Allogeneic mesenchymal precursor cell therapy to limit remodeling after myocardial infarction: the effect of cell dosage

Abstract

Background: This experiment assessed the dose-dependent effect of a unique allogeneic STRO-3-positive mesenchymal precursor cell (MPC) on postinfarction left ventricular (LV) remodeling. The MPCs were administered in a manner that would simulate an off-the-self, early postinfarction, preventative approach to cardiac cell therapy in a sheep transmural myocardial infarct (MI) model.

Methods: Allogeneic MPCs were isolated from male crossbred sheep. Forty-six female sheep underwent coronary ligation to produce a transmural LV anteroapical infarction. One hour after infarction, the borderzone myocardium received an injection of 25, 75, 225, or 450 x 10(6) MPCs, or cell medium. Echocardiography was performed at 4 and 8 weeks after MI to quantify LV end-diastolic (LVEDV) and end-systolic volumes (LVESV), ejection fraction (EF), and infarct expansion. CD31 and smooth muscle actin (SMA) immunohistochemical staining was performed on infarct and borderzone specimens to quantify vascular density.

Results: Compared with controls, low-dose (25 and 75 x 10(6) cells) MPC treatment significantly attenuated infarct expansion and increases in LVEDV and LVESV. EF was improved at all cell doses. CD31 and SMA immunohistochemical staining demonstrated increased vascular density in the borderzone only at the lower cell doses. There was no evidence of myocardial regeneration within the infarct.

Conclusion: Allogeneic STRO-3 positive MPCs attenuate the remodeling response to transmural MI in a clinically relevant large-animal model. This effect is associated with vasculogenesis and arteriogenesis within the borderzone and infarct and is most pronounced at lower cell doses.

Fig 1

(A) In this postinfarction view of the ovine heart as seen through a left thoracotomy, the size and location of the anteroapical infarct are clearly depicted. Cells were injected in the normally perfused borderzone region within 1 cm of the readily identifiable infarction demarcation line (shaded area). (B) Postmortem view of the left ventricle shows ovine left ventricle opened through the intraventricular septum demonstrating the standard infarct size and location as well as the injection site and tissue sampling sites. (APM = anterior papillary muscle; PPM = posterior papillary muscle.)

Fig 2

(A) Smooth muscle actin vs (B) CD31 immunohistochemical staining of the same section of sheep infarct tissue. In sheep, CD31 effectively stains the endothelium of thin-walled venules and capillaries but does not reliably stain arterioles and small muscular arteries. Smooth muscle actin staining readily identifies arterioles and small muscular arteries in sheep.

Fig 3

Photomicrographs (original magnification ×40) show results of smooth muscle actin (SMA) immunohistochemical staining of borderzone myocardium 8 weeks after infarction in representative sheep treated with (A) 25 million mesenchymal precursor cells (MPCs), (B) 75 million MPCs, (C) 225 million MPCs, and (D) 450 million MPCs, and the (E) untreated controls. Lower-dose animals had a higher density of muscular arterioles compared with higher-dose animals and control sheep. (MPC = mesenchymal precursor cells.)

Fig 4

Graphs show quantitative assessment of (A, B) borderzone (BZ) vascular density and (C, D) infarct vascular density using smooth muscle actin (SMA) and CD31 immunohistochemical staining in control sheep and animals treated with mesenchymal precursor cells (MPCs) 8 weeks after infarction (M = 10⁶). *p < 0.05 vs control; †p < 0.05 vs 25 × 10⁶ MPCs; ‡p < 0.05 vs 75 × 10⁶ MPCs. Range bars represent the standard error of the mean.

Fig 5

Polymerase chain reaction (PCR) amplification of myocardial DNA from the borderzone (BZ) of representative treatment animals that died 1 hour after cell injection (225 million cells), 4 weeks after infarction (25 million cells), and 8 weeks after infarction (75 million cells) using the (A) SRY2 and (B) SRY3 primer pairs to identify the presence of male DNA. PCR products of expected size were produced in the 1-hour animal with both the SRY2 and SRY3 primers pairs. PCR products from treated animals 4 weeks and 8 weeks after infarction (Inf) were below the detection limit of 500-pg template DNA. Male sheep myocardial DNA was used as the positive control, and female sheep myocardial DNA was used as the negative control.

Fig 6

(A) Masson trichrome stain at original magnification ×10 and (B)×40 of sheep infarct tissue 8 weeks after infarction demonstrates an island of myofibrillarlytic myocytes within the scar. These isolated nests of myocytes are commonly seen in transmural infarcts and were not observed with increased frequency in animals treated with mesenchymal precursor cells at any cell dose.