title: Intracellular thiol redox status regulates lymphangiogenesis and dictates corneal limbal graft survival.
abstract: OBJECTIVE
  Compounds regulating intracellular thiol redox status, such as N,N-diacetyl-L-cystine dimethylester (NM(2)), were shown to prolong corneal graft survival in a penetrating keratoplasty (PKP) model. However, the effect of NM(2) on hemangiogenesis and lymphangiogenesis has not been investigated. The effect of manipulating ambient thiol redox status on riskier (higher rejection rate) transplantation models, such as limbal graft survival and hemangiogenesis and lymphangiogenesis in a corneal suture model, were investigated.
  
  
  METHODS
  C57BL/10 mice that received BALB/c corneas were treated by subconjunctival injection of NM(2), and limbal graft survival was assessed. Sutured C57BL/6 received daily intraperitoneal injections of NM(2), glutathione diethylester (GSH-OEt), or PBS. Lymphatic endothelial cell (LEC) and peritoneal mps were treated with NM(2) or GSHOEt, and then VEGFR3, neuropilin-2, podoplanin, and LYVE-1 expression were analyzed. Supernatants were collected for analysis of TNF-alpha and VEGF-A levels by ELISA.
  
  
  RESULTS
  Significantly less cellular infiltration was detected in mice with corneal limbal transplant-treated NM(2)-treated mice. Hemangiogenesis and lymphangiogenesis were suppressed in the NM(2)-treated mice. NM(2) treatment of mps led to reduced levels of VEGFR3, neuropilin-2, podoplanin, and LYVE-1 expression compared with PBS- or GSHOEt- treated mps, lower levels of TNF-alpha, and increased secretion of VEGF. Moreover, NM(2)-treated LECs had reduced levels of LYVE-1 and Prox-1.
  
  
  CONCLUSIONS
  Reduction of ambient redox status reduced inflammatory cell infiltrates. Consequently, reduced inflammatory response might have contributed to both the observed prolonged corneal limbal graft survival and the attenuated hemangiogenesis and lymphangiogenesis in cornea.
authors.affiliation: Department of Ophthalmology, Kyoto Prefectural University of Medicine, Kamigyo-ku, Kyoto, Japan.
authors.forename: Akiko
authors.initials: A
authors.lastname: Fukumoto
authors.forename: Kazuichi
authors.initials: K
authors.lastname: Maruyama
authors.forename: Tony
authors.initials: T
authors.lastname: Walsh
authors.forename: Kentaro
authors.initials: K
authors.lastname: Kajiya
authors.forename: Junji
authors.initials: J
authors.lastname: Hamuro
authors.forename: Patricia A
authors.initials: PA
authors.lastname: D'Amore
authors.forename: Shigeru
authors.initials: S
authors.lastname: Kinoshita
mesh_terms: D000111:Acetylcysteine / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D015153:Blotting, Western; D016510:Corneal Neovascularization / Q000517:prevention & control; D016039:Corneal Transplantation; D004195:Disease Models, Animal; D004729:Endothelium, Lymphatic / Q000187:drug effects / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005978:Glutathione / Q000031:analogs & derivatives / Q000494:pharmacology; D006023:Glycoproteins / Q000378:metabolism; D006085:Graft Survival / Q000187:drug effects*; D016850:Limbus Corneae; D042583:Lymphangiogenesis / Q000187:drug effects*; D017737:Macrophages, Peritoneal; D008297:Male; D008562:Membrane Glycoproteins / Q000378:metabolism; D026901:Membrane Transport Proteins; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D039943:Neuropilin-2 / Q000378:metabolism; D010084:Oxidation-Reduction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013438:Sulfhydryl Compounds; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism; D040321:Vascular Endothelial Growth Factor Receptor-3 / Q000378:metabolism; D000098853:Podoplanin
chemical_list: D000111:Acetylcysteine; D005978:Glutathione; D006023:Glycoproteins; D008562:Membrane Glycoproteins; D026901:Membrane Transport Proteins; D039943:Neuropilin-2; D013438:Sulfhydryl Compounds; D014409:Tumor Necrosis Factor-alpha; D042461:Vascular Endothelial Growth Factor A; D040321:Vascular Endothelial Growth Factor Receptor-3; C490441:Pdpn protein, mouse; D000098853:Podoplanin; C492273:LYVE1 protein, mouse; C467480:vascular endothelial growth factor A, mouse; C030889:N,N-diacetylcysteine; C083627:glutathione diethyl ester