Survival and maturation of human embryonic stem cell-derived cardiomyocytes in rat hearts

Wangde Dai, Loren Field, Michael Rubart-von der Lohe, Sean Reuter, Sharon L. Hale, Robert Zweigerdt, Ralph E. Graichen, Gregory L. Kay, Aarne J. Jyrala, Alan Colman, Bruce P. Davidson, Martin Pera, Robert A. Kloner

Research output: Contribution to journalArticle

133 Citations (Scopus)

Abstract

Human embryonic stem cell (hESC)-derived cardiomyocytes are a promising cell source for cardiac repair. Whether these cells can be transported long distance, survive, and mature in hearts subjected to ischemia/reperfusion with minimal infarction is unknown. Taking advantage of a constitutively GFP-expressing hESC line we investigated whether hESC-derived cardiomyocytes could be shipped and subsequently form grafts when transplanted into the left ventricular wall of athymic nude rats subjected to ischemia/reperfusion with minimal infarction. Co-localization of GFP-epifluorescence and cardiomyocyte-specific marker staining was utilized to analyze hESC-derived cardiomyocyte fate in a rat ischemia/reperfused myocardium. Differentiated, constitutively green fluorescent protein (GFP)-expressing hESCs (hES3-GFP; Envy) containing about 13% cardiomyocytes were differentiated in Singapore, and shipped in culture medium at 4 °C to Los Angeles (shipping time ∼ 3 days). The cells were dissociated and a cell suspension (2 × 106 cells for each rat, n = 10) or medium (n = 10) was injected directly into the myocardium within the ischemic risk area 5 min after left coronary artery occlusion in athymic nude rats. After 15 min of ischemia, the coronary artery was reperfused. The hearts were harvested at various time points later and processed for histology, immunohistochemical staining, and fluorescence microscopy. In order to assess whether the hESC-derived cardiomyocytes might evade immune surveillance, 2 × 106 cells were injected into immune competent Sprague-Dawley rat hearts (n = 2), and the hearts were harvested at 4 weeks after cell injection and examined as in the previous procedures. Even following 3 days of shipping, the hESC-derived cardiomyocytes within embryoid bodies (EBs) showed active and rhythmic contraction after incubation in the presence of 5% CO2 at 37 °C. In the nude rats, following cell implantation, H&E, immunohistochemical staining and GFP epifluorescence demonstrated grafts in 9 out of 10 hearts. Cells that demonstrated GFP epifluorescence also stained positive (co-localized) for the muscle marker alpha-actinin and exhibited cross striations (sarcomeres). Furthermore, cells that stained positive for the antibody to GFP (immunohistochemistry) also stained positive for the muscle marker sarcomeric actin and demonstrated cross striations. At 4 weeks engrafted hESCs expressed connexin 43, suggesting the presence of nascent gap junctions between donor and host cells. No evidence of rejection was observed in nude rats as determined by inspection for lymphocytic infiltrate and/or giant cells. In contrast, hESC-derived cardiomyocytes injected into immune competent Sprague-Dawley rats resulted in an overt lymphocytic infiltrate. hESCs-derived cardiomyocytes can survive several days of shipping. Grafted cells survived up to 4 weeks after transplantation in hearts of nude rats subjected to ischemia/reperfusion with minimal infarction. They continued to express cardiac muscle markers and exhibit sarcomeric structure and they were well interspersed with the endogenous myocardium. However, hESC-derived cells did not escape immune surveillance in the xenograft setting in that they elicited a rejection phenomenon in immune competent rats.

Original languageEnglish
Pages (from-to)504-516
Number of pages13
JournalJournal of Molecular and Cellular Cardiology
Volume43
Issue number4
DOIs
StatePublished - Oct 2007

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Cardiac Myocytes
Nude Rats
Green Fluorescent Proteins
Ischemia
Myocardium
Infarction
Reperfusion
Staining and Labeling
Human Embryonic Stem Cells
Sprague Dawley Rats
Coronary Vessels
Embryoid Bodies
Transplants
Actinin
Muscles
Connexin 43
Sarcomeres
Los Angeles
Coronary Occlusion
Gap Junctions

Keywords

  • Cardiomyocytes
  • Cell transplantation
  • Human embryonic stem cell
  • Immunology
  • Myocardial regeneration

ASJC Scopus subject areas

  • Molecular Biology
  • Cardiology and Cardiovascular Medicine

Cite this

Survival and maturation of human embryonic stem cell-derived cardiomyocytes in rat hearts. / Dai, Wangde; Field, Loren; Rubart-von der Lohe, Michael; Reuter, Sean; Hale, Sharon L.; Zweigerdt, Robert; Graichen, Ralph E.; Kay, Gregory L.; Jyrala, Aarne J.; Colman, Alan; Davidson, Bruce P.; Pera, Martin; Kloner, Robert A.

In: Journal of Molecular and Cellular Cardiology, Vol. 43, No. 4, 10.2007, p. 504-516.

Research output: Contribution to journalArticle

Dai, W, Field, L, Rubart-von der Lohe, M, Reuter, S, Hale, SL, Zweigerdt, R, Graichen, RE, Kay, GL, Jyrala, AJ, Colman, A, Davidson, BP, Pera, M & Kloner, RA 2007, 'Survival and maturation of human embryonic stem cell-derived cardiomyocytes in rat hearts', Journal of Molecular and Cellular Cardiology, vol. 43, no. 4, pp. 504-516. https://doi.org/10.1016/j.yjmcc.2007.07.001
Dai, Wangde ; Field, Loren ; Rubart-von der Lohe, Michael ; Reuter, Sean ; Hale, Sharon L. ; Zweigerdt, Robert ; Graichen, Ralph E. ; Kay, Gregory L. ; Jyrala, Aarne J. ; Colman, Alan ; Davidson, Bruce P. ; Pera, Martin ; Kloner, Robert A. / Survival and maturation of human embryonic stem cell-derived cardiomyocytes in rat hearts. In: Journal of Molecular and Cellular Cardiology. 2007 ; Vol. 43, No. 4. pp. 504-516.
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AU - Dai, Wangde

AU - Field, Loren

AU - Rubart-von der Lohe, Michael

AU - Reuter, Sean

AU - Hale, Sharon L.

AU - Zweigerdt, Robert

AU - Graichen, Ralph E.

AU - Kay, Gregory L.

AU - Jyrala, Aarne J.

AU - Colman, Alan

AU - Davidson, Bruce P.

AU - Pera, Martin

AU - Kloner, Robert A.

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