title: Rnd3 as a Novel Target to Ameliorate Microvascular Leakage.
abstract: BACKGROUND
  Microvascular leakage of plasma proteins is a hallmark of inflammation that leads to tissue dysfunction. There are no current therapeutic strategies to reduce microvascular permeability. The purpose of this study was to identify the role of Rnd3, an atypical Rho family GTPase, in the control of endothelial barrier integrity. The potential therapeutic benefit of Rnd3 protein delivery to ameliorate microvascular leakage was also investigated.
  
  
  RESULTS
  Using immunofluorescence microscopy, Rnd3 was observed primarily in cytoplasmic areas around the nuclei of human umbilical vein endothelial cells. Permeability to fluorescein isothiocyanate-albumin and transendothelial electrical resistance of human umbilical vein endothelial cell monolayers served as indices of barrier function, and RhoA, Rac1, and Cdc42 activities were determined using G-LISA assays. Overexpression of Rnd3 significantly reduced the magnitude of thrombin-induced barrier dysfunction, and abolished thrombin-induced Rac1 inactivation. Depleting Rnd3 expression with siRNA significantly extended the time course of thrombin-induced barrier dysfunction and Rac1 inactivation. Time-lapse microscopy of human umbilical vein endothelial cells expressing GFP-actin showed that co-expression of mCherry-Rnd3 attenuated thrombin-induced reductions in local lamellipodia that accompany endothelial barrier dysfunction. Lastly, a novel Rnd3 protein delivery method reduced microvascular leakage in a rat model of hemorrhagic shock and resuscitation, assessed by both intravital microscopic observation of extravasation of fluorescein isothiocyanate-albumin from the mesenteric microcirculation, and direct determination of solute permeability in intact isolated venules.
  
  
  CONCLUSIONS
  The data suggest that Rnd3 can shift the balance of RhoA and Rac1 signaling in endothelial cells. In addition, our findings suggest the therapeutic, anti-inflammatory potential of delivering Rnd3 to promote endothelial barrier recovery during inflammatory challenge.
authors.affiliation: Department of Molecular Pharmacology and Physiology, Morsani College of Medicine, University of South Florida, Tampa, FL jbreslin@health.usf.edu.
authors.forename: Jerome W
authors.initials: JW
authors.lastname: Breslin
authors.affiliation: Department of Biological Sciences, Old Dominion University, Norfolk, VA.
authors.forename: Dayle A
authors.initials: DA
authors.lastname: Daines
authors.affiliation: Department of Molecular Pharmacology and Physiology, Morsani College of Medicine, University of South Florida, Tampa, FL.
authors.forename: Travis M
authors.initials: TM
authors.lastname: Doggett
authors.affiliation: Department of Physiology, School of Medicine, Louisiana State University Health Sciences Center, New Orleans, LA.
authors.forename: Kristine H
authors.initials: KH
authors.lastname: Kurtz
authors.affiliation: Department of Physiology, School of Medicine, Louisiana State University Health Sciences Center, New Orleans, LA.
authors.forename: Flavia M
authors.initials: FM
authors.lastname: Souza-Smith
authors.affiliation: Department of Molecular Pharmacology and Physiology, Morsani College of Medicine, University of South Florida, Tampa, FL.
authors.forename: Xun E
authors.initials: XE
authors.lastname: Zhang
authors.affiliation: Department of Surgery, Morsani College of Medicine, University of South Florida, Tampa, FL.
authors.forename: Mack H
authors.initials: MH
authors.lastname: Wu
authors.affiliation: Department of Molecular Pharmacology and Physiology, Morsani College of Medicine, University of South Florida, Tampa, FL.
authors.forename: Sarah Y
authors.initials: SY
authors.lastname: Yuan
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002199:Capillary Permeability / Q000502:physiology*; D004730:Endothelium, Vascular / Q000166:cytology / Q000000981:diagnostic imaging / Q000502:physiology; D006801:Humans; D007249:Inflammation / Q000503:physiopathology; D008297:Male; D008856:Microscopy, Fluorescence; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000502:physiology; D020830:rac1 GTP-Binding Protein / Q000502:physiology; D020741:rho GTP-Binding Proteins / Q000502:physiology*; D020742:rhoA GTP-Binding Protein / Q000502:physiology
chemical_list: C107953:RAC1 protein, human; C066737:RHOA protein, human; C508705:RND3 protein, human; D020830:rac1 GTP-Binding Protein; D020741:rho GTP-Binding Proteins; D020742:rhoA GTP-Binding Protein
keywords: actin cytoskeleton; inflammation; lamellipodia; microcirculation; microvascular permeability