Abstract
Purpose: :
To evaluate the distribution of CFH, CFB, and C3 immunoreactivity within age-related extramacular drusen for comparison with our previous descriptions of apolipoproteins and cholesterol in drusen 1,2. These proteins are encoded by genes believed important in modifying risk for age-related maculopathy (ARM).
Methods: :
Drusen were isolated manually from 8 eyes of 8 donors (66-93 years) with grossly normal maculas preserved in 4% paraformaldehyde within 6 hr of death. In cryosections of druse-enriched pellets (6-75 drusen per section), CFH, CFB and C3 were detected by indirect immunofluorescence using polyclonal antibodies directed against the human proteins. Sections were examined with differential interference contrast and wide-field epi-fluorescence microscopy in order to generate a druse census.
Results: :
All proteins examined were detectable, the prevalence are: CFH (94.6%, 314 of 331 drusen), C3 (84.1%, 238 of 283), and CFB (63.8%, 178 of 279). 6 out of 8 eyes were 100% stained by CFH, and 4 out of 8 were 100% stained by both C3 and CFB. In 4 eyes, all drusen stained for all 3 proteins. Immunoreactivity patterns included a diffuse distribution throughout each druse, an intensely fluorescent rim around the druse exterior or core3 in the center, or a heterogeneous distribution. Diffuse staining occurred less frequently for CFH (62.7% of total stained drusen) than for CFB (72.5%) or C3 (75.6%). Rims and cores occurred more frequently for CFH (17.8%, 16.2%, respectively) than for CFB (6.7%, 10.7%) and C3 (10.0%, 9.6%). Heterogeneous distributions were found in a small fraction of drusen (CFH, 15.2%; CFB, 12.9%; C3, 15.1%).
Conclusions: :
High druse prevalence for CFH, CFB, and C3 supports roles for these proteins in ARM. CFH joins apolipoproteins E, J, and cholesterol (unesterified and esterified) as ubiquitous druse components. Lower prevalence of CFB and C3 relative to CFH may be antibody-related. Greater overall heterogeneity of CFH immunoreactivity may signify greater involvement in druse remodeling or turnover. Different patterns of complement and lipid components(previously published) may signify different processes or time courses for their deposition.1. Li et al, Invest Ophthalmol Vis Sci. 2006 47:3119.2. Li et al, Exp Eye Res. 2007 85:192.3. Mullins, J Histochem Cytochem. 1999 47:1533.
Keywords: age-related macular degeneration • drusen • immunohistochemistry