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Am J Physiol Cell Physiol 282: C59-C66, 2002. First published September 5, 2001; doi:10.1152/ajpcell.00256.2001
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Vol. 282, Issue 1, C59-C66, January 2002

Vascular endothelial cells express isoforms of protein kinase A inhibitor

Hazel Lum1, Zengping Hao1, Dave Gayle1, Priyadarsini Kumar2, Carolyn E. Patterson3, and Michael D. Uhler4

1 Department of Pharmacology, Rush Presbyterian St. Luke's Medical Center, Chicago, Illinois 60612; 2 Department of Biological Chemistry, University of California, Davis, California 95616; 3 Department of Medicine, Indiana University School of Medicine, Indianapolis, Indiana 46202; and 4 Department of Biological Chemistry, University of Michigan, Ann Arbor, Michigan 48104-1687

First published September 5, 2001; 10.1152/ ajpcell.00256.2001.---The expression and function of the endogenous inhibitor of cAMP-dependent protein kinase (PKI) in endothelial cells are unknown. In this study, overexpression of rabbit muscle PKI gene into endothelial cells inhibited the cAMP-mediated increase and exacerbated thrombin-induced decrease in endothelial barrier function. We investigated PKI expression in human pulmonary artery (HPAECs), foreskin microvessel (HMECs), and brain microvessel endothelial cells (HBMECs). RT-PCR using specific primers for human PKIalpha , human PKIgamma , and mouse PKIbeta sequences detected PKIalpha and PKIgamma mRNA in all three cell types. Sequencing and BLAST analysis indicated that forward and reverse DNA strands for PKIalpha and PKIgamma were of >96% identity with database sequences. RNase protection assays showed protection of the 542 nucleotides in HBMEC and HPAEC PKIalpha mRNA and 240 nucleotides in HBMEC, HPAEC, and HMEC PKIgamma mRNA. Western blot analysis indicated that PKIgamma protein was detected in all three cell types, whereas PKIalpha was found in HBMECs. In summary, endothelial cells from three different vascular beds express PKIalpha and PKIgamma , which may be physiologically important in endothelial barrier function.

cAMP; protein kinase inhibitor; endothelial resistance; barrier function


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