title: Quantitative in vivo and ex vivo confocal microscopy analysis of corneal cystine crystals in the Ctns knockout mouse.
abstract: OBJECTIVE
  The purpose of this study was to assess the ability of quantitative in vivo confocal microscopy to characterize the natural history and detect changes in crystal volume in corneas from a novel animal model of cystinosis, the cystinosin (Ctns(-/-)) mouse.
  
  
  METHODS
  Two Ctns(-/-) mice and one C57Bl/6 mouse were examined at each of the following time points: 2, 3, 5, 7, 10, 12, and 14 months of age. In vivo confocal microscopy scans were performed in 4 different regions of the cornea per eye. After, animals were sacrificed and cornea blocks evaluated for cell morphology using phalloidin and lymphocytic infiltration using CD45 antibodies by ex vivo confocal microscopy. Cystine crystal content in the cornea was measured by calculating the pixel intensity of the crystals divided by the stromal volume using Metamorph Image Processing Software.
  
  
  RESULTS
  Corneal crystals were identified in Ctns(-/-) eyes beginning at 3 months of age and increased in density until 7-12 months, at which time animals begin to succumb to the disease and corneas become scarred and neovascularized. Older Ctns(-/-) mice (7 months and older) showed the presence of cell infiltrates that stained positively for CD45 associated with progressive keratocyte disruption. Finally, at 12 months of age, decreased cell density and endothelial distortion were detected.
  
  
  CONCLUSIONS
  Confocal microscopy identified corneal crystals starting at 3 month old Ctns(-/-) eyes. Cystine crystals induce inflammatory and immune response with aging associated with loss of keratocyte and endothelial cells. These findings suggest that the Ctns(-/-) mouse can be used as a model for developing and evaluating potential alternative therapies for corneal cystinosis.
authors: Department of Biochemistry, Jawaharlal Nehru Medical College, Maharashtra, India. jens@uci.edu
authors: Jennifer
authors: J
authors: Simpson
authors: Chyong Jy
authors: CJ
authors: Nien
authors: Kevin
authors: K
authors: Flynn
authors: Brian
authors: B
authors: Jester
authors: Stephanie
authors: S
authors: Cherqui
authors: James
authors: J
authors: Jester
mesh_terms: D000367:Age Factors; D026921:Amino Acid Transport Systems, Neutral / Q000172:deficiency* / Q000235:genetics; D000818:Animals; D002921:Cicatrix / Q000378:metabolism / Q000473:pathology*; D003315:Cornea / Q000098:blood supply / Q000378:metabolism / Q000473:pathology*; D003460:Crystallization; D003553:Cystine / Q000378:metabolism*; D003554:Cystinosis / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D004195:Disease Models, Animal; D042783:Endothelial Cells / Q000378:metabolism / Q000473:pathology; D005260:Female; D015603:Keratinocytes / Q000378:metabolism / Q000473:pathology; D017493:Leukocyte Common Antigens / Q000032:analysis; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D018613:Microscopy, Confocal / Q000379:methods*; D009389:Neovascularization, Pathologic / Q000378:metabolism*; D010590:Phalloidine
chemical_list: D026921:Amino Acid Transport Systems, Neutral; C534463:cystinosin protein, mouse; D010590:Phalloidine; D003553:Cystine; D017493:Leukocyte Common Antigens