Abstract
Purpose: :
To determine the role(s) of decorin and biglycan, 2 closely related class I small leucine–rich proteoglycans (SLRP), in the regulation of corneal fibril and matrix assembly. Our hypothesis is that stromal matrix assembly involves coordinate regulation by pairs of SLRPs.
Methods: :
Expression patterns were analyzed in wild type and deficient corneas using semi–quantitative RT–PCR and immuno–blots as well as immuno–localization. Fibril structure and stromal organization was analyzed using transmission electron microscopy.
Results: :
Decorin and biglycan mRNA demonstrated stable expression in newborn to mature corneas. Biglycan demonstrated a consistent decrease in expression around eye opening and then maintained constant expression. Both protein cores had an overlapping expression patterns throughout the stroma. This pattern was constant throughout postnatal development and protein core expression was relatively stable. There was a consistent decrease in biglycan at P90. The data indicate that decorin is the major class I SLRP with biglycan present at constant low levels. Ultrastructural analyses demonstrated comparable fibril structure and organization in the wild type, decorin– and biglycan–deficient corneas. However, our data indicate an up–regulation of biglycan in decorin–deficient mice, but not the reverse. This prompted us to further investigate the relationship between decorin and biglycan. We crossed the decorin–deficient and biglycan–deficient mice to produce a double–deficient, compound mutant. The stroma of 2 month double–deficient mice demonstrated a severe disruption in fibril structure and stromal organization. This phenotype was observed throughout the stroma, however, differences between the anterior and posterior stroma were observed. The posterior stroma contained larger, more irregular abnormal fibrils than in the anterior stroma. Fibril packing/organization was disrupted in both regions, but very severe focal disruption was observed only in the posterior stroma.
Conclusions: :
These data indicate that decorin has a key role in the regulation of stromal fibril assembly. This role is masked in the decorin–deficient stroma by the up–regulation of biglycan. These SLRPs interact with the same fibril binding site, but with differing affinities. The up–regulation of biglycan in decorin–deficient corneas compensates for loss of decorin resulting in normal fibrillogenesis. However, when compensation was prevented in double–deficient mice, the decorin–deficient phenotype was dominant.
Keywords: cornea: stroma and keratocytes • extracellular matrix • development