Cardiac stem cells delivered intravascularly traverse the vessel barrier, regenerate infarcted myocardium, and improve cardiac function

Abstract

The ability of cardiac stem cells (CSCs) to promote myocardial repair under clinically relevant conditions (i.e., when delivered intravascularly after reperfusion) is unknown. Thus, rats were subjected to a 90-min coronary occlusion; at 4 h after reperfusion, CSCs were delivered to the coronary arteries via a catheter positioned into the aortic root. Echocardiographic analysis showed that injection of CSCs attenuated the increase in left ventricular (LV) end-diastolic dimensions and impairment in LV systolic performance at 5 weeks after myocardial infarction. Pathologic analysis showed that treated hearts exhibited a smaller increase in LV chamber diameter and volume and a higher wall thickness-to-chamber radius ratio and LV mass-to-chamber volume ratio. CSCs induced myocardial regeneration, decreasing infarct size by 29%. A diploid DNA content and only two chromosomes 12 were found in new cardiomyocytes, indicating that cell fusion did not contribute to tissue reconstitution. In conclusion, intravascular injection of CSCs after reperfusion limits infarct size, attenuates LV remodeling, and ameliorates LV function. This study demonstrates that CSCs are effective when delivered in a clinically relevant manner, a clear prerequisite for clinical translation, and that these beneficial effects are independent of cell fusion. The results establish CSCs as candidates for cardiac regeneration and support an approach in which the heart's own stem cells could be collected, expanded, and stored for subsequent therapeutic repair.

Fig. 1.

Administration of CSCs improves LV systolic function. Illustrated are echocardiographic measurements in untreated and treated animals. *, P < 0.05 between untreated and treated rats. Data are mean ± SEM.

Fig. 2.

Administration of CSCs improves postinfarction ventricular remodeling. Illustrated are multiple anatomical parameters obtained in hearts arrested in diastole. *, P < 0.05 between untreated and treated rats. Data are mean ± SEM.

Fig. 3.

Administration of CSCs promotes myocardial regeneration. (A) Large transverse section halfway between the base and the apex of the left ventricle, showing an infarct in a CSC-treated rat. (Inset) Regenerated infarcted myocardium (EGFP and α-sarcomeric actin, yellow-green). (Bars, 1 mm and 100 μm.) (B) Another example of regenerated infarcted myocardium (arrows) is shown first by α-sarcomeric actin staining (red), then by EGFP labeling (green) and then by the combination of EGFP and α-sarcomeric actin (yellow-green). (C) EGFP^POS (green), α-sarcomeric actin^POS (red), small newly formed myocytes within the infarcted region express in their nuclei GATA-4 (white) and MEF2C (magenta). (D) EGFP^POS (green), cardiac myosin heavy chain^POS (red) small newly formed myocytes within the infarcted region express in their plasma membrane connexin 43 (magenta, arrowheads). (E) Newly formed myocytes in the surviving noninfarcted LV myocardium (arrows) are shown first by EGFP labeling (green) and then by the combination of EGFP and α-sarcomeric actin (yellow-green) [Bar (B–E), 10 μm.]

Fig. 4.

Administration of CSCs promotes regeneration in the absence of cell fusion. (A and B) Nuclear localization of chromosome 12 (white dots) is shown in newly formed EGFP^POS-α-sarcomeric actin^POS (yellow-green) myocytes in the infarcted (A) and spared (B) myocardium.

Fig. 5.

CSCs traverse the wall of coronary vessels migrating to the myocardium. (A) Detection by two-photon microscopy of numerous EGFP^POS clonogenic CSCs (green) located within the lumen of coronary vessels (rhodamine-labeled dextran; red) 20 min after cell injection. (A–E) Transcoronary migration of EGFP^POS CSCs to the myocardium; images of the same field (A–E) were taken at 30-min intervals, starting at 30 min after the intravascular delivery of these cells. Arrowheads point to the cells that translocated through the vessel wall (C–E). Arrowheads point to the same EGFP^POS CSCs detected in the living tissue by two-photon microscopy (E, green) and after fixation, embedding, and staining of the same region of the ventricular wall by confocal microscopy (F, green). Myocytes are stained by cardiac myosin (magenta), and nuclei are labeled by DAPI (blue). Red fluorescence in the confocal microscopic image illustrates the coronary circulation (rhodamine-labeled dextran, red). (Inset) Superimposition of E and F. Two-photon microscopy: focal depth, 20 μm. Confocal microscopy: section thickness, 10 μm. (Bars, 20 μm.)