title: Ocular surface extracellular DNA and nuclease activity imbalance: a new paradigm for inflammation in dry eye disease.
abstract: OBJECTIVE
  We determined whether nucleases are deficient in the tear fluid of dry eye disease (DED) patients, and whether this causes extracellular DNA (eDNA) and neutrophil extracellular trap (NET) accumulation in the precorneal tear film, thus causing ocular surface inflammation.
  
  
  METHODS
  Exfoliated cells adhered to Schirmer test strips were collected on glass slides, and immunofluorescence confocal microscopy was used to evaluate neutrophils, eDNA, NETs, and their molecular components. Similar experiments were performed with mucoid films collected from the inferior conjunctival fornix or bulbar conjunctiva. We used quantitative PCR to evaluate eDNA signaling pathways and inflammatory cytokine expression. We also determined the amount of ocular surface eDNA and evaluated tear fluid nuclease activity.
  
  
  RESULTS
  eDNA, NETs, and neutrophils were present on the ocular surface in DED patients and abundant in mucoid films. NETs consisted of eDNA, histones, cathelicidin, and neutrophil elastase. Tear fluid nuclease activity was decreased significantly in DED patients, whereas the amount of eDNA on the ocular surface was increased significantly. Expression of genes downstream of eDNA signaling, such as TLR9, MyD88, and type I interferon, as well as the inflammatory cytokines interleukin-6 and tumor necrosis factor-α, was significantly increased in DED patients.
  
  
  CONCLUSIONS
  Extracellular DNA production and clearance mechanisms are dysregulated in DED. Nuclease deficiency in tear fluid allows eDNA and NETs to accumulate in precorneal tear film, and results in ocular surface inflammation. These findings point to novel therapeutic interventions in severe DED based on clearance of eDNA, NETs, and other molecular components from the ocular surface.
authors.affiliation: Corneal Neurobiology Laboratory, Department of Ophthalmology and Visual Sciences, University of Illinois at Chicago, College of Medicine, Chicago, Illinois 60612, USA.
authors.forename: Snehal
authors.initials: S
authors.lastname: Sonawane
authors.forename: Vishakha
authors.initials: V
authors.lastname: Khanolkar
authors.forename: Abed
authors.initials: A
authors.lastname: Namavari
authors.forename: Shweta
authors.initials: S
authors.lastname: Chaudhary
authors.forename: Sonal
authors.initials: S
authors.lastname: Gandhi
authors.forename: Sapna
authors.initials: S
authors.lastname: Tibrewal
authors.forename: Sarmad H
authors.initials: SH
authors.lastname: Jassim
authors.forename: Brittany
authors.initials: B
authors.lastname: Shaheen
authors.forename: Joelle
authors.initials: J
authors.lastname: Hallak
authors.forename: John H
authors.initials: JH
authors.lastname: Horner
authors.forename: Martin
authors.initials: M
authors.lastname: Newcomb
authors.forename: Joy
authors.initials: J
authors.lastname: Sarkar
authors.forename: Sandeep
authors.initials: S
authors.lastname: Jain
mesh_terms: D023181:Antimicrobial Cationic Peptides / Q000378:metabolism*; D003228:Conjunctiva / Q000378:metabolism; D004247:DNA / Q000378:metabolism*; D003850:Deoxyribonuclease I / Q000378:metabolism*; D015352:Dry Eye Syndromes / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D031541:Fluorescence Resonance Energy Transfer; D019084:Fluorescent Antibody Technique, Indirect; D015870:Gene Expression; D006657:Histones / Q000378:metabolism; D006801:Humans; D019272:Leukocyte Elastase / Q000378:metabolism*; D054835:Lipocalin 1 / Q000378:metabolism*; D018613:Microscopy, Confocal; D009504:Neutrophils / Q000502:physiology; D016133:Polymerase Chain Reaction; D012463:Saliva / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D013666:Tears / Q000201:enzymology*; D054804:Cathelicidins
chemical_list: D023181:Antimicrobial Cationic Peptides; D006657:Histones; D054835:Lipocalin 1; D004247:DNA; D003850:Deoxyribonuclease I; D019272:Leukocyte Elastase; D054804:Cathelicidins