Protein phosphatase 2B inhibitor potentiates endothelial PKC activity and barrier dysfunction.
Serine/threonine (Ser/Thr) protein phosphatases (PPs) are implicated in the recovery from endothelial barrier dysfunction caused by inflammatory mediators. We hypothesized that Ser/Thr PPs may regulate protein kinase C (PKC), a critical signaling molecule in barrier dysfunction, in the promotion of barrier recovery. Western analysis indicated that bovine pulmonary microvascular endothelial cells (BPMECs) expressed the three major Ser/Thr PPs, PP1, PP2A, and PP2B. Pretreatment with 100 ng/ml of FK506 (a PP2B inhibitor) but not with the PP1 and PP2A inhibitors calyculin A or okadaic acid potentiated the thrombin-induced increase in PKC phosphotransferase activity. FK506 also potentiated thrombin-induced PKC-alpha but not PKC-beta phosphorylation. FK506 but not calyculin A or okadaic acid inhibited recovery from the thrombin-induced decrease in transendothelial resistance. Neither FK506 nor okadaic acid altered the thrombin-induced resistance decrease, whereas calyculin A potentiated the decrease. Downregulation of PKC with phorbol 12-myristate 13-acetate rescued the FK506-mediated inhibition of recovery, which was consistent with the finding that the thrombin-induced phosphorylation of PKC-alpha was reduced during the recovery phase. These results indicated that PP2B may play a physiologically important role in returning endothelial barrier dysfunction to normal through the regulation of PKC.
Lum
H
H
Department of Pharmacology, Rush-Presbyterian-St. Luke's Medical Center, Chicago, Illinois 60612, USA. hlum@rush.edu
Podolski
J L
JL
Gurnack
M E
ME
Schulz
I T
IT
Huang
F
F
Holian
O
O
D000818:Animals; D002199:Capillary Permeability / Q000502:physiology*; D002417:Cattle; D002478:Cells, Cultured; D017097:Electric Impedance; D004730:Endothelium, Vascular / Q000502:physiology* / Q000503:physiopathology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000378:metabolism* / Q000502:physiology*; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors / Q000502:physiology*; D010766:Phosphorylation / Q000187:drug effects; D011493:Protein Kinase C / Q000378:metabolism*; D051571:Protein Kinase C-alpha; D016559:Tacrolimus / Q000494:pharmacology; D013917:Thrombin / Q000494:pharmacology
D004791:Enzyme Inhibitors; D007527:Isoenzymes; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D010749:Phosphoprotein Phosphatases; D013917:Thrombin; D016559:Tacrolimus