ALTERED EXPRESSION OF APOLIPOPROTEIN E, AMYLOID PRECURSOR PROTEIN AND PRESENILIN-1 IS ASSOCIATED WITH CHRONIC REACTIVE GLIOSIS IN RAT CORTICAL TISSUE R. N. MARTINS, a * K. TADDEI, a C. KENDALL, b G. EVIN, c K. A. BATES b and A. R. HARVEY b a Sir James McCusker Alzheimer Research Unit and University Department of Surgery, The University of Western Australia and Hollywood Private Hospital, Monash Avenue, Nedlands, WA 6009, Australia b Department of Anatomy and Human Biology, The University of Western Australia, Crawley, Perth, WA 6009, Australia c Department of Pathology, The University of Melbourne, Parkville, Vic. 3050, Australia AbstractöA major characteristic feature of Alzheimer's disease is the formation of compact, extracellular deposits of L-amyloid (senile plaques). These deposits are surrounded by reactive astrocytes, microglia and dystrophic neurites. Mutations in three genes have been implicated in early-onset familial Alzheimer's disease. However, in£ammatory changes and astrogliosis are also believed to play a role in Alzheimer's pathology. What is unclear is the extent to which these factors initiate or contribute to the disease progression. Previous rat studies demonstrated that heterotopic transplantation of foetal cortical tissue onto the midbrain of neonatal hosts resulted in sustained glial reactivity for many months. Similar changes were not seen in cortex-to-cortex grafts. Using this model of chronic cortical gliosis, we have now measured reactive changes in the levels of the key Alzheimer's disease proteins, namely the amyloid precursor protein, apolipoprotein E and presenilin-1. These changes were visualised immunohistochemically and were quanti¢ed by western blot analysis. We report here that chronic cortical gliosis in the rat results in a sustained increase in the levels of apolipoprotein E and total amyloid precursor protein. Reactive astrocytes in heterotopic cortical grafts were immuno- positive for both of these proteins. Using a panel of amyloid precursor protein antibodies we demonstrate that chronic reactive gliosis is associated with alternative cleavage of the peptide. No signi¢cant changes in apolipoprotein E or amyloid precursor protein expression were seen in non-gliotic cortex-to-cortex transplants. Compared to host cortex, the levels of both N-terminal and C-terminal fragments of presenilin-1 were signi¢cantly lower in gliotic heterotopic grafts. The changes described here largely mirror those seen in the cerebral cortex of humans with Alzheimer's disease and are consistent with the proposal that astrogliosis may be an important factor in the pathogenesis of this disease. ß 2001 IBRO. Published by Elsevier Science Ltd. All rights reserved. Key words: Alzheimer's disease, astrocytes, amyloid, cerebral cortex, in£ammation, foetal transplants. Alzheimer's disease (AD) is characterised neuropatholog- ically by the presence of insoluble cerebral amyloid deposits found intracellularly and extracellularly. The major protein component of the extracellular deposits is a 4-kDa peptide termed L-amyloid (AL) (Glenner and Wong, 1984; Masters et al., 1985), which is a pro- teolytic product of the amyloid precursor protein (APP). Although the aetiology of AD in most cases is unclear, mutations in the presenilin-1 (PS1), presenilin-2 (PS2), and APP genes have been demonstrated to account for 50% of early-onset familial AD cases (Goate et al., 1991; Rogaev et al., 1995; Sherrington et al., 1995; Kwok et al., 1997). Mutations in these three AD genes are all associated with an increased production of AL (Lendon et al., 1997; Storey and Cappai, 1999; Emilien et al., 2000). In the more common sporadic form of AD, while no pathogenic mutations in any genes have yet been identi¢ed, the frequency of the O4 allele of the apo- lipoprotein E (apoE) gene is considered to be the major genetic risk factor for this devastating disease (Corder et al., 1993; Martins et al., 1995). The brains of AD patients who are homozygous for the O4 allele exhibit an increased AL load compared to AD cases who lack an O4 allele (Rebeck et al., 1993). This ¢nding suggests that the apoE4 protein alters AL metabolism and this e¡ect may be exerted by its action on the APP and PS proteins that are both integral to the production of AL. Indeed, recent ¢ndings demonstrate that APP and PS1 557 *Corresponding author. Tel. : +61-8-9346-6656 ; fax : +61-8-9346- 6666. E-mail address : rmartins@cyllene.uwa.edu.au (R. N. Martins). Abbreviations : AL, L-amyloid; ABC, avidin^biotin^peroxidase complex ; AD, Alzheimer's disease ; apoE, apolipoprotein E; APP, amyloid precursor protein ; BSA, bovine serum albumin ; CSPGs, chondroitin sulphate proteoglycans ; DPBS, Dulbecco's `A' phosphate-bu¡ered saline ; ECL, enhanced chemilumines- cence ; E, embryonic day; FITC, £uorescein isothiocyanate ; GFAP, glial ¢brillary acidic protein; HRP, horseradish peroxi- dase ; IL, interleukin ; KPI, Kunitz protease inhibitor ; NSAIDs, non-steroidal anti-in£ammatory drugs ; PAGE, polyacrylamide gel electrophoresis ; PS1/2, presenilin-1 or 2 ; SDS, sodium dode- cylsulphate; TBS, Tris-bu¡ered saline; TBST, TBS with Tween 20 ; TRITC, tetramethyl rhodamine isothiocyanate. NSC 5132 26-9-01 Cyaan Magenta Geel Zwart www.elsevier.com/locate/neuroscience Neuroscience Vol. 106, No. 3, pp. 557^569, 2001 ß 2001 IBRO. Published by Elsevier Science Ltd Printed in Great Britain. All rights reserved PII:S0306-4522(01)00289-5 0306-4522 / 01 $20.00+0.00