Accumulation of Amyloid  Protein in Transgenic Mice M. SHOJI,* 1 T. KAWARABAYASHI,* M. SATO,† A. SASAKI,‡ E. MATSUBARA,* Y. IGETA,* M. KANAI,* Y. TOMIDOKORO,* M. SHIZUKA,* K. ISHIGURO,* Y. HARIGAYA,* K. OKAMOTO,* AND S. HIRAI§ *Department of Neurology and ‡Pathology, Gunma University School of Medicine, 3-39-15 Showamachi, Maebashi, Gunma 371, Japan †Molecular Medicine Research Center, The Institute of Medical Sciences, Tokai University, Bohseidai, Isehara, Kanagawa 259-11, Japan §Tokyo Metropolitan Neurological Hospital, 2-6-1 Musashidai, Futyu, Tokyo 183, Japan SHOJI, M., T. KAWARABAYASHI, M. SATO, A. SASAKI, E. MATSUBARA, Y. IGETA, M. KANAI, Y. TOMIDOKORO, M. SHIZUKA, K. ISHIGURO, Y. HARIGAYA, K. OKAMOTO, AND S. HIRAI. Accumulation of amyloid  protein in transgenic mice. NEUROBIOL AGING 19(1S) S59 –S63, 1998.—Carboxyl-terminal fragments of  amyloid precursor protein (APP) were expressed in mice under the transcriptional control of an ubiquitous promoter system, based upon a chicken -actin (A) promoter combined with cytomegalovirus (CMV) enhancer to obtain a systemic overproduction of amyloid  protein (A). Three transgene constructs were designed to encode signal peptide and carboxyl-terminal 99 amino acid residues to APP (NOR-), methionine and C-terminal 103 amino acid residues of APP (NOR-), and methionine and C-terminal 103 amino acid residues with KM-NL substitution of APP (NL-). Although the transcriptional mRNA level and post-translational protein level from transgenes showed the same expression pattern, both the expression of A and distribution of A deposits were completely different among these strains. In NOR- mice, considerable amounts of A were detected in plasma and A deposits were observed in the pancreas. Brain A deposits and small amounts of plasma A were recognized in NL-. These findings indicate that tissue specific processing and transgene constructs are major facters to determine the distribution of A deposits. © 1998 Elsevier Science Inc. A APP Transgenic mice Tissue specific processing Amyloid deposits ALZHEIMER’S disease (AD), one of the most devastating brain diseases, is a medical, sociological, and economic problem in modern society. About 600,000 AD patients and 60 AD families are present in Japan. The progress of AD research in the last decade identified two major accumulated materials, amyloid  protein (A) and hyperphosphorylated tau. The deposition of A, a 4-kD peptide (7,18) derived from the A precursor (APP) of 120 kD (15), is a specific, early event in the development of AD. Then after several decades, neurofibrillary tangles and dementia appear. Genetic studies have revealed some cosegregated genes with familial AD. They are APP (8), presenilin 1 (21), 2 (17), and a major risk factor gene ApoE4 (3). Recent studies have suggested the meaning of mutation of APP and presenilins are overproduc- tion of A1-42 (1,2,4,23). Based on these findings, A1-42 is considered to be a common initiating factor of AD (9,14,20). This is a reason to make AD model mice expressing considerable amounts of A to analyze how A accumulates and progresses, and finally how to prevent this pathological development (6,11). Because the native APP is ubiquitously expressed, and the cultured cells transfected with carboxyl-terminal fragment (CTF) of APP (5) release A effectively (12,22), we generated trans- genic mouse lines overexpressing CTF under the transcriptional control of an ubiquitous promoter system, based upon a chicken -actin (A) promoter combined with cytomegalovirus (CMV) enhancer to obtain a possible overproduction of A (16). Three transgene constructs were designed to code signal peptide and C-terminal 99 amino acid residues of APP (NOR-) (16), methionine plus C-terminal 103 amino acid residues of APP (NOR-), and methionine plus C-terminal 103 amino acid residues with KM-NL substitution (19) of APP (NL-) for analyzing A production and accumulation from expressed pro- teins in vivo. METHODS The NOR- gene construct was inserted into the EcoRI site of the third exon of the rabbit -globin gene in pBsCAG-2. The transgene consisted of a 2.35-kb SalI-BamHI fragment contain- ing A, the first intron of the chicken -actin gene, NOR gene, and a part of the rabbit -globin gene (16). The NOR- gene encoding methionine + C-terminal 103 amino acid residues of APP and NL- gene encoding methionine + C-terminal 103 amino acid residues with KM-NL substitution at APP595/596 were generated by using PCR from cDNA of APP695 with primer sets of 5'-GGTCTAGAGATGGAAGTGAAGATG-3',5'- GGTCTAGAGATGGAAGTGAATCTGGAT-3', and 5'- GGAGATCTCGATCAAGACGTA-3'. These gene constructs were inserted into the XbaI site of the third exon of the rabbit -globin gene in pBsCAG-2. Approximately 2000 copies of the 1 To whom correspondence should be addressed. Neurobiology of Aging, Vol. 19, No. 1S, pp. S59 –S63, 1998 Copyright © 1998 Elsevier Science Inc. Printed in the USA. All rights reserved 0197-4580/98 $19.00 + .00 PII:S0197-4580(98)00043-8 S59