Our findings provide the first examples of amyloid pathology formed by rodent A? peptides. Unlike other mammalian species, aged rodents have not been reported to develop A? deposits unless they express APP genes encoding human A? sequences [12, 38] or harbor modified APP genes in which the exon encoding the A? domain has been modified to encode human A? [10, 36]. The lack of amyloid pathology in aged rodents cannot be fully explained by the inherent abilities of mouse and human A? peptides to form fibrillar aggregates since both peptides show similar propensity to aggregate in vitro [11]. However, it is now well recognized that the processing of APP by its three secretases is heavily influenced by the sequence of the peptide and adjacent sequences in the APP holoprotein (for review see [8, 16]), with the processing by BACE1 having a major influence on whether A?1-40/42 or 11–40/42 is generated [6]. For wild-type moAPP, cleavage by murine BACE favors the production of the non-amyloidogenic A?11-40/42 peptides. Thus, whether murine A?1-40/42 is really incapable of fibrillogenesis had not rigorously been tested in vivo. Our study now provides the rigorous test by introducing moAPP transgenes harboring mutations linked to FAD that shift cleavage of APP to favor the production of moA?1-40/42. Our findings indicate the moA? is fully capable of forming both diffuse and compact amyloid plaques in vivo. For reasons that we have yet to elucidate, the morphology of the deposited amyloid in these models was heavily influenced by the mode of transgene expression and whether human PS1dE9 was co-expressed. Nevertheless, the data provide the first definitive proof that A? peptides of murine amino acid sequence can produce amyloid plaques that are morphologically similar to human amyloid deposits.
The present report also documents, for the first time, the frequency and severity of cerebellar amyloid deposits in the mice that co-express mutant APP and mutant PS1. In the initial iterations of bigenic APP/PS1dE9 mice, generated by crossing 2 independent lines of mice, cerebellar amyloid deposits were inconsistently observed, and present at a low frequency when observed (see Table 1). However, in the more commonly used bigenic APPswe/PS1dE9 mice, generated by co-injection of the transgenes [20, 24], the levels of cerebellar amyloid are considerably higher. The more often used APPswe/PS1dE9 mice Line 85 mice (Jax Strains 005864 and 004462) were created at the same time as a second line, designated line 57. Mice from line 57 were first described by Jankowsky et al. [20], whereas mice from Line 85, which initially bred poorly, were not described until later [24]. For enigmatic reasons, the Line 85 mice emerged as the more used line mice that is deposited in the Jackson Laboratories. We have routinely aged mice from Line 85 out to advanced ages and have never observed obvious gait abnormalities. However, mice of this line have been tested on the rotarod device and have been reported to show deficits [40]. Thus, the presence of amyloid pathology in the cerebellum of the APPswe/PS1dE9 mice may produce moderate deficits in motor performance.
Although cerebellar amyloid pathology is rare in sporadic AD, studies of Finnish patients with the PS1dE9 mutation have documented frequent amyloid deposits in the cerebellum [47]. This unusual pathology correlated to unusual symptoms of paraparesis in this pedigree [9]. In this same pedigree, pathologic descriptions of patients with the PS1dE9 mutation demonstrated unusual amyloid plaque pathology with structures termed cotton wool plaques along with abundant non-cored senile plaques in the cerebral cortex [9, 47]. Thus, the presence of cerebellar amyloid pathology in mice that co-express mutant APP and PS1dE9 may represent a partial reproduction of the human pathology. However, we also observed amyloid pathology in the pia surrounding the cerebellum of PrP.HuA?(GFP) mice, which do not express PS1dE9. Moreover, McGowen et al observed cerebellar amyloid in mice that express fusion proteins of Bri-A?42 via the MoPrP.Xho vector [30]. Thus, there may also be influences of the vector used to produce the transgenic expression on the distribution of amyloid deposition.
Presenilins are integral components of the ?-secretase complex, which is one of two primary enzymes involved in cleaving APP to produce A?40 and A?42 [for review see [8, 16]. In regard to the generation of A? peptide, the major consequence of AD-linked mutations in PS1 and PS2 on APP processing is to shift the processivity of ?-secretase cleavage such that more A?42 is produced relative to shorter A? peptides such as A?38 (for review see [8, 16]). This shift in abundance promotes the deposition of A?40 and 42 into senile plaques. Multiple laboratories have demonstrated that the PS1dE9 mutation was among those that produce the most robust increase in the production of A?42 [5, 41]. Additionally, when mutant human PS1dE9 is overexpressed, it competes for other cofactors of ?-secretase (nicastrin, Pen2, Aph1) causing a displacement of the endogenous PS1 from this complex [26, 44]. Thus, we can reasonably expect mice that co-express APPswe with PS1dE9 may produce a slightly different spectrum of A? peptides than mice in which ?-secretase contains only endogenous mouse PS1. This difference in the spectrum of peptides produced could modulate the location and architecture of deposited amyloid.
Both of our lines of mice depositing MoA? peptides displayed novel patterns of amyloid deposition and distinct plaque morphologies (Table 2). Notably, regardless of distribution or morphology, all types of deposits were found to include ubiquitin-immunoreactive profiles indicative of neuritic pathology. Collectively, we compare 3 lines of mice that use the MoPrP.Xho vectors to 3 lines that use CamKII-tTA + tetPrP.Xho vectors (Table 2). The PrP.HuA?/PS1 (Line 85) mice are representative of all lines of mice expressing MoHuAPPswe and PS1dE9, developing cored amyloid deposits in both the cortex and hippocampus that show a wide distribution in the parenchyma (Table 2). The PrP.MoA?/PS1 mice developed amyloid deposits in the meninges surrounding the cortex and cerebellum with additional deposits in the white-matter tracts. Since the identical vector (MoPrP.Xho) was used in generating these lines of mice, the difference in patterns of deposition cannot be easily explained by transgene expression patterns. Although we cannot rule out the possibility that the site of transgene integration modulates expression levels in some population of cells to produce these distinct patterns of deposition, the simplest explanation is that the amino acid sequence differences between human and mouse A?, in some manner, modulate the distribution of amyloid plaques.
Characteristics of HuA? and MoA? mouse models
P Parenchymal, M Meningeal, WM White matter tract distribution, C Cored, D Diffuse morphology
The strikingly distinct pattern of deposition and morphology of deposits in the tet.MoA?(GFP) mice is also remarkable. The distribution of the moA? deposits shifts to the parenchyma of the cortex, and the morphology is quite distinct (Table 2). The tet.MoA?(GFP) mice stand alone as the only one in which the amyloid deposits are primarily diffuse. However, we did observe that the PrP.HuA?(GFP) mice, which also lack co-expressed PS1dE9, displayed a higher level of diffuse amyloid, particularly in the cortex. From these direct comparisons, it would seem that the amino acid sequence of the A? peptides that are deposited may have some influence on the morphology of deposited amyloid, but the co-expression of human PS1dE9 may also have some influence.
The unexpected question is: Why do the PrP.MoA?/PS1 and tet.MoA?(GFP) mice differ in both distribution and morphology of deposition? The two vectors systems used will be expressed in different, but overlapping populations of neurons. MoPrP.Xho is expressed in all neurons and astrocytes [4, 27] whereas the CamKII-tTA is expressed only in forebrain neurons [29]. The final processed A? derivatives of the APP transgenes in these mice are expected to have identical sequences. It is possible that the co-expression of PS1dE9 changes the mixture of A?1-40/42 and other smaller A? peptide derivatives, such as A?38, relative to the population of A? derivatives produced by ?-secretase containing endogenous presenilin, and that these differences in relative levels of the different derivatives underlie our observations. However, these new questions that arise from our findings do not diminish the overall conclusions of our study, which demonstrates for the first time that murine A? peptides possess the capacity to form amyloid deposits in vivo.
The degree to which murine A? deposits may influence cognitive behavior in mice is a topic for future study, but the late onset and distribution of amyloid in the PrP.MoA?/PS1 mice is not indicative of a high probability of having a major impact on cognition. The tet.MoA?(GFP) mice may offer an opportunity to examine the effects of diffusely deposited A?42 on cognitive function, but again the late onset of deposition creates challenges in assessing cognitive behavior.