title: Effects of ranibizumab on TGF-β1 and TGF-β2 production by human Tenon's fibroblasts: An in vitro study.
abstract: OBJECTIVE
  Inhibiting exaggerated wound healing responses, which are primarily mediated by human Tenon's fibroblast (HTF) migration and proliferation, has become the major determining factor for a successful trabeculectomy. Antivascular endothelial growth factor (anti-VEGF) has showed promising results as a potential antifibrotic candidate for use concurrently in trabeculectomy. Preliminary cohort studies have revealed improved bleb morphology following trabeculectomy augmented with ranibizumab. However, the effects on HTFs remain unclear. This study was conducted to understand the effects of ranibizumab on transforming growth factor (TGF)-β1 and transforming growth factor (TGF)-β2 expression by HTFs.
  
  
  METHODS
  The effect of ranibizumab on HTF proliferation and cell viability was determined using 3-(4,5-dimethylthiazone-2-yl)-2,5-diphenyl tetrazolium (MTT) assay. Ranibizumab at concentrations ranging from 0.01 to 0.5 mg/ml were administered for 24, 48, and 72 h in serum and serum-free conditions. Supernatants and cell lysates from samples were assessed for TGF-β1 and TGF-β2 mRNA and protein levels using quantitative real-time polymerase chain reaction (qRT-PCR) and enzyme-linked immunosorbent assay (ELISA).
  
  
  RESULTS
  At 48 h, 0.5 mg/ml of ranibizumab significantly induced cell death under serum-free culture conditions (p<0.05). Ranibizumab caused a significant reduction in TGF-β1 mRNA, but not for TGF-β2. However, the total protein production of TGF-β1 and TGF-β2 was unaffected by this anti-VEGF treatment.
  
  
  CONCLUSIONS
  Exposure of HTFs to an intravitreal dose of ranibizumab significantly suppresses cell viability in vitro; however, the application seemed unable to affect the ultimate production of TGF-β. Therefore, we highlighted ranibizumab as a potential antiscarring agent that acts via a different mechanism when used synergistically with another antifibrotic agent. Understanding the mechanism of actions of ranibizumab offers an additional view of a possible new rational therapeutic strategy.
authors.affiliation: Faculty of Medicine, Brain and Neuroscience Communities of Research, Universiti Teknologi MARA (UiTM) Shah Alam, Selangor, Malaysia.
authors.forename: Siti Munirah Md
authors.initials: SM
authors.lastname: Noh
authors.affiliation: Faculty of Medicine, Brain and Neuroscience Communities of Research, Universiti Teknologi MARA (UiTM) Shah Alam, Selangor, Malaysia.
authors.forename: Siti H Sheikh
authors.initials: SH
authors.lastname: Abdul Kadir
authors.affiliation: Centre for Eye Research Australia, the University Of Melbourne, Melbourne, VIC, Australia.
authors.forename: Jonathan G
authors.initials: JG
authors.lastname: Crowston
authors.affiliation: Department of Ophthalmology, Faculty of Medicine, University of Malaya, Malaysia.
authors.forename: Visvaraja
authors.initials: V
authors.lastname: Subrayan
authors.affiliation: Faculty of Medicine, Brain and Neuroscience Communities of Research, Universiti Teknologi MARA (UiTM) Shah Alam, Selangor, Malaysia.
authors.forename: Sushil
authors.initials: S
authors.lastname: Vasudevan
mesh_terms: D020533:Angiogenesis Inhibitors / Q000494:pharmacology*; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002921:Cicatrix / Q000209:etiology / Q000473:pathology / Q000517:prevention & control; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D005786:Gene Expression Regulation; D006801:Humans; D061251:Primary Cell Culture; D012333:RNA, Messenger / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D000069579:Ranibizumab / Q000494:pharmacology*; D015398:Signal Transduction; D058475:Tenon Capsule / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D014130:Trabeculectomy / Q000009:adverse effects; D053773:Transforming Growth Factor beta1 / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000235:genetics; D053781:Transforming Growth Factor beta2 / Q000096:biosynthesis / Q000235:genetics; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000378:metabolism
chemical_list: D020533:Angiogenesis Inhibitors; D012333:RNA, Messenger; D053773:Transforming Growth Factor beta1; D053781:Transforming Growth Factor beta2; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; D000069579:Ranibizumab