title: Targeted expression of a lumican transgene rescues corneal deficiencies in lumican-null mice.
abstract: OBJECTIVE
  To investigate whether targeted expression of lumican in the mouse cornea rescued the Lum(-/-) phenotype.
  
  
  METHODS
  Lum(-/-)/Kera-Lum mice were generated by crossing Lum(-/-) mice with Kera-Lum transgenic mice that overexpressed lumican under the control of the keratocan promoter. Mouse eyes were analyzed in vivo by confocal microscopy through focusing (CMTF) to determine corneal sublayer thickness and haze. Subsequently, one cornea from each mouse was processed for SDS-PAGE/western blotting while the other was used for either electron microscopy (EM) or real-time polymerase chain reaction (RT-PCR).
  
  
  RESULTS
  Overall, corneas of Lum(-/-)/Kera-Lum mice showed significant improvement over Lum(-/-) but were still deficient when compared to wildtype (WT) mice. Specifically, analysis of Lum(-/-)/Kera-Lum mouse eyes by CMTF showed a similar stromal but slightly increased epithelial thickness compared to matching Lum(-/-) mice. Analysis of the CMTF scans for light backscattering revealed a small yet significant reduction in corneal haze in Lum(-/-)/Kera-Lum mice as compared to Lum(-/-) mice. At the EM level, the pronounced disarray of the posterior fibrillar matrix seen in Lum(-/-) mice was not observed in Lum(-/-)/Kera-Lum mice. Moreover, analyses of collagen fibril diameter distributions showed a significant reduction in the number of large-diameter (>40 nm) fibrils in Lum(-/-)/Kera-Lum mice as compared to Lum(-/-) mice. No significant differences in keratocan expression were found at the mRNA level, but western blot analysis detected an approximately twofold increase in keratocan protein levels in Lum(-/-)/Kera-Lum over Lum(-/-) mice.
  
  
  CONCLUSIONS
  Together these data suggest that despite the low keratocan promoter activity driving the transgene in Lum(-/-) cornea, transgenic lumican expression was sufficient to partially rescue corneal phenotypic deficiencies.
authors.affiliation: Department of Ophthalmology, University of Cincinnati, Cincinnati, OH, USA.
authors.forename: Johanna T A
authors.initials: JT
authors.lastname: Meij
authors.forename: Eric C
authors.initials: EC
authors.lastname: Carlson
authors.forename: Li
authors.initials: L
authors.lastname: Wang
authors.forename: Chia-Yang
authors.initials: CY
authors.lastname: Liu
authors.forename: James V
authors.initials: JV
authors.lastname: Jester
authors.forename: David E
authors.initials: DE
authors.lastname: Birk
authors.forename: Winston W Y
authors.initials: WW
authors.lastname: Kao
mesh_terms: D000818:Animals; D011508:Chondroitin Sulfate Proteoglycans / Q000172:deficiency* / Q000235:genetics* / Q000378:metabolism; D003315:Cornea / Q000473:pathology; D003316:Corneal Diseases / Q000209:etiology / Q000378:metabolism* / Q000473:pathology*; D015870:Gene Expression; D018390:Gene Targeting; D007632:Keratan Sulfate / Q000172:deficiency* / Q000235:genetics* / Q000378:metabolism; D000071234:Lumican; D051379:Mice; D018345:Mice, Knockout; D008822:Mice, Transgenic; D018613:Microscopy, Confocal; D008854:Microscopy, Electron; D010641:Phenotype; D011401:Promoter Regions, Genetic; D011509:Proteoglycans / Q000235:genetics; D019076:Transgenes
chemical_list: D011508:Chondroitin Sulfate Proteoglycans; C099280:Kera protein, mouse; C000604307:Lum protein, mouse; D000071234:Lumican; D011509:Proteoglycans; D007632:Keratan Sulfate