Can Cytoprotective Cobalt Protoporphyrin Protect Skeletal Muscle and Muscle-derived Stem Cells From Ischemic Injury?

Background: Extremity trauma is the most common injury seen in combat hospitals as well as in civilian trauma centers. Major skeletal muscle injuries that are complicated by ischemia often result in substantial muscle loss, residual disability, or even amputation, yet few treatment options are available. A therapy that would increase skeletal muscle tolerance to hypoxic damage could reduce acute myocyte loss and enhance preservation of muscle mass in these situations.

Human embryonic stem cell-derived cardiomyocytes migrate in response to gradients of fibronectin and Wnt5a.

An improved understanding of the factors that regulate the migration of human embryonic stem cell-derived cardiomyocytes (hESC-CMs) would provide new insights into human heart development and suggest novel strategies to improve their electromechanical integration after intracardiac transplantation. Since nothing has been reported as to the factors controlling hESC-CM migration, we hypothesized that hESC-CMs would migrate in response to the extracellular matrix and soluble signaling molecules previously implicated in heart morphogenesis. To test this, we screened candidate factors by transwell assay for effects on hESC-CM motility, followed by validation via live-cell imaging and/or gap-closure assays.

Monocyte ADAM17 promotes diapedesis during transendothelial migration: identification of steps and substrates targeted by metalloproteinases.

Despite expanded definition of the leukocyte adhesion cascade and mechanisms underlying individual steps, very little is known about regulatory mechanisms controlling sequential shifts between steps. We tested the hypothesis that metalloproteinases provide a mechanism to rapidly transition monocytes between different steps. Our study identifies diapedesis as a step targeted by metalloproteinase activity.

Targeted genomic integration of a selectable floxed dual fluorescence reporter in human embryonic stem cells.

The differentiation of pluripotent stem cells involves transition through a series of specific cell states. To understand these cell fate decisions, the field needs improved genetic tools for the labeling, lineage tracing and selection of specific cell types from heterogeneous differentiating populations, particularly in the human embryonic stem cell (hESC) system. We used zinc finger nuclease technology to stably insert a unique, selectable, floxed dual-fluorescence reporter transgene into the AAVS1 locus of RUES2 hESCs.

Differentiation and fiber type-specific activity of a muscle creatine kinase intronic enhancer.

Background: Hundreds of genes, including muscle creatine kinase (MCK), are differentially expressed in fast- and slow-twitch muscle fibers, but the fiber type-specific regulatory mechanisms are not well understood.

Results: Modulatory region 1 (MR1) is a 1-kb regulatory region within MCK intron 1 that is highly active in terminally differentiating skeletal myocytes in vitro. A MCK small intronic enhancer (MCK-SIE) containing a paired E-box/myocyte enhancer factor 2 (MEF2) regulatory motif resides within MR1.

Embryonic cardiomyocyte expression of endothelial genes.

Vertebrate precardiac mesoderm contains cells destined to become cardiomyocyte or endothelial cells. To determine the stability of these phenotypes freshly isolated embryonic day (E) 2.5-E6 chicken hearts were immunostained for myosin heavy chain (MyHC) to identify cardiomyocytes, and von Willebrand factor (vWF) and Flk-1 to identify endothelial cells.

BMP induction of cardiogenesis in P19 cells requires prior cell-cell interaction(s).

Mouse P19 embryonal carcinoma cells undergo cardiogenesis in response to high density and DMSO. We have derived a clonal subline that undergoes cardiogenesis in response to high density, but without requiring exposure to DMSO. The new subline retains the capacity to differentiate into skeletal muscle and neuronal cells in response to DMSO and retinoic acid.

Human umbilical vein endothelial cells fuse with cardiomyocytes but do not activate cardiac gene expression.

It was recently reported that human umbilical endothelial vein cells (HUVECs) transdifferentiate and express cardiac genes when co-cultured with rat neonatal cardiomyocytes (Condorelli et al. (2001)). If substantiated and optimized, this phenomenon could have many therapeutic applications.

The C-terminal IgI domains of myosin-binding proteins C and H (MyBP-C and MyBP-H) are both necessary and sufficient for the intracellular crosslinking of sarcomeric myosin in transfected non-muscle cells.

Using the COS cell transfection assay developed previously, we examined which domains of myosin-binding proteins C and H (MyBP-C and MyBP-H) are involved in intracellular interactions with sarcomeric myosin heavy chain (MyHC). Earlier studies demonstrated that overexpression of sarcomeric MyHC in COS cells results in the cytoplasmic assembly of anisotropic, spindle-like aggregates of myosin-containing filaments in the absence of other myofibrillar proteins. When the same sarcomeric MyHC was co-expressed with either MyBP-C or MyBP-H, prominent cable-like co-polymers of MyHC and the MyBPs formed in the cytoplasm instead of the spindle-like aggregates formed by MyHC alone.