Abnormal serine phosphorylation of insulin receptor substrate 1 is associated with tau pathology in Alzheimer's disease and tauopathies

Abstract

Neuronal insulin signaling abnormalities have been associated with Alzheimer's disease (AD). However, the specificity of this association and its underlying mechanisms have been unclear. This study investigated the expression of abnormal serine phosphorylation of insulin receptor substrate 1 (IRS1) in 157 human brain autopsy cases that included AD, tauopathies, α-synucleinopathies, TDP-43 proteinopathies, and normal aging. IRS1-pS(616), IRS1-pS(312) and downstream target Akt-pS(473) measures were most elevated in AD but were also significantly increased in the tauopathies: Pick's disease, corticobasal degeneration and progressive supranuclear palsy. Double immunofluorescence labeling showed frequent co-expression of IRS1-pS(616) with pathologic tau in neurons and dystrophic neurites. To further investigate an association between tau and abnormal serine phosphorylation of IRS1, we examined the presence of abnormal IRS1-pS(616) expression in pathological tau-expressing transgenic mice and demonstrated that abnormal IRS1-pS(616) frequently co-localizes in tangle-bearing neurons. Conversely, we observed increased levels of hyperphosphorylated tau in the high-fat diet-fed mouse, a model of insulin resistance. These results provide confirmation and specificity that abnormal phosphorylation of IRS1 is a pathological feature of AD and other tauopathies, and provide support for an association between insulin resistance and abnormal tau as well as amyloid-β.

Fig. 1

Serine-phosphorylated IRS-1 (IRS1-pS⁶¹⁶) percent area measures in neurodegenerative diseases with: AD, tauopathies (including Pick's disease, corticobasal degeneration and progressive supranu-clear palsy), α-synucleinopathies (dementia with Lewy bodies, Parkinson disease, Parkinson dementia and multiple system atrophy) and TDP-43 proteinopathies (frontotemporal dementia, amyotrophic lateral sclerosis), and normal aging. As compared to the normal aging group, IRS1-pS⁶¹⁶ measures are most severely elevated in AD but were also significantly elevated in other tauopathies. Error bars represent standard error of the mean

Fig. 2

Regression plots of IRS1-pS⁶¹⁶ and PHF-Tau percent area of immunolabeled cells and neurites in AD and other tauopathies in (a) mid-frontal gyrus cortex, (b) hippocampal CA1/subiculum and (c) angular gyrus cortex in AD and other tauopathies. All p values were <0.0001

Fig. 3

Double immunofluorescence images of IRS1-pS⁶¹⁶ (red) and neurodegenerative disease lesion proteins (green) in AD (a–d), tauopathies (e–h), α-synucleinopathies (i, j) and TDP43 proteinopathies (k, l). Yellow indicates proteins’ co-localization. a 72-year-old female with AD double labeled for IRS1-pS⁶¹⁶ and amyloid-β shows pathological IRS1-pS⁶¹⁶ expression in neuronal cytoplasm and neu-rites interspersed among diffuse amyloid-β plaque deposits in mid-frontal cortex. In this and all images except (d), magnification is 400× and scale bar = 50 μm. b 58-year-old male with AD showing IRS1-pS⁶¹⁶ immunoreactive neurites enveloped in neuritic amyloid-β plaque and surrounding IRS1-pS⁶¹⁶ immunoreactive neurons in mid-frontal cortex; c Same case as in (a) double labeled for IRS1-pS⁶¹⁶ and abnormally phosphorylated tau (p-tau) in midfrontal cortex. Note the frequent co-localization (yellow) of perikaryal and neuritic IRS1-pS⁶¹⁶ and tau in neurofibrillary tangle-bearing neurons as well as p-tau-immunoreactive neurites; d High magnification (1,000×) image of midfrontal cortex from 68-year-old male with AD shows neurofibrillary tangle that is intensely double labeled for IRS1-pS⁶¹⁶ and p-tau neurofibrillary tangle. e 57-year-old male with PiD midfrontal cortex labeled for IRS1-pS⁶¹⁶ and p-tau shows double-labeled fibrillar perikaryon of neuron as well as extensive p-tau-immunoreactive neuritic pathology, with some double labeling for IRS1-pS⁶¹⁶. f 52-year-old male with CBD shows IRS1-pS⁶¹⁶ and p-tau double-labeled neurons. g 68-year-old female with PSP midfrontal cortex double labeled for pS-IRS1 and p-tau shows co-localization in several cells. Such abnormal IRS-1 pS⁶¹⁶ expression and p-tau pathology was relatively rare in cortical regions in our sample of PSP cases, prompting us to evaluate IRS-1 pS⁶¹⁶ in subcortical regions known to be especially vulnerable in PSP (see text for details). h Pons in 63-year-old female with PSP exhibits extensive IRS1-pS⁶¹⁶ and tau co-localization. i Midfrontal cortex in 70-year-old male with PDD immunolabeled for IRS1-pS⁶¹⁶ and α-synuclein. Intracellular Lewy inclusions did not co-localize with abnormal IRS1-pS⁶¹⁶. j Pons locus coeruleus in 81-year-old male with PD double labeled for IRS-1 pS⁶¹⁶ and α-synuclein shows Lewy pathology and no abnormal IRS1-pS⁶¹⁶ expression or double labeling. k Midfrontal cortex in 66-year-old female with FTD-TDP immunolabeled for IRS1-pS⁶¹⁶ and TDP-43 shows normal-appearing IRS1-pS⁶¹⁶ nuclear labeling amidst frequent cytoplasmic and neuritic TDP-43 lesions without co-localization. l Cervical spinal cord from 82-year-old female with ALS shows motor neuron with cytoplasmic TDP-43 surrounding normal IRS1-pS⁶¹⁶ labeled nucleus, without co-localization

Fig. 4

Abnormal IRS1-pS⁶¹⁶ expression in hippocampus tangle-bearing neurons of PS19 tau transgenic mice. Double immunofluorescence for tau (a–e) and IRS-1 pS⁶¹⁶ (a′–e′) and merged images (a″–e″) in 19-month-old PS19 human tau Tg mouse without pff inoculation (a), 3 PS19 human tau Tg mice (b–d) injected with syn thetic tau pffs at 3 months of age and surviving until 5 months (b, c) or 7 months (d) or WT control (e). Note frequent abnormal IRS1-pS⁶¹⁶ expression in many, but not all tangle-bearing neurons of the transgenic animals

Fig. 5

Phosphorylated tau (T231) in high-fat diet mice. Immunob-lots of prefrontal cortex from mice fed a normal control diet (NCD) or high-fat diet (HFD, 60 % kcal by fat) for 17 days. Total tau was slightly increased in the HFD mice. Average phosphorylated tau (normalized to total tau) was increased approximately 60 % in the HFD group compared to NCD mice