The origin of the
heparan sulfate proteoglycan (PG),
perlecan, in
beta-amyloid protein (A beta)-containing
amyloid deposits in
Alzheimer's disease (AD) brain is not known. In the present investigation we used indirect immunofluorescence, SDS-PAGE, and Western blotting with a specific
perlecan core
protein antibody to identify possible cell candidates of
perlecan production in both primary cell cultures and in a rat infusion model. Double and triple-labeled indirect immunofluorescence was performed on dissociated primary rat septal cultures using
antibodies for specific identification of cell types and for
perlecan core
protein. In mixed cultures of both embryonic day 18 (containing neurons and glia) and postnatal day 2-3 (devoid of neurons), microglia identified by labeling with OX-42 or anti-ED1 were the only cell type also double labeled with an affinity-purified polyclonal antibody against
perlecan core
protein. Similar immunolabeling of microglia with the anti-
perlecan antibody was also observed in purified cultures of post-natal rat microglia. Analyses of PGs from cultured postnatal rat microglia by Western blotting using a polyclonal antibody against
perlecan core
protein revealed an approximately 400 kDa band in cell layer, which was intensified following
heparitinase/
heparinase digestion, suggestive of
perlecan core
protein. Other lower Mr bands were also found implicating either degradation of the 400 kDa core
protein or the presence of separate and distinct gene products immunologically related to
perlecan. Reverse transcription followed by polymerase chain reaction using human
perlecan domain I specific primers demonstrated
perlecan mRNA in cultured human microglia derived from postmortem normal aged and AD brain. Following a 1-week continuous infusion of
A beta (1-40) into rodent hippocampus, immunoperoxidase immunocytochemistry and double-labeled immunofluorescent studies revealed
perlecan accumulation primarily localized to microglia/macrophages within the A beta infusion site. These studies have identified microglia/macrophages as one potential source of
perlecan (or a
perlecan-related macromolecule) which may be important for the ongoing accumulation of both
perlecan and A beta in the
amyloid deposits of AD.