RÉSUMÉ
Tissue damage in diabetes is at least partly due to elevated reactive oxygen species production by the mitochondrial respiratory chain during hyperglycemia. Sustained hyperglycemia results in mitochondrial dysfunction and the abnormal expression of mitochondrial genes, such as NADH: Ubiquinone oxidoreductase subunit A13 (NDUFA13). Metformin, an AMPactivated protein kinase (AMPK) activator, protects cardiomyocytes from oxidative stress by improving mitochondrial function; however, the exact underlying mechanisms are not completely understood. The aim of the present study was to investigated the molecular changes and related regulatory mechanisms in the response of H9C2 cardiomyocytes to metformin under high glucose conditions. H9C2 cells were subjected to CCK8 assay to assess cell viability. Reactive oxygen species generation was measured with DCFHDA assay. Western blotting was used to analyze the expression levels of NDUFA13, AMPK, pAMPK and GAPDH. Reverse transcriptionquantitative PCR was used to evaluate the expression levels of mitochondrial genes and transcription factors. It was observed that metformin protected H9C2 cardiomyocytes by suppressing high glucose (HG)induced elevated oxidative stress. In addition, metformin stimulated mitochondrial biogenesis, as indicated by increased expression levels of mitochondrial genes (NDUFA1, NDUFA2, NDUFA13 and manganese superoxide dismutase) and mitochondrial biogenesisrelated transcription factors [peroxisome proliferatoractivated receptorgamma coactivator1α, nuclear respiratory factor (NRF)1, and NRF2] in the metformin + HG group compared with the HG group. Moreover, metformin promoted mitochondrial NDUFA13 protein expression via the AMPK signaling pathway, which was abolished by pretreatment with the AMPK inhibitor, Compound C. The results suggested that metformin protected cardiomyocytes against HGinduced oxidative stress via a mechanism involving AMPK, NDUFA13 and mitochondrial biogenesis.
Sujet(s)
Complexe I de la chaîne respiratoire/métabolisme , Metformine/pharmacologie , Chaperons moléculaires/métabolisme , Myocytes cardiaques/métabolisme , AMP-Activated Protein Kinases/métabolisme , Animaux , Lignée cellulaire , Survie cellulaire/effets des médicaments et des substances chimiques , Chine , Complexe I de la chaîne respiratoire/effets des médicaments et des substances chimiques , Glucose/métabolisme , Hyperglycémie/métabolisme , Metformine/métabolisme , Mitochondries/effets des médicaments et des substances chimiques , Mitochondries/métabolisme , Protéines mitochondriales/métabolisme , Chaperons moléculaires/effets des médicaments et des substances chimiques , Myocytes cardiaques/effets des médicaments et des substances chimiques , Biogenèse des organelles , Stress oxydatif/effets des médicaments et des substances chimiques , Coactivateur 1-alpha du récepteur gamma activé par les proliférateurs de peroxysomes/métabolisme , Protein-Serine-Threonine Kinases , Rats , Espèces réactives de l'oxygène/métabolisme , Transduction du signal/effets des médicaments et des substances chimiques , Superoxide dismutase/métabolisme , Facteurs de transcription/génétiqueRÉSUMÉ
High-throughput sequencing of 16S rDNA and culture-dependent methods were applied to determine the bacterial communities of braised chicken during processing and storage. Environmental microorganisms were also evaluated using a sedimentation plate method. The results showed that airborne microbial counts in the braising room were higher than those in the control room (25°C, a space to lower the temperature of the chicken products) and storage rooms (4°C). The microbial identification technique 16S rDNA sequences has indicated that more than 229 operational bacterial species were associated with the microbiota present in braised chicken, largely involving Pseudomonas, Psychrobacter, Weissella, Kurthia, Brochothrix, and Lactobacillus in modified-atmosphere packing (MAP) products. The storage place and temperature during processing has great impact on the shelf life of the chicken. The microbes in MAP were significantly higher (P < 0.05) in 0 and 7th day, while the microbial activity in vacuum packaging (VP) was lower because the VP products were treated at higher temperature (100°C for 20 min). Within chicken products, Pseudomonas, Brochothrix, and Lactobacillus were most prevalent in MAP products. According to this research, in order to prolong the shelf life of meat products, proper storage places and packaging conditions are necessary to be improved to reduce the microbial load in the food products.
Sujet(s)
Bactéries/isolement et purification , Microbiologie alimentaire , Microbiote , Produits de basse-cour/microbiologie , Animaux , Poulets , Cuisine (activité) , Emballage alimentaire/méthodesRÉSUMÉ
Diaphragm dysfunction is an important clinical problem worldwide. Hydrogen sulfide (H2S) is involved in many physiological and pathological processes in mammals. However, the effect and mechanism of H2S in diaphragm dysfunction have not been fully elucidated. In this study, we detected that the level of H2S was decreased in lipopolysaccharide- (LPS-) treated L6 cells. Treatment with H2S increased the proliferation and viability of LPS-treated L6 cells. We found that H2S decreased reactive oxygen species- (ROS-) induced apoptosis through the mitogen-activated protein kinase (MAPK) signaling pathway in LPS-treated L6 cells. Administration of H2S alleviated LPS-induced inflammation by mediating the toll-like receptor-4 (TLR-4)/nuclear factor-kappa B (NF-κB) signaling pathway in L6 cells. Furthermore, H2S improved diaphragmatic function and structure through the reduction of inflammation and apoptosis in the diaphragm of septic rats. In conclusion, these findings indicate that H2S ameliorates LPS-induced diaphragm dysfunction in rats by reducing apoptosis and inflammation through ROS/MAPK and TLR4/NF-κB signaling pathways. Novel slow-releasing H2S donors can be designed and applied for the treatment of diaphragm dysfunction.
Sujet(s)
Apoptose/effets des médicaments et des substances chimiques , Muscle diaphragme/effets des médicaments et des substances chimiques , Sulfure d'hydrogène/pharmacologie , Inflammation/métabolisme , Transduction du signal/effets des médicaments et des substances chimiques , Animaux , Apoptose/physiologie , Muscle diaphragme/métabolisme , Lipopolysaccharides/toxicité , Système de signalisation des MAP kinases/effets des médicaments et des substances chimiques , Système de signalisation des MAP kinases/physiologie , Facteur de transcription NF-kappa B/effets des médicaments et des substances chimiques , Facteur de transcription NF-kappa B/métabolisme , Rats , Espèces réactives de l'oxygène/métabolisme , Transduction du signal/physiologie , Récepteur de type Toll-4/effets des médicaments et des substances chimiques , Récepteur de type Toll-4/métabolismeRÉSUMÉ
Background: Increased cardiomyocyte apoptosis under high glucose condition contributes to diabetic cardiomyopathy. Degradation of cardiac Connexin43 (Cx43) has been associated with cardiac dysfunction in diabetic heart. Clinical and experimental studies suggested that metformin (Met) exhibits cardioprotective properties against diabetes. Aim: The aim of this study was to investigate the effect and underlying signaling mechanisms of metformin on apoptosis and Cx43 expression in H9c2 cells presenting with hyperglycemia conditions. Methods: In the present study, H9c2 cardiac cells were incubated with 5.5 mM glucose, 33.3 mM glucose, 33.3 mM glucose with metformin at two dose (100 µM, 1 mM) for 96 hours, and 1 mM metformin with chloroquine (50 µM) in 33.3 mM glucose medium. Cell viability was determined by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) cell survival assay. Cytotoxicity was determined by the release of lactate dehydrogenase (LDH). The expression of Cx43, autophagic maker protein (LAMP-1, Beclin-1, p62 and LC3) and apoptosis maker protein (Bcl-2 and Bax) were determined by western blot. Results: The results showed that high glucose increased apoptosis and decreased Cx43 expression. Interestingly, metformin attenuated hyperglycemia-increased apoptosis and restored Cx43 expression. Moreover, this treatment caused autophagy as well, which indicated by up-regulation of autophagy-related proteins LAMP-1, Beclin-1, p62 and reduction in the ratio of LC3-II/LC3-I. In addition, administration autophagy inhibitor chloroquine (CQ) did not block the effect of metformin on Cx43 expression while increasing Cx43 content, together with an increased apoptosis. Conclusion: Administration metformin can protect the H9c2 cells against hyperglycemia-induced apoptosis and Cx43 down-regulation, in part, mediated through the induction of autophagy pathway.
Sujet(s)
Autophagie/effets des médicaments et des substances chimiques , Connexine 43/génétique , Diabète/traitement médicamenteux , Hyperglycémie/traitement médicamenteux , Metformine/administration et posologie , Apoptose/effets des médicaments et des substances chimiques , Protéines régulatrices de l'apoptose/génétique , Autophagie/génétique , Lignée cellulaire , Chloroquine/administration et posologie , Diabète/anatomopathologie , Régulation de l'expression des gènes/effets des médicaments et des substances chimiques , Humains , Hyperglycémie/anatomopathologie , Myocytes cardiaques/effets des médicaments et des substances chimiques , Myocytes cardiaques/anatomopathologie , Transduction du signal/effets des médicaments et des substances chimiquesRÉSUMÉ
The expression of connexin43 (Cx43) protein and the apoptotic rate of cardiomyocytes may be regulated by autophagy and associated with diabetic cardiomyopathy. It is possible that the beneficial effect of resveratrol on diabetic cardiomyocytes occurs via the autophagy pathway. However, it remains to be elucidated whether resveratrol treatment may attenuate the hyperglycemiainduced remodeling of Cx43 and apoptosis through the regulation of autophagy. H9c2 cardiac cells were incubated with 5.5 and 25 mM glucose, 25 mM glucose with chloroquine (50 µM), and 25 mM glucose with or without resveratrol (10, 25 µM) for 24 h. H9c2 cells were also incubated with 25 µM resveratrol in the presence of chloroquine (50 µM). Cell viability was determined using an MTT cell survival assay. Cytotoxicity was determined by quantification of the release of lactate dehydrogenase. The expression of Cx43, autophagic maker proteins [Beclin1, p62 and microtubuleassociated protein 1 light chain 3 (LC3)], apoptosis maker proteins (Bcell lymphoma2 and Bcl2 associated X protein), AMPactivated protein kinase (AMPK) and mammalian target of rapamycin (mTOR) were determined using western blotting. Resveratrol treatment led to reduced Cx43 expression levels compared with the 25 mM glucose treatment and significantly reduced the expression of apoptosisassociated proteins in H9c2 cells under hyperglycemic conditions. Autophagy was increased as indicated by the upregulation of Beclin1 and p62 expression and the reduced LC3II/LC3I ratio. AMPK expression was increased, whereas mTOR expression was reduced in the resveratrol treatment groups. Treatment with chloroquine reversed effect of resveratrol. In conclusion, administration resveratrol may protect H9c2 cells against hyperglycemiainduced Cx43 upregulation and apoptosis, which may be mediated through the induction of the autophagy signaling pathway.
Sujet(s)
Apoptose/effets des médicaments et des substances chimiques , Autophagie/effets des médicaments et des substances chimiques , Connexine 43/métabolisme , Glucose/toxicité , Transduction du signal/effets des médicaments et des substances chimiques , Stilbènes/pharmacologie , Régulation positive/effets des médicaments et des substances chimiques , AMP-Activated Protein Kinases/métabolisme , Animaux , Lignée cellulaire , Survie cellulaire/effets des médicaments et des substances chimiques , Chloroquine/pharmacologie , Régulation négative/effets des médicaments et des substances chimiques , Hyperglycémie/anatomopathologie , L-Lactate dehydrogenase/métabolisme , Souris , Resvératrol , Sérine-thréonine kinases TOR/métabolismeRÉSUMÉ
The objective of this study was to determine the combination of left ventricular ejection fraction (LVEF) and individual electrocardiographic parameters related to abnormal depolarization/repolarization or baroreceptor sensitivity that had the best predictive value for major adverse cardiac events (MACE) in patients with acute coronary syndrome (ACS). Patients with ACS who underwent coronary angiography and percutaneous coronary intervention (PCI) were included in this prospective study. Ventricular late potential (VLP), heart rate turbulence (HRT), heart rate variability (HRV), and T wave alternans (TWA) parameters were measured using 24 h Holter monitoring 2-4 weeks after onset of ACS. Initial and follow-up LVEF was measured by ultrasound. Patients were followed for at least 6 months to record the occurrence of MACE. Models using combinations of the individual independent prognostic factors found by multivariate analysis were then constructed to use for estimation of risk of MACE. In multivariate analysis, VLP measured as QRS duration, HRV measured as standard deviation of normal RR intervals, and followup LVEF, but none of the other parameters studied, were independent risk factors for MACE. Areas under ROC curve (AUCs) for combinations of 2 or all 3 factors ranged from 0.73 to 0.76. Combinations of any of the three independent risk factors for MACE in ACS patients with PCI improved prediction and, because these risk factors were obtained non-invasively, may have future clinical usefulness.
RÉSUMÉ
The normal thymus contributes to T lymphocytes differentiation and induction of tolerance to self-antigens. Myasthenia gravis (MG) is characterized by abnormal thymic hyperplasia. To assess the potential influence of MG-thymus on the differentiation of T lymphocytes differentiation, we used the MG-thymus transplanted severe combined immunodeficiency (SCID) mice model to evaluate the human cord blood stem cells differentiation. Thymus fragments from MG patient and human cord blood stem cells were transplanted into SCID mice successively. SCID mice were observed to develop sustained human T lymphocytes and a functional anti-tumor immune. The levels of various T cell subsets in SCID mice with MG-thymus were different from that of control group. Among that, the frequency of CD4(+)CD25(+) T cells was significant lower in SCID mice with MG-thymus. The deficiency of CD4(+)CD25(+) T cells seens to contribute to the pathogenesis of MG.
Sujet(s)
Cellules souches hématopoïétiques/physiologie , Lymphopoïèse , Myasthénie/physiopathologie , Lymphocytes T/physiologie , Thymus (glande)/physiopathologie , Animaux , Antigènes CD34/métabolisme , Lymphocytes T CD4+/physiologie , Lignée cellulaire tumorale , Sang foetal , Cytométrie en flux , Transplantation de cellules souches hématopoïétiques , Humains , Immunohistochimie , Mâle , Souris , Souris SCID , Transplantation tumorale , Tumeurs expérimentales/immunologie , Thymus (glande)/transplantation , Transplantation hétérologueRÉSUMÉ
The aim of this study was to discuss the new methods of clinical classification and staging of patients with myasthenia gravis (MG) proposed by our group and to summarize the experiences of surgical treatment of MG with a novel incision by cutting the sternum cross-sectionally at the second intercostal level. A retrospective analysis was made for the clinical data from the patients with MG who underwent thymectomy from July 1988 to May 2009. The surgical procedures were designed into three groups, a group with Osserman classification and median incision of the sternum (Group 1), a group with MGFA typing (Myasthenia Gravis Foundation of America) and a small transverse sternal incision at the second intercostal level (Group 2), and a group with new typing and a smaller transverse sternal incision at the second intercostal level (Group 3). Observation of the clinical typing and staging was made in the patients with myasthenia crisis. The parameters such as procedure duration in Group 2 and 3 was significantly lower than those in Group 1 (P < 0.05). The incidence of myasthenia crisis in Group 3 was significantly lower than that in Groups 2 and 3 (P < 0.05). The procedure with a smaller transverse sternal incision at the second intercostal level (Group 3) is a safer method for patients with MG. The combination of this procedure with the new typing and staging methods proposed by our group could facilitate the selection of operation indications and opportunity, resulting in the lower incidence of myasthenia crisis and mortality. Our new procedure is well deserved to be a preferential selection by other hospitals.
Sujet(s)
Myasthénie/classification , Myasthénie/chirurgie , Sternum/chirurgie , Thymectomie/méthodes , Adolescent , Adulte , Perte sanguine peropératoire , Études de cohortes , Femelle , Études de suivi , Humains , Durée du séjour , Mâle , Adulte d'âge moyen , Interventions chirurgicales mini-invasives/méthodes , Myasthénie/diagnostic , Durée opératoire , Soins postopératoires/méthodes , Soins préopératoires/méthodes , Études rétrospectives , Appréciation des risques , Thymome/chirurgie , Tumeurs du thymus/chirurgie , Résultat thérapeutique , Jeune adulteRÉSUMÉ
AIM: To study the effects of CD74 gene on IFN-γR expression in myasthenia gravis thymic epithelial cell(TEC). METHODS: PCMV-CD74 transiently transfect to primary cultured TEC mediated with lipofectamine, the result of transfection and some autoimmune related genes such as IFN-γR were detected by real-time RT-PCR. RESULTS: Real-time quantitative RT-PCR results show that the mRNA expression level of IL-4R, IL-32, IFN-γR, ECGF1, HLA-A, HLA-DR increased significantly, with the high express ion of CD74 gene in thymic epithelial cells. CONCLUSION: CD74 gene over-expression in TEC can increase IFN-γR mRNA expression.
Sujet(s)
Antigènes de différenciation des lymphocytes B/génétique , Cellules épithéliales/métabolisme , Régulation de l'expression des gènes/génétique , Antigènes d'histocompatibilité de classe II/génétique , Myasthénie/génétique , Récepteur interféron/génétique , Thymus (glande)/anatomopathologie , Humains , Immunomodulation/génétique , Mâle , Myasthénie/immunologie , Plasmides/génétique , ARN messager/génétique , ARN messager/métabolisme , Transfection , Jeune adulte ,RÉSUMÉ
OBJECTIVE: To investigate potential pathophysiological role of cardiac mast cells accumulation and degranulation on the collagen deposition after coronary microembolization (CME). METHODS: CME was induced in miniswine by selective infusion of 15X10(4) microspheres (diameter, 45 mum) into the left anterior descending artery groups (CME group, n=8). Some CME-induced animals were pretreated with the MC stabilizer tranilast (50 mg/kg, twice daily), beginning 2 weeks before CME and thereafter throughout the experimental period (CME +tranilast group, n=8), while some animals received tranilast without CME (tranilast group, n=8). Eight sham-operated animals without CME served as controls. After 30 days, the total number of MC and degranulating MCs and collagen deposition was assessed by histological and electronic microscopy studies. RESULTS: The numbers of total and degranulating MCs and collagen volume fraction (CVF) at day 30 in CME group were significantly higher than those in controls (P <0.01). Treatment with tranilast significantly reduced the numbers of total and degranulating MCs and CVF at day 30 (all P <0.01). There was a significant positive correlation of the CVF with the number of total MCs (r=0.91, P <0.001) and degranulating MCs (r=0.92, P <0.001) over the CME myocardium. CONCLUSION: MCs accumulation and degranulating contribute to myocardial fibrosis collagen deposition.
Sujet(s)
Dégranulation cellulaire , Collagène/métabolisme , Vaisseaux coronaires/anatomopathologie , Embolie/anatomopathologie , Mastocytes/physiologie , Myocarde/anatomopathologie , Animaux , Vaisseaux coronaires/physiopathologie , Embolie/physiopathologie , Mastocytes/anatomopathologie , Myocarde/métabolisme , Suidae , Porc miniatureRÉSUMÉ
1. Coronary microembolisation (CME) is associated with progressive myocardial dysfunction, and mast cells (MC) might have an important role in myocardial apoptosis after CME. We investigated whether the MC stabilizer tranilast suppresses the accumulation and degranulation of MC while reducing cardiomyocyte apoptosis after CME. 2. We induced CME in miniswine by selective infusion of 15 x 10(4) microspheres (diameter 45 microm) into the left anterior descending artery. Some CME-induced miniswine were treated with the MC stabilizer tranilast (50 mg/kg, p.o., b.d.) beginning 2 weeks before CME, and thereafter throughout the experimental period; others received tranilast without CME; and sham-operated animals without CME served as controls. After 30 days, we assessed cardiomyocyte apoptosis by TUNEL assay and by the total number of MC and the number of degranulating MC using histology and transmission electron microscopy. The wall motion score index and left ventricular ejection fraction were studied by dobutamine stress echocardiography. 3. Coronary microembolisation was associated with increases in the total number of MC, the number of degranulating MC, and myocyte apoptosis. The number of total MC and degranulating MC and apoptotic cardiomyocytes over the anterior embolized myocardium after CME were significantly higher than those over the posterior control myocardium and anterior segments per animal without CME (P < 0.01). Tranilast administration to CME miniswine suppressed cardiomyocyte apoptosis while maintaining regional and global function, which was associated with reductions in the accumulation and degranulation of MC. 4. These findings suggest that tranilast suppresses the accumulation and degranulation of MC while reducing cardiomyocyte apoptosis after CME.
Sujet(s)
Apoptose/effets des médicaments et des substances chimiques , Dégranulation cellulaire/effets des médicaments et des substances chimiques , Vaisseaux coronaires/effets des médicaments et des substances chimiques , Embolie/traitement médicamenteux , Mastocytes/effets des médicaments et des substances chimiques , Myocytes cardiaques/effets des médicaments et des substances chimiques , ortho-Aminobenzoates/usage thérapeutique , Animaux , Vaisseaux coronaires/ultrastructure , Modèles animaux de maladie humaine , Échocardiographie de stress , Embolie/imagerie diagnostique , Embolie/anatomopathologie , Femelle , Méthode TUNEL , Mâle , Mastocytes/physiologie , Mastocytes/ultrastructure , Microscopie électronique à transmission , Myocytes cardiaques/ultrastructure , Suidae , Porc miniature , ortho-Aminobenzoates/administration et posologie , ortho-Aminobenzoates/pharmacologieRÉSUMÉ
AIM: To study the relationship between the abnormal differentiation thymocytes and the pathogenesis of myasthenia gravis (MG). METHODS: The gene expression of some ILs, IFN and their receptors was examined by cDNA array. The percentage of CD45RO(+) and D45RA(+) thymocytes was measured by flow cytometry (FCM). The expression and distribution of CD45RA and CD45RO in the samples of thymus tissues were observed by immunohistochemistry. RESULTS: The levels of IL-1R, IL-4R, IFNgammaR1, IFNgammaR2, IL-6 and IL-8 in the patients with MG were lower than those of normal controls and there were significant differences between them. But the levels of IL-1, IL-2, IL-4, IL-10, IL-7, IFN-alpha, IFN-beta and IFN-gamma showed no significant changes in patients with MG in comparison with normal controls. The percentage of CD56(+) thymocytes in MG patients 0.56+/-0.33 was significantly lower than that in normal controls(1.78+/-0.69).The CD45RO(+) and CD1a(+) in MG patients increased more significantly those in normal controls. The immunohistochemistry and RT-PCR showed the same result. CONCLUSION: Aberrant transformation from CD45R0(+)T to CD45RA(+)T has been found in the development of thymocytes of the patients with MG.
Sujet(s)
Cytokines/génétique , Antigènes CD45/métabolisme , Myasthénie/métabolisme , Thymus (glande)/métabolisme , Adolescent , Adulte , Enfant , Enfant d'âge préscolaire , Femelle , Cytométrie en flux , Humains , Immunohistochimie , Interféron alpha/génétique , Interféron bêta/génétique , Interféron gamma/génétique , Interleukine-1/génétique , Interleukine-10/génétique , Interleukine-2/génétique , Interleukine-4/génétique , Interleukine-6/génétique , Interleukine-7/génétique , Interleukine-8/génétique , Mâle , Adulte d'âge moyen , Myasthénie/génétique , Séquençage par oligonucléotides en batterie , Récepteur interféron/génétique , Récepteurs à l'interleukine-1/génétique , Récepteurs à l'interleukine-4/génétique , RT-PCR , Jeune adulte ,RÉSUMÉ
OBJECTIVE: To determine the optimal bone marrow (BM) cell types, and their potential mechanisms of action for neovascularization in chronic ischaemic myocardium. METHODS AND RESULTS: The functional effects, angiogenic potential and cytokine expression of direct intramyocardial implantation of autologous BM CD31-positive endothelial progenitor cells (EPC, n=9), BM mononuclear cells (MNCs, n=9), and saline (n=9) were compared in a swine model of chronic ischaemic myocardium. Autologous BM cells were harvested and catheter-based electromechanical mapping-guided direct intramyocardial injection was performed to target ischaemic myocardium. After 12 weeks, injection of BM-MNC resulted in significant improvements in left ventricular dP/dt (+21+/-8%, P=0.032), left ventricular pressure (+17+/-4%, P=0.048) and regional microsphere myocardial perfusion over ischaemic endocardium (+74+/-28%, P<0.05) and epicardium (+73+/-29%, P<0.05). No significant effects were observed following injection of BM-EPC or saline. Capillary density (1132+/-69 versus 903+/-44 per mm(2), P=0.047) and expression of mRNA of vascular endothelial growth factor (VEGF, 32.3+/-5.6 versus 13.1+/-3.7, P<0.05,) and angiopoietin-2 (23.9+/-3.6 versus 13.7+/-3.1, P<0.05) in ischaemic myocardium was significantly greater in the BM-MNC group than the saline group. The capillary density in ischaemic myocardium demonstrated a significant positive correlation with VEGF expression (r=0.61, P<0.001). CONCLUSION: Catheter-based direct intramyocardial injection of BM-MNC enhanced angiogenesis more effectively than BM-EPC or saline, possibly via a paracrine effect, with increased expression of VEGF that subsequently improved cardiac performance of ischaemic myocardium.
Sujet(s)
Transplantation de moelle osseuse/méthodes , Transplantation de moelle osseuse/physiologie , Agranulocytes/physiologie , Ischémie myocardique/physiopathologie , Revascularisation myocardique/méthodes , Cellules souches/physiologie , Animaux , Numération cellulaire , Différenciation cellulaire , Maladie chronique , Techniques électrophysiologiques cardiaques , Cytométrie en flux , Immunohistochimie , Microsphères , Myocarde/métabolisme , Néovascularisation physiologique , Communication paracrine , Suidae , Porc miniature , Facteur de croissance endothéliale vasculaire de type A/métabolisme , Fonction ventriculaire gaucheRÉSUMÉ
OBJECTIVE: To investigate the relationship between endothelin-1 (ET-1) and tumor necrosis factor-alpha and myocardial microcircular dysfunction during coronary microembolization (CME). METHODS: CME was induced in 10 miniswine by selective infusion of microspheres (45 microm) into left anterior descending artery (LAD). We measured (1) coronary sinus level of ET-1, TNF-alpha using radioimmunoassay; (2) CFR, a measure of microvascular integrity, using Doppler flow wire in LAD at baseline and different doses of microspheres. RESULTS: CFR decrease significantly with different doses of microspheres (vs. baseline, P < 0.05). Level of ET-1, TNF-alpha increased significantly with doses of 5 x 10(4) and peaked with 10 x 10(4). Interestingly, ET-1 progressively decrease while TNF-alpha persistently elevated from doses of 12 x 10(4) to 15 x 10(4). There are reverse correlations between ET-1 and CFR (r = -0.31, P < 0.05). CONCLUSIONS: The extent of microvascular injury wasn't linearly related to the extent of ME, where, it closely associated with myocardial ET-1.
Sujet(s)
Thrombose coronarienne/physiopathologie , Endothéline-1/sang , Microcirculation/physiopathologie , Facteur de nécrose tumorale alpha/sang , Animaux , Modèles animaux de maladie humaine , Suidae , Porc miniatureRÉSUMÉ
BACKGROUND: The normal cardiac rhythm originates in the sinoatrial (SA) node that anatomically resides in the right atrium. Malfunction of the SA node leads to various forms of arrhythmias that necessitate the implantation of electronic pacemakers. We hypothesized that overexpression of an engineered HCN construct via somatic gene transfer offers a flexible approach for fine-tuning cardiac pacing in vivo. METHODS AND RESULTS: Using various electrophysiological and mapping techniques, we examined the effects of in situ focal expression of HCN1-DeltaDeltaDelta, the S3-S4 linker of which has been shortened to favor channel opening, on impulse generation and conduction. Single left ventricular cardiomyocytes isolated from guinea pig hearts preinjected with the recombinant adenovirus Ad-CMV-GFP-IRES-HCN1-DeltaDeltaDelta in vivo uniquely exhibited automaticity with a normal firing rate (237+/-12 bpm). High-resolution ex vivo optical mapping of Ad-CGI-HCN1-DeltaDeltaDelta-injected Langendorff-perfused hearts revealed the generation of spontaneous action potentials from the transduced region in the left ventricle. To evaluate the efficacy of our approach for reliable atrial pacing, we generated a porcine model of sick-sinus syndrome by guided radiofrequency ablation of the native SA node, followed by implantation of a dual-chamber electronic pacemaker to prevent bradycardia-induced hemodynamic collapse. Interestingly, focal transduction of Ad-CGI-HCN1-DeltaDeltaDelta in the left atrium of animals with sick-sinus syndrome reproducibly induced a stable, catecholamine-responsive in vivo "bioartificial node" that exhibited a physiological heart rate and was capable of reliably pacing the myocardium, substantially reducing electronic pacing. CONCLUSIONS: The results of the present study provide important functional and mechanistic insights into cardiac automaticity and have further refined an HCN gene-based therapy for correcting defects in cardiac impulse generation.
Sujet(s)
Troubles du rythme cardiaque/chirurgie , Organes bioartificiels , Canaux ioniques/génétique , Pacemaker , Maladie du sinus/chirurgie , Animaux , Troubles du rythme cardiaque/physiopathologie , Canaux cationiques contrôlés par les nucléotides cycliques , Modèles animaux de maladie humaine , Électrophysiologie , Techniques de transfert de gènes , Cochons d'Inde , Rythme cardiaque , Canaux contrôlés par les nucléotides cycliques et activés par l'hyperpolarisation , Souris , Canaux potassiques , Maladie du sinus/physiopathologie , Noeud sinuatrial/physiologie , Suidae , Porc miniatureRÉSUMÉ
Coronary microembolization (CME) is associated with progressive myocardial dysfunction despite restoration of coronary flow reserve (CFR). The potential pathophysiological role of mast cells (MCs) remains unclear. Therefore, we induced CME in 18 miniswines and determined whether MC accumulation occurs and their effects on local cytokine secretion [interleukin (IL)-6, IL-8, tumor necrosis factor-alpha (TNF-alpha)]; cardiomyocyte apoptosis; and collagen formation at day 1 (D1), day 7 (D7), and day 30 (D30) after CME. Four sham-operated animals without CME (controls) and six animals treated with a MC stabilization agent (tranilast) for 30 days after CME were also studied. CFR decreased at D1 but returned to baseline level at D7 and D30. Coronary sinus levels of IL-6, IL-8, and TNF-alpha increased significantly at D1 and D7 (p<0.01 vs baseline). Levels of IL-6 and IL-8 at D30 returned to baseline level, but not those of TNF-alpha. The numbers of total and degranulating MCs, % apoptotic cardiomyocytes, and collagen volume fraction (CVF) over CME myocardium at D1, D7, and D30 were significantly higher than controls (p<0.01). Treatment with tranilast significantly reduced the serum level of TNF-alpha, numbers of total and degranulating MCs, % apoptotic cardiomyocytes, and CVF at D30 (all p<0.05). There was a significant positive correlation between the numbers of MCs with % apoptotic cardiomyocytes (r = 0.77, p<0.001) and CVF (r = 0.75, p<0.001) over the CME myocardium. Despite restoration of CFR, cardiomyocyte apoptosis persisted after CME and was positively correlated with the number of MCs but was prevented with tranilast treatment. These findings suggest that MCs contribute to cardiomyocyte apoptosis after CME.
Sujet(s)
Apoptose , Vaisseaux coronaires/anatomopathologie , Embolie/anatomopathologie , Mastocytes/physiologie , Myocytes cardiaques/physiologie , Animaux , Collagène/métabolisme , Circulation coronarienne , Vaisseaux coronaires/physiopathologie , Embolie/physiopathologie , Femelle , Interleukine-6/métabolisme , Interleukine-8/métabolisme , Mâle , Mastocytes/effets des médicaments et des substances chimiques , Microscopie électronique à transmission , Myocarde/métabolisme , Myocarde/anatomopathologie , Myocarde/ultrastructure , Suidae , Porc miniature , Facteur de nécrose tumorale alpha/métabolisme , ortho-Aminobenzoates/pharmacologieRÉSUMÉ
OBJECTIVE: To investigate the effect of coronary microemboliation (CME) on coronary microvascular injury and myocardial endothelin-1 (ET-1) level. METHODS: CME was induced in 10 miniswines by selective infusion of microspheres (45 microm) into left anterior descending artery (LAD). The ET-1 level in coronary sinus was measured with radioimmunoassay. The microvascular integrity indicator CFR was measured by Doppler flow wire in LAD at baseline and after infusion of microspheres. RESULT: Compared to the baseline, CFR decreased significantly with different doses of microspheres. ET-1 level increased significantly with doses of 5 x 10(4) and peaked with 10 x 10(4), and progressively decreased from doses of 12 x 10(4) to 15 x 10(4) microspheres. There was negative correlation between ET-1 and CFR (r=-0.31, P=0.02). CONCLUSION: The extent of microvascular injury is not linearly related to the extent of microembolization, but it is closely associated with myocardial ET-1 level.
Sujet(s)
Vaisseaux coronaires/anatomopathologie , Endothéline-1/sang , Infarctus du myocarde , Animaux , Modèles animaux de maladie humaine , Embolie , Femelle , Mâle , Infarctus du myocarde/sang , Infarctus du myocarde/anatomopathologie , Suidae , Porc miniatureRÉSUMÉ
OBJECTIVE: To assess the microvascular function of coronary artery after intracoronary microembolization using coronary resistance system. METHODS: The left anterior descending coronary artery (LAD) of 10 pigs weighing 21 kg-25 kg were embolized by repetitive injection of microspheres 45 micro m in diameter through a 2.8F Tracker catheter. Intra-vascular ultrasound (IVUS) images, intracoronary Doppler and pressure signals in the middle segment of LAD were acquired by use of intracoronary ultrasound imaging catheter, Doppler flow wire and pressure wire separately. Intracoronary bolus injection of 18 micro g adenosine was administered to maximally vasodilate the coronary arterial bed through the 2.8F Tracker catheter. The resting and hyperemic signals were acquired respectively before microembolization and in different levels of microembolization. Coronary resistance system reflecting the resistance to pulsatile coronary flow was established by a self-made software of PC system. The resting and hyperemic CR parameters included average resting coronary resistance (rCR) and average minimal coronary resistance (min-CR), the first-harmonic rCR and min-CR, the first-harmonic rCR orientation and min-CR orientation, and so on. Factor analysis was performed to extract the best coronary parameter from the coronary resistance parameters. RESULTS: Factor analysis showed that the first-harmonic rCR and first-harmonic min-CR were correlated better with the first component extracted from the resting and hyperemic CR parameters than rCR and min-CR, with the correlation coefficient being 0.913 and 0.950 in the first-harmonic CR and first-harmonic min-CR respectively. No significant difference in min-CR was found between the value at the dosage of 5 x 10(4) microspheres and that before microembolization. The min-CR value increased markedly from 271 mm Hg.ml(-1).s(-1) +/- 99 mm Hg.ml(-1).s(-1) at the dosage of injecting 5 x 10(4) microspheres to 361 mm Hg.ml(-1).s(-1) +/- 158 mm Hg.ml(-1).s(-1) at the dosage of injecting 10 x 10(4) microspheres (P < 0.05). The min-CR value remained almost unchanged from the dosage of 10 x 10(4) to 15 x 10(4) microspheres. There was no significant difference concerning the first-harmonic min-CR between the value at the dosage of 5 x 10(4) microspheres and that before microembolization. Along with the increase of number of microspheres injected the min-CR value increased gradually. The min-CR value was increased significantly than that before microembolization since the number of microspheres injected surpassed 14 x 10(4). CONCLUSION: The first-harmonic min-CR reflected the coronary microvascular dysfunction in different extents of microembolization better than min-CR. The extent of coronary microvascular dysfunction wasn't linearly related to the extent of microembolization.
Sujet(s)
Vaisseaux coronaires/physiopathologie , Embolie/anatomopathologie , Résistance vasculaire , Animaux , Vaisseaux coronaires/anatomopathologie , Modèles animaux de maladie humaine , Embolie/étiologie , Femelle , Mâle , Suidae , Échographie-dopplerRÉSUMÉ
AIM: To explore the roles of abnormalities of thymocytic abnormal apoptosis and its Fas expression in patients with myasthenia gravis (MG) in generation of autoimmune response. METHODS: The thymocytes were isolated and thymus extractive was prepared from fresh thymic tissue excised from MG patients. The proliferation of thymocyes was evaluated by MTT colorimetry. The apoptosis of thymocytes were examined by DNA electrophoresis. Expression of Fas was analyzed by a FACScan flow cytometry. Transcription and mutation of Fas gene were detected by RT-PCR and single-stranded conformation polymorphism ( SSCP) analysis. RESULTS: Thymus extractive could inhibit proliferation of normal thymocytes, while had no influence on the thymocytic proliferation of MG patients. The proliferation of thymocytes of both normal persons and MG patients could be inhibited by thymus extractive in the presence of dexamethasone. The inhibition rates were 58. 33% and 54. 26% respectively. The cocultured thymocytes with dexamethasone showed characteristic apoptosis pattern and the DNA electrophoresis exhibited 'ladder' appearance. The abnormal bands in Fas mRNA of partial MG patients' thymocytes were found by RT-PCR-SSCP analysis. CONCLUSION: Abnormalities of apoptosis and Fas gene expression of MG patients' thymocytes, and Fas gene mutation may be related to the pathogenesis and progression of MG.