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Engineered endothelial progenitor cells that overexpress prostacyclin protect vascular cells
Author(s) -
Liu Qi,
Xi Yutao,
Terry Toya,
So ShuiPing,
Mohite Anita,
Zhang Jia,
Wu Geru,
Liu Xiaobing,
Cheng Jie,
Ruan KeHe,
Willerson James T.,
Dixon Richard A.F.
Publication year - 2012
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/jcp.23035
Subject(s) - prostacyclin , progenitor cell , microbiology and biotechnology , endothelial stem cell , stem cell , biology , chemistry , biochemistry , in vitro
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. J. Cell. Physiol. 227: 2907–2916, 2012. © 2011 Wiley Periodicals, Inc.