Bone marrow-derived stromal cells home to and remain in the infarcted rat heart but fail to improve function: an in vivo cine-MRI study

Abstract

Basic and clinical studies have shown that bone marrow cell therapy can improve cardiac function following infarction. In experimental animals, reported stem cell-mediated changes range from no measurable improvement to the complete restoration of function. In the clinic, however, the average improvement in left ventricular ejection fraction is around 2% to 3%. A possible explanation for the discrepancy between basic and clinical results is that few basic studies have used the magnetic resonance (MR) imaging (MRI) methods that were used in clinical trials for measuring cardiac function. Consequently, we employed cine-MR to determine the effect of bone marrow stromal cells (BMSCs) on cardiac function in rats. Cultured rat BMSCs were characterized using flow cytometry and labeled with iron oxide particles and a fluorescent marker to allow in vivo cell tracking and ex vivo cell identification, respectively. Neither label affected in vitro cell proliferation or differentiation. Rat hearts were infarcted, and BMSCs or control media were injected into the infarct periphery (n = 34) or infused systemically (n = 30). MRI was used to measure cardiac morphology and function and to determine cell distribution for 10 wk after infarction and cell therapy. In vivo MRI, histology, and cell reisolation confirmed successful BMSC delivery and retention within the myocardium throughout the experiment. However, no significant improvement in any measure of cardiac function was observed at any time. We conclude that cultured BMSCs are not the optimal cell population to treat the infarcted heart.

Fig. 1.

Rat bone marrow stromal cell (BMSC) characterization, labeling, proliferation, and differentiation. A: BMSCs (n = 3) characterized using flow cytometry were positive for CD90, as shown in the representative histogram and scatter plot, and negative for CD4, CD11b/c, CD31, CD45, CD45R, and CD49d. B: more than 95% of cells took up micron-sized particles of iron oxide (MPIO) in culture, as shown by the representative images and flow cytometric histograms of labeled and unlabeled cells. C: MPIO-labeled BMSCs proliferated at similar rates to those of unlabeled BMSCs. D: MPIO-labeled BMSCs differentiated into adipocytes and osteoblasts. PE, phycoerythrin.

Fig. 2.

Magnetic resonance imaging (MRI) tracking. Representative in vivo (A) and ex vivo (B) MRI of hearts 10 wk after intramuscular administration of 5 × 10⁵ MPIO-labeled BMSCs (Fe-BMSCs) showing Fe-BMSCs tracking from the point of injection into the scar and in vivo (C) and ex vivo (D) MRI of hearts 4 wk after intravenous administration of 4 × 10⁶ Fe-BMSCs 1 day after myocardial infarction (MI) showing Fe-BMSCs located through the scar but not in the viable myocardium. RV, right ventricle; LV, left ventricle.

Fig. 3.

Immunohistochemistry and recovery of grafted cells. A–D: light microscopy of 5-μm sections stained for green fluorescent protein (GFP), using peroxidise-based GFP signal amplification, in a heart that received Fe-BMSCs 10 wk previously. Expanded regions show GFP-labeled donor cells within the myocardium. Fe-BMSCs before administration (E) and phase contrast (F) and fluorescence microscopy (G) of chloromethyl-benzamidodialkylcarbocyanine (CM-DiI)-Fe-BMSCs reisolated from heart digests 6 wk after intravenous infusion (n = 3). H: representative examples of alkaline phosphatase staining of normal LV in sham-operated rat heart or peri-infarct area (magnification, ×40). I: quantitative analysis of capillary density in peri-infarct area.

Fig. 4.

In vivo MRI measurements of cardiac morphology and function. Ejection fraction, end systolic volume, and area of akinetic myocardium measured using in vivo MRI at 1, 4, and 10 wk after BMSCs or Fe-BMSCs were administered by direct myocardial injection and at 1 and 4 wk after systemic infusion of Fe-BMSCs 1 or 3 days after MI. There were no significant differences between cell- and saline-treated groups. For clarity, significant differences between shams and infarcted hearts are not shown.