Detailed analysis of bone marrow from patients with ischemic heart disease and left ventricular dysfunction: BM CD34, CD11b, and clonogenic capacity as biomarkers for clinical outcomes.

RATIONALE: Bone marrow (BM) cell therapy for ischemic heart disease (IHD) has shown mixed results. Before the full potency of BM cell therapy can be realized, it is essential to understand the BM niche after acute myocardial infarction (AMI). OBJECTIVE: To study the BM composition in patients with IHD and severe left ventricular (LV) dysfunction. METHODS AND RESULTS: BM from 280 patients with IHD and LV dysfunction were analyzed for cell subsets by flow cytometry and colony assays. BM CD34(+) cell percentage was decreased 7 days after AMI (mean of 1.9% versus 2.3%-2.7% in other cohorts; P<0.05). BM-derived endothelial colonies were significantly decreased (P<0.05). Increased BM CD11b(+) cells associated with worse LV ejection fraction (LVEF) after AMI (P<0.05). Increased BM CD34(+) percentage associated with greater improvement in LVEF (+9.9% versus +2.3%; P=0.03, for patients with AMI and +6.6% versus -0.02%; P=0.021 for patients with chronic IHD). In addition, decreased BM CD34(+) percentage in patients with chronic IHD correlated with decrement in LVEF (-2.9% versus +0.7%; P=0.0355). CONCLUSIONS: In this study, we show a heterogeneous mixture of BM cell subsets, decreased endothelial colony capacity, a CD34+ cell nadir 7 days after AMI, a negative correlation between CD11b percentage and postinfarct LVEF, and positive correlation of CD34 percentage with change in LVEF after cell therapy. These results serve as a possible basis for the small clinical improvement seen in autologous BM cell therapy trials and support selection of potent cell subsets and reversal of comorbid BM impairment. CLINICAL TRIAL REGISTRATIONS URL: http://www.clinicaltrials.gov. Unique identifiers: NCT00684021, NCT00684060, and NCT00824005.

Manufacturing mesenchymal stromal cells for phase I clinical trials.

Mesenchymal stromal cells (MSCs) are multipotent progenitor cells capable of differentiating into adipocytes, osteoblasts and chondroblasts as well as secreting a vast array of soluble mediators. This potentially makes MSCs important mediators of a variety of therapeutic applications. They are actively under evaluation for immunomodulatory purposes such as graft-versus-host disease and Crohn's disease as well as regenerative applications such as stroke and congestive heart failure. We report our method of generating clinical-grade MSCs together with suggestions gathered from manufacturing experience in our Good Manufacturing Practices facility.

Nuclear plakoglobin is essential for differentiation of cardiac progenitor cells to adipocytes in arrhythmogenic right ventricular cardiomyopathy.

RATIONALE: Arrhythmogenic right ventricular cardiomyopathy (ARVC) is a disease of desmosome proteins characterized by fibroadipogenesis in the myocardium. We have implicated signaling properties of junction protein plakoglobin (PG) in the pathogenesis of ARVC. OBJECTIVE: To delineate the pathogenic role of PG in adipogenesis in ARVC. METHODS AND RESULTS: We generated mice overexpressing PG, either a wildtype (PG(WT)) or a truncated (PG(TR)), known to cause ARVC, in the heart; and PG null (PG⁻/⁻) embryos. PG(WT) and PG(TR) mice exhibited fibro-adiposis, cardiac dysfunction, and premature death. Subcellular protein fractionation and immunofluorescence showed nuclear localization of PG(WT) and PG(TR) and reduced membrane localization of PG(TR). Coimmunoprecipitation showed reduced binding of PG(TR) but not PG(WT) to desmosome proteins DSP and DSG2. Transgene PG(WT) and PG(TR) were expressed in c-Kit+:Sca1+ cardiac progenitor cells (CPCs) isolated from the hearts of PG(WT) and PG(TR) by fluorescence activated cell sorting. CPCs isolated from the transgenic hearts showed enhanced adipogenesis, increased levels of adipogenic factors KLF15, C/EBP-α and noncanonical Wnt5b, and reduced level of CTGF, an inhibitor of adipogenesis. Treatment with BIO activated the canonical Wnt signaling, reversed the proadipogenic transcriptional switch and prevented adipogenesis in a dose-dependent manner. Moreover, c-Kit+ CPCs, isolated from PG⁻/⁻ embryos, were resistant to adipogenesis, expressed high mRNA levels of CTGF and other canonical Wnt signaling targets. CONCLUSIONS: Nuclear PG provokes adipogenesis in c-Kit+ CPCs by repressing the canonical Wnt signaling and inducing a proadipogenic gene expression. The findings suggest that adipocytes in ARVC, at least in part, originate from c-Kit+ CPCs.

Histopathological validation of electromechanical mapping in assessing myocardial viability in a porcine model of chronic ischemia.

BACKGROUND AND OBJECTIVE: Left ventricular electromechanical mapping (EMM) determines myocardial viability on the basis of endocardial electrograms. The aim of the present study was to validate EMM in differentiating infarcted myocardium from viable myocardium by histopathological analysis. METHODS: Sixty days after implanting an ameroid constrictor over the left circumflex artery to create chronic ischemia in 19 pigs, EMM was performed to construct unipolar voltage (UPV), bipolar voltage (BPV) and linear local shortening (LLS) maps. Noninfarcted and infarcted myocardium were identified by histopathology. Threshold determinations comparing noninfarcted tissue with scarred tissue were made by measuring the area under the receiver operating characteristic curves. RESULTS: From the 19 hearts, 149 myocardial segments were divided into noninfarcted myocardium (n=128) and transmural infarct (n=21). UPV, BPV and LLS values (4.7+/-1.2 mV, 2.8+/-2.5 mV and 10.0+/-5.1%, respectively) of infarcted segments were significantly lower than those in noninfarcted myocardium (10.9+/-3.4 mV, 4.5+/-2.4 mV and 15.7+/-9.5%, respectively; P<0.01 for each comparison). The threshold values of UPV, BPV and LLS differentiating noninfarcted from infarcted myocardium were 6.2 mV (98% sensitivity, 95% specificity, 97% accuracy), 2.8 mV (80% sensitivity, 72% specificity, 79% accuracy) and 12.3% (68% sensitivity, 67% specificity, 68% accuracy), respectively. The relative dispersion of voltage was lower for UPV versus BPV. CONCLUSION: UPV can accurately differentiate infarcted from noninfarcted tissue in the chronic ischemic heart of pigs; however, BPV and LLS results were less accurate.

A New High-Performance Gadonanotube-Polymer Hybrid Material for Stem Cell Labeling and Tracking by MRI.

A gentle, rapid method has been developed to introduce a polyacrylic acid (PAA) polymer coating on the surface of gadonanotubes (GNTs) which significantly increases their dispersibility in water without the need of a surfactant. As a result, the polymer, with its many carboxylic acid groups, coats the surface of the GNTs to form a new GNT-polymer hybrid material (PAA-GNT) which can be highly dispersed in water (ca. 20 mg·mL-1) at physiological pH. When dispersed in water, the new PAA-GNT material is a powerful MRI contrast agent with an extremely short water proton spin-lattice relaxation time (T1) which results in a T1-weighted relaxivity of 150 mM-1·s-1 per Gd3+ ion at 1.5 T. Furthermore, the PAA-GNTs have been used to safely label porcine bone-marrow-derived mesenchymal stem cells for magnetic resonance imaging. The labeled cells display excellent image contrast in phantom imaging experiments, and transmission electron microscopy images of the labeled cells reveal the presence of highly dispersed PAA-GNTs within the cytoplasm with 1014 Gd3+ ions per cell.