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1.
Panminerva Med ; 50(2): 129-37, 2008 Jun.
Article in English | MEDLINE | ID: mdl-18607336

ABSTRACT

Cardiovascular diseases remain the leading cause of morbidity and mortality in Western society. Among these diseases, congestive heart failure continues to be a significant health care burden. Recent medical and surgical advances in therapy have improved the quality and quantity of life for patients with heart failure. However, none of these therapies address the fundamental problem of loss of functional cardiomyocytes. Cell regeneration therapies have become an exciting potential treatment for heart failure and other cardiovascular diseases. This emerging therapeutic field has been pursued experimentally with both embryonic-derived stem cells and adult-derived progenitor cells. The identification of resident cardiac progenitor cells has propelled the field of cardiac regenerative biology forward at astonishing rates. This review will examine current findings of various stem and progenitor cells that have been proposed as potential sources for cardiac regeneration, and the recent therapeutic findings from preliminary clinical trials using some of these cell types for cardiovascular repair.


Subject(s)
Adult Stem Cells/transplantation , Coronary Artery Disease/therapy , Embryonic Stem Cells/transplantation , Myocytes, Cardiac/cytology , Stem Cell Transplantation , Animals , Humans
2.
Wound Repair Regen ; 9(4): 287-96, 2001.
Article in English | MEDLINE | ID: mdl-11679137

ABSTRACT

Biolistic transmission of mRNA provides transient gene therapy to in vivo organs. This study documents particle mediated mRNA transmission to a solid organ and wound healing model using the mRNA of Green Fluorescent Protein to determine optimal delivery parameters. Renal function, bullet penetration, cellular injury, and Green Fluorescent Protein synthesis were quantified. Chimeric human epidermal growth factor-FLAG epitope cDNA or mRNA was transmitted to wounds in normal or steroid treated animals. Wound bursting strength, human epidermal growth factor-FLAG, and collagen synthesis were determined. Injury and bullet penetration correlated with the delivery velocity and bullet size. Optimal delivery parameters were established which provided widespread Green Fluorescent Protein synthesis. Human epidermal growth factor-FLAG treatment significantly increased collagen content and wound breaking strength in normal and steroid treated animals. FLAG protein synthesis was evident in mRNA treated fascia following treatment. We found the gene gun provides a novel method for efficient, in vivo delivery of mRNA-based therapeutic strategies to mammalian organs with minimal histologic damage allowing transient expression of protein in in vivo target tissues. Co-delivery of Green Fluorescent Protein mRNA may provide a useful positive control to determine effective transmission. Biolistic transmission of human epidermal growth factor-FLAG mRNA provides increased tissue epidermal growth factor levels and accelerates wound healing in normal and steroid exposed animals.


Subject(s)
Genetic Therapy/methods , Kidney/physiology , Luminescent Proteins , RNA, Messenger/pharmacology , Wounds and Injuries/therapy , Animals , Biological Availability , Disease Models, Animal , Drug Delivery Systems , Epidermal Growth Factor/pharmacology , Gene Transfer Techniques , Green Fluorescent Proteins , Humans , Immunohistochemistry , Male , Rats , Rats, Sprague-Dawley , Regeneration , Sensitivity and Specificity , Wound Healing/physiology , Wounds and Injuries/genetics , Wounds and Injuries/pathology
3.
Am Surg ; 66(3): 302-6, 2000 Mar.
Article in English | MEDLINE | ID: mdl-10759204

ABSTRACT

Surgical glove integrity is essential for universal precautions; glove safety is verified by the water load test (WLT). Concerns regarding glove injury have prompted newer testing methodologies, including electrical conductance testing (ECT); however, the sensitivities of these tests are not known. We compared the sensitivity of WLT and ECT in detecting glove needle-stick injury in two commonly used brands of surgical gloves. Punctures were made with hollow-bore and solid surgical needles of various configurations. The WLT failed to detect glove holes from the smallest-caliber needles and only detected the injury in 60 per cent for the largest caliber. The ECT provided a graded index of glove injury in all holes made by both solid surgical needles and hollow-bore needles. The WLT is a poor test for clinical defects in latex surgical gloves; the ECT is significantly more sensitive and provides a gauge of the cross-sectional area of the defect. Interbrand differences in self-sealing properties of surgical gloves were evidenced and may be clinically relevant after glove perforation.


Subject(s)
Gloves, Surgical/standards , Electric Conductivity , Humans , Methods , Needlestick Injuries/prevention & control , Safety
4.
Cell Struct Funct ; 22(1): 123-9, 1997 Feb.
Article in English | MEDLINE | ID: mdl-9113398

ABSTRACT

The vertebrate sarcomere is a complex structure composed of numerous proteins arranged in an exquisitely precise manner. Sarcomeric proteins are organized into interdigitating thick or thin filaments and the sliding of these filaments relative to one another constitutes muscle contraction at the sarcomere level. Consequently, an understanding of sarcomeric structure and function requires a thorough knowledge of the individual components of the thick and thin filaments, as well as their associations. Thick filaments are comprised of myosin, which provides the force required to drive muscle contraction and also plays a major structural role in thick filament formation. In addition, a family of thick filament-associated proteins plays a role in organization of the thick filament. We have used both molecular genetic and cell biological approaches to define the diversity of the myosin heavy chain gene family and to analyze the assembly of myosin and it's associated proteins into thick filaments.


Subject(s)
Myosin Heavy Chains/genetics , Animals , Carrier Proteins/metabolism , Carrier Proteins/ultrastructure , Humans , Myofibrils/ultrastructure , Myosin Heavy Chains/ultrastructure
5.
J Mol Biol ; 266(2): 317-30, 1997 Feb 21.
Article in English | MEDLINE | ID: mdl-9047366

ABSTRACT

Myosin is a motor protein whose functional unit in the sarcomere is the thick filament. The myosin molecule is capable of self-assembly into thick filaments through its alpha-helical coiled-coil rod domain. To define more precisely the sequence requirements for this assembly, segments of the human fast IId skeletal myosin rod were expressed in Escherichia coli and examined differential solubility and the formation of ordered paracrystals. We show that both properties appear to require a 29 residue sequence (residues 1874 to 1902) near the C terminus of the rod region. To test further the role of this region in assembly, a protein was constructed which consisted of this assembly competence domain (ACD) fused to the carboxy terminus of an assembly-incompetent myosin rod fragment. This chimeric fragment exhibited myosin's characteristic solubility properties and formed ordered paracrystals. To complement these in vitro experiments, both a full-length myosin heavy chain (MYH) and one from which the 29 residues were deleted were transfected into cultured mammalian cells. While the full-length construct formed the spindle-shaped structures characteristic of arrays of thick filaments, the deleted MYH showed only diffuse staining throughout the cytoplasm by light microscopy. Thus, there appears to be a specific sequence in the C-terminal region of the myosin heavy chain rod which is necessary for ordered paracrystal formation and is sufficient to confer assembly properties to an assembly-incompetent rod fragment.


Subject(s)
Myosins/chemistry , Myosins/ultrastructure , Amino Acid Sequence , Animals , Binding Sites , COS Cells/metabolism , Crystallization , Escherichia coli/genetics , Escherichia coli/metabolism , Humans , Microscopy, Electron , Models, Molecular , Molecular Sequence Data , Muscle, Skeletal/ultrastructure , Myosin Heavy Chains/chemistry , Myosin Heavy Chains/genetics , Myosin Heavy Chains/ultrastructure , Myosin Light Chains/chemistry , Myosin Light Chains/genetics , Myosin Light Chains/ultrastructure , Myosins/genetics , Protein Conformation , Recombinant Proteins/chemistry , Recombinant Proteins/metabolism , Recombinant Proteins/ultrastructure , Sequence Deletion , Structure-Activity Relationship
6.
Hum Mol Genet ; 5(10): 1667-72, 1996 Oct.
Article in English | MEDLINE | ID: mdl-8894705

ABSTRACT

Schnyder's crystalline corneal dystrophy (SCCD) is an autosomal dominant eye disease characterized by a bilateral clouding of the central cornea, arcus lipoides and/or visible crystalline deposits of cholesterol in the stroma. There is accumulation of phospholipid, unesterified cholesterol and cholesterol ester in the corneal stroma; this is believed to be due to an imbalance in the local factors affecting lipid/cholesterol transport or metabolism. The cellular mechanism of abnormal lipid transport and metabolism in SCCD is of interest due to its potential involvement in atherosclerosis, and its implications for the pathogenesis of cerebrovascular, coronary and peripheral vascular disease as well as corneal opacification. To determine the chromosomal location of the SCCD locus, genome-wide linkage analysis has been performed in two large Swede-Finn kindreds recently identified in central Massachusetts. After analysing 300 microsatellite markers > 90% of the genome was excluded from linkage to the SCCD locus. We now report the chromosomal assignment of the gene for SCCD in both families to be 1p34.1-p36; the maximum multipoint lod-score was 8.48 in the interval between D1S214 and D1S503. From haplotype analysis, the SCCD locus lies in the 16 cM interval between markers D1S2663 and D1S228. Several candidate genes for SCCD have been localized to the 1p34.1-p36 interval.


Subject(s)
Chromosomes, Human, Pair 1 , Corneal Dystrophies, Hereditary/genetics , Chromosome Mapping , Haplotypes , Humans , Pedigree
7.
Cell Motil Cytoskeleton ; 27(4): 313-26, 1994.
Article in English | MEDLINE | ID: mdl-8069939

ABSTRACT

Manipulation of the single conventional myosin heavy chain (mhc) gene in Dictyostelium discoideum (Dd) has delineated an essential role for the filament-forming, or light meromyosin (LMM) domain of the myosin molecule in cytokinesis, development, and in the capping of cell surface receptors (see Spudich: Cell Regulation 1:1-11, 1989; Egelhoff et al.: Journal of Cell Biology, 112:677-688, 1991a). In order to assess the functional relationship between sarcomeric and cytoplasmic myosins, a chimeric gene encoding the Dd myosin head and subfragment 2 fused to rat beta cardiac LMM was transfected into both wild-type and Dd mhc null cells. Chimeric myosin was organized into dense cortical patches in the cytoplasm of both wild-type and Dd mhc null cells. Although null cells expressing chimeric mhc at approximately 10% of Dd mhc levels were unable to grow in shaking suspension or to complete development, chimeric myosin was able to rescue capping of cell surface receptors, to associate with filamentous actin, and to localize to the correct subcellular position during aggregation. Deletion of 29 amino acids in the rod corresponding to a previously defined filament assembly competent region eliminated the cortical patches and the posterior localization during chemotaxis. Taken together, these observations suggest that sarcomeric and cytoplasmic myosin rods are functionally interchangeable in several aspects of nonmuscle motility.


Subject(s)
Dictyostelium/chemistry , Frameshift Mutation , Myosin Subfragments/ultrastructure , Actins/analysis , Animals , Base Sequence , Chemotaxis/physiology , Chromosome Deletion , Molecular Sequence Data , Myocardium/chemistry , Myosin Subfragments/analysis , Myosin Subfragments/genetics , Rats , Recombinant Fusion Proteins/analysis , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/ultrastructure , Transfection
8.
Proc Natl Acad Sci U S A ; 88(21): 9618-22, 1991 Nov 01.
Article in English | MEDLINE | ID: mdl-1658787

ABSTRACT

The chromosome 11p13 Wilms tumor susceptibility gene WT1 appears to play a crucial role in regulating the proliferation and differentiation of nephroblasts and gonadal tissue. The WT1 gene consists of 10 exons, encoding a complex pattern of mRNA species: four distinct transcripts are expressed, reflecting the presence or absence of two alternative splices. Splice I consists of a separate exon, encoding 17 amino acids, which is inserted between the proline-rich amino terminus and the zinc finger domains. Splice II arises from the use of an alternative 5' splice junction and results in the insertion of 3 amino acids between zinc fingers 3 and 4. RNase protection analysis demonstrates that the most prevalent splice variant in both human and mouse is that which contains both alternative splices, whereas the least common is the transcript missing both splices. The relative distribution of splice variants is highly conserved between normal fetal kidney tissue and Wilms tumors that have intact WT1 transcripts. The ratio of these different WT1 mRNA species is also maintained as a function of development in the mouse kidney and in various mouse tissues expressing WT1. The conservation in structure and relative levels of each of the four WT1 mRNA species suggests that each encoded polypeptide makes a significant contribution to normal gene function. The control of cellular proliferation and differentiation exerted by the WT1 gene products may involve interactions between four polypeptides with distinct targets and functions.


Subject(s)
DNA-Binding Proteins/genetics , Kidney/physiology , RNA Splicing , RNA, Messenger/genetics , Wilms Tumor/genetics , Amino Acid Sequence , Animals , Base Sequence , Chromosomes, Human, Pair 11 , Genes , Genes, Tumor Suppressor , Humans , Mice , Molecular Sequence Data , Oligonucleotides/chemistry , Polymerase Chain Reaction , Zinc Fingers
9.
Cell ; 61(7): 1257-69, 1990 Jun 29.
Article in English | MEDLINE | ID: mdl-2163761

ABSTRACT

We have recently described the isolation of a candidate for the Wilms' tumor susceptibility gene mapping to band p13 of human chromosome 11. This gene, primarily expressed in fetal kidney, appears to encode a DNA binding protein. We now describe a sporadic, unilateral Wilms' tumor in which one allele of this gene contains a 25 bp deletion spanning an exon-intron junction and leading to aberrant mRNA splicing and loss of one of the four zinc finger consensus domains in the protein. The mutation is absent in the affected individual's germline, consistent with the somatic inactivation of a tumor suppressor gene. In addition to this intragenic deletion affecting one allele, loss of heterozygosity at loci along the entire chromosome 11 points to an earlier chromosomal nondisjunction and reduplication. We conclude that inactivation of this gene, which we call WT1, is part of a series of events leading to the development of Wilms' tumor.


Subject(s)
Chromosome Deletion , Chromosomes, Human, Pair 11 , DNA-Binding Proteins/genetics , Kidney Neoplasms/genetics , Metalloproteins/genetics , Wilms Tumor/genetics , Zinc/metabolism , Alleles , Base Sequence , Chromosome Mapping , DNA, Neoplasm/genetics , Exons , Genetic Predisposition to Disease , Humans , Introns , Molecular Sequence Data , Nucleic Acid Hybridization , Oligonucleotide Probes , Polymerase Chain Reaction , RNA Splicing , RNA, Messenger/genetics , RNA, Messenger/isolation & purification , RNA, Neoplasm/genetics , RNA, Neoplasm/isolation & purification
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