Bone marrow-derived B cells preserve ventricular function after acute myocardial infarction.

OBJECTIVES: In view of evidence that mature cells play a role in modulating the stem cell niche and thereby stem cell potential and proliferation, we hypothesized that a mature bone marrow (BM) mononuclear cell (MNC) infusion subfraction may have particular potency in promoting hematopoietic or resident stem cell-induced cardiac repair post-infarction. BACKGROUND: Treatment of acute myocardial infarction (MI) with BM MNC infusion has shown promise for improving patient outcomes. However, clinical data are conflicting, and demonstrate modest improvements. BM MNCs consist of different subpopulations including stem cells, progenitors, and differentiated leukocytes. METHODS: Stem cells (c-kit+) and subsets of mature cells including myeloid lineage, B and T-cells were isolated from bone marrow harvested from isogeneic donor rats. Recipient rats had baseline echocardiography then coronary artery ligation; 1 x 10(6) cells (enriched subpopulations or combinations of subpopulations of BM MNC) or saline was injected into ischemic and ischemic border zones. Cell subpopulations were either injected fresh or after overnight culture. After 2 weeks, animals underwent follow-up echocardiography. Cardiac tissue was assayed for cardiomyocyte proliferation and apoptosis. RESULTS: Fractional ventricular diameter shortening was significantly improved compared with saline (38 +/- 3.2%) when B cells alone were injected fresh (44 +/- 3.0%, p = 0.035), or after overnight culture (51 +/- 2.9%, p < 0.001), or after culture with c-kit+ cells (44 +/- 2.4%, p = 0.062). B cells reduced apoptosis at 48 h after injection compared with control cells (5.7 +/- 1.2% vs. 12.6 +/- 2.0%, p = 0.005). CONCLUSIONS: Intramyocardial injection of B cells into early post-ischemic myocardium preserved cardiac function by cardiomyocyte salvage. Other BM MNC subtypes were either ineffective or suppressed cardioprotection conferred by an enriched B cell population.

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

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 (10(7) cells) or high-dose BMC (10(8)). 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.

Characterization of primitive marrow CD34+ cells that persist after a sublethal dose of total body irradiation.

Knowledge of the molecular events that occur during hematopoietic stem/progenitor cell (HSPC) development is vital to our understanding of blood cell production. To study the functional groups of genes characteristic of HSPCs we isolated a subpopulation of CD34+ bone marrow (BM) cells from nonhuman primates that persisted in vivo after a sublethal dose of total body irradiation (TBI). CD34+ cells isolated during the phase of maximal hematopoietic suppression show a transcriptional profile characteristic of metabolically inactive cells, with strong coordinate downregulation of a large number of genes required for protein production and processing. Consistent with this profile, these CD34+ cells were not able to generate hematopoietic colonies. Transcriptional profiling of these CD34+ cells in conjunction with a pathway analysis method reveals several classes of functionally related genes that are upregulated in comparison to the CD34+ cells obtained prior to TBI. These families included genes known to be associated with self-renewal and maintenance of HSPCs (including bone morphogenetic proteins), resistance to apoptosis (Bcl-2) as well as genes characteristic of a variety of nonhematopoietic tissues (gamma-aminobutyric acid/glycine receptor, complement receptor C1qRp). In contrast, during the period of hematopoietic recovery, the CD34+ cells expressed higher level of genes encoding factors regulating maturation and differentiation of HSPCs. Our data indicate that the primitive BM CD34+ cell population that persists after radiation possesses a transcriptional profile suggestive of pluripotency.

Studies of the route of administration and role of conditioning with radiation on unrelated allogeneic mismatched mesenchymal stem cell engraftment in a nonhuman primate model.

OBJECTIVE: The aim of this study was to examine the effects of the route of administration [intrabone marrow (IBM) vs intravenous (IV)] and the role of conditioning with irradiation in optimizing mesenchymal stem cell (MSC) transplantation. MATERIALS AND METHODS: To determine if irradiation resulted in depletion of colony-forming unit fibroblasts (CFU-F), which might favor the engraftment of donor MSC, the number of CFU-Fs was assayed from animals receiving either hemibody irradiation (HBI) or total body irradiation (TBI). RESULTS: TBI resulted in a marked reduction of CFU-F numbers that spontaneously resolved, whereas animals receiving HBI did not experience depletion of CFU-F. Animals receiving MSC grafts by the IV route had higher numbers of marrow CFU-F. MSC were transduced using retroviral vectors encoding the neomycin resistance gene (Neo(R)) and a second gene encoding either the human soluble tumor necrosis factor receptor (hsTNFRII) or beta-galactosidase (beta-Gal). MSCs were administered by either the IV or IBM route to animals receiving HBI. The Neo(R) transgene was detectable in hematopoietic tissues of all animals and nonhematopoietic tissues in a single animal. Evidence of transgene expression was documented by detection of beta-Gal(+) cells in BM smears and transiently elevated serum levels of hsTNFRII. CONCLUSION: These studies indicate that 1) MSC possess the ability to engraft and persist in an unrelated mismatched allogeneic hosts; 2) 250-cGy HBI did not favor engraftment of MSC; 3) the IBM route was not more effective than the IV route in delivering MSC grafts; and 4) transplanted MSC preferentially localized to the marrow rather than nonhematopoietic tissues.

Comparative study of stromal cell lines derived from embryonic, fetal, and postnatal mouse blood-forming tissues.

OBJECTIVE: To better understand the differentiation of stromal cells of the hematopoietic microenvironment, we set out to characterize stromal cells from the different developmental sites of hematopoiesis in the mouse (30 bone marrow, 7 spleen, 3 embryonic and 15 fetal liver, 6 yolk sac, and 6 aorta-gonad-mesonephros lines) for expression of 22 cytoskeletal, membrane, and extracellular matrix proteins. MATERIALS AND METHODS: Western blotting, immunofluorescence, and flow cytometry were used. Statistical methods included principal components analysis and analysis of variance. RESULTS: Stromal cells from 11 dpc mouse embryos express mesenchymal and vascular smooth muscle cell (VSMC) markers. Principal components analysis on the 70 stromal cell lines isolated from different anatomic sites and developmental stages allows classification of stromal lines along a mesenchymal to VSMC differentiation pathway. Stromal cells do not express endothelial and hematopoietic differentiation membrane antigens, but they do express integrin alpha(5), alpha(6), and beta(1) subunits, vascular cell adhesion molecule-1, CD44, stem cell antigen-1, Thy-1, CD34, and endoglin. The intensity of expression of certain markers differs between lines according to the anatomic site of origin. CONCLUSIONS: This study indicates that stromal cells, whatever their anatomic site of origin, follow a VSMC differentiation pathway, suggesting a blood-forming tissue-specific differentiation of mesenchymal stem cells. Differential quantitative expression of distinct sets of markers appears to be correlated with the anatomic sites of origin of the stromal cells.