"triplet" polycistronic vectors encoding Gata4, Mef2c, and Tbx5 enhances postinfarct ventricular functional improvement compared with singlet vectors

Megumi Mathison, Vivek P. Singh, Robert P. Gersch, Maricela O. Ramirez, Austin Cooney, Stephen M. Kaminsky, Maria J. Chiuchiolo, Ahmed Nasser, Jianchang Yang, Ronald Crystal, Todd K. Rosengart

Research output: Contribution to journalArticle

25 Citations (Scopus)

Abstract

Objective: The in situ reprogramming of cardiac fibroblasts into induced cardiomyocytes by the administration of gene transfer vectors encoding Gata4 (G), Mef2c (M), and Tbx5 (T) has been shown to improve ventricular function in myocardial infarction models. The efficacy of this strategy could, however, be limited by the need for fibroblast targets to be infected 3 times-once by each of the 3 transgene vectors. We hypothesized that a polycistronic "triplet" vector encoding all 3 transgenes would enhance post infarct ventricular function compared with use of "singlet" vectors.

Methods: After validation of the polycistronic vector expression in vitro, adult male Fischer 344 rats (n = 6) underwent coronary ligation with or without intramyocardial administration of an adenovirus encoding all 3 major vascular endothelial growth factor (VEGF) isoforms (AdVEGF-All6A positive), followed 3 weeks later by the administration to AdVEGF-All6A-positive treated rats of singlet lentivirus encoding G, M, or T (1 × 105 transducing units each) or the same total dose of a GMT "triplet" lentivirus vector.

Results: Western blots demonstrated that triplet and singlet vectors yielded equivalentGMTtransgene expression, and fluorescence activated cell sorting demonstrated that triplet vectors were nearly twice as potent as singlet vectors in generating induced cardiomyocytes from cardiac fibroblasts. Echocardiography demonstrated that GMT triplet vectors were more effective than the 3 combined singlet vectors in enhancing ventricular function from postinfarct baselines (triplet, 37% ± 10%; singlet, 13% ± 7%; negative control, 9% ± 5%; P < .05).

Conclusions: These data have confirmed that the in situ administration of G, M, and T induces postinfarct ventricular functional improvement and that GMT polycistronic vectors enhance the efficacy of this strategy.

Original languageEnglish
Pages (from-to)1656-1664.e2
JournalJournal of Thoracic and Cardiovascular Surgery
Volume148
Issue number4
DOIs
Publication statusPublished - 1 Jan 2014
Externally publishedYes

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Ventricular Function
Lentivirus
Fibroblasts
Transgenes
Cardiac Myocytes
Inbred F344 Rats
Adenoviridae
Vascular Endothelial Growth Factor A
Ligation
Echocardiography
Flow Cytometry
Protein Isoforms
Western Blotting
Myocardial Infarction
Genes

ASJC Scopus subject areas

  • Surgery
  • Pulmonary and Respiratory Medicine
  • Cardiology and Cardiovascular Medicine

Cite this

"triplet" polycistronic vectors encoding Gata4, Mef2c, and Tbx5 enhances postinfarct ventricular functional improvement compared with singlet vectors. / Mathison, Megumi; Singh, Vivek P.; Gersch, Robert P.; Ramirez, Maricela O.; Cooney, Austin; Kaminsky, Stephen M.; Chiuchiolo, Maria J.; Nasser, Ahmed; Yang, Jianchang; Crystal, Ronald; Rosengart, Todd K.

In: Journal of Thoracic and Cardiovascular Surgery, Vol. 148, No. 4, 01.01.2014, p. 1656-1664.e2.

Research output: Contribution to journalArticle

Mathison, M, Singh, VP, Gersch, RP, Ramirez, MO, Cooney, A, Kaminsky, SM, Chiuchiolo, MJ, Nasser, A, Yang, J, Crystal, R & Rosengart, TK 2014, '"triplet" polycistronic vectors encoding Gata4, Mef2c, and Tbx5 enhances postinfarct ventricular functional improvement compared with singlet vectors', Journal of Thoracic and Cardiovascular Surgery, vol. 148, no. 4, pp. 1656-1664.e2. https://doi.org/10.1016/j.jtcvs.2014.03.033
Mathison, Megumi ; Singh, Vivek P. ; Gersch, Robert P. ; Ramirez, Maricela O. ; Cooney, Austin ; Kaminsky, Stephen M. ; Chiuchiolo, Maria J. ; Nasser, Ahmed ; Yang, Jianchang ; Crystal, Ronald ; Rosengart, Todd K. / "triplet" polycistronic vectors encoding Gata4, Mef2c, and Tbx5 enhances postinfarct ventricular functional improvement compared with singlet vectors. In: Journal of Thoracic and Cardiovascular Surgery. 2014 ; Vol. 148, No. 4. pp. 1656-1664.e2.
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abstract = "Objective: The in situ reprogramming of cardiac fibroblasts into induced cardiomyocytes by the administration of gene transfer vectors encoding Gata4 (G), Mef2c (M), and Tbx5 (T) has been shown to improve ventricular function in myocardial infarction models. The efficacy of this strategy could, however, be limited by the need for fibroblast targets to be infected 3 times-once by each of the 3 transgene vectors. We hypothesized that a polycistronic {"}triplet{"} vector encoding all 3 transgenes would enhance post infarct ventricular function compared with use of {"}singlet{"} vectors.Methods: After validation of the polycistronic vector expression in vitro, adult male Fischer 344 rats (n = 6) underwent coronary ligation with or without intramyocardial administration of an adenovirus encoding all 3 major vascular endothelial growth factor (VEGF) isoforms (AdVEGF-All6A positive), followed 3 weeks later by the administration to AdVEGF-All6A-positive treated rats of singlet lentivirus encoding G, M, or T (1 × 105 transducing units each) or the same total dose of a GMT {"}triplet{"} lentivirus vector.Results: Western blots demonstrated that triplet and singlet vectors yielded equivalentGMTtransgene expression, and fluorescence activated cell sorting demonstrated that triplet vectors were nearly twice as potent as singlet vectors in generating induced cardiomyocytes from cardiac fibroblasts. Echocardiography demonstrated that GMT triplet vectors were more effective than the 3 combined singlet vectors in enhancing ventricular function from postinfarct baselines (triplet, 37{\%} ± 10{\%}; singlet, 13{\%} ± 7{\%}; negative control, 9{\%} ± 5{\%}; P < .05).Conclusions: These data have confirmed that the in situ administration of G, M, and T induces postinfarct ventricular functional improvement and that GMT polycistronic vectors enhance the efficacy of this strategy.",
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T1 - "triplet" polycistronic vectors encoding Gata4, Mef2c, and Tbx5 enhances postinfarct ventricular functional improvement compared with singlet vectors

AU - Mathison, Megumi

AU - Singh, Vivek P.

AU - Gersch, Robert P.

AU - Ramirez, Maricela O.

AU - Cooney, Austin

AU - Kaminsky, Stephen M.

AU - Chiuchiolo, Maria J.

AU - Nasser, Ahmed

AU - Yang, Jianchang

AU - Crystal, Ronald

AU - Rosengart, Todd K.

PY - 2014/1/1

Y1 - 2014/1/1

N2 - Objective: The in situ reprogramming of cardiac fibroblasts into induced cardiomyocytes by the administration of gene transfer vectors encoding Gata4 (G), Mef2c (M), and Tbx5 (T) has been shown to improve ventricular function in myocardial infarction models. The efficacy of this strategy could, however, be limited by the need for fibroblast targets to be infected 3 times-once by each of the 3 transgene vectors. We hypothesized that a polycistronic "triplet" vector encoding all 3 transgenes would enhance post infarct ventricular function compared with use of "singlet" vectors.Methods: After validation of the polycistronic vector expression in vitro, adult male Fischer 344 rats (n = 6) underwent coronary ligation with or without intramyocardial administration of an adenovirus encoding all 3 major vascular endothelial growth factor (VEGF) isoforms (AdVEGF-All6A positive), followed 3 weeks later by the administration to AdVEGF-All6A-positive treated rats of singlet lentivirus encoding G, M, or T (1 × 105 transducing units each) or the same total dose of a GMT "triplet" lentivirus vector.Results: Western blots demonstrated that triplet and singlet vectors yielded equivalentGMTtransgene expression, and fluorescence activated cell sorting demonstrated that triplet vectors were nearly twice as potent as singlet vectors in generating induced cardiomyocytes from cardiac fibroblasts. Echocardiography demonstrated that GMT triplet vectors were more effective than the 3 combined singlet vectors in enhancing ventricular function from postinfarct baselines (triplet, 37% ± 10%; singlet, 13% ± 7%; negative control, 9% ± 5%; P < .05).Conclusions: These data have confirmed that the in situ administration of G, M, and T induces postinfarct ventricular functional improvement and that GMT polycistronic vectors enhance the efficacy of this strategy.

AB - Objective: The in situ reprogramming of cardiac fibroblasts into induced cardiomyocytes by the administration of gene transfer vectors encoding Gata4 (G), Mef2c (M), and Tbx5 (T) has been shown to improve ventricular function in myocardial infarction models. The efficacy of this strategy could, however, be limited by the need for fibroblast targets to be infected 3 times-once by each of the 3 transgene vectors. We hypothesized that a polycistronic "triplet" vector encoding all 3 transgenes would enhance post infarct ventricular function compared with use of "singlet" vectors.Methods: After validation of the polycistronic vector expression in vitro, adult male Fischer 344 rats (n = 6) underwent coronary ligation with or without intramyocardial administration of an adenovirus encoding all 3 major vascular endothelial growth factor (VEGF) isoforms (AdVEGF-All6A positive), followed 3 weeks later by the administration to AdVEGF-All6A-positive treated rats of singlet lentivirus encoding G, M, or T (1 × 105 transducing units each) or the same total dose of a GMT "triplet" lentivirus vector.Results: Western blots demonstrated that triplet and singlet vectors yielded equivalentGMTtransgene expression, and fluorescence activated cell sorting demonstrated that triplet vectors were nearly twice as potent as singlet vectors in generating induced cardiomyocytes from cardiac fibroblasts. Echocardiography demonstrated that GMT triplet vectors were more effective than the 3 combined singlet vectors in enhancing ventricular function from postinfarct baselines (triplet, 37% ± 10%; singlet, 13% ± 7%; negative control, 9% ± 5%; P < .05).Conclusions: These data have confirmed that the in situ administration of G, M, and T induces postinfarct ventricular functional improvement and that GMT polycistronic vectors enhance the efficacy of this strategy.

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