Corneal opacity in lumican-null mice: Defects in collagen fibril structureand packing in the posterior stroma

PURPOSE. Gene targeted lumican-null mutants (lum(tm1sc)/lum(tm1sc)) have cl oudy corneas with abnormally thick collagen fibrils. The purpose of the pre sent study was to analyze the loss of transparency quantitatively and to de fine the associated corneal collagen fibril and stromal defects. METHODS. Backscattering of light, a function of corneal haze and opacificat ion, was determined regionally using in vivo confocal microscopy in lumican -deficient and wild-type control mice. Fibril organization and structure we re analyzed using transmission electron microscopy. Biochemical approaches were used to quantify glycosaminoglycan contents. Lumican distribution in t he cornea was elucidated immunohistochemically. RESULTS. Compared with control stromas, lumican-deficient stromas displayed a threefold increase in backscattered light with maximal increase confined to the posterior stroma. Confocal microscopy through-focusing (CMTF) measu rement profiles also indicated a 40% reduction in stromal thickness in the lumican-null mice. Transmission electron microscopy indicated significant c ollagen fibril abnormalities in the posterior stroma, with the anterior str oma remaining relatively unremarkable. The lumican-deficient posterior stro ma displayed a pronounced increase in fibril diameter, large fibril aggrega tes, altered fibril packing, and poor lamellar organization. Immunostaining of wild-type corneas demonstrated high concentrations of lumican in the po sterior stroma. Biochemical assessment of keratan sulfate (KS) content of w hole eyes revealed a 25% reduction in KS content in the lumican-deficient m ice. CONCLUSIONS. The structural defects and maximum backscattering of Light cle arly localized to the posterior stroma of lumican-deficient mice. In normal mice, an enrichment of lumican was observed in the posterior stroma compar ed with that in the anterior stroma. Taken together, these observations ind icate a key role for lumican in the posterior stroma in maintaining normal fibril architecture, most likely by regulating fibril assembly and maintain ing optimal KS content required for transparency.