title: Elevated expression of matrix metalloproteinase-9 and inflammatory cytokines in keratoconus patients is inhibited by cyclosporine A.
abstract: OBJECTIVE
  The present study was designed to understand the role of inflammatory cytokines secreted by corneal epithelial cells in keratoconus (KC) and the response to treatment with cyclosporine A (CyA).
  
  
  METHODS
  The study involved 129 Indian KC patients clinically graded according to Amsler-Krumeich classification and 20 healthy, nonectatic subjects as controls. Tear levels of matrix metalloproteinase-9 (MMP9), interleukin-6 (IL6), and tumor necrosis factor-α (TNFα) were measured using ELISA kits. Gene expression was measured by qPCR in corneal epithelial cells obtained by debridement from subjects undergoing ocular surface surgeries. In addition, epithelial cells were stimulated with TNFα and treated with CyA to study its role on MMP9 expression. Finally, 20 KC patients (27 eyes) with inflammatory symptoms were treated with topical CyA application.
  
  
  RESULTS
  We observed that MMP9, TNFα, and IL6 levels were strongly upregulated at the mRNA level in KC patient epithelia. Similarly, tears collected from KC patients exhibited high levels of MMP9 and IL6 protein. Cyclosporine A treatment significantly reduced the mRNA expression levels of IL6 and TNFα in both short- and long-term treatments; however, it reduced MMP9 levels only in long-term treatment in cultured corneal epithelial cells. Subsequent treatment of KC patients with CyA for approximately 6 months reduced tear MMP9 levels and led to local reduction in corneal curvatures as determined by corneal topography maps.
  
  
  CONCLUSIONS
  The data indicate that corneal epithelium contributes to elevated MMP9 and inflammatory cytokine expression in tears of KC patients. Cyclosporine A treatment reduced MMP9 and inflammatory cytokine levels in an in vitro inflammation model system. In KC patients, CyA treatment reduced MMP9 levels measured in tears with concomitant arrest of disease progression. Therefore, CyA might be a novel treatment strategy in KC patients but requires additional evaluation in larger cohorts. (ClinicalTrials.gov number, NCT01746823.).
authors: Cornea Department, Narayana Nethralaya, Bangalore, India.
authors: Rohit
authors: R
authors: Shetty
authors: GROW Research Laboratory, Narayana Nethralaya Foundation, Bangalore, India.
authors: Anuprita
authors: A
authors: Ghosh
authors: Singapore Eye Research Institute, Singapore.
authors: Rayne R
authors: RR
authors: Lim
authors: GROW Research Laboratory, Narayana Nethralaya Foundation, Bangalore, India.
authors: Murali
authors: M
authors: Subramani
authors: GROW Research Laboratory, Narayana Nethralaya Foundation, Bangalore, India.
authors: Krina
authors: K
authors: Mihir
authors: GROW Research Laboratory, Narayana Nethralaya Foundation, Bangalore, India.
authors: A R
authors: AR
authors: Reshma
authors: Cornea Department, Narayana Nethralaya, Bangalore, India.
authors: Ashwini
authors: A
authors: Ranganath
authors: Cornea Department, Narayana Nethralaya, Bangalore, India.
authors: Sriharsha
authors: S
authors: Nagaraj
authors: Cornea Clinic, Department of Ophthalmology, Maastricht University Medical Center, The Netherlands.
authors: Rudy M M A
authors: RM
authors: Nuijts
authors: Singapore Eye Research Institute, Singapore.
authors: Roger
authors: R
authors: Beuerman
authors: GROW Research Laboratory, Narayana Nethralaya Foundation, Bangalore, India.
authors: Reshma
authors: R
authors: Shetty
authors: GROW Research Laboratory, Narayana Nethralaya Foundation, Bangalore, India.
authors: Debashish
authors: D
authors: Das
authors: Singapore Eye Research Institute, Singapore Signature Research Program in Neuroscience and Behavioral Disorders, Duke-NUS Graduate Medical School, Singapore Department of Ophthalmology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.
authors: Shyam S
authors: SS
authors: Chaurasia
authors: GROW Research Laboratory, Narayana Nethralaya Foundation, Bangalore, India.
authors: Abhijit
authors: A
authors: Sinha-Roy
authors: GROW Research Laboratory, Narayana Nethralaya Foundation, Bangalore, India Singapore Eye Research Institute, Singapore.
authors: Arkasubhra
authors: A
authors: Ghosh
mesh_terms: D000328:Adult; D002478:Cells, Cultured; D019781:Corneal Topography; D016572:Cyclosporine / Q000627:therapeutic use*; D016207:Cytokines / Q000096:biosynthesis / Q000187:drug effects / Q000235:genetics*; D004791:Enzyme Inhibitors / Q000627:therapeutic use; D004797:Enzyme-Linked Immunosorbent Assay; D019573:Epithelium, Corneal / Q000378:metabolism / Q000473:pathology; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects / Q000235:genetics*; D006801:Humans; D007640:Keratoconus / Q000188:drug therapy / Q000235:genetics* / Q000378:metabolism; D008297:Male; D020780:Matrix Metalloproteinase 9 / Q000096:biosynthesis / Q000187:drug effects / Q000235:genetics*; D012313:RNA / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D013666:Tears / Q000378:metabolism; D016896:Treatment Outcome; D055815:Young Adult
chemical_list: D016207:Cytokines; D004791:Enzyme Inhibitors; D012313:RNA; D016572:Cyclosporine; D020780:Matrix Metalloproteinase 9
keywords: IL6; MMP9; TNFα; cornea; cyclosporine A; ectasia; epithelial cells; keratoconus