Immunochemical identification of ubiquitin and heat-shock proteins in corpora amylacea from normal aged and Alzheimer's disease brains

Abstract

Corpora amylacea (CA) accumulation in the central nervous system (CNS) is associated with both normal aging and neurodegenerative conditions such as Alzheimer's disease (AD). CA is reported to be primarily composed of glucose polymers, but approximately 4% of the total weight of CA is consistently composed of protein. CA protein resolved on sodium dodecylsulfate-polyacrylamide gel electrophoresis showed a broad range of polypeptides ranging from 24 to 133 kDa, with four abundant bands. Immunoblots of the profile of polypeptides solubilized from purified CA, showed positive ubiquitin (Ub) immunoreactivity for all the bands. Antisera to heat-shock proteins (hsp) 28 and 70 reacted selectively with bands of 30 and 67 kDa. These results show that Ub is associated with the primary protein components of CA and that the polypeptides are likely to be Ub conjugates. Immunostaining experiments were performed to specifically characterize the protein components of CA in brain tissue sections as well as those of CA purified from both AD and normal aged brains. In all cases CA showed positive reactions with antibodies to Ub, with antibodies raised against either paired helical filaments or hsp 28 or 70, the most prominent staining being with antibodies to Ub, hsp 28 or hsp 70. The presence of Ub and hsp 28 and 70, which are actively induced after stress, suggests that accumulation of altered proteins, possibly attributed to an increased frequency of unusual post-translational modifications or to a sustained physiological stress (related to both normal aging and neurodegenerative process), may be involved in the pathogenesis of CA.