Document Detail


Engineered endothelial progenitor cells that overexpress prostacyclin protect vascular cells.
MedLine Citation:
PMID:  21938725     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Prostacyclin (PGI2) is a potent vasodilator and important mediator of vascular homeostasis; however, its clinical use is limited because of its short (<2-min) half-life. Thus, we hypothesize that the use of engineered endothelial progenitor cells (EPCs) that constitutively secrete high levels of PGI2 may overcome this limitation of PGI2 therapy. A cDNA encoding COX-1-10aa-PGIS, which links human cyclooxygenase-1 (COX-1) to prostacyclin synthase (PGIS), was delivered via nucleofection into outgrowth EPCs derived from rat bone marrow mononuclear cells. PGI2-secreting strains (PGI2-EPCs) were established by continuous subculturing of transfected cells under G418 selection. Genomic PCR, RT-PCR, and Western blot analyses confirmed the overexpression of COX-1-10aa-PGIS in PGI2-EPCs. PGI2-EPCs secreted significantly higher levels of PGI2 in vitro than native EPCs (P < 0.05) and showed higher intrinsic angiogenic capability; conditioned medium (CM) from PGI2-EPCs promoted better tube formation than CM from native EPCs (P < 0.05). Cell- and paracrine-mediated in vitro angiogenesis was attenuated when COX-1-10aa-PGIS protein expression was knocked down. Whole-cell patch-clamp studies showed that 4-aminopyridine-sensitive K(+) current density was increased significantly in rat smooth muscle cells (rSMCs) cocultured under hypoxia with PGI2-EPCs (7.50 ± 1.59 pA/pF; P < 0.05) compared with rSMCs cocultured with native EPCs (3.99 ± 1.26 pA/pF). In conclusion, we successfully created EPC strains that overexpress an active novel enzyme resulting in consistent secretion of PGI2. PGI2-EPCs showed enhanced intrinsic proangiogenic properties and provided favorable paracrine-mediated cellular protections, including promoting in vitro angiogenesis of native EPCs and hyperpolarization of SMCs under hypoxia.
Authors:
Qi Liu; Yutao Xi; Toya Terry; Shui-Ping So; Anita Mohite; Jia Zhang; Geru Wu; Xiaobing Liu; Jie Cheng; Ke-He Ruan; James T Willerson; Richard A F Dixon
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Publication Detail:
Type:  Journal Article; Research Support, N.I.H., Extramural    
Journal Detail:
Title:  Journal of cellular physiology     Volume:  227     ISSN:  1097-4652     ISO Abbreviation:  J. Cell. Physiol.     Publication Date:  2012 Jul 
Date Detail:
Created Date:  2012-03-21     Completed Date:  2012-07-19     Revised Date:  2013-07-03    
Medline Journal Info:
Nlm Unique ID:  0050222     Medline TA:  J Cell Physiol     Country:  United States    
Other Details:
Languages:  eng     Pagination:  2907-16     Citation Subset:  IM    
Copyright Information:
Copyright © 2011 Wiley Periodicals, Inc.
Affiliation:
The Texas Heart Institute at St. Luke's Episcopal Hospital, Houston, TX, USA.
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MeSH Terms
Descriptor/Qualifier:
4-Aminopyridine / metabolism
Animals
Anoxia / genetics,  metabolism
Apoptosis / genetics
Cell Engineering / methods*
Cell Growth Processes / genetics
Culture Media, Conditioned / metabolism
Cyclooxygenase 1 / genetics
Cytochrome P-450 Enzyme System / genetics
DNA, Complementary / genetics
Endothelium, Vascular / cytology,  metabolism*
Epoprostenol / biosynthesis*,  genetics*,  metabolism
Half-Life
Intramolecular Oxidoreductases / genetics
Membrane Proteins / genetics
Muscle, Smooth, Vascular / cytology,  metabolism*
Myocytes, Smooth Muscle / metabolism*
Neovascularization, Physiologic
Phenotype
Potassium Channels / metabolism
Rats
Recombinant Fusion Proteins / genetics,  metabolism
Stem Cells / metabolism*
Transfection / methods
Grant Support
ID/Acronym/Agency:
RC1 HL100807-01/HL/NHLBI NIH HHS; RC1HL100807/HL/NHLBI NIH HHS; UL1 RR024148/RR/NCRR NIH HHS
Chemical
Reg. No./Substance:
0/Culture Media, Conditioned; 0/DNA, Complementary; 0/Membrane Proteins; 0/Potassium Channels; 0/Recombinant Fusion Proteins; 35121-78-9/Epoprostenol; 504-24-5/4-Aminopyridine; 9035-51-2/Cytochrome P-450 Enzyme System; EC 1.14.99.1/Cyclooxygenase 1; EC 1.14.99.1/Ptgs1 protein, rat; EC 5.3.-/Intramolecular Oxidoreductases; EC 5.3.99.4/prostacyclin synthetase
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