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We report a novel gene, designated Humanin (HN) cDNA, that suppresses neuronal cell death by K595N/M596L-APP (NL-APP), a mutant causing familial Alzheimer's disease (FAD), termed Swedish mutant. Transfection of neuronal cells with HN cDNA or treatment with the coding HN polypeptide abrogated cytotoxicity by NL-APP. HN suppressed neurotoxicity by Abeta1-43 in the absence of N2 supplement, but could not inhibit Abeta secretion from NL-APP. HN could also protect neuronal cells from death by NL-APP lacking the 41st and 42nd residues of the Abeta region. Therefore, HN suppressed neuronal cell death by NL-APP not through inhibition of Abeta42 secretion, but with two targets for its inhibitory action: (i) the intracellular toxic mechanism directly triggered by NL-APP and (ii) neurotoxicity by Abeta. HN will contribute to the development of curative therapy of AD, especially as a novel reagent that could mechanistically supplement Abeta-production inhibitors.

Mutations in the presenilin genes PS1 and PS2 cause the most common form of early-onset familial Alzheimer's disease. The influence of PS1 mutations on the generation of endogenous intracellular amyloid beta-protein (A beta) species was assessed using a highly sensitive immunoblotting technique with inducible mouse neuroblastoma (Neuro 2a) cell lines expressing the human wild-type (wt) or mutated PS1 (M146L or delta exon 10). The induction of mutated PS1 increased the intracellular levels of two distinct A beta species ending at residue 42 that were likely to be A beta1-42 and its N-terminally truncated variant(s) A beta x-42. The induction of mutated PS1 resulted in a higher level of intracellular A beta1-42 than of intracellular A beta x-42, whereas extracellular levels of A beta1-42 and A beta x-42 were increased proportionally. In addition, the intracellular generation of these A beta42 species in wt and mutated PS1-induced cells was completely blocked by brefeldin A, whereas it exhibited differential sensitivities to monensin: the increased accumulation of intracellular A beta x-42 versus inhibition of intracellular A beta1-42 generation. These data strongly suggest that A beta x-42 is generated in a proximal Golgi, whereas A beta1-42 is generated in a distal Golgi and/or a post-Golgi compartment. Thus, it appears that PS1 mutations enhance the degree of 42-specific gamma-secretase cleavage that occurs in the normal beta-amyloid precursor protein processing pathway (a) in the endoplasmic reticulum or the early Golgi apparatus prior to beta-secretase cleavage or (b) in the distinct sites where A beta x-42 and A beta1-42 are generated.

We have found that amorphous tau deposits in chloroquine myopathy (CM), a vacuolar myopathy induced by the administration of chloroquine, a well-known lysosomotropic agent. The dynamics of tau in CM and immunocytochemistry strongly suggest that the accumulation of tau is due to defective tau degradation in the lysosomal compartment in the muscle. This observation may offer a new view on the formation of paired helical filaments in Alzheimer's disease: this selective protein degradation pathway may be defective and result in intracellular accumulation of tau, thereby forming the unusual filaments.

Transferrin (Tf), an iron-transporting protein, has many variants, but C1 and C2 variants account for the majority of the population in all races. Since Tf is reported to be immunocytochemically detectable in senile plaques in Alzheimer's disease (AD), we have examined the Tf allele frequency among AD patients. The C2 allele frequency in late-onset AD patients is significantly higher than that in age-matched controls. Unexpectedly, the C2 allele frequency in AD patients homozygous for the ApoE epsilon 4 allele is markedly increased, i.e., it is twice as high as that in the remaining AD patients carrying zero or one copy of the epsilon 4 allele.

We have investigated the effect of genotypes of apolipoprotein E (ApoE) on the pathologies found in Alzheimer's disease (AD) and its related gene expression in 38 aged human brains obtained from consecutive autopsied cases. ApoE2/3, -3/3, -3/4, and -4/4 were typed in those aged brains, with ApoE3/3 being most prevalent. The AD pathologies were undetectable in ApoE2/3 brains, but were frequently observed in the other ApoE groups. In ApoE3/3 brains, 55%, 34%, and 24% of the cortical sections examined showed senile plaques (SPs), neurofibrillary tangles (NFTs), and cerebral amyloid angiopathy (CAA), respectively. In ApoE4/4 brains, the SP formation was significantly higher. The ApoE genotype neither affected ApoE, APP, or tau mRNA level, nor the differential expression of the latter two. These results suggest that ApoE4/4 accelerates and ApoE2/3 decelerates the development of the AD pathologies in the aged brain, but this is not through alterations of the APP and tau gene expression.

To learn whether or not the levels of beta-amyloid protein precursor (APP) and tau mRNAs are related to the formation of beta-amyloid and neurofibrillary tangles, we quantified these mRNA levels in three cortical regions of 38 aged human brains, which were examined immunocytochemically for beta-amyloid and tangles. Marked individual variabilities were noted in APP and tau mRNA levels among elderly individuals. The mean APP mRNA level was slightly reduced in the beta-amyloid plaque (+2) group, but not in the plaque (+) group, compared to the plaque (-) group. Some brains in the plaque (-) group showed increased APP expression, the extent of which was not seen in the plaque (+) or (+2) group. The differences in the mean tau mRNA levels were not statistically significant among the tangle (-), (+), and (+2) groups. These results show that beta-protein and tau deposition do not accompany increased expression of the APP and tau genes, respectively, and thus suggest that factors other than gene expression may be at work in the progression of beta-amyloid and/or tangle formation in the aged human brain.