2004年 · 第117卷第10期

Background The infarct size determines the long-term prognosis of patients with acute myocardial infarction (AMI). There is a growing interest in repairing scar area by transplanting bone marrow stem cells. However, effectiveness of intracoronary injection of bone marrow mesenchymal stem cells (BMSCs) in patients with AMI still remains unclear. Methods Sixty-nine patients with AMI after percutaneous coronary intervention (PCI) were randomly divided into intracoronary injection of BMSCs (n = 34) and saline (control group, n = 35) groups. Serial single positron emission computer tomography (SPECT), cardiac echo and cardiac electromechanical mapping were done at the designed time intervals until six months after transplantation of BMSCs or injection of saline. Results The proportion with functional defect decreased significantly in the BMSCs patients after three months [ (13 ±5) %] compared with that pre-transplantation [ (32 ±11) %] and the control group [ (28 ± 10) %] at three month follow-up (P < 0. 05, respectively). Wall movement velocity over the infracted region increased significantly in the BMSCs group [ (4. 2 ±2. 5) cm/ s vs (2. 2 ±1. 3) cm/s, P < 0. 05 ], but not in the control group [ (2. 2 ±1. 5) cm/ s vs (2. 7 ±1. 7) cm/ s, P > 0. 05 ]. Left ventricular ejection fraction (LVEF) three months after transplantation in BMSCs group increased significantly compared with that pre-implantation and with that of the control group at three months post-injection [ (67 ±11) % vs (49 ± 9) % and (53 ±8) %, P < 0. 05 respectively ]. SPECT scan results showed that perfusion defect was improved significantly in BMSCs group at three-month follow-up compared with that in the control group [ (134 ±66) cm2 vs (185 ±87) cm2, P <0. 01]. At the same time, left ventricular end-diastolic volume [(136 ±31) ml vs (162 ±27) ml, P < 0.05] and end-systolic volume [(63 ±20) ml vs (88 ±19) ml, P < 0.05] decreased synchronously. The ratio of end-systolic pressure to end-systolic volume [Psyst/ESV, (2. 84 ±1.30) mmHg/ ml vs (1.72 ±1.23) mmHg/ ml, P < 0.05 ] increased significantly. Cardiac electromechnical mapping demonstrated significant improvement at three months after implantation of BMSCs compared with that pre-injection in both cardiac mechanical capability as left line local shorting [LLS, (11.29 ±1.64) % vs (7.32 ± 1.86)%, P < 0. 05] and electrical property as left ventricular endocardial unipolar voltage [UV, (10.38 ±1.12) mV vs (7.61 ±1.09) mV, P < 0. 01 ]; perfusion defect decreased from (36. 2 ± 6. 2) % to (20. 3 ±5.31) % (P < 0.01). Twenty-four-hour electrocardiographic monitoring demonstrated no arrhythmias occurred at three-months follow-up. Conclusions The transplantation of BMSCs might improve the cardiac function and it is safe and feasible with no deaths or malignant arrhythmias. Chin Med J 2004; 117(10):1443-1448

Background Autoimmune mechanisms are likely to participate in the pathogenesis of subgroup of idiopathic dilated cardiomyopathy (IDC), and components of the major histocompatibility complex may serve as markers for the propensity to develop immune-mediated myocardial damage. Human leukocyte antigen (HLA) class Ⅱ genes, especially highly polymorphic HLA-DQ genes, play an important role in the activation of immune responses, and thus control the predisposition for or protect from IDC. This study was conducted to investigate the HLA-DQA1 allele polymorphisms in IDC patients and to explore the underlying immunological mechanism and the hereditary susceptibility to IDC. Methods The polymerase chain reaction sequence-specific primers (PCR-SSP) technique was used to analyze the polymorphisms in the second exon of DQA1 in three groups: 72 IDC patients diagnosed according to the criteria of World Health Organization (IDC group); 100 end-stage heart failure patients suffering from a disease of known etiology (HF group); and 100 healthy subjects enrolled for the study during a routine health survey (control group). Patients in the IDC group were stratified according to ejection fraction (EF). Those with EF values were higher than 35 % were placed into subgroup 1; subgroup 2 included patients with an EF value of 15% - 35%; and subgroup 3 consisted of those whose EF values less than 15%. Results The frequency of HLA-DQA1* 0501 alleles was significantly higher in the IDC group (0. 39) than that in the HF group (0. 12) and the control group (0. 09) (both P < 0. 05). Further analysis of the three IDC subgroups showed a higher frequency of DQA1* 0501 among patients with lower EF values (both P < 0. 05, compared with subgroups 1 and 2). The frequency of DQA1*0201 was higher in the control group than that in the IDC group (P<0.05). Conclusions The HLA-DQA1* 0501 allele confers susceptibility to IDC, while the DQA1* 0201 allele confers protection against it, which indicates that genetic background involved in IDC and heart failure is different. HLA-DQA1 genes are involved in the regulation of specific immune responses by auto- or foreign anti-myocardium antibody. Chin Med J 2004; 117(10):1449-1452

Background Time-intensity curves derived from microbubble destruction/refilling sequences and recorded using myocardial contrast echocardiography (MCE) can provide parameters that correlate with coronary blood flow. The response of these parameters to adenosine vasodilatation correlates with coronary flow reserve (CFR) measured by fluorescent microsphere techniques (FMT). Currently, no data exist regarding the effect of physiological variables, such as hypoxia, on the determination of CFR by MCE The purpose of this study was to define the effects of decreases in blood partial pressure of oxygen (PO2) on CFR as measured by MCE. Methods Studies were performed in 9 closed chest swine. Low-energy, real-time MCE was performed with commercial instruments in short axis view at papillary muscle level while infusing BR1 at 30 ml/ h. High-energy ultrasound bursts (referred to as FLASH frames) destroyed the bubbles every 15 cardiac cycles, and resultant time-intensity curves derived from these sequences were fitted to the exponential function y = A (1-e-bt) +c, from which the rate of signal rise (b) was obtained. CFR was calculated as the ratio of b values after adenosine infusion to baseline and was obtained during the control period and after decreasing blood PO2 by giving nitrogen via a respirator to create artificial hypoxic conditions. CFR was independently determined by FMT. Results Nitrogen led to significant decreases in mean PO2, from (120. 6 ±18. 9) mmHg to (51. 8 ±15. 9) mmHg (P <0. 01). Adenosine produced a similar increase in CFR (2. 5 fold vs 3. 1 fold) as assessed by MCE and FMT during the control period. The decrease in PO2 post nitrogen resulted in a slight increase in values at rest: 0. 46 ±0. 15 to 0. 53 ±0. 18 for b and (1. 39 ±0. 66) ml· min-1· g-1 to (1. 72 ±0. 30) ml ·min-1 ·g-1 for myocardial blood flow (MBF) (both P < 0. 05). In addition, values decreased in response to adenosine using both techniques: 1. 05 ±0. 35 to 0. 82 ±0. 27 for b and (4. 30 ±3. 16) ml ·min-1 ·g-1 to (3. 93 ±1. 27) ml ·min-1 ·g-1 for MBF (both P < 0. 05). Thus, CFR was markedly reduced under hypoxic conditions, to 1. 4 by MCE (P < 0. 05 compared with the baseline), and to 2. 5 by FMT (P > 0. 05 compared with the baseline). Conclusions CFR values diminish under hypoxic conditions according to both MCE and FMT. The reductions in CFR involve both an increase in resting values and a decrease in post adenosine measurements, as determined by both techniques. The reduction in CFR under hypoxia is slightly greater using MCE than using FMT. Physiological variables, such as hypoxia, must be taken into consideration when assessing CFR by MCE. Chin Med J 2004; 117(10):1453-1458

Background The results of clinical trials of rapamycin-eluting stents reduce restenosis have been quite promising. The main purpose of this study was to characterize the in vivo pharmacokinetics of high dose rapamycin (Rapa)-eluting stents in a miniswine coronary model. Methods Ten mimiswines underwent placement of 18 high dose Rapa-eluting stents in the left anterior descending and right coronary arteries. At the planned times of the 1. 5th, 12th, 24th hour, 3th, 7th and 28th day, the animals (n = 1, 1, 2, 2, 2, and 2, respectively) were euthanized after completion of coronary angiography. Blood samples were obtained at 0, 10, 20, 30 minutes; 1, 2, 6, 24 hours; and 3, 7, 28 days to determine systemic Rapa levels. Rap a levels in whole blood, arterial wall, heart, renal and liver tissues were determined by high-performance liquid chromatography/ mass spectroscopy. Results Peak whole blood concentration (Cmax), time to peak concentration (tmax), elimination half-life (t1/2β), area under the curve (AUC), and apparent systemic clearance (Cl/F) were (10. 91 ±1. 28) ng/ml, (2.0 ±0. 2) hours, (7.25 ±0. 63) hours, (1. 15 ±0. 11) ng· h · ml-1, and (180 ±12) ml · h-1· kg-1, respectively. More than 95 % Rapa detected is localized in the coronary artery surrounding the stent and heart. Conclusion Stent-based delivery of Rap a via a copolymer stent is feasible and safe. This strategy holds promise for the prevention of stent restenosis. Chin Med J 2004; 117(10):1459-1463

Background Vascular smooth muscle cell (VSMC) proliferation following arterial injury plays a critical role in a variety of vascular proliferative disorders, such as atherosclerosis and restenosis after balloon angioplasty. Herpes singlex virus-thymidine kinase (HSV-TK)/ ganciclovir (GCV) and E. coli cytosine deaminase (CD)/5-fluorocytosine (5-Fc) suicide gene systems have been successfully employed in cardiovascular gene therapy, respectively. We reasoned that coexpression of both HSV-TK with CD suicide genes would lead to increased cell killing. To test this imagine, the adenoviral vectors expressing TK and/or CD genes were developed and tested on vascular smooth muscle cells. Methods Adenoviral vectors, including Ad-EF1α-CD-cytomegolovirus (CMV)-TK coexpressing both CD and TK double suicide genes, Ad-EF1α-CD and Ad-CMV-TK expressing CD and TK respectively, and control vector Ad-CMV-LacZ, were constructed and prepared with homologous recombination in RecA+ E. coli cells. Integration and expression of CD and/or TK gene were identified by PCR and Western blot. Primary cultured VSMCs were infected at a multiplicity of infection (MOI) of 20 with exposure to their matching prodrugs 5-Fc and GCV. Cell mortality was measured by methyl thiazolyl tetrazolium (MTT) assays. Flow cytometry analysis was used to detect cell death, Apoptotic cells were analyzed using Hoechst 33342 fluorescence dye as a DNA probe. Genomic DNA cleavage of apoptotic VSMCs was tested by agarose gel electrophoresis. Results Recombinant adenovirus expressing CD and/or TK suicide genes were successfully constructed. Both single and double suicide genes could be integrated into adenoviral genome and expressed. Cytotoxic effects of Ad-EF1α-CD-CMV-TK double suicide genes combined with 5-Fc and GCV were higher than those of Ad-CMV-TK and Ad-EF1α-CD single gene groups. The rate of cell survival was only (9 ±3) %in the Ad-EF1α-CD-CMV-TK group, but (37 ±3) % in the Ad-CMV-TK and (46 ±4) % in the Ad-EF1α-CD groups (P < 0. 05). Flow cytometry analysis indicated that the killing mechanisms of the groups were different. Necrosis and apoptosis were involved in the mechanism of the double gene group. Based on the DNA stainability with Hoechst 33342, the apoptotic rates of VSMCs in the Ad-EF1α-CD-CMV-TK [(11.0 ± 2. 1) %] and Ad-CMV-TK [(12. 0 ±2.2) %] groups were higher than those in Ad-CMV-LacZ [(1. 2 ± 0. 11) %] and Ad-EF1α-CD [(5.0 ±1.8) %] groups (P < 0. 05, respctively). DNA smear could be observed in both Ad-CMV-TK and Ad-EF1α-CD-CMV-TK groups after administration of prodrugs. Conclusions The killing effect on rat VSMCs mediated by adenoviral CD/ TK double suicide genes is superior to that of single suicide gene. The killing mechanism of recombinant adenovirus coexpressing CD/ TK double suicide genes is mainly through cytotoxic effect and apoptosis. Chin Med J 2004; 117(10):1464-1470

Background SMAD proteins have recently been identified as the first family of putative transforming growth factor-β1 (TGF-β1) signal transducers. This study was to investigate the effects of TGF-β1 and signal protein Smad3 on rat cardiac hypertrophy. Methods The incorporation of [3H]-leucine was measured to determine the hypertrophy of cardiomyocyte incubated with different doses of TGF-β1 in cultured neonatal cardiomyocytes. The model of rat cardiac hypertrophy was produced with constriction of the abdominal aorta. At different times after the operation, rats were killed, and their left ventricular mass index (LVMI) determined. The mRNA expression of TGF-β1 and Smad3 of cultured cells and hypertrophic left ventricles were assessed by RT-PCR. The protein expression of Smad3 was assessed by Western blot. Results In cultured neonatal cardiomyocytes, TGF-β1 significantly promoted incorporation of [3H]-leucine. With the concentration of 3 pg/L, it increased the expression of Smad3 in mRNA and protein levels after 15 minutes, and continued for up to 8 hours of cultured cardiomyocytes. The LVMI and the expression of TGF-β1 (mRNA) and Smad3 (mRNA and protein) of hypertrophic left ventricle were increased by day 3 after the operation and continued to the 4th week. The peak expression of these was in the second week after operation. Conclusion TGF-β1 has positive effects on rat cardiomyocyte hypertrophy. Signal protein Smad3 could be related to the pathologic progression of rat cardiac hypertrophy. Chin Med J 2004; 117(10):1471-1475

Background Low-power helium-neon (He-Ne) lasers have been increasingly widely applied in the treatment of cardiovascular diseases, and its vasodilation effect has been proven. The aim of this study was to determine the effects of low-power He-Ne laser irradiation directed at the precardial region of Wistar rats on capillary permeability in the myocardium and the expression of myocardial vascular endothelial growth factor (VEGF). Methods Sixteen rats were divided randomly into control and irradiated groups (n = 8, each). A He-Ne laser (632. 8 nm) was applied to the irradiated group with a dose of 60. 5 J/cm2. Ferritin was perfused into the left femoral vein and capillary permeability was examined under an electron microscope. VEGF expression in the myocardium was investigated by immunohistochemical methods, RT-PCR, and image analysis. Results The ultra structures of the myocardial capillaries were examined. Compared to the control group, more high-density granules (ferritin), which were present within the capillary endothelium and the mitochondrions of myocardial cells in the internal layer of the myocardium, were observed in the irradiated group. VEGF staining of the myocardium was stronger in the irradiated group than that in the control group. The optic density of the irradiated group (0. 246 ±0. 015) was significantly higher than that of the control group (0. 218 ±0. 012, P < 0. 05). Finally, the levels of RT-PCR products of VEGF165 mRNA were 2. 79 times higher in irradiated rats than in the control rats. Conclusions Our study demonstrates that He-Ne laser irradiation (in doses of 60. 5 J/ cm2) increases myocardial capillary permeability and the production of VEGF in myocardial microvessels and in myocardium. Our study provides experimental morphological evidence that myocardial microcirculation can be improved using He-Ne laser irradiation. Chin Med J 2004; 117(10):1476-1480

Background There are many candidate genes for chronic obstructive pulmonary disease (COPD). Matrix metalloproteinase-9 (MMP-9) plays an essential role in tissue remodeling and repair associated with development of COPD. In this study we investigated the correlation between MMP-9 gene polymorphism and COPD susceptibility in the Han population of South China. Methods We examined the frequency of polymorphic genotypes of the MMP-9 promoter (- 1562C/ T) in 100 COPD patients and 98 healthy smokers by restriction fragment length polymorphism. Results The frequencies of polymorphic genotypes in promoters of MMP-9 were C/ C 86 %, C/ T 14 % in COPD group; and C/ C 98 %, C/ T 2 % in the control group. There were significant differences between the two groups (P < 0. 01). The allele frequencies were also significantly different between the COPD group and the control group (C allele frequency: 93 % vs 99 %, T allele frequency: 7 % vs 1 %, P < 0. 05 respectively). Conclusion The genetic polymorphism in promoters of MMP-9 gene is associated with the susceptibility to COPD in the Han population of South China. Chin Med J 2004; 117(10):1481-1484

Background Recent studies suggest that circulating DNA may be a potential tumor marker for lung cancer, but most of these studies are conducted between healthy controls and lung cancer patients, with few or no benign lung disease patients included. The objective of this study was to evaluate the performance of plasma DNA quantification in discriminating lung cancer from the healthy and benign lung disease. Methods Plasma DNA was extracted with a QIAamp DNA Blood Midi kit and quantified by a Pico Green9 dsDNA quantitation kit in 44 healthy individuals, 36 benign lung disease patients and 67 lung cancer patients. Discrimination power was evaluated by the receiver operating characteristic curve. Results Plasma DNA values were significantly increased in lung cancer patients, especially in those with metastases, and in benign lung disease patients compared with that in the healthy individuals (P <0. 001, respectively). The values in lung cancer patients were significantly increased compared with that in the benign lung disease patients (P < 0. 001). The area under the curve was 0. 96 [95 % confidence interval (CI) 0. 92 - 0. 99] for the healthy versus lung cancer, 0. 73 (95 % CI 0. 64 - 0. 83) for lung cancer versus benign lung disease, and 0. 86 (95 % CI 0. 80 - 0. 91) for lung cancer versus the healthy and benign lung disease. Conclusions Plasma DNA quantification has a strong power to discriminate lung cancer from the healthy and from the healthy and benign lung disease, less power to discriminate lung cancer from benign lung disease. Plasma DNA quantification may be useful as a screening tool for lung cancer. Chin Med J 2004; 117(10):1485-1488

Background The assessment of regional pulmonary ventilation and perfusion is essential for the evaluation of a variety of lung disorders. Pulmonary ventilation MRI using inhaled oxygen as a contrast medium can be obtained with a clinical MR scanner, without additional equipment, and has been demonstrated to be a feasible means of assessing ventilation in animal models and some clinical patients. However, few studies have reported on MR ventilation-perfusion imaging. In this study, we evaluated the usefulness of oxygen-enhanced ventilation in combination with first-pass Gd-DTPA-enhanced perfusion MRI in a canine model of pulmonary embolism and airway obstruction. Methods Peripheral pulmonary embolisms were produced in eight dogs by intravenous injection of gelfoam strips at the pulmonary segmental arterial level, and airway obstructions were created in five of the dogs by inserting a self-designed balloon catheter into a secondary bronchus. Oxygen-enhanced MR ventilation images were produced by subtracting images from before and after inhalation of pure oxygen. Pulmonary perfusion MR images were acquired with a dynamic three-dimensional fast gradient-echo sequence. MR ventilation and perfusion images were read and contrasted with results from general examinations of pathological anatomy, ventilation-perfusion scintigraphy, and pulmonary angiography. Results Regions identified as having airway obstructions matched using both MR ventilation and perfusion imaging, but regions of pulmonary embolisms were mismatched. The area of airway obstruction defects was smaller using MR ventilation imagery than that using ventilation scintigraphy. Abnormal perfusion regions due to pulmonary embolisms were divided into defective regions and reduced regions based on the time course of signal intensity changes. In the diagnosis of pulmonary embolisms with the technique of ventilation and perfusion MRI, sensitivity and specificity were 75. 0 % and 98. 1 %, respectively, and the diagnostic results of this MRI technique were in agreement with the results of ventilation-perfusion scintigraphy and pulmonary angiography (K: 0. 899, 0. 743). Conclusions Oxygen-enhanced ventilation in combination with pulmonary perfusion MRI can be used to diagnose abnormalities of airways and blood vessels in the lungs, and can provide regional functional information with high spatial and temporal resolution. This method possesses great potential value for clinical applications. Chin Med J 2004; 117(10):1489-1496