title: Sirolimus-FKBP12.6 impairs endothelial barrier function through protein kinase C-α activation and disruption of the p120-vascular endothelial cadherin interaction.
abstract: OBJECTIVE
  Sirolimus (SRL) is an immunosuppressant drug used to prevent rejection in organ transplantation and neointimal hyperplasia when delivered from drug-eluting stents. Major side effects of SRL include edema and local collection of intimal lipid deposits at drug-eluting stent sites, suggesting that SRL impairs endothelial barrier function (EBF). The aim of this study was to address the role of SRL on impaired EBF and the potential mechanisms involved.
  
  
  RESULTS
  Cultured human aortic endothelial cells (HAECs) and intact human and mouse endothelium was examined to determine the effect of SRL, which binds FKBP12.6 to inhibit the mammalian target of rapamycin, on EBF. EBF, measured by transendothelial electrical resistance, was impaired in HAECs when treated with SRL or small interfering RNA for FKBP12.6 and reversed when pretreated with ryanodine, a stabilizer of ryanodine receptor 2 intracellular calcium release channels. Intracellular calcium increased in HAECs treated with SRL and normalized with ryanodine pretreatment. SRL-treated HAECs demonstrated increases in protein kinase C-α phosphorylation, a calcium sensitive serine/threonine kinase important in vascular endothelial (VE) cadherin barrier function through its interaction with p120-catenin (p120). Immunostaining of HAECs, human coronary and mouse aortic endothelium treated with SRL showed disruption of p120-VE cadherin interaction treated with SRL. SRL impairment of HAEC EBF was reduced with protein kinase C-α small interfering RNA. Mice treated with SRL demonstrated increased vascular permeability by Evans blue albumin extravasation in the lungs, heart, and aorta.
  
  
  CONCLUSIONS
  SRL-FKBP12.6 impairs EBF by activation of protein kinase C-α and downstream disruption of the p120-VE cadherin interaction in vascular endothelium. These data suggest this mechanism may be an important contributor of SRL side effects related to impaired EBF.
authors.affiliation: From the Department of Medicine, Emory University School of Medicine, Atlanta, GA (A.H., V.K., R.P., H.A., K.P., A.V.F.); and CV Path Institute, Inc, Gaithersburg, MD (Q.C., F.D.K.).
authors.forename: Anwer
authors.initials: A
authors.lastname: Habib
authors.forename: Vinit
authors.initials: V
authors.lastname: Karmali
authors.forename: Rohini
authors.initials: R
authors.lastname: Polavarapu
authors.forename: Hirokuni
authors.initials: H
authors.lastname: Akahori
authors.forename: Qi
authors.initials: Q
authors.lastname: Cheng
authors.forename: Kim
authors.initials: K
authors.lastname: Pachura
authors.forename: Frank D
authors.initials: FD
authors.lastname: Kolodgie
authors.forename: Aloke V
authors.initials: AV
authors.lastname: Finn
mesh_terms: D000818:Animals; D015703:Antigens, CD / Q000378:metabolism*; D015820:Cadherins / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects; D002199:Capillary Permeability / Q000187:drug effects*; D051177:Catenins / Q000378:metabolism*; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000187:drug effects* / Q000201:enzymology; D004789:Enzyme Activation; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D010766:Phosphorylation; D011485:Protein Binding; D051571:Protein Kinase C-alpha / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D019837:Ryanodine Receptor Calcium Release Channel / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D020123:Sirolimus / Q000633:toxicity*; D022021:Tacrolimus Binding Proteins / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D013997:Time Factors; D014162:Transfection; D000095002:Delta Catenin; D000099152:Cadherin 5
chemical_list: D015703:Antigens, CD; D015820:Cadherins; D051177:Catenins; D051571:Protein Kinase C-alpha; D019837:Ryanodine Receptor Calcium Release Channel; D020123:Sirolimus; D022021:Tacrolimus Binding Proteins; D000095002:Delta Catenin; D000099152:Cadherin 5; C496628:PRKCA protein, human; C496630:Prkca protein, mouse; C403484:tacrolimus binding protein 1B
keywords: PKC; endothelium; sirolimus