Magnesium Lithospermate B Derived from Salvia miltiorrhiza Ameliorates Right Ventricle Remodeling in Pulmonary Hypertensive Rats via Inhibition of NOX/VPO1 Pathway.

Right ventricle (RV) remodeling is a major pathological feature in pulmonary arterial hypertension (PAH). Magnesium lithospermate B (MLB) is a compound isolated from the roots of Salvia miltiorrhiza and it possesses multiple pharmacological activities such as anti-inflammation and antioxidation. This study aims to investigate whether MLB is able to prevent RV remodeling in PAH and the underlying mechanisms. In vivo, SD rats were exposed to 10% O2 for 21 d to induce RV remodeling, which showed hypertrophic features (increases in the ratio of RV weight to tibia length, cellular size, and hypertrophic marker expression), accompanied by upregulation in expression of NADPH oxidases (NOX2 and NOX4) and vascular peroxidase 1 (VPO1), increases in hydrogen peroxide (H2O2) and hypochlorous acid (HOCl) production and elevation in phosphorylation levels of ERK; these changes were attenuated by treating rats with MLB. In vitro, the cultured H9c2 cells were exposed to 3% O2 for 24 h to induce hypertrophy, which showed hypertrophic features (increases in cellular size and hypertrophic marker expression). Administration of MLB or VAS2870 (a positive control for NOX inhibitor) could prevent cardiomyocyte hypertrophy concomitant with decreases in NOX (NOX2 and NOX4) and VPO1 expression, H2O2 and HOCl production, and ERK phosphorylation. Based on these observations, we conclude that MLB is able to prevent RV remodeling in hypoxic PAH rats through a mechanism involving a suppression of NOX/VPO1 pathway as well as ERK signaling pathway. MLB may possess the potential clinical value for PAH therapy.

Suppression of NADPH oxidase attenuates hypoxia-induced dysfunctions of endothelial progenitor cells.

NADPH oxidases (NOX) - derived reactive oxygen species (ROS) contribute to oxidative injury in hypoxia-induced pulmonary arterial hypertension. This study aims to evaluate the status of NOX in endothelial progenitor cells (EPCs) under hypoxic condition and to determine whether NOX inhibitors could attenuate hypoxia-induced dysfunctions of EPCs. EPCs were isolated from peripheral blood of SD rats and subjected to hypoxia (O2/N2/CO2, 1/94/5) for 24 h. The cells were collected for ß-galactosidase or Hoechst staining, or for functional analysis (migration, adhesion and tube formation). The NOX expression, activity and H2O2 content in EPCs were measured. The results showed that hypoxia treatment promoted EPC senescence and apoptosis, accompanied by the deteriorated functions of EPCs (the reduced abilities in adhesion, migration and tube formation), as well as an increase in NOX2 and NOX4 expression, NOX activity and H2O2 production, these phenomena were attenuated by NOX inhibitors. Furthermore, administration of catalase could also improve the functions of hypoxia-treated EPCs. Based on these observations, we conclude that NOX-derived ROS contributes to the dysfunctions of EPCs under hypoxic condition. Thus, suppression of NOX may provide a novel strategy to improve endothelial functions in hypoxia-relevant diseases.

Duplication of 10q22.3-q23.3 encompassing BMPR1A and NGR3 associated with congenital heart disease, microcephaly, and mild intellectual disability.

Chromosome region 10q22.3-q23.3 contains several low copy repeats (LCRs) and is prone to recombination. Deletions with breakpoints within LCR3 and LCR4 have been described to be associated with intellectual disability and dysmorphic features, while the reciprocal duplications are rarely reported. We present an additional case with multiple congenital anomalies that include microcephaly, cardiac defect, and mild intellectual disability, in which a de novo interstitial 8.2-Mb duplication of 10q22.3-q23.3, including BMPR1A and NGR3, was identified by Illumina SNP array platform. Our study is consistent with the hypothesis that the BMPR1A is a plausible candidate gene for congenital heart disease (CHD) and should contribute to the diagnosis and treatment of these genomic diseases.

Microduplication of 3p25.2 encompassing RAF1 associated with congenital heart disease suggestive of Noonan syndrome.

Noonan syndrome (NS) is a clinically variable and genetically heterogeneous disorder with congenital heart defects (CHD), short stature, and craniofacial dysmorphisms. Gain-of-function mutations in RAF1 can cause NS and the highly related NS with multiple lentigines (previously known as LEOPARD syndrome). Here we report on a 15-year-old male with NS phenotype: short stature, heart defects, low posterior hairline, facial malformations, malformed left ear with sensorineural hearing loss, widely spaced nipples, and unilateral upper limb anomaly. Using high-resolution SNP array technology, we identified in this patient a 0.25 Mb microduplication at 3p25.2 in which RAF1 is located. Sequence analysis did not identify mutations in genes associated with Holt-Oram syndrome. These findings suggest that duplications of genomic regions encompassing RAF1 could cause NS and are consistent with the notion that rare copy number variations encompassing causative genes may underlie a small percentage of patients with syndromic CHD like NS.

[Experimental study of rat mesenchymal stem cells transfected with Sonic Hedgehog gene].

OBJECTIVE: To evaluate the feasibility of genetic modification of mesenchymal stem cells (MSC) with Sonic Hedgehog (Shh) gene. METHODS: The pcDNA3.1-Shh eukaryotic expression plasmid was constructed and its correctness evaluated by the restriction enzyme analysis and sequencing. MSC were isolated from Wistar rats by density gradient centrifugation and purified, transfected with pcDNA3.1-Shh, blank plasmid pcDNA3.1(-) or pmaxGFP respectively by Nucleofector(TM). The protein expression of Shh in MSC was detected by Western blot after 48 hours. RESULTS: Correct construction of pcDNA3.1-Shh was identified by the methods of restriction enzyme analysis and nucleotide sequence determination. The expression of green fluorescent protein (GFP) could be observed by fluorescence microscopy after 48 hours. The expression of Shh gene was detected by Western blot. But the MSC transfected with empty plasmid expression was not detected. CONCLUSIONS: Recombinant Eukaryotic expression plasmid pcDNA3.1-Shh is successfully detected in rat MSC. It may provide experimental rationales for the future gene therapy.

[Efficacy comparison between micro invasive occlusion procedure and extracorporeal circulation procedure for treating patients with simple ventricular septal defect].

OBJECTIVE: To compare the efficacy between micro invasive occlusion procedure and extracorporeal circulation procedure for treating patients with simple ventricular septal defect. METHODS: Two hundred and twenty patients with simple ventricular septal defect (except subarterial ventricular septal defect) were randomly divided into micro invasive group (n = 116) and traditional cardiopulmonary bypass surgery group (n = 104). Clinical data were collected and compared at baseline and at 3, 30 and 180 days after surgery. RESULTS: Age, gender, body weight and ventricular septal defect type were similar between the two groups (all P > 0.05). Operation time and hospitalization duration were significantly shorter in the micro invasive group than the traditional cardiopulmonary bypass surgery group (all P < 0.05). The proportion of blood transfusion was less in micro invasive group than the traditional cardiopulmonary bypass surgery group [2.59% (3/116) vs. 72.12% (75/104), P < 0.01]. Three days after surgery, incidence of mild and above tricuspid insufficiency was less in micro invasive group than the traditional cardiopulmonary bypass surgery group [0.86% (1/116) vs. 2.88% (3/104), P < 0.05]. There was one patient developed mild pericardial effusion at 30 days post surgery in micro invasive group. There was no intracardiac infection in the two groups during follow-up. At 30 and 180 days post surgery, incidence of residual shunt was also less in micro invasive group than the traditional cardiopulmonary bypass surgery group [1.72% (2/116) vs. 7.69 (8/104) and 0(0/116) vs. 7.69(8/104), all P < 0.05]. After surgery for 3, 30 and 180 days, transthoracic echocardiography derived chamber size, left ventricular end-diastolic volume index and left ventricular ejection fraction were similar between the two groups (all P > 0.05). CONCLUSIONS: The efficacy is similar for patients with simple ventricular septal defect (except subarterial ventricular septal defect) using micro invasive occlusion therapy or extracorporeal circulation surgery. Micro invasive occlusion procedure can shorten operation and hospitalization time, and reduce the need for blood transfusion and risk of developing procedural-related tricuspid insufficiency and post-procedural residual shunt.

Automatic feature extraction and fusion recognition of motor imagery EEG using multilevel multiscale CNN.

A motor imagery EEG (MI-EEG) signal is often selected as the driving signal in an active brain computer interface (BCI) system, and it has been a popular field to recognize MI-EEG images via convolutional neural network (CNN), which poses a potential problem for maintaining the integrity of the time-frequency-space information in MI-EEG images and exploring the feature fusion mechanism in the CNN. However, information is excessively compressed in the present MI-EEG image, and the sequential CNN is unfavorable for the comprehensive utilization of local features. In this paper, a multidimensional MI-EEG imaging method is proposed, which is based on time-frequency analysis and the Clough-Tocher (CT) interpolation algorithm. The time-frequency matrix of each electrode is generated via continuous wavelet transform (WT), and the relevant section of frequency is extracted and divided into nine submatrices, the longitudinal sums and lengths of which are calculated along the directions of frequency and time successively to produce a 3 × 3 feature matrix for each electrode. Then, feature matrix of each electrode is interpolated to coincide with their corresponding coordinates, thereby yielding a WT-based multidimensional image, called WTMI. Meanwhile, a multilevel and multiscale feature fusion convolutional neural network (MLMSFFCNN) is designed for WTMI, which has dense information, low signal-to-noise ratio, and strong spatial distribution. Extensive experiments are conducted on the BCI Competition IV 2a and 2b datasets, and accuracies of 92.95% and 97.03% are yielded based on 10-fold cross-validation, respectively, which exceed those of the state-of-the-art imaging methods. The kappa values and p values demonstrate that our method has lower class skew and error costs. The experimental results demonstrate that WTMI can fully represent the time-frequency-space features of MI-EEG and that MLMSFFCNN is beneficial for improving the collection of multiscale features and the fusion recognition of general and abstract features for WTMI.

Magnesium lithospermate B prevents phenotypic transformation of pulmonary arteries in rats with hypoxic pulmonary hypertension through suppression of NADPH oxidase.

Magnesium lithospermate B (MLB) shows multiple biological activities including anti-oxidation and anti-proliferation in various diseases. However, the function of MLB in pulmonary arterial hypertension (PAH) is still unknown. This study aims to investigate the effect of MLB on hypoxia-induced phenotypic transformation of pulmonary arterial smooth muscle cells (PASMCs) and the underlying mechanisms. SD rats (or PASMCs) were exposed to 10% O2 for 3 weeks (or 3% O2 for 48 h) along with MLB or NADPH oxidase （NOX） inhibitor intervention. The effects of MLB on hemodynamics, pulmonary vascular remodeling and phenotypic transformation of PASMCs were observed first. Then, its effects on the protein levels of NOX (NOX2 and NOX4), ERK and p-ERK were examined. The results showed that MLB prevented the elevation in right ventricular systolic pressure and the increase in ratio of wall thickness to vessel external diameter of pulmonary arteries in PAH rats, and attenuated phenotypic transformation of PASMCs (decrease in α-smooth muscle actin while increase in osteopontin), accompanied by downregulation of NOX (NOX2 and NOX4) protein levels, decrease of ROS and H2O2 production, and suppression of the phosphorylation of ERK. NOX inhibitor (VAS2870) achieved similar results to that of MLB did in the hypoxia-treated PASMCs. Based on the observations, we conclude that MLB is able to prevent phenotypic transformation of pulmonary arteries in hypoxic PAH rats through suppression of NOX/ROS/ERK pathway, and MLB might have the potentials in PAH therapy.

Coordination between NADPH oxidase and vascular peroxidase 1 promotes dysfunctions of endothelial progenitor cells in hypoxia-induced pulmonary hypertensive rats.

Previous studies have demonstrated that NADPH oxidase (NOX)/vascular peroxidase (VPO1) pathway - mediated oxidative stress plays an important role in the pathogenesis of multiple cardiovascular diseases. This study aims to evaluate the correlation between NOX/VPO1 pathway and endothelial progenitor cells (EPCs) dysfunctions in hypoxia-induced pulmonary hypertension (PH). The rats were exposed to 10% hypoxia for 3 weeks to establish a PH model, which showed increases in right ventricle systolic pressure, right ventricular and pulmonary vascular remodeling, acceleration in apoptosis and impairment in functions of the peripheral blood derived - EPCs (the reduced abilities in adhesion, migration and tube formation), accompanied by up-regulation of NOX (NOX2 and NOX4) and VPO1. Next, normal EPCs were cultured under hypoxia to induce apoptosis in vitro. Consistent with the in vivo findings, hypoxia enhanced the apoptosis and dysfunctions of EPCs concomitant with an increase in NOX and VPO1 expression, hydrogen peroxide (H2O2) and hypochlorous acid (HOCl) production; these phenomena were attenuated by NOX2 or NOX4 siRNA. Knockdown of VPO1 showed similar results to that of NOX siRNA except no effect on NOX expression and H2O2 production. Based on these observations, we conclude that NOX/VPO1 pathway-derived reactive oxygen species promote the oxidative injury and dysfunctions of EPCs in PH, which may contribute to endothelial dysfunctions in PH.

Taurine inhibits osteoblastic differentiation of vascular smooth muscle cells via the ERK pathway.

Vascular calcification develops within atherosclerotic lesions and results from a process similar to osteogenesis. Taurine is a free beta-amino acid and plays an important physiological role in mammals. We have recently demonstrated that vascular smooth muscle cells (VSMCs) express a functional taurine transporter. To evaluate the possible role of taurine in vascular calcification, we assessed its effects on osteoblastic differentiation of VSMCs in vitro. The results showed that taurine inhibited the beta-glycerophosphate-induced osteoblastic differentiation of VSMCs as evidenced by both the decreasing alkaline phosphate (ALP) activity and expression of the core binding factor alpha1 (Cbfalpha1). Taurine also activated the extracellular signal-regulated protein kinase (ERK) pathway. Inhibition of ERK pathway reversed the effect of taurine on ALP activity and Cbfalpha1 expression. These results suggested that taurine inhibited osteoblastic differentiation of vascular cells via the ERK pathway.

[Relationship between the expression of heat shock protein 25 and selenium binding protein 1 with ischemia/reperfusion lung injury in vivo: experiment with rats].

OBJECTIVE: To analyze the differential expression proteins of rat ischemia/reperfusion (I/R) lung tissues in vivo and normal lung tissues by comparative proteome analysis, and to study the mechanism of donor lung I/R injury. METHODS: Forty male SD rats were randomly divided into 2 equal groups: I/R group undergoing mimic orthotopic left lung auto-grafting and harvesting of the left lung five hours after the operation, and control group undergoing isolation of the left hilus of lung and then harvesting of the left lung. The differential proteins in the left ventricle of transplanted heart were separated by means of immobilized pH gradient-based two-dimensional gel electrophoresis (2-DE), identified by matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS), and searched through Matrix Science software system. Western blotting was used to verify part of the differentially expressed proteins. RESULTS: Well-resolved and reproducible 2-DE profile of rat I/R lung tissues and normal lung tissues were obtained. In the I/R lung tissue profile, the average spot number from 3 gels was 489 +/- 52 spots (P > 0.05) with an average matching rate of 89.28% (P > 0.05), and in the control group, the average spot number from 3 gels was 511 +/- 83 spots (P > 0.05) with an average matching rate of 91.22% (P > 0.05). Fourteen differential proteins were identified by peptide mass fingerprinting (PMF) searched in Matrix Science (P < 0.05). Western blotting confirmed that the protein expression of selenium binding protein 1 (SBP-1) and heat shock protein 25 (HSP25) increased at the early stage of I/R group. CONCLUSION: The protein expression of HSP25 and SBP-1 with stress protection function is upregulated in the early stage of lung I/R injury. Other differentially expressed proteins identified may have important functions in energy metabolism, tissue stress, cell apoptosis, and signal transduction.

[Retrospective study on clinical application of biovalves in 52 cases].

OBJECTIVE: To summarize the clinical application of bioprosthetic valve replacement. METHODS: Fifty two patients, aged 13-73(52.4+/-14.0) years, underwent cardiac valve replacement with biovalves from June 2002 to June 2006 in our hospital. Fifty three tissue valve replacements combined with 8 mechanical valve replacements were performed. Other procedures were also carried out if appropriate, including bidirectional Glenn shunt in 1 patient with double outlet of right ventricle and pulmonary artery stenosis; interruption of accessory pathway in 1 patient with Wolff-Parkinson-White syndrome; coronary artery bypass graft in 5 with severe coronary artery disease; and atrial or ventricular repair in 4 with congenital septum defects. RESULTS: One patient died in hospital due to multiple organ failure. The hospital mortality rate was 1.9%. The mean follow-up was 25 months. Two patients lost follow-up and the follow-up rate was 96%. Forty one patients returned to NYHA class I, 9 to class II, and 1 to class III. Echocardiography showed the implanted bioprosthesis functioned well without stenosis or regurgitation. There was no structural deterioration with freedom from thromboembolism and anticoagulant hemorrhage in all patients. CONCLUSION: The efficacy of biovalves in heart valve replacement is satisfactory.

[Early surgical treatment for infants with large atrial septal defects or ventricular septal defects complicated by pneumonia: experience of 39 cases].

OBJECTIVE: This research reported the experience of early surgical treatment for infants with large atrial septal defects (ASD) or ventricular septal defects (VSD) complicated by pneumonia. METHODS: Between January 2003 and January 2008, 39 infants with large ASD or VSD complicated by pneumonia were admitted to the Second Xiangya Hospital. Thirty-six patients underwent surgical repair within 7-10 days after pneumonia had been controlled. Mean age was 5.4+/-3.4 months and mean weight was 4.7+/-1.6 kg in the 36 patients. Three patients received conservative treatment due to uncontrolled lung infections. RESULTS: Of the 36 patients, 33 had successful surgery and 3 (8.3%) died of serious low cardiac output (n=1) or respiratory failure due to congenital tracheostenosis (n=2). The 33 survivors showed normal growth and development in a 6 month-5 year follow-up. Of the 3 patients receiving conservative treatment, 1 died of cardiopulmonary failure and 2 were discharged after the symptoms had been improved. CONCLUSIONS: With increasing medical experience and technique, early surgical operation may be performed with good outcomes in infants with large ASD or VSD complicated by pneumonia.

The time-dependent effects of St John's wort on cytochrome P450, uridine diphosphate-glucuronosyltransferase, glutathione S-transferase, and NAD(P)H-quinone oxidoreductase in mice.

Hypericum perforatum [St. John's wort (SJW)] is known to cause a drug interaction with the substrates of cytochrome P450 (P450, CYP) isoforms, mainly CYP3A. This study aims to determine the dose response and time course of the effects of SJW extract on P450s, UDP-glucuronosyltransferase (UGT), glutathione S-transferase (GST), and NAD(P)H-quinone oxidoreductase (NQO) in mice. The oral administration of SJW extract to male mice at 0.6 g/kg/d for 21 days increased hepatic oxidation activity toward a Cyp3a substrate nifedipine. By extending the SJW treatment to 28 days, hepatic nifedipine oxidation (NFO) and warfarin 7-hydroxylation (WOH) (Cyp2c) activities were increased by 95% and 34%, respectively. Immunoblot analysis of liver microsomal proteins revealed that the Cyp2c protein level was elevated by the 28-day treatment. However, the liver microsomal activities of the oxidation of the respective substrates of Cyp1a, Cyp2a, Cyp2b, Cyp2d, and Cyp2e1 remained unchanged. In the kidney, SJW increased the NFO, but not the WOH activity. The extended 28-day treatment did not alter mouse hepatic and renal UGT, GST, and NQO activities. These findings demonstrate that SJW stimulates hepatic and renal Cyp3a activity and hepatic Cyp2c activity and expression. The induction of hepatic Cyp2c requires repeated treatment for a period longer than the initial induction of Cyp3a.

[Experience in treatment of Eisenmenger's syndrome by heart-lung transplantation].

OBJECTIVE: To summarize the experience in treatment of Eisenmenger's syndrome by heart-lung transplantation (HLT). METHODS: Two cases of congenital heart disease with Eisenmenger's syndrome, aged 20 and 22, underwent bicaval orthotopic HLT. Modified St. Thomas cardioplegic solution and modified LPD solution were used to preserve the donor heart and lung. After removing the heart and lung of the recipient and thorough hemostasis in thoracic cavity and mediastinal septum, the donor heart and lung was implanted, and trachea, superior/inferior vena cava and aortic artery were anastomosed gradually. Intensive care against infection, rejection and other complications were performed after operation. RESULTS: The operations were successful. Tracheal anastomotic stenosis happened in one case seven months after HLT and was cured by sleeve resection of the stenosed trachea segment. Acute rejection happened in one case 10 days after HLT and was cured by stoss therapy. Both patients recovered to normal life and work. CONCLUSION: The success of HLT is related to perfect organ preservation, precise surgical performance and proper peri-operative treatment.

[Studies of spectra classification based on kernel covering algorithm].

A kernel based covering algorithm, called the kernel covering algorithm (KCA), is proposed for the classification of celestial spectra. This algorithm is a combination of kernel trick with the covering algorithm, and is used to extract the support vectors in feature space. The experiments show that the classification result based on KCA is a little less than that based on SVM. However, KCA only involves the distance computation without the need to solve the quadratic programming problem. Also, KCA is insensitive to the width of gauss window. Although KCA has a comparable classification performance with the covering algorithm, it changes the distance between samples in feature space by the nonlinear mapping such that the distribution of samples is more adaptable to classify. Therefore, the number of KCA's resulting support vectors is significantly smaller than that of the covering algorithm.

[Mechanical ventilation treatment on acute respiratory distress syndrome in operations by cardiopulmonary bypass].

OBJECTIVE: To explore the early diagnosis of acute respiratory distress syndrome (ARDS) and the mechanical ventilation treatment in operations of cardiopulmonary bypass. METHODS: Thirty-four patients with acute respiratory distress syndrome were divided into the traditional ventilation strategy plus positive end expiratory pressure (PEEP) group (n=13) and the protective ventilation strategy group (n=21), and their diagnosis and treatment were reviewed. RESULTS: The average ventilator-carrying time in the protective ventilation strategy group and the traditional ventilation strategy plus PEEP group was (4.8+/-2.1) and (7.2+/-3.6) days,and the mortality was 23.8% and 61.5% respectively. There was statistical significance in the 2 groups (P<0.05). CONCLUSION: The key to ARDS is the early diagnosis and ventilation strategy. Protective ventilation strategy in treating ARDS is more effective than the traditional ventilation strategy plus PEEP, and it can obviously shorten the ventilator-carrying time and reduce the mortality of ARDS.

Surgical treatment of total anomalous pulmonary venous connection.

OBJECTIVE: To summarize the surgical treatment of total anomalous pulmonary venous connection (TAPVC). METHODS: We retrospectively analyzed 49 patients with TAPVC as follows: 37 patients with supra-cardiac type in which 35 received anastomoses between the rear wall of the left atrium and the common pulmonary vein and the other 2 received anastomoses between the rear edge of the left atrium roof and the common pulmonary vein; another 12 patients with cardiac type who were incised the upper edge of coronary sinus, connected the common junction of pulmonary veins with the left atrium to form a new left atrium, and repaired the atrial septal defect with a pericardial patch. RESULTS: Forty-six patients recovered and 3 patients died. Thirty-eight patients were followed-up from 3 months to 8 years. The rest patients got fluent pulmonary vein drainage and their heart function resumed to NYHA I. CONCLUSION: TAPVC patients should be operated on immediately at definite diagnosis. The fluency of the common pulmonary vein-left atrium anastomoses and proper postoperative care can ensure a satisfactory outcome.

[Effects of myocardial transplantation of mesenchymal stem cells transfected with vascular endothelial factor gene on improvement of heart function and angiogenesis after myocardial infarction: experiment with rats].

UNLABELLED: To establish a method to transfect vascular endothelial factor (VEGF) gene into mesenchymal stem cells ( MSCs) , to investigate the effects of the gene-transfected MSCs on heart function restoration and angiogenesis after myocardial infarction, and to compare the differences among cell therapy, gene therapy, and combined therapy. METHODS: Seventy-one Wistar rats underwent ligation of the left anterior descending coronary artery so as to establish heart ischemia models. Fifteen rats underwent sham operation. MSCs were isolated from several Wistar rats by density gradient centrifugation, purified, and transfected with pcDNA3.1-hVEGF165 or blank plasmid pcDNA3.1 respectively using the liposome mediated method. ELISA, Western blotting, and RT-PCR were used to detect the protein and mRNA expression of hVEG in these MSCs Forty-eight surviving rats that underwent ligation were randomly divided into 4 equal groups: combination group (Combo group) to be injected into the heart infarct zone with suspension of hVEGF165-transfected MSCs 2 weeks after the establishment of the model, cell group to be injected with suspension of MSCs not transfected with VEGF, gene group to be injected with suspension of DNA-liposome containing pcDNA3.1-VEGF165 and control group to be injected with cold culture fluid only. Twelve surviving rats that underwent sham operation were used as normal non-ischemic group. Four weeks after the injection the surviving rats underwent examination of heart functions by the Buxco system. The rats were killed and their hearts were taken out to undergo immunohistochemistry with 5-bromodeoxyuridine (Brdu) and troponin T and factor VIII to measure the area of cardiac infarction and the capillary density. RT-PCR was used to examine the mRNA expression of VEGF. The heart infarcted size was calculated by Evan's blue staining. RESULTS: (1) MSCs can be successfully isolated and cultured by density gradient centrifugation followed by adherence-separation. The expression of hVEGF165 in the transfected MSCs was demonstrated with ELISA, RT-PCR and Western Blot Assay. (2) Four weeks after the cells were transplanted, among all groups but the nonischemic group, the heart infarcted size of the Combo group was 27.8% +/- 3. 0% ,significantly less than those of the cell group (37.0% +/- 10. 1% ) and gene group (37.1% +/- 5.2%, both P <0.05). The heart function of the Combo group was better than those of other groups. (3) The capillary density of the Combo group was 40. 2 +/- 5.5/visual field, significantly greater than those of both the cell group (27.2 +/- 6. 3/visual field, P <0. 01) and that of the control group (18.5 +/- 5.8/visual field, P <0. 01) , and greater to some degree than that of the gene group (35. 8 +/-7.7/visual field, P =0. 189). (4)The heart infarcted size, heart function and capillary density of the cell and gene groups were similar and were smaller, better and greater than those of the control group. (5) Brdu and troponin T double staining detected a varied increase in the number of surviving cardiomyocytes at the heart infarcted area, some of which were double stain positive. RT-PCR showed mRNA expression of hVEGF165 in the Combo and gene groups, that in the Combo group being higher than that in the gene group. CONCLUSION: Eukaryotic expression vector pcDNA3.1-hVEGF165 can effectively be expressed in MSCs. Transplantation of VEGF gene by means of transfected MSCs brings better improvement in myocardial perfusion and in restoration of heart function than either cellular or gene therapy alone.