Vascular endothelial cells express isoforms of protein kinase A inhibitor.
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 PKI alpha, human PKI gamma, and mouse PKI beta sequences detected PKI alpha and PKI gamma mRNA in all three cell types. Sequencing and BLAST analysis indicated that forward and reverse DNA strands for PKI alpha and PKI gamma were of >96% identity with database sequences. RNase protection assays showed protection of the 542 nucleotides in HBMEC and HPAEC PKI alpha mRNA and 240 nucleotides in HBMEC, HPAEC, and HMEC PKI gamma mRNA. Western blot analysis indicated that PKI gamma protein was detected in all three cell types, whereas PKI alpha was found in HBMECs. In summary, endothelial cells from three different vascular beds express PKI alpha and PKI gamma, which may be physiologically important in endothelial barrier function.
Lum
Hazel
H
Department of Pharmacology, Rush Presbyterian St. Luke's Medical Center, Chicago, Illinois 60612, USA. hlum@rush.edu
Hao
Zengping
Z
Gayle
Dave
D
Kumar
Priyadarsini
P
Patterson
Carolyn E
CE
Uhler
Michael D
MD
D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D001921:Brain / Q000098:blood supply; D002199:Capillary Permeability / Q000502:physiology; D002352:Carrier Proteins / Q000235:genetics*; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors*; D020405:Dermis / Q000098:blood supply; D004730:Endothelium, Vascular / Q000166:cytology / Q000201:enzymology*; D015870:Gene Expression / Q000502:physiology; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D007527:Isoenzymes / Q000235:genetics*; D018482:Muscle, Skeletal / Q000201:enzymology; D011651:Pulmonary Artery / Q000166:cytology; D012333:RNA, Messenger / Q000032:analysis; D011817:Rabbits; D014655:Vascular Resistance / Q000502:physiology
D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D047908:Intracellular Signaling Peptides and Proteins; D007527:Isoenzymes; C491526:PKIA protein, human; C107591:PKIB protein, human; C107592:PKIG protein, human; D012333:RNA, Messenger; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases