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Rationale: Catecholamines increase cardiac contractility, but exposure to high concentrations or prolonged exposures can cause cardiac injury. A recent study demonstrated that a single subcutaneous injection of isoproterenol (ISO; 200 mg/kg) in mice causes acute myocyte death (8%-10%) with complete cardiac repair within a month. Cardiac regeneration was via endogenous cKit(+) cardiac stem cell-mediated new myocyte formation., Objective: Our goal was to validate this simple injury/regeneration system and use it to study the biology of newly forming adult cardiac myocytes., Methods and Results: C57BL/6 mice (n=173) were treated with single injections of vehicle, 200 or 300 mg/kg ISO, or 2 daily doses of 200 mg/kg ISO for 6 days. Echocardiography revealed transiently increased systolic function and unaltered diastolic function 1 day after single ISO injection. Single ISO injections also caused membrane injury in ≈10% of myocytes, but few of these myocytes appeared to be necrotic. Circulating troponin I levels after ISO were elevated, further documenting myocyte damage. However, myocyte apoptosis was not increased after ISO injury. Heart weight to body weight ratio and fibrosis were also not altered 28 days after ISO injection. Single- or multiple-dose ISO injury was not associated with an increase in the percentage of 5-ethynyl-2'-deoxyuridine-labeled myocytes. Furthermore, ISO injections did not increase new myocytes in cKit(+/Cre)×R-GFP transgenic mice., Conclusions: A single dose of ISO causes injury in ≈10% of the cardiomyocytes. However, most of these myocytes seem to recover and do not elicit cKit(+) cardiac stem cell-derived myocyte regeneration., (© 2016 American Heart Association, Inc.)

Determination of fundamental mechanisms of disease often hinges on histopathology visualization and quantitative image analysis. Currently, the analysis of multi-channel fluorescence tissue images is primarily achieved by manual measurements of tissue cellular content and sub-cellular compartments. Since the current manual methodology for image analysis is a tedious and subjective approach, there is clearly a need for an automated analytical technique to process large-scale image datasets. Here, we introduce Nuquantus (Nuclei quantification utility software) - a novel machine learning-based analytical method, which identifies, quantifies and classifies nuclei based on cells of interest in composite fluorescent tissue images, in which cell borders are not visible. Nuquantus is an adaptive framework that learns the morphological attributes of intact tissue in the presence of anatomical variability and pathological processes. Nuquantus allowed us to robustly perform quantitative image analysis on remodeling cardiac tissue after myocardial infarction. Nuquantus reliably classifies cardiomyocyte versus non-cardiomyocyte nuclei and detects cell proliferation, as well as cell death in different cell classes. Broadly, Nuquantus provides innovative computerized methodology to analyze complex tissue images that significantly facilitates image analysis and minimizes human bias.

Background: Cardiac- (CSC) and mesenchymal-derived (MSC) CD117+ isolated stem cells improve cardiac function after injury. However, no study has compared the therapeutic benefit of these cells when used autologously., Methods: MSCs and CSCs were isolated on day 0. Cardiomyopathy was induced (day 28) by infusion of L-isoproterenol (1,100 ug/kg/hour) from Alzet minipumps for 10 days. Bromodeoxyuridine (BrdU) was infused via minipumps (50 mg/mL) to identify proliferative cells during the injury phase. Following injury (day 38), autologous CSC (n = 7) and MSC (n = 4) were delivered by intracoronary injection. These animals were compared to those receiving sham injections by echocardiography, invasive hemodynamics, and immunohistochemistry., Results: Fractional shortening improved with CSC (26.9 ± 1.1% vs. 16.1 ± 0.2%, p = 0.01) and MSC (25.1 ± 0.2% vs. 12.1 ± 0.5%, p = 0.01) as compared to shams. MSC were superior to CSC in improving left ventricle end-diastolic (LVED) volume (37.7 ± 3.1% vs. 19.9 ± 9.4%, p = 0.03) and ejection fraction (27.7 ± 0.1% vs. 19.9 ± 0.4%, p = 0.02). LVED pressure was less in MSC (6.3 ± 1.3 mmHg) as compared to CSC (9.3 ± 0.7 mmHg) and sham (13.3 ± 0.7); p = 0.01. LV BrdU+ myocytes were higher in MSC (0.17 ± 0.03%) than CSC (0.09 ± 0.01%) and sham (0.06 ± 01%); p < 0.001., Conclusions: Both CD117+ isolated CSC and MSC therapy improve cardiac function and attenuate pathological remodeling. However, MSC appear to confer additional benefit., (© 2015 Wiley Periodicals, Inc.)

Rationale: Autologous bone marrow-derived or cardiac-derived stem cell therapy for heart disease has demonstrated safety and efficacy in clinical trials, but functional improvements have been limited. Finding the optimal stem cell type best suited for cardiac regeneration is the key toward improving clinical outcomes., Objective: To determine the mechanism by which novel bone-derived stem cells support the injured heart., Methods and Results: Cortical bone-derived stem cells (CBSCs) and cardiac-derived stem cells were isolated from enhanced green fluorescent protein (EGFP+) transgenic mice and were shown to express c-kit and Sca-1 as well as 8 paracrine factors involved in cardioprotection, angiogenesis, and stem cell function. Wild-type C57BL/6 mice underwent sham operation (n=21) or myocardial infarction with injection of CBSCs (n=67), cardiac-derived stem cells (n=36), or saline (n=60). Cardiac function was monitored using echocardiography. Only 2/8 paracrine factors were detected in EGFP+ CBSCs in vivo (basic fibroblast growth factor and vascular endothelial growth factor), and this expression was associated with increased neovascularization of the infarct border zone. CBSC therapy improved survival, cardiac function, regional strain, attenuated remodeling, and decreased infarct size relative to cardiac-derived stem cells- or saline-treated myocardial infarction controls. By 6 weeks, EGFP+ cardiomyocytes, vascular smooth muscle, and endothelial cells could be identified in CBSC-treated, but not in cardiac-derived stem cells-treated, animals. EGFP+ CBSC-derived isolated myocytes were smaller and more frequently mononucleated, but were functionally indistinguishable from EGFP- myocytes., Conclusions: CBSCs improve survival, cardiac function, and attenuate remodeling through the following 2 mechanisms: (1) secretion of proangiogenic factors that stimulate endogenous neovascularization, and (2) differentiation into functional adult myocytes and vascular cells.