Cell Therapy
Safety of intramyocardial injection of autologous bone marrow cells to treat myocardial ischemia in pigs

https://doi.org/10.1016/j.carrev.2006.04.003Get rights and content

Abstract

Objective

The purpose of this study is to determine the potential adverse consequences of intracardiac injections of bone marrow mononuclear cells (BMCs) to facilitate the revascularization of ischemic myocardium.

Background

Bone marrow mononuclear cells are used to treat heart failure, though there are few studies that evaluated the safety of BMC transplantation for chronic myocardial ischemia.

Methods

The pigs received coronary ameroid constrictors to induce chronic myocardial ischemia and left ventricular dysfunction. At 4 weeks, autologous BMCs were injected intramyocardially by Boston Scientific Stiletto catheter with low-dose (107 cells) or high-dose BMC (108). Control animals received saline. Blood samples were collected for hematological and chemical indices, including cardiac enzyme levels at regular time intervals postinfarction. At 7 weeks, animals underwent electrophysiological study to evaluate the arrhythmic potential of transplanted BMC, followed by necropsy and histopathology.

Results

No mortalities were associated with intramyocardial delivery of BMC or saline. At Day 0, the total creatine phosphokinase (CPK) was in the normal range in all groups. All groups had significant elevations in CPK after ameroid placement, with no significant differences between groups. At 7 weeks, CPK in all groups had returned to pretreatment levels. Electrophysiological assessment revealed that one control animal had an inducible arrhythmia. No arrhythmias were induced in low- or high-dose BMC-treated pigs. There were no histopathological changes associated with BMC injection.

Conclusion

This study showed, in a clinically relevant large-animal model, that catheter-based intramyocardial injection of autologous BMC into ischemic myocardium is safe.

Introduction

Cardiovascular disease accounted for 40% of all deaths or 1 in every 2.5 deaths in the United States in 2000 [1]. Ischemic heart disease accounts for nearly half of all cardiovascular deaths and is the leading cause of congestive heart failure. Chronic myocardial ischemia results in cardiac contractile dysfunction. If the ischemic episode is limited in severity or duration, then cardiomyocytes survive and are protected from further ischemic insult through several preconditioning mechanisms. However, with prolonged severe periods of ischemia, cardiomyocyte death occurs [2]. Adult human cardiomyocytes lack the capability to regenerate, and over time, damaged myocardial cells are replaced by connective scar tissue [3] along with a compensatory hypertrophy of the remaining viable cardiomyocytes. This replacement of infarct by scar tissue leads to a loss of functional myocardium within the ischemic area, a progressive remodeling of the nonischemic area, or border zone, and an overall reduction in cardiac performance.

It has been demonstrated that bone marrow mononuclear cells (BMCs) can differentiate into cardiomyocytes [4], [5], [6]. In one study, transplanted isogenic bone marrow cells were introduced into damaged rat hearts by coronary perfusion. Surviving cells were formed in the majority of animals with distribution throughout the myocardium [6]. This study also revealed that the BMCs that infiltrated the myocardial scar were fibroblast-like, whereas BMCs within the surrounding tissue expressed cardiomyocyte markers. Other studies have shown that BMCs transplanted into the damaged myocardium of mice survive, express cardiomyocyte-specific genes, and may participate in neovascularization [4], [5]. We and others have conducted studies demonstrating improvements in myocardial blood flow and cardiac function following intramyocardial injection of bone marrow or bone marrow-derived cells [7], [8]. Additionally, clinical studies have documented the feasibility of harvesting and transplanting circulating endothelial progenitor cells or BMCs in patients suffering from ischemic heart disease.

Because of the compelling preclinical data illustrating the ability of BMCs to improve cardiac function, clinical trials were initiated. These trials are currently being performed as single-center studies using BMCs or skeletal myoblasts. Safety and feasibility have initially been favorable in trials utilizing human adult bone marrow stem cells, but unresolved issues remain regarding the initial arrhythmogenic potential of myoblasts [9]. In the published reports of human stem cell trials thus far, the use of BMCs delivered in patients with previous myocardial infarction was shown to improve function and perfusion in a small number of patients [10]. It appears therefore that stem cells, whether derived from bone marrow or isolated from the blood circulation, can help improve cardiac dysfunction and repair damaged cardiac tissue. However, there are few studies [11] that systematically evaluated the safety of bone marrow-derived cell transplantation for chronic ischemic conditions of the myocardium. The purpose of this study is to evaluate the safety of autologous stem cell transplantation into chronically ischemic myocardium as a potential therapy for the repair of cardiac tissue. By demonstrating that catheter-based BMC injection was safe, we provided a basis for developing, evaluating, and improving the therapeutic potential of bone marrow-derived cells.

Section snippets

Chronic ischemia surgical procedure (ameroid constrictor based)

Animal care and handling conformed to the National Institutes of Health and American Heart Association guidelines and were approved by the Institutional Animal Care and Use Committee of the American Cardiovascular Research Institute. Adult Yucatan miniswines (30–60 kg) were sedated with intramuscular injection of Telazol (2–4 mg/kg), intubated, and anesthetized. While under anesthesia and using mechanical ventilatory support, a thoracotomy was performed, the pericardium was opened, and an

Mortalities

Of the 14 animals enrolled, 2 animals died (14% mortality rate). Both mortalities occurred prior to treatment: one on the surgical table after ameroid placement and the other 3 days after ameroid placement surgery. No casualties occurred during or immediately after intracardiac BMC or saline injections.

Bone marrow cell characterization

The majority of BMCs were positive for the common leukocyte marker CD45 (63±6%) (Fig. 1, Fig. 2). The percentage of T cells (CD3+ cells) isolated from the BMC injectate was 29.97±1.4%. T-helper

Discussion

This study is the first to report the safety of intramyocardial bone marrow cell injection with full characterization of the bone marrow cell injectate. Injection of autologous bone marrow cells into ischemic myocardium was not associated with any serious side effects; specifically, there were no casualties due to cell implantation, arrhythmias were unable to be induced even during electrophysiological stimulation, there was no evidence of local infection as assessed by histopathological

Conclusions

In conclusion, intramyocardial injection of freshly isolated bone marrow cells is safe and feasible for the clinical setting. Many other groups, including our own, have previously shown the efficacy of the cellular therapy by the ability of these cells to induce angiogenesis and improve cardiac function. This study shows, in a clinically relevant large-animal model, that catheter-based intramyocardial injection of autologous bone marrow cells into chronically ischemic myocardium is safe.

Acknowledgments

We would like to thank Dr. Roger Broderson, DVM, MS, PhD, for his help during the necropsy procedures and Mr. Todd Dennis for acquiring the echocardiographic images. We would like to thank Dr. Alka Redkar for providing laboratory space and equipment for the cell characterization studies. We would also like to thank Mr. Jesse Rios, Ms. Melissa Fowlkes, RVT, and Mr. David Robinson for their support during the procedures and Ms. Molly Jones for all her administrative assistance.

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Funding for this project was provided by AC Therapeutics, 3155 Northwoods Place, Norcross, GA 30071, USA.

1

These authors contributed equally to this manuscript.

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