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The Effects of Bone Marrow Stromal Cell Infusion after Myocardial Infarction Through Cardiac Vein in Swine
The 9th Annual Scientific Meeting
•
JHFS
S291
O-065
O-067
New Tissue Stem Cell Source for Cardiomyocyte Regeneration
Regulation of Left Ventricular Remodeling and Regeneration by ACE Inhibitor Following Donor Heart Myocardial Infarction with Heterotopic Transplant-Coronary Ligation Model HIROSHI NAKAMURA, ATSUSHI MATSUSHIMA, YASUHIRO IKEDA, MASUNORI MATSUZAKI Department of Cardiovascular Medicine, Yamaguchi University School of Medicine, Yamaguchi, Japan
MASAKI SHIBUYA, TOSHIRO MIURA, RUIJUAN WANG, MASUNORI MATSUZAKI Department of Cardiovascular Medicine, Yamaguchi University Graduate School of Medicine, Ube, Japan Background: Recently, tissue stem cells which can differentiate into cardiomyocyte were identified from heart, bone marrow, skeletal muscle, and so on. However, the efficiency of differentiation into cardiomyocyte is very low. Skeletal myoblast is a promising cell source,but it may be arrhythmogenic because of the lack of gap junction protein. Thus new tissue stem cells which can express gap junction protein and beat spontaneously are desired. Purpose: We search new tissue stem cells which can develop into cardiomyocyte. Method: Heart, thigh muscle and tongue muscle of 6-8W C57/BL6 mouse were dissociated with 0.2% collagenase and the SCA-1 positive cells were selected by the magnetic cell sorting system. These cells were cultured in DMEM⫹10%FBS with growth factors. The gene expression of cardiomyocyte specific markers, NKX2.5, connexin 43, ANP were examined by RT-PCR and immunostaining at 1 and 2 weeks. Result: The Sca-1(⫹) cells only from tongue muscle can developed into beating cells in 1-2 weeks with the gene expression of cardiomyocyte specific markers. Conclusion: The tongue derived tissue stem cells may be a novel source for cardiomyocyte regeneration.
The present study investigated the new insights into the pathologic role of ACE inhibitor on cardiac remodeling and regeneration at the late stage of myocardial infarction (MI). Methods: We performed isogenic heterotopic cardiac transplantation in the GFP mice (C57BL/6 background) and simultaneous coronary ligation to produce MI in the C57BL/6 donor heart, and to evaluate the donor heart at the 60 days after the transplantation. To examine the effects of the ACE inhibitor on LV remodeling and regeneration, five mice in the ligation group were orally treated with perindopril(PE) at a dose of 1mg/kg/day after the operation. On days 60, GFP⫹ myosin⫹ large cells were evaluated. Results: Although a significant increase in donor heart/body weight ratio compared with the sham group were observed in the ligation group on day 60. GFP⫹ myosin⫹ large cells were mainly localized around the site of pericardium of the donor heart. Maximum GFP⫹ myosin⫹ large cells counts treated with PE was significantly higher among 3 groups (sham vs. ligation vs. PE: 0.3⫾0.5 vs. 3.0⫾0.8 vs. 9.8⫾5.5 cells/LPF, P⬍0.05). These results suggested that PE might promote cardiac regeneration in the donor heart and prevent the LV remodeling. Conclusions: Modification of cardiac regeneration might be a beneficial strategy for the treatment of cardiac dysfunction and subsequent cardiac remodeling at the late phase of MI.
O-066
O-068
The Effects of Bone Marrow Stromal Cell Infusion after Myocardial Infarction Through Cardiac Vein in Swine
Transcriptional Control of the Mitochondrial Gene Determines the Cardiac Contractile: Possible Involvement of SERCA2 Gene Activation by Mitochondrial Transcription Factors ATAI WATANABE, MASASHI ARAI, NORIMICHI KOITABASHI, KAZUO NIWANO, MASAHIKO KURABAYASHI Department of Medicine and Biological Science, Gunma University Graduate School of Medicine, Maebashi, Japan
TAKATOSHI SATO1, HIROSHI SUZUKI1, TAROU KUSUYAMA1, YASUTOSHI OHMORI1, TERUKO SODA1, FUMIYOSHI TSUNODA1, MAKOTO SHOJI1, YOSHITAKA ISO1, SHINJI KOBA1, EIICHI GESHI1, TAKASHI KATAGIRI1, KOHEI WAKABAYASHI2, YOUICHI TAKEYAMA2, TAROU UYAMA3, AKIHIRO UMESAWA3 1 Third Department of Internal Medicine, Showa University School of Medicine, Tokyo, Japan, 2Division of Cardiology, Department of Internal Medicine, Showa University Fujigaoka Hospital, Kanagawa, Japan, 3Department of Reproductive Biology and Pathology, National Research Institute for Child Health and Development, Tokyo, Japan Purpose: Recently, bone marrow stromal cell (BMSC) is focused on as the useful source of cell transplantation even in cardiovascular field. However, there is a few reports examined the effects in large animal, and the useful transplantation method has not been explored. In this study, we investigated the effects of BMSC infusion from coronary vein in swine myocardial infarction model. Methods: To generate myocardial infarction, beads were placed in left anterior descending coronary artery of domestic swine. Bone marrow cells were aspirated, and were cultivated in BMSC culture medium. Four weeks later, BMSC, labeled with dye were infused retrogradely from anterior interventricular vein. Medium was infused in control group. Left ventriculography was performed just before and 4 weeks after cell infusion before sacrifice. In immunohistochemistry, anti-alpha smooth muscle actin antibody was used, and vessel number was calculated in border area. Immunofluorescent staining was performed to detect BMSC. Results: In left ventriculography, % change of ejection fraction was significantly improved in BMSC group as compared with control group (p⬍0.05). At 4 weeks, BMSC detected by red fluorescence were observed in myocardium. In immunohistochemistry, number of alpha smooth muscle actin positive vessels was significantly larger in BMSC group in border area (p⬍0.05). Conclusions: BMSC infusion from coronary vein may improve cardiac function after myocardial infarction through inducing angiogenesis.
Background: Expression of mitochondrial (Mt) proteins is controlled by Mt-transcriptional machinery composed of Tfam, Tfb2m and Mt-RNA polymerase. Correlations among Mt-transcription factors, each Mt respiratory complex, SERCA2 mRNA level, and systolic and diastolic function were determined in rat failing heart. We next tested the hypothesis that Mt-transcription factors regulate the transcription of the SERCA2 gene as well as Mt-genes. Results: Seven days after myocardial infarction in rat, the amount of Mtcomplex I and III was significantly correlated with fractional shortening (I, r⫽0.62; III, r⫽0.47), LVEDP (⫺0.50, ⫺0.77), ⫹dP/dt (0.48, 0.60) and tau (⫺0.61, ⫺0.80). Tfam, Tfb2m and Mt-RNA polymerase mRNAs were correlated with mRNAs for complex I and III. Interestingly, SERCA2 mRNA level was significantly correlated with Tfam (r⫽0.53), Tfb2m (r⫽0.77) and Mt-RNA polymerase (r⫽0.74) and with systolic and diastolic function. Robust expression of Tfam, Tfb2m and Mt-RNA polymerase in cultured myocytes activated SERCA2 gene transcription by 2.43 fold followed by 1.32 times increase of SERCA2 protein. Additionally, serial deletion of the SERCA2 promoter revealed that binding sites of these transcription factors located between ⫺284 and ⫺72 region. Conclusions: Our study suggested that the transcriptional activity of the Mt-gene determines the systolic and diastolic function in the failing heart. More importantly, Mt-transcription factors may regulate E-C coupling not only by producing ATP-generating Mt-proteins but also by increasing SERCA2 protein.
•
JHFS
S291
O-065
O-067
New Tissue Stem Cell Source for Cardiomyocyte Regeneration
Regulation of Left Ventricular Remodeling and Regeneration by ACE Inhibitor Following Donor Heart Myocardial Infarction with Heterotopic Transplant-Coronary Ligation Model HIROSHI NAKAMURA, ATSUSHI MATSUSHIMA, YASUHIRO IKEDA, MASUNORI MATSUZAKI Department of Cardiovascular Medicine, Yamaguchi University School of Medicine, Yamaguchi, Japan
MASAKI SHIBUYA, TOSHIRO MIURA, RUIJUAN WANG, MASUNORI MATSUZAKI Department of Cardiovascular Medicine, Yamaguchi University Graduate School of Medicine, Ube, Japan Background: Recently, tissue stem cells which can differentiate into cardiomyocyte were identified from heart, bone marrow, skeletal muscle, and so on. However, the efficiency of differentiation into cardiomyocyte is very low. Skeletal myoblast is a promising cell source,but it may be arrhythmogenic because of the lack of gap junction protein. Thus new tissue stem cells which can express gap junction protein and beat spontaneously are desired. Purpose: We search new tissue stem cells which can develop into cardiomyocyte. Method: Heart, thigh muscle and tongue muscle of 6-8W C57/BL6 mouse were dissociated with 0.2% collagenase and the SCA-1 positive cells were selected by the magnetic cell sorting system. These cells were cultured in DMEM⫹10%FBS with growth factors. The gene expression of cardiomyocyte specific markers, NKX2.5, connexin 43, ANP were examined by RT-PCR and immunostaining at 1 and 2 weeks. Result: The Sca-1(⫹) cells only from tongue muscle can developed into beating cells in 1-2 weeks with the gene expression of cardiomyocyte specific markers. Conclusion: The tongue derived tissue stem cells may be a novel source for cardiomyocyte regeneration.
The present study investigated the new insights into the pathologic role of ACE inhibitor on cardiac remodeling and regeneration at the late stage of myocardial infarction (MI). Methods: We performed isogenic heterotopic cardiac transplantation in the GFP mice (C57BL/6 background) and simultaneous coronary ligation to produce MI in the C57BL/6 donor heart, and to evaluate the donor heart at the 60 days after the transplantation. To examine the effects of the ACE inhibitor on LV remodeling and regeneration, five mice in the ligation group were orally treated with perindopril(PE) at a dose of 1mg/kg/day after the operation. On days 60, GFP⫹ myosin⫹ large cells were evaluated. Results: Although a significant increase in donor heart/body weight ratio compared with the sham group were observed in the ligation group on day 60. GFP⫹ myosin⫹ large cells were mainly localized around the site of pericardium of the donor heart. Maximum GFP⫹ myosin⫹ large cells counts treated with PE was significantly higher among 3 groups (sham vs. ligation vs. PE: 0.3⫾0.5 vs. 3.0⫾0.8 vs. 9.8⫾5.5 cells/LPF, P⬍0.05). These results suggested that PE might promote cardiac regeneration in the donor heart and prevent the LV remodeling. Conclusions: Modification of cardiac regeneration might be a beneficial strategy for the treatment of cardiac dysfunction and subsequent cardiac remodeling at the late phase of MI.
O-066
O-068
The Effects of Bone Marrow Stromal Cell Infusion after Myocardial Infarction Through Cardiac Vein in Swine
Transcriptional Control of the Mitochondrial Gene Determines the Cardiac Contractile: Possible Involvement of SERCA2 Gene Activation by Mitochondrial Transcription Factors ATAI WATANABE, MASASHI ARAI, NORIMICHI KOITABASHI, KAZUO NIWANO, MASAHIKO KURABAYASHI Department of Medicine and Biological Science, Gunma University Graduate School of Medicine, Maebashi, Japan
TAKATOSHI SATO1, HIROSHI SUZUKI1, TAROU KUSUYAMA1, YASUTOSHI OHMORI1, TERUKO SODA1, FUMIYOSHI TSUNODA1, MAKOTO SHOJI1, YOSHITAKA ISO1, SHINJI KOBA1, EIICHI GESHI1, TAKASHI KATAGIRI1, KOHEI WAKABAYASHI2, YOUICHI TAKEYAMA2, TAROU UYAMA3, AKIHIRO UMESAWA3 1 Third Department of Internal Medicine, Showa University School of Medicine, Tokyo, Japan, 2Division of Cardiology, Department of Internal Medicine, Showa University Fujigaoka Hospital, Kanagawa, Japan, 3Department of Reproductive Biology and Pathology, National Research Institute for Child Health and Development, Tokyo, Japan Purpose: Recently, bone marrow stromal cell (BMSC) is focused on as the useful source of cell transplantation even in cardiovascular field. However, there is a few reports examined the effects in large animal, and the useful transplantation method has not been explored. In this study, we investigated the effects of BMSC infusion from coronary vein in swine myocardial infarction model. Methods: To generate myocardial infarction, beads were placed in left anterior descending coronary artery of domestic swine. Bone marrow cells were aspirated, and were cultivated in BMSC culture medium. Four weeks later, BMSC, labeled with dye were infused retrogradely from anterior interventricular vein. Medium was infused in control group. Left ventriculography was performed just before and 4 weeks after cell infusion before sacrifice. In immunohistochemistry, anti-alpha smooth muscle actin antibody was used, and vessel number was calculated in border area. Immunofluorescent staining was performed to detect BMSC. Results: In left ventriculography, % change of ejection fraction was significantly improved in BMSC group as compared with control group (p⬍0.05). At 4 weeks, BMSC detected by red fluorescence were observed in myocardium. In immunohistochemistry, number of alpha smooth muscle actin positive vessels was significantly larger in BMSC group in border area (p⬍0.05). Conclusions: BMSC infusion from coronary vein may improve cardiac function after myocardial infarction through inducing angiogenesis.
Background: Expression of mitochondrial (Mt) proteins is controlled by Mt-transcriptional machinery composed of Tfam, Tfb2m and Mt-RNA polymerase. Correlations among Mt-transcription factors, each Mt respiratory complex, SERCA2 mRNA level, and systolic and diastolic function were determined in rat failing heart. We next tested the hypothesis that Mt-transcription factors regulate the transcription of the SERCA2 gene as well as Mt-genes. Results: Seven days after myocardial infarction in rat, the amount of Mtcomplex I and III was significantly correlated with fractional shortening (I, r⫽0.62; III, r⫽0.47), LVEDP (⫺0.50, ⫺0.77), ⫹dP/dt (0.48, 0.60) and tau (⫺0.61, ⫺0.80). Tfam, Tfb2m and Mt-RNA polymerase mRNAs were correlated with mRNAs for complex I and III. Interestingly, SERCA2 mRNA level was significantly correlated with Tfam (r⫽0.53), Tfb2m (r⫽0.77) and Mt-RNA polymerase (r⫽0.74) and with systolic and diastolic function. Robust expression of Tfam, Tfb2m and Mt-RNA polymerase in cultured myocytes activated SERCA2 gene transcription by 2.43 fold followed by 1.32 times increase of SERCA2 protein. Additionally, serial deletion of the SERCA2 promoter revealed that binding sites of these transcription factors located between ⫺284 and ⫺72 region. Conclusions: Our study suggested that the transcriptional activity of the Mt-gene determines the systolic and diastolic function in the failing heart. More importantly, Mt-transcription factors may regulate E-C coupling not only by producing ATP-generating Mt-proteins but also by increasing SERCA2 protein.