Transthyretin protects Alzheimer’s mice from the
behavioral and biochemical effects of A? toxicity
Joel N. Buxbaum*†, Zhengyi Ye*, Nata `lia Reixach*, Linsey Friske*, Coree Levy‡, Pritam Das§, Todd Golde§,
Eliezer Masliah¶, Amanda R. Roberts‡, and Tamas Bartfai‡
*Division of Rheumatology Research, W. M. Keck Autoimmune Disease Center, and Department of Molecular and Experimental Medicine, and‡Department
of Molecular and Integrative Neurosciences, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037;§Department of
Neurosciences, Mayo Clinic College of Medicine, Birdsall 210, 4500 San Pablo Road, Jacksonville, FL 32224; and¶Department of Pathology, University of
California at San Diego, La Jolla, CA 92093
Communicated by Ernest Beutler, The Scripps Research Institute, La Jolla, CA, December 24, 2007 (received for review October 3, 2007)
Cells that have evolved to produce large quantities of secreted
proteins to serve the integrated functions of complex multicellular
organisms are equipped to compensate for protein misfolding.
Hepatocytes and plasma cells have well developed chaperone and
proteasome systems to ensure that secreted proteins transit the
cell efficiently. The number of neurodegenerative disorders asso-
ciated with protein misfolding suggests that neurons are particu-
larly sensitive to the pathogenic effects of aggregates of misfolded
molecules because those systems are less well developed in this
lineage. Aggregates of the amyloidogenic (A?1–42) peptide play a
major role in the pathogenesis of Alzheimer’s disease (AD), al-
though the precise mechanism is unclear. In genetic studies exam-
ining protein–protein interactions that could constitute native
mechanisms of neuroprotection in vivo, overexpression of a WT
human transthyretin (TTR) transgene was ameliorative in the
APP23 transgenic murine model of human AD. Targeted silencing
of the endogenous TTR gene accelerated the development of the
neuropathologic phenotype. Intraneuronal TTR was seen in the
brains of normal humans and mice and in AD patients and APP23
mice. The APP23 brains showed colocalization of extracellular TTR
with A? in plaques. Using surface plasmon resonance we obtained
in vitro evidence of direct protein–protein interaction between TTR
and A? aggregates. These findings suggest that TTR is protective
because of its capacity to bind toxic or pretoxic A? aggregates in
both the intracellular and extracellular environment in a chaper-
one-like manner. The interaction may represent a unique normal
host defense mechanism, enhancement of which could be thera-
protein interaction ? protein misfolding ? amyloidosis ? dementia
reports of physical interaction between amyloidogenic (A?)
peptides and TTR proteins in vitro, prevention of A? aggrega-
tion in Caenorhabditis elegans transgenic for both mutant A? and
AD models, immunohistochemically detectable TTR in the
vicinity of A? plaques in A? transgenic mice, and more aggres-
sive histologic disease in such mice after local treatment with
anti-TTR antibody (1–5). However, none of those studies dem-
onstrated functional effects of the putative TTR–A? interaction.
We performed genetic experiments designed to determine
whether TTR has an effect on the development of the neuro-
pathologic and behavioral phenotypes in a well characterized
murine model of human AD.
relationship between Alzheimer’s disease (AD) and tran-
sthyretin (TTR) has been hypothesized on the basis of
APP23 mice, carrying the Swedish autosomal dominant AD
mutation and displaying the neuropathologic (Congophilic
plaques, gliosis, neuronal death, Congophilic angiopathy) and
behavioral (defined cognitive deficits) features associated with
human AD, were mated with mice overexpressing WT human
TTR (hTTR) and animals in which both copies of the endog-
enous TTR gene had been silenced by targeted disruption (6–8).
The transgenics carried ?90 copies of the human gene with
serum concentrations of hTTR between 1 and 3 mg/ml and
cerebrospinal fluid concentrations between 0.007 and 0.019
Results of Barnes maze testing (to assess cognitive function and
spatial learning) of 15-month-old APP23, APP23 overexpressing
APP23/hTTR and control mice the number of errors decreased
across blocks [F (2,64) ? 5.2, P ? 0.01], demonstrating the effect
of training. There was a significant effect of group (genotype) on
the number of errors across the three blocks [(F(2,32) ? 4.1, P ?
0.05] caused by group differences in blocks 2 [F(2,32) ? 3.0, P ?
in these blocks relative to control mice (P ? 0.05). APP23/ hTTR
mice were not significantly different from age-matched WT con-
trols, whereas the performance of APP23/hTTR animals was
superior to that of APP23 mice lacking the hTTR transgene (P ?
0.05). The percentage of trials in each block in which mice used a
[F(2,64) ? 17.7, P ? 0.001], but no significant group by block
interaction. Spatial strategy utilization increased across blocks.
Both control and APP23/ hTTR mice used this strategy more than
APP23 mice in block 3 (P ? 0.05). There was a significant effect of
APP23 on numbers of errors made and strategy utilization in the
15-month-old animals. The presence of hTTR resulted in virtually
normal performance in both measures, indicating amelioration of
the APP23 behavioral phenotype.
There were no significant group differences in anxiety-like
behavior, rearing, or locomotor activity. There were trends for the
older APP23 mice to show increased anxiety-like behavior and
decreased activity (horizontal and vertical). Visual cliff analysis
are consistent with previous analyses of APP23 mice (9).
To determine whether absence of the endogenous murine
TTR (mTTR) gene would have an effect on the development of
disease in APP23 mice we carried out behavioral studies on
5.5-month-old mice expressing the APP23 gene in the presence
(APP23) or absence of mTTR (APP23/mTTR?/?) and in mice
gene (mTTR?/?). The number of errors is shown in Fig. 1c and
the percent spatial strategy is shown in Fig. 1d. Because we had
behavior of APP23 mice, we performed three general analyses.
Author contributions: J.N.B. and T.B. designed research; Z.Y., L.F., C.L., P.D., T.G., E.M., and
A.R.R. performed research; N.R. contributed new reagents/analytic tools; J.N.B., Z.Y., L.F.,
T.G., E.M., A.R.R., and T.B. analyzed data; and J.N.B. wrote the paper.
The authors declare no conflict of interest.
†To whom correspondence should be addressed. E-mail: email@example.com.
© 2008 by The National Academy of Sciences of the USA
February 19, 2008 ?
vol. 105 ?
no. 7 ?
min) (31). The concentration of protein was determined by UV absorption at
with aqueous NaOH (pH 10.5) and used immediately. We noted that approx-
imately half of the A? sample was retained on the 10-kDa filter. After
pretreatment, the final peptide solution was seed-free by atomic force mi-
croscopy analysis. A? fibrils were formed by incubating the monomer in PBS
buffer for 7 days at 37°C with constant shaking.
hTTR and mTTR were overexpressed in Escherichia coli and purified as
described (17, 30).
ACKNOWLEDGMENTS. This work was supported by National Institute on
Aging Grants R01 AG15916 (to J.N.B.) and R01 AG18440 (to E.M.), the W. M.
Keck Foundation (J.N.B.), The Skaggs Foundation (Z.Y.), the Fidelity Founda-
tion (J.N.B.), and The Stein Fund.
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www.pnas.org?cgi?doi?10.1073?pnas.0712197105Buxbaum et al.