This model also appears to be supported by the fact that the 180 different PSEN1 mutations are scattered selleck inhibitor over the entire molecule without any apparent hot spots, which is most compatible with the loss-of-function hypothesis. A small number of knock-in mouse strains for FAD PSEN1 mutations have been created, in which the mutant alleles are expressed under control of the endogenous mouse PSEN1 promoter [66-69,84]. These studies have either not provided evidence for substantially diminished ??-secretase activity (PSEN1-P264L, PSEN1-R278I) [66,68] or have produced inconclusive results (PSEN1-I213T) [67,84]. The PSEN1-R278I mutation has been particularly well studied in knock-in mice [68] (Table ?(Table2).2). Homozygous knock-in mice for this mutation (R278I/R278I) were embryonic lethal and displayed a phenotype similar to NOTCH knockout mice.

In the brain of these mice, accumulation of APP and N-cadherin CTFs was observed, and AICD and NICD fragments were undetectable. This was confirmed in kinetic in vitro studies using solubilized membrane preparations from heterozygous (WT/R278I) or homozygous knock-in mice, which showed reduced A?? and AICD generation from recombinant APP-CTF substrates in a gene-dose-dependent manner. Taken together, these findings indicate a substantial loss of ??- secretase activity of the mutant allele [68]. The decrease in enzymatic activity of the PSEN1-R278I mutant appeared to be particularly severe as other PSEN1 mutations have not caused embryonic lethality in homozygous knock-in mice or were able to rescue the phenotype in PSEN1-/- knockout mice [66,67,69,85,86].

Importantly, no developmental defects were observed in heterozygous knock-in mice, and the brain morphology of 3- to 24-month-old mice was indistinguishable from that of WT mice. In brain tissue from heterozygous knock-in mice, no changes in the Carfilzomib steady-state levels of APP and N-cadherin CTFs, AICD and NICD were observed, indicating that the ??-secretase-mediated release of intracellular domains is not affected by heterozygous expression of the PSEN1-R278I mutation [68]. A slight decrease in endogenous mouse A??40 was detected when brain tissue of 3-monthold heterozygous mice was extracted in guanidine-HCL, but not when these mice were crossed to APP-transgenic mice. Very similar results have been reported for PSEN1-M146V knock-in mice [69].

Interestingly, in APP/PSEN1-R278I double transgenic mice, increased brain levels of A??43 were detected that correlated with enhanced amyloid pathology and cognitive deficits, and A??43 appeared to induce the formation of Thioflavin T-positive aggregates in vitro even more efficiently than A??42 [68]. This suggests that A??43 might be an overlooked A?? species that contributes though to the formation of neurotoxic A?? oligomers and plaque pathology.