Our study of in vivo
CNS Aβ isoform kinetics in participants with PSEN
mutations confirms the hypothesis that the production rate of Aβ42 is increased relative to Aβ40 in humans with PSEN
mutations associated with ADAD, and also revealed increased exchange of soluble Aβ42 and increased irreversible loss of soluble Aβ42 associated with the presence of fibrillar amyloid plaques. The finding of increased Aβ42 production was predicted by some in vitro
) and evidence of altered γ-secretase function in carriers of PSEN
1 or PSEN
2 ADAD mutations (24
). Here, we directly quantify increased Aβ42 production in vivo
in humans due to mutations that cause ADAD, and confirm that such mutations increase CNS Aβ42 production rates in vivo
. The ratio of Aβ42 to Aβ40 production rates was greater than 15% for almost all mutation carriers (average 17.4%), whereas non-carriers were less than 15% (average 14%), suggesting tight physiological control that is disrupted in the presence of a mutation. There was a ~25% relative increase in Aβ42 to Aβ40 production rate in mutation carriers, which correlates with the 40% decrease in Aβ production due to a mutation that is protective for the development of AD (7
). Changes in the Aβ42 to Aβ40 production rate ratio are likely to have a large impact on the rate of Aβ42 deposition into plaques because in vitro
studies have demonstrated that even relatively small changes in the Aβ42:Aβ40 ratio significantly impact aggregation kinetics and morphology (15
) that could promote plaque formation. The Aβ42:Aβ40 ratio and the total amount of Aβ42 have been hypothesized to affect the age of onset of ADAD(25
). Some PSEN
mutations in cultured cells in vitro
have been reported to decrease Aβ40 production or loss of overall Aβ production (16
), however, our in vivo
SILK studies did not confirm decreased Aβ40 or total Aβ production.
Our finding of a faster irreversible FTR of soluble Aβ42 relative to Aβ38 or Aβ40 in the presence of PSEN
mutations or plaques was not anticipated. This faster relative FTR was a primary factor causing the Aβ42 isotopic enrichment curve to peak earlier than Aβ38 or Aβ40 in the presence of amyloidosis (). We cannot determine the physiological mechanisms that underlie this irreversible FTR, but it includes all losses of soluble Aβ peptides including transport into CSF or plasma, proteolytic degradation, and deposition into amyloid plaques. The rate constant for transport to CSF is likely to be similar for all Aβ isoforms since the transport of soluble Aβ isoforms to CSF is driven by fluid flow (26
). Therefore, the faster relative FTR of Aβ42 in the presence of plaques implies a greater loss of Aβ42 from the system to fates other than CSF. Although we cannot identify the specific fates of peptides permanently lost from the system, our observations are consistent with increased deposition of Aβ42 into amyloid plaques (27
) relative to Aβ38 and Aβ40, which subsequently causes increased Aβ42 in brain plaques(29
) as shown in Video 1
, and a reduction in the amount of Aβ42 recovered in CSF (30
). Our finding of increased irreversible FTR of Aβ42 in subjects with PSEN
mutations but no amyloid load suggests increased Aβ42 deposition may be occurring in these subjects, and SILK studies of Aβ42 turnover kinetics may provide a sensitive window into altered processes before amyloidosis is detectable by PET PIB.
Another unanticipated finding was a reversible exchange process whereby a portion of newly synthesized (i.e. isotopically labeled) peptides exchange with pre-existing unlabeled Aβ. The structure of the pre-existing unlabeled Aβ structures cannot be determined from this tracer study, but they could potentially consist of oligomers or aggregates (31
) or may reflect reversible binding of Aβ42 to the surface of amyloid plaques before structural alterations cause permanent assimilation into plaques (33
We previously reported that late onset Alzheimer’s disease was associated with a slower clearance rate for Aβ40 and Aβ42 based on a monoexponential slope applied to the terminal phase (24–36 h) of the SILK enrichment time course (18
). The present study utilized a compartmental model to fit the SILK enrichment time courses over the full experimental period (0–36 h) in a manner consistent with known physiological processes (35
). This compartmental model confirms the previous report that the FTR is slower for Aβ40 with increased insoluble amyloid plaque deposition (). The exchange process described by our compartmental model was minimal for Aβ38 and Aβ40 in the presence or absence of amyloid plaques (), and thus the terminal tail approximates a monoexponential shape for these isoforms. The observation that the irreversible FTRs of Aβ38 and Aβ40 are negatively correlated with PIB MCBP score () suggests that the presence of plaques substantially alters physiological processes (e.g. fluid flow perfusion through brain tissue) that retards the transfer of soluble Aβ peptide from the brain to CSF. An outcome of this decreased transport to the CSF may be loss of the normal physiological diurnal pattern of CSF Aβ concentrations (36
The present study also confirms that the terminal slope of the Aβ42 enrichment curve is slower in the presence of plaques. We observed that Aβ42 irreversible FTR was increased in the presence of PSEN
mutations independent of plaque load. The SILK tracer technique thus reveals abnormalities in Aβ42 before plaques are detectable by PET PIB. Indeed, 3 of the 4 mutation carriers who did not have detectable amyloid plaques by PET PIB had features of Aβ42 kinetics that were more similar to mutation carriers with amyloid plaques than to non-carriers, including Aβ42:Aβ40 FTR ratio > 1.1, and evidence of Aβ42 exchange (Fig. S2
). These features strongly suggest that plaque development or some form of Aβ aggregation has been initiated in these individuals, but fibrillar plaque load has not reached the threshold of PET PIB detection. In contrast, only one mutation carrier who was PET PIB negative had no evidence of Aβ42 exchange. This participant also had the highest Aβ42:Aβ40 CSF concentration ratio (0.215; Fig. S4
) of all participants in our study (compare to Figs. S2 and S3
; this ratio was 27% higher than the next highest concentration ratio observed in individual participants, 0.169), demonstrating that elevated Aβ42:Aβ40 CSF concentration ratios are only observed in ADAD persons in the absence of amyloid plaques. As amyloid deposition in the caudate may be one of the earliest deposition regions, we evaluated caudate PIB as a sensitive measure of amyloid deposition. The caudate PIB and MCBP PIB were highly correlated with each other with a Pearson correlation coefficient of 0.94, p=5×10−11
; no kinetic parameters changed significance with amyloidosis as measured by caudate PIB.
Our findings offer a context for the recent report that CSF Aβ42 concentration is elevated in ADAD individuals decades prior to predicted age for onset of clinical symptoms of dementia, and then drops below normal as plaques develop (2
) (see for schematic of proposed process). Our finding of increased Aβ42 production in mutation carriers regardless of the amount of amyloid deposition indicates that increased Aβ42 production precedes amyloid deposition, and likely occurs decades before the onset of dementia and is possibly present throughout life. The long delay in the emergence of plaque deposits of Aβ42 even in the presence of overproduction of Aβ42 suggests the presence of an initial slow process (e.g. initial Aβ nucleation event in which monomeric Aβ forms small aggregates) followed by a growth phase of Aβ polymerization (39
). We therefore hypothesize that Aβ42 production rate (solid gold line, ) remains above “normal” (solid black line) throughout life due to their PSEN
genotype. ADAD individuals have an increased irreversible loss of soluble Aβ42 (FTR solid red line, ) that tracks or precedes PET-detectable amyloid deposition, consistent with a faster removal of soluble Aβ42 as it is deposited into plaques. This results in a decreased recovery of Aβ42 in the CSF, accounting for the decrease in CSF Aβ42 concentration (solid blue line, ) as plaque development proceeds (solid green line, ). The physiological identity of the pool(s) in exchange with newly labeled soluble Aβ42 identified by SILK (dotted black lines, ) cannot be determined in this study. If exchange occurs with micelles or oligomers, exchange may precede PET-detectable amyloid deposition (dotted line 1, ). If exchange is with plaque surfaces, it may track or even lag behind amyloid deposition (dotted lines 2&3, ). Future studies with greater numbers of participants or longitudinal designs will be required to resolve this question.
Potential scheme for the time course of plaque deposition in ADAD.
Although ADAD represents less than one percent of all AD cases, it has been informative for elucidating the pathophysiology of AD, and can manifest as late-onset AD indistinguishable from the more idiopathic forms (40
). The findings of this study further support the amyloid hypothesis and provide quantitative estimates of life-long increases in Aβ42 production that may cause AD in humans. Further, these results indicate profound changes in Aβ kinetics in the presence of amyloid plaques. With increased understanding of the pathogenic causes of AD and the quantitative changes associated with AD pathology, it is hoped that better tests and directed therapeutics can be developed in the future.