCs. The purpose of this study was to identify the factors and signaling pathways involved in the proliferation of stem cell antigen-1(+) (Sca-1(+)) BMMSCs. Among the cytokines and growth factors examined in this study, fibroblast growth factor-2 (FGF-2) and FGF-4 significantly stimulated the proliferation of Sca-1(+) BMMSCs, as determined by bromodeoxyuridine incorporation. PI3K-Akt, ERK1/2, and JAK/STAT3 pathways were investigated after stimulation with FGF-2 or FGF-4 via Western blot analysis. No changes were observed in the total ERK1/2 and Akt; however, the pERK1/2 and pAkt levels were upregulated early within 15 min in the FGF-2- or FGF-4-treated Sca-1(+) BMMSCs. Moreover, the pERK1/2 and pAkt upregulation induced by FGF-2 and -4 were completely abolished by treatment with the MEK1/2 inhibitor, U0126 and the PI3K inhibitor, LY294002. However, no change in pJAK2 or total JAK2 levels was observed in the Sca-1(+) BMMSCs induced by FGF-2 or FGF-4. As a consequence of PI3K-Akt and ERK1/2, the upregulation of c-Jun in the Sca-1(+) BMMSCs, after stimulation with FGF-2 or FGF-4, was observed after 12 and 24 h. Moreover, the activation of c-Jun in FGF-2- and FGF-4-treated Sca-1(+) BMMSCs was significantly reduced by U0126. Taken together, these data suggest that FGF-2 and -4 promote the proliferation of Sca-1(+) BMMSCs by activation of the ERK1/2 and PI3K-Akt signaling pathways.

Multiple predictors of coronary restenosis after drug-eluting stent implantation in patients with diabetes

Objectives: To identify parameters influencing the likelihood of restenosis after implantation of drug-eluting stents (DES) in patients with diabetes. Methods: Stented patients (n  =  840) with DES were retrospectively reviewed for inclusion in the study from the Multicenter PCI Database Registry. From this database, 211 (25.1%) of 840 patients with six-month angiographic follow up had diabetes. Predictors of coronary restenosis were identified with univariate and multivariate logistic regression analyses. Results: Restenosis occurred in 92 of 629 (14.6%) patients without diabetes and in 44 (20.9%) of 211 patients with diabetes (p < 0.001). Multivariate parameters for predicting restenosis in the diabetic group were current smoking (odds ratio (OR) 1.923, 95% confidence interval (CI) 1.055 to 4.725, p  =  0.036), higher C reactive protein concentration (OR 1.031, 95% CI 1.011 to 1.075, p  =  0.043), use of the paclitaxel-eluting stent (OR 2.638, 95% CI 1.338 to 5.200, p  =  0.005), longer stent length (OR 1.065, 95% CI 1.021 to 1.119, p  =  0.033), smaller reference diameter before DES implantation (OR 0.501, 95% CI 0.110 to 0.965, p  =  0.040), smaller reference diameter (OR 0.455, 95% CI 0.120 to 0.814, p  =  0.026) and minimum lumen diameter (OR 0.447, 95% CI 0.068 to 0.876, p  =  0.039) after DES implantation. Conclusion: Even with the introduction of DES, diabetes remains a significant predictor of coronary restenosis, especially in cases of a small baseline vessel size, small vessel size after percutaneous coronary intervention, longer stent length, use of the paclitaxel-eluting stent, current smoking and high C reactive protein concentration.

Percutaneous coronary intervention with drug-eluting stent implantation vs. minimally invasive direct coronary artery bypass (MIDCAB) in patients with left anterior descending coronary artery stenosis

The aim of this study was to assess the effects of percutaneous coronary intervention with drug‐eluting stents (DESs) versus minimally invasive direct coronary artery bypass (MIDCAB) surgery in the management of patients with proximal left anterior descending (LAD) coronary artery stenosis. Until recent years, despite the advantages of percutaneous transluminal coronary angioplasty (PTCA) with bare metal stent implantation, such as shorter hospital stays and recovery time, MIDCAB showed better results with regard to the need for repeated intervention in the target vessel than PTCA with proximal LAD lesions. Symptomatic patients (n = 189) were randomly assigned to DES group (n = 119) and MIDCAB group (n = 70). Patients with an isolated high‐grade lesion (stenosis of ≥ 70% of the luminal diameter) in the proximal LAD coronary artery (from the ostium to the first diagonal branch) were included in this study. During the 6‐month follow‐up period, 1.7% (n = 2) in the DES group needed repeated revascularization procedures for target lesion revascularization compared with 5.9% (n = 4) in the MIDCAB group (P = 0.196). The rates of death and myocardial infarction were similar in both groups [DES 0.0% (n = 0) vs. MIDCAB 2.9% (n = 2), P = 0.135; DES 1.7% (n = 2) vs. MIDCAB 2.9% (n = 2), P = 0.627; respectively] during 6 months of follow‐up. In‐hospital length of stay was significantly shorter in the DES group compared with the MIDCAB group (5.8 ± 2.1 days vs. 8.9 ± 2.6 days; P = 0.001). DES implantation and MIDCAB surgery showed similar rates of myocardial infarction, the need for repeated revascularization, and death during 6 months of follow‐up. However, DES implantation resulted in lower average number of hospital stays and similar postoperative complications. Catheter Cardiovasc Interv 2005;64:75–81.

Differentiation, engraftment and functional effects of pre-treated mesenchymal stem cells in a rat myocardial infarct model.

Background — Mesenchymal stem cells (MSCs) offer a novel therapeutic option in the treatment of acute myocardial infarction. MSCs are able to differentiate into myogenic cells after 5-azacytitdine treatment. However, 5-azacytidine might have genotoxic effects. Recently, it was reported that combined treatment with bone morphogenetic protein-2(BMP-2) and fibroblast growth factor-4(FGF-4) caused cardiac differentiation in non-precardiac mesoderm explants. Therefore, we investigated whether MSCs treated with combined BMP-2 and FGF-4 showed evidence of myogenic differentiation in vitro, and whether these cells resulted in sustained engraftment, myogenic differentiation, and improved cardiac function after implantation in infarcted myocardium. Methods and results — In vitro study: MSCs were treated with BMP-2 + FGF-4 (GF-MSCs) and myogenic phenotype was evaluated immunohistochemically. Cell growth curve was used to compare MSC proliferative capacity between the growth factors and 5-azacytidine treatments. In vivo study: two weeks after coronary artery occlusion, GF-MSCs (n = 15), MSCs (n = 5) labelled with PKH26 were injected into infarcted myocardium. Control animals (n = 5) received a culture medium into the infarcted myocardium.Two weeks after implantation, some engrafted GF-MSCs or MSCs expressed sarcomeric-a-actinin and cardiac myosin heavy chain, as was observed in culture. Echocardiography showed that the GF-MSC group had a better (p < 0.05) left ventricular performance than the other groups. Conclusion — GF-MSCs induced myogenic differentiation in vitro. Moreover, GF-MSCs engrafted into the infarcted myocardium increased myogenic differentiation, prevented dilation of the infarcted region, and eventually improved heart function.

Impact of obstructive sleep apnea on left ventricular diastolic function.

The aim of this study was to investigate the impact of obstructive sleep apnea (OSA) on left ventricular (LV) functional changes by using tissue Doppler imaging-derived indexes in patients with OSA. We studied 62 patients classified into 3 groups, namely 18 with mild to moderate OSA, 24 with severe OSA, and 20 control subjects without OSA according to the apnea-hypopnea index (AHI) on complete overnight polysomnogram. All underwent conventional and tissue Doppler echocardiographies. Only early diastolic velocity (Ea; -6.2 +/- 0.3 vs -7.1 +/- 0.3 vs -7.3 +/- 0.3 cm/s, respectively, for the 3 groups, p = 0.023) was significantly decreased in the severe OSA group. Other echocardiographic parameters of diastolic function such as isovolumic relaxation time, deceleration time, mitral inflow early/late wave velocity ratio, and pulmonary vein systolic/diastolic pulmonary vein velocity ratio were comparable among the 3 groups. AHI was correlated only with tissue Doppler imaging-derived indexes of LV diastolic function (Ea r = -0.382, p = 0.002; Ea/late diastolic velocity r = -0.329, p = 0.009), but not with conventional Doppler indexes. AHI remained a significant predictor of Ea after adjusting for age, heart rate, fasting glucose level, blood pressure, body mass index, and LV mass index in a multiple stepwise linear regression model (p = 0.007). In conclusion, only patients with severe OSA showed a greater impairment of LV diastolic function. Of all echocardiographic parameters of diastolic dysfunction investigated, only Ea was identified as the best index to demonstrate an association between LV diastolic dysfunction and severity of OSA independently of body mass index, diabetes mellitus, and hypertension.

Left Atrial Electromechanical Conduction Time Can Predict Six-Month Maintenance of Sinus Rhythm After Electrical Cardioversion in Persistent Atrial Fibrillation by Doppler Tissue Echocardiography

BACKGROUND The purpose of this study was to determine whether atrial electromechanical conduction time (EMT) measured by echocardiography can predict 6-month maintenance of sinus rhythm (SR) after electrical cardioversion in patients with atrial fibrillation (AF). METHODS Fifty-three patients with persistent AF (>1 month) who had successful cardioversion and 30 controls with SR were prospectively enrolled. SR maintenance was assessed during 6-month follow-up. EMT was measured as the time interval from the onset of the P wave on electrocardiography to the peak of the late diastolic wave from the septal and lateral mitral annulus (EMT-S and EMT-L, respectively) and the lateral tricuspid annulus (EMT-T) on tissue Doppler echocardiography. RESULTS Compared with controls, left atrial (LA) volume index, P-wave duration, and EMT were significantly larger in patients with AF (all P values<.001). In patients with AF, the duration of AF (P=.71) and P-wave duration (P=.24) were not different between the SR maintenance group (n=23) and the AF recurrence group (n=30), and there was a trend toward increased LA volume index in the AF recurrence group (47.0+/-12.4 vs 45.3+/-12.6 mL/m2, P=.07). EMT-S and EMT-L were significantly larger in the AF recurrence group (131.4+/-20.9 vs 116.3+/-15.5 ms, P=.005, and 152.2+/-15.7 vs 128.9+/-13.8 ms, P<.001, respectively), but not EMT-T. EMT-S and EMT-L were related to LA volume index (r=.36, P=.008, and r=.33, P=.02, respectively). On multivariate logistic regression analysis, only EMT-L was an independent predictor of identifying patients who remained in SR (P<.001), and the sensitivity and specificity for the prediction of 6-month maintenance of restored SR were 82.6% and 83.3% using a cutoff value of EMT-L<or=138.0 ms (odds ratio, 0.862; 95% confidence interval, 0.788-0.942; P=.001). CONCLUSION LA EMT was significantly prolonged in patients with recurring AF, indicating significantly depressed atrial conduction in enlarged LA, and can predict 6-month maintenance of SR after electrical cardioversion.

Epicardial ablation of ventricular tachycardia associated with isolated ventricular noncompaction.

A 52‐year‐old man presented with sudden onset of palpitations and dizziness. Echocardiogram confirmed the diagnosis of isolated noncompaction of ventricular myocardium with moderated systolic dysfunction, and the electrocardiogram (ECG) revealed ventricular tachycardia (VT), of which the focus seemed to match an area of prominent left ventricular noncompaction on the 12‐lead surface ECG. Through the activation mapping from the endo‐ and epicardium, simultaneously, a discrete potential preceding the QRS during VT was observed at the anterolateral epicardial wall. He subsequently underwent radiofrequency ablation, and VT was successfully eliminated.