Amyloid precursor protein dosage normalization rescues neurogenesis and Alzheimer’s Disease phenotypes associated with Down Syndrome

Deepika Patel; Karen Rakowiecki; Orly Lazarov

doi:10.18103/mra.2026.0302

. Author manuscript; available in PMC: 2026 Sep 1.

Published in final edited form as: Med Res Arch. 2026 Jun;14(6):10.18103/mra.2026.0302. doi: 10.18103/mra.2026.0302

Amyloid precursor protein dosage normalization rescues neurogenesis and Alzheimer’s Disease phenotypes associated with Down Syndrome

Deepika Patel ^1,^#, Karen Rakowiecki ^1,^#, Orly Lazarov ^1,^*

PMCID: PMC13529314 NIHMSID: NIHMS2203834 PMID: 42677302

Abstract

Down Syndrome (DS) is the most abundant genetic form of mental retardation. It is caused by the triplication of partial or complete human chromosome 21 (HSA21). The molecular mechanisms of DS are not fully understood. Amyloid precursor protein (APP) resides on HSA21 and is triplicated in DS. While it is not thought to be a part of the “Down syndrome Critical Region”, APP plays a role in developmental and post-natal neurogenesis and synaptic plasticity, thus, its triplication may affect cortical development in DS. Further, mutations in APP cause familial Alzheimer’s disease (AD). However, whether APP overdose is sufficient or required for the development of Alzheimer’s disease in DS is not fully elucidated. Here, we addressed the role of APP overdose in neuronal development and AD pathology. Using DS patient-derived induced pluripotent stem cells in which one copy of APP was silenced using CRISPR-Cas9, we examined developmenta neurogenesis, AD-related pathology and the expression levels of genes on HSA21 that are implicated in DS, neurodegeneration and inflammation. Amyloid precursor protein triplication, in DS induced pluripotent stem cells, led to reduced stem cell proliferation, enhanced differentiation into neurons, increased amyloid pathology, and altered protein levels of selected genes on HSA21. Correction of APP gene dosage rescued the altered neurogenesis phenotype and reduced pathological amyloidogenic processing. These data highlight APP dosage as a key regulator of neuronal maturation and AD pathology in DS, suggesting therapeutic value in targeting APP expression to mitigate neurodevelopmental and neurodegenerative features of the disorder.

Keywords: Down Syndrome, Amyloid precursor protein, Neurogenesis, Alzheimer’s disease

Introduction

Down Syndrome (DS) is a genetic disorder that is caused by the triplication of human chromosome 21 (HSA21)¹. It is manifested by alterations in brain development, leading to brain dysfunction and intellectual disability^2,3. However, how triplication of HSA21 induces these alterations is not fully understood. Previous attempts to narrow down genes that are essential or sufficient for the development of DS yielded several versions of the “Down syndrome Critical Region”⁴. These studies remain largely inconclusive, in part because they depend on a limited number of epidemiological cases. Amyloid precursor protein (APP) resides on HSA21 and is triplicated in DS, but was not thought to be part of the Down syndrome Critical Region.

Amyloid precursor protein is a ubiquitously expressed, single-pass transmembrane protein⁵. A multitude of binding partners and downstream molecular pathways have been identified precluding a discrete role for APP in cellular functions^6–9. In the nervous system, APP has been implicated in development, neuromuscular junction function, synaptic plasticity, and neuroprotection^10–14. Notably, previous reports suggested a role of APP in neurogenesis^15–17, including the regulation of neural progenitor cell proliferation in the post-natal mouse brain^18,19 and in human progenitors²⁰. Moreover, limited studies exist attempting to isolate the effect of increased APP gene copy number on neuronal development in DS from additional genes located on HSA21. In one, transcriptome profiling of induced pluripotent stem cells-derived neuronal cultures at day 65 of differentiation shows that APP gene dosage by itself cannot explain the full spectrum of amyloid and tau pathology as manifested in AD²¹. In the second, siRNA mediated knockdown of APP reduced enlarged, rapidly migrating growth cones observed in DS fetal cells²².

Mutations in APP cause familial Alzheimer’s disease²³. A large proportion of DS patients develop Alzheimer’s disease (AD). Approximately 50% to 70% of DS patients experience dementia by the time they reach 60 to 70 years of age^24,25. Previous studies suggested that APP overdose underlies increased β-amyloid (Aβ) production, altered Aβ_42/40 in neurons and deposition of the pyroglutamate (E3)-containing amyloid aggregates, but conflicting evidence was reported concerning tau-linked AD phenotypes^26,27. Together, this suggests that further studies are warranted to determine whether APP overdose is sufficient or required for the induction of brain malformation or the development of Alzheimer’s disease (AD) in DS.

Here, we addressed the role of APP overdose in neuronal development in DS. Using Down Syndrome-derived induced pluripotent stem cells in which one APP allele was silenced using CRISPR/Cas9 [DS APP(+/+/−)], we examined neurogenesis and AD-related pathology. We observed that the correction of APP dose rescues impairments in proliferation of neural progenitor cells and in differentiation of neural precursors and neurons. In addition to amyloid processing, we show that APP overdose increased tau expression and the correction of APP rescued this increase. Notably, correction of APP dose rescued overdose of other genes on HSA21, suggesting that it acts upstream of these genes. Together, this study suggests that APP overdosage plays a role in altered neuronal development and in amyloidosis.

Materials and Methods

HUMAN-INDUCED PLURIPOTENT STEM CELLS CULTURE, NEURAL PROGENITOR CELLS DIFFERENTIATION AND NEURON MATURATION

The induced pluripotent stem cells (iPSCs) lines used in this study were obtained from repositories and collaborating laboratories. Control line Ctrl #1 (UWWC1-DS2U) and Down syndrome line DS #1 (UWWC1-DS1) were obtained from WiCell. The DS mock-corrected (DS MC) and APP gene-corrected (APP+/+/−) lines were graciously supplied by the Young-Pearse laboratory. The CRISPR targeting of iPSC lines was previously described in²⁷. Schematic representation of the experimental groups are depicted in Figure 1A.

Frozen vials from master stocks were thawed at 37°C and plated onto Matrigel hESC-Qualified Matrix-coated plates (Corning, 354277) in mTeSR Plus medium (STEMCELL Technologies, 100-0276), supplemented with 10 μM Y-27632 (ROCK inhibitor) during the first 24 hours post-thaw to enhance survival. The medium was refreshed daily. Individual colonies were mechanically dissected and passaged weekly. Unless otherwise noted, all reagents were purchased from STEMCELL Technologies.

Neural differentiation was performed following STEMCELL Technologies’ protocols, with key stages and validation illustrated in Figure 1B. To generate CNS-type neural progenitor cells (NPCs) from iPSCs, we employed the STEMdiff Neural Induction Medium with SMAD pathway inhibitors using an embryoid body (EB) formation approach. Briefly, confluent iPSCs were dissociated with Gentle Cell Dissociation Reagent and seeded at 3 x 10^6 cells per well into AggreWell 800 plates (preconditioned with Anti-Adherence Rinsing Solution), resulting in approximately 10,000 cells per microwell. The medium was partially replaced daily over five days. The EBs were subsequently collected and plated onto poly-L-ornithine (Millipore sigma, A-004-C)/laminin (Fischer scientific, 23-017-015)-coated 6-well plates to induce neural rosette formation. After 7 days, neural rosettes were isolated using STEMdiff Neural Rosette Selection Reagent and replated on poly-L-ornithine/laminin-coated plates for further NPCs expansion. The NPCs were either banked or used directly for differentiation and expanded up to 4 passages.

For differentiation into neural precursors, NPCs were cultured for 7 days in STEMdiff Neural Progenitor Medium, then dissociated with accutase and plated in STEMdiff Forebrain Neuron Differentiation Medium for an additional week. Neural precursors were matured using the STEMdiff Forebrain Neuron Maturation Kit. All culture steps were performed on tissue culture-treated plastic plates, with 24-well glass-bottom plates used for imaging and immunocytochemical analyses. This differentiation protocol reliably produces validated NPCs, precursor cells, and 10 weeks old forebrain neurons suitable for downstream analyses.

IMMUNOCYTOCHEMISTRY

For immunocytochemistry (ICC) on cultured cells, culture media was removed, and cells were washed twice with 1X PBS. The cells were then fixed in 4% paraformaldehyde prepared in 1X PBS for 15 minutes at room temperature (RT). Following fixation, cells were washed twice with 1X PBS and blocked in blocking solution consisting of 1X PBS with 0.3 M glycine, 0.2% Triton X-100, and 5% normal donkey serum for 60 minutes at RT.

For 5-ethynyl-2’-deoxyuridine (EdU) detection, cells were processed prior to the blocking step following the manufacturer’s instructions provided in the Click-iT EdU Cell Proliferation Kit (Invitrogen). Primary antibodies were diluted in blocking solution and cells were incubated for overnight at 4°C. Antibodies used in experiments are listed in Table S1. After primary antibody incubation, cells were washed three times with TBST (PBS with 0.1% Tween-20) for 5 minutes each and incubated with fluorescently labeled secondary antibodies diluted in TBST solution for 2h at RT. Cells were then washed three times with TBST, counterstained with DAPI, and mounted using ProLong Gold Antifade Mountant (Invitrogen). Images were acquired at 20x magnification using a Keyence BZ-X800 microscope.

WESTERN BLOTTING

Cells were washed three times with ice-cold 1X PBS and lysed on ice for 10 minutes using RIPA buffer supplemented with protease and phosphatase inhibitor cocktails (Thermo Fisher Scientific). Lysates were sonicated on ice at 20% power for three cycles of 15 seconds each, with 5-second rests between sonication. Following sonication, samples were centrifuged at 10,000 x g for 15 minutes at 4°C to pellet insoluble debris, and the clarified supernatant was collected. Protein concentrations were determined using the Pierce BCA Protein Assay Kit (Thermo Fisher Scientific).

For electrophoresis, equal amounts of protein were mixed with NuPAGE LDS Sample Buffer and NuPAGE Reducing Agent (Invitrogen), then denatured by boiling at 95°C for 5 minutes. Samples were resolved on Bolt Bis-Tris Plus SDS-PAGE gels (Thermo Fisher Scientific) using Bolt MES SDS Running Buffer. Proteins were transferred to nitrocellulose membranes using the iBlot 3 Dry Blotting System (Invitrogen). Membranes were blocked for 1 hour at room temperature in 5% non-fat dry milk prepared in Tris-buffered saline with 0.1% Tween-20 (TBST). Primary antibodies diluted in 5% milk/TBST were incubated with membranes overnight at 4°C. Antibodies used in experiments are listed in Table S1. After washing membranes three times for 15 minutes each in TBST, membranes were incubated with horseradish peroxidase-conjugated secondary antibodies diluted in 5% milk/TBST for 2 hours at room temperature. Following secondary antibody incubation, membranes were washed three times as before and developed using SuperSignal West Pico PLUS Chemiluminescent Substrate (Thermo Fisher Scientific). Blots were imaged with the Azure Biosystems 300q Image Western Blot Imaging System.

Densitometric quantification of band intensities was performed using Fiji. Protein expression levels were normalized to housekeeping controls such as GAPDH.

ENZYME-LINKED IMMUNOSORBENT ASSAY FOR AMYLOID-BETA IN NEURONAL CONDITIONED MEDIA

Conditioned media were collected from 10 weeks old neuronal cultures derived from iPSCs. For conditioned media samples, cells were rinsed three times with Dulbecco’s phosphate-buffered saline (DPBS) to remove residual media, then cultured in 1 mL of fresh media for 48 hours. Following incubation, conditioned media were harvested and snap-frozen in liquid nitrogen and stored until analysis. To quantify amyloid-beta isoforms, Human β Amyloid (1-40) Enzyme-Linked Immunosorbent Assay (ELISA) Kit Wako II (Fujifilm Irving, 298-64601) and Human β Amyloid (1-42) ELISA Kit Wako (Fujifilm Irving, 298-62401) were used according to the manufacturer’s protocols. Conditioned media samples were diluted 1:4 for Aβ40 measurement and 1:2 for Aβ42 measurement before loading. All ELISA plates were read on an microplate reader at the 450nm wavelength, and data were processed following kit guidelines. Resulting amyloid-beta concentrations were normalized to the total protein extracted from the corresponding wells.

STATISTICS

In all graphs, data is shown as mean ± SEM. Prism (Graphpad) was used for statistical analysis with tests indicated in figure legends. The following was used for p values: *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001.

Results

DOWN SYNDROME INDUCED PLURIPOTENT STEM CELLS-DERIVED NEURAL CELLS EXHIBIT REDUCED PROLIFERATION AND PREMATURE NEURONAL DIFFERENTIATION.

We first examined the course of neuronal differentiation using DS patient- derived iPSC (DS) and isogenic (Ctrl) lines (Figure 1A). The iPSC lines were differentiated into NPCs followed by neural precursor cells and forebrain neurons using neural induction method (Figure 1B). We assessed cell proliferation of NPCs using Click-It EdU assay. We observed a decrease in the proliferative capacity of DS NPCs compared to Ctrl, as indicated by a significantly reduced percentage of EdU+ cells in DS NPCs (Figure 1C–D). Further, the trajectory of neuronal differentiation progression was assessed by quantifying the expression of SOX2, DCX, and βIII-Tubulin in the cultures. As expected, immunofluorescence studies assessing SOX2 expression in NPCs, precursor and forebrain neurons revealed a decrease in SOX2+ cells across differentiation stages in both DS and Ctrl. However, DS lines showed a moderate but significant increase in SOX2+ cells compared to Ctrl at each differentiation stage. (Figure 1E–F). In addition, a significant increase in the number of DCX+ cells was observed at each stage of neurogenesis in DS compared to Ctrl (Figure 1G–H). Lastly, a significant increase in the number of β-III-tubulin+ neurons was observed in the DS lines compared to Ctrl (Figure 1I–J).

To further validate the neuronal differentiation phenotypes, we examined the expression levels of these proxies by Western blot analysis (Figure 2A). Results revealed significant alterations in the expression of neuronal proxies during the differentiation of DS iPSCs-derived neural cells compared to Ctrl. Specifically, and in agreement with the immunofluorescence analysis, we observed increased expression of SOX2 in NPCs and precursors in the DS lines compared to Ctrl (Figure 2A,B). In addition, we observed increased DCX (Figure 2A,C) and β-III-tubulin (Figure 2A,D) expression in NPCs, precursors and 10-week-old neurons in the DS lines. Finally, there was an increase in neurofilament heavy chain in the 10-week-old DS neurons (Figure 2A,E). Together, this is indicative of disruptive neurogenesis that includes prolonged stemness along with enhanced neuronal maturation. Collectively, these results indicate that DS iPSCs-derived neural cells display reduced proliferative output with altered neurogenic state progression reflected by persistently increased SOX2+ cells and elevated neuronal lineage markers, mirroring neurodevelopmental abnormalities observed in Down Syndrome^27,28.

Figure 2. — (A) Expression levels of SOX2, DCX (doublecortin), β-III-Tubulin, neurofilament heavy chain (NFH), and APP full-length (APP FL) in NPCs, PRE, and 10-week-old neurons (MN) derived from Control (Ctrl) and Down Syndrome (DS) iPSCs, as detected by Western blot analysis. (B, C, D, E, F) Quantification of SOX2, DCX, β-III-Tubulin, NFH, and APP FL expression normalized to GAPDH in NPCs, precursor cells (left) and neurons differentiated from control and DS groups (right) supports increased neuronal differentiation in the DS group (n=3 replicates). Data represented as mean ± SEM with individual data points shown. Data analyzed by unpaired t test (neurons), one-way ANOVA with Tukey’s multiple comparisons test (NPCs and precursor cells data), *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001.

ENHANCED AMYLOIDOGENIC AMYLOID PRECURSOR PROTEIN PROCESSING AND TAU ACCUMULATION IN DOWN SYNDROME INDUCED PLURIPOTENT STEM CELLS-DERIVED NEURONS.

To start to address the role of APP in this developmental trajectory, we first assessed the protein levels of FL-APP (full length APP) at differentiation stages of cortical neurogenesis. Gene dosage effect of HSA21 triplication was apparent at the precursor stage; FL-APP was upregulated in DS precursors and neurons compared to Ctrl counterparts (Figure 2A, F). Examination of FAD-linked signalling revealed elevated levels of BACE-1 (Figure 3B), which was associated with significantly higher levels of APP C-terminal fragments (APP CTFs, Figure 3C), suggesting enhanced BACE-1 activity in DS relative to Ctrl. Down syndrome neurons also exhibited increased total tau levels (Figure 3E), with a trend toward higher phosphorylated tau (AT8, Figure 3D). The ratio of phosphorylated to total tau (AT8/Tau5) did not differ significantly (Figure 3F), suggesting that the increase in p-tau is due to increased tau, may be due to enhanced neuronal differentiation in DS, rather than AD pathology. A marked increase in secretion of both Aβ₄₀ (Figure 3G) and Aβ₄₂ (Figure 3H) peptides was observed in conditioned media of 10-weeks old DS neurons. Notably, the Aβ₄₂/Aβ₄₀ ratio remained unchanged between groups, supporting an increase in Aβ due to enhanced BACE-1 activity (Figure 3I). Together, these results suggest that trisomy 21 drives a coordinated shift toward amyloidogenic APP processing in human DS neurons, even at an early maturation stage.

Figure 3. — (A) Increased expression of BACE-1, APP C-terminal fragments (CTFs), phosphorylated tau (AT8), total tau (Tau5), DYRK1A, BACE-2, SOD1, and S100β in protein lysate of 10-week-old neurons derived from DS iPSCs compared to Ctrl, as detected by Western blot analysis. (B-E, J-N) Quantification of BACE-1, total APP CTFs, AT8, Tau5, DYRK1A, BACE-2, SOD1, and S100β levels normalized to GAPDH in Ctrl and DS groups. (n = 3 replicates). (F) AT8/Tau5 ratio indicates no significant difference between DS and Ctrl groups. (G-H) Quantification of secreted Aβ₄₀ and Aβ₄₂ in DS neuronal cultures compared to controls using ELISA. (n = 3 replicates). (I) The Aβ₄₂/Aβ₄₀ ratio is comparable between genotypes. Data represented as mean ± SEM with individual data points shown. Data analysed by unpaired t test, *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001.

UPREGULATION OF CHROMOSOME 21 ENCODED PROTEINS IN DOWN SYNDROME NEURONS.

Next, we asked whether alterations in expression of additional genes on HSA21 would be apparent in our iPSC-derived DS neurons. Particularly, selected genes that are implicated in inflammation, neurodegeneration and neurogenesis, DYRK1A, BACE2, SOD1 and S100β. Quantitative analysis of protein expression in 10-week-old iPSCs-derived neurons revealed the upregulation of DYRK1A (Figure 3J), BACE-2 (Figure 3K) and SOD1 (Figure 3L) in the DS neurons compared to Ctrl. Notably, DS neurons showed a robust increase in S100β dimer levels (Figure 3M), while monomer levels remained unchanged between groups (Figure 3N). This suggests a shift in the oligomeric state of S100β in DS neurons and may indicate altered S100β signaling or stress responses without a change in total S100β production^30,31. Oligomeric S100β appeared as a prominent dimer band, consistent with formation of disulfide-linked S100β dimers that are preserved in the mildly reducing, oxidizing environment of DS neurons^32–34. Together, these findings highlight the pervasive impact of chromosome 21 trisomy on gene and protein expression profiles, implicating these molecular changes in disrupted cellular homeostasis and heightened stress responses in DS-derived neural populations.

NORMALIZATION OF AMYLOID PRECURSOR PROTEIN COPY NUMBER RESCUES NEUROGENESIS DEFICITS IN DOWN SYNDROME INDUCED PLURIPOTENT STEM CELLS-DERIVED NEURONS.

Having established the phenotypes of DS iPSC-derived neurogenesis and neurons, we examined the hypothesis that increased APP gene dosage contributes to deficits in neurogenesis and the development of AD pathology in DS. For that, we used a DS patient-derived iPSCs in which one APP allele was silenced by CRISPR/Cas9 (DS APP +/+/−), and a mock CRISPR-Cas9 DS control (DS MC) (Figure 4A). Driven by this hypothesis, we investigated whether the restoration of APP gene dosage in DS iPSCs would reverse neurogenesis deficits. Our results showed that DS APP+/+/− cells exhibited a marked increase in the percentage of proliferating NPCs, as shown by greater EdU incorporation compared to DS MC (Figure 4B–C). Immunofluorescence quantification further demonstrated restoration of SOX2+ cell number at all three stages of differentiation (Figure 4D–E) and normalization of the number of DCX+ cells at neuronal precursor and neuronal stages in DS APP+/+/− group (Figure 4F–G). Moreover, the number of β-III-tubulin+ neurons was normalized in DS APP+/+/− compared to DS MC (Figure 4H–I).

Figure 4. — (A) Schematic depicting the manipulation of DS iPSCs with either mock CRISPR editing (DS MC) or APP gene copy correction (DS APP+/+/−). (B) Representative immunofluorescence images showing EdU incorporation (red) and DAPI (blue) in NPCs from DS MC and DS APP+/+/− lines. Yellow arrowheads indicate EdU+ proliferating cells. Scale bar: 20μm. (C) Quantification of EdU+ cells as a percentage of total DAPI-labelled nuclei, suggesting significantly increased proliferation in DSAPP+/+/− NPCs compared to DS MC. (D, F, H) Representative images of SOX2 (magenta), DCX (magenta), β-III-Tubulin (red), and DAPI (blue) immunostaining in NPCs, precursor cells, and 10 weeks old neurons derived from DS MC and DS APP+/+/− iPSCs. Scale bars: 20μm. (E, G, I) Quantification of SOX2+, DCX+, and β-III-Tubulin+ cells (% of total DAPI) across differentiation stages. n=9 images per group. Data represented as mean ± SEM with individual points shown. Data analyzed by unpaired t-test (C), two-way ANOVA with Sidak’s multiple comparisons test (E, G, I), *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001.

To further validate these findings, we examined the expression levels of these proxies in protein lysates of these lines. Normalization of APP gene dosage in DS iPSCs-derived neural cultures validated the robust rescue of aberrant neurogenesis proxies (Figure 5A). The DS APP+/+/− NPCs, precursors and neurons exhibited markedly reduced levels of SOX2 (Figure 5A, B), DCX (Figure 5A, C), and β-III-tubulin (Figure 5A, D) compared to DS MC controls. Western blot analysis demonstrated no significant change in NFH protein levels in both the groups (Figure 5A, E). Collectively, these results indicate that restoring APP gene dosage corrects the detected proliferative phenotype and to some extent differentiation abnormalities in DS iPSCs-derived neural stem cells.

Figure 5. — (A) Expression levels of SOX2, DCX (doublecortin), β-III-Tubulin, neurofilament heavy chain (NFH), and APP full-length (APP FL) across NPCs, precursor cells, and 10 week-old-neurons (MN) derived from DS iPSCs with DS MC and DS APP+/+/− groups, as detected by Western blot. (B, C, D, E, F) Quantification of SOX2, DCX, β-III-Tubulin, NFH, and APP FL expression normalized to GAPDH in NPCs and precursor stages (left) and in neurons differentiated from APP protein levels in DS APP+/+/− and DS MC iPSC lines, (n = 3 replicates). Data represented as mean ± SEM with individual data points shown. Data analyzed by unpaired t- test (neurons), one-way ANOVA with Tukey’s multiple comparisons test (NPCs and precursor cells data), *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001.

RESCUE OF ALZHEIMER’S DISEASE MOLECULAR PHENOTYPES BY AMYLOID PRECURSOR PROTEIN DOSAGE NORMALIZATION IN DOWN SYNDROME NEURONS.

Next, we asked whether the normalization of APP copy number and dose, rescues the increase in APP processing and subsequent AD-linked pathology. We observed that FL-APP protein levels were reduced at all stages in DS APP+/+/− lines compared to DS MC (Figure 5A, F), confirming successful genetic correction. Furthermore, normalization of APP gene dosage in DS iPSCs-derived neurons profoundly attenuates alterations in AD-related signals. Specifically, we observed significantly reduced levels of BACE-1 in DS APP+/+/− neurons compared to DS MC (Figure 6A–B). Levels of APP-CTFs were comparable between the genotypes (Figure 6A, C). Both AT8 (Figure 6A, D) and Tau5 (Figure 6A, E) were markedly diminished in DS APP+/+/− neurons compared to DS MC, suggesting normalization of tau metabolism. The ratio of AT8/Tau5 remained unchanged (Figure 6F), suggesting that APP correction primarily reduces overall tau levels rather than alters phosphorylation stoichiometry. Levels of secreted Aβ₄₀ (Figure 6G) and Aβ₄₂ (Figure 6H) were robustly decreased in DS APP+/+/− neurons. Aβ₄₂/Aβ₄₀ ratio showed trend, albeit not statistically significant (Figure 6I). Collectively, these results demonstrate that restoring APP dosage levels in DS iPSCs-derived neurons rescues key AD-associated biochemical phenotypes.

Figure 6. — (A) Expression levels of BACE-1, APP C-terminal fragments (APP CTFs), phosphorylated tau (AT8), total tau (Tau5), DYRK1A, BACE-2, SOD1, and S100β in protein lysate of 10 week old neurons (MN) derived from DS MC and DS APP+/+/− iPSC lines, as examined by Western blot. (B-E, J-N) Quantification of BACE-1, APP CTFs, AT8, Tau5, DYRK1A, BACE-2, SOD1, and S100β levels normalized to GAPDH in DS MC and DS APP+/+/− groups. (n=3 replicates). (F) The ratio of phosphorylated- to total- tau (AT8/Tau5) is comparable between genotypes. (G, H) Quantification of secreted Aβ₄₀ and Aβ₄₂ in conditioned media of DS APP^+/+/− vs DS MC neuronal cultures using ELISA. (n=3 replicates). (I) Aβ₄₂/Aβ₄₀ ratio following APP dosage correction. Data represented as mean ± SEM with individual data points shown. Data analyzed by unpaired t-test, *p < 0.05, **p < 0.01, and ****p < 0.0001.

AMYLOID PRECURSOR PROTEIN GENE NORMALIZATION DIMINISHES CHROMOSOME 21 GENE DOSAGE EFFECTS AND STRESS PATHWAYS IN DOWN SYNDROME NEURONS.

Finally, we examined whether the normalization of APP gene dosage in DS iPSCs-derived neurons had an effect on HSA21 genes that were observed to be upregulated (Figure 3). Examination of protein lysate of DS APP+/+/− and DS MC neurons revealed no significant change in DYRK1A, but significantly lower levels of BACE2 in DS APP+/+/− neurons compared to DS MC (Figure 6A, J–K). Levels of SOD1 did not change (Figure 6A, L). Notably, S100β dimer and monomer levels were comparable in both genotypes (Figure 6A, M,N). These data indicate that APP gene dosage in DS neurons partially affects the dysregulation of genes on HSA21, underscoring the central regulatory role of APP in DS neuropathology.

Discussion

This study reveals several novel aspects. First, we show that APP overdose impairs DS NPC proliferation. Interestingly, we observed upregulation of SOX2 expression in NPCs and precursor cells and increased number of cells expressing SOX2. Together, this may suggest reduced pools of progenitor cells with enhanced stemness due to APP overdose. In turn, APP overdose induced premature differentiation, based on the expression of neurogenesis proxies in DS forebrain neurons. This may suggest that APP regulates more than one checkpoint along the neurogenic path. This central role of APP in DS brain development aligns with previous reports showing increased APP expression in DS fetal brain, leading to impaired neural progenitor proliferation and a gliocentric shift^34,35. Recent evidence demonstrates that APP is broadly expressed in human cortical progenitors throughout neurogenesis and is required for maintaining the temporal balance between self-renewal and differentiation of human neural stem cells. Loss of APP in human neural progenitors drives premature cell cycle exit, and increased generation of human cortical neurons. Importantly, APP modulates neurogenesis through proneurogenic activator protein-1 transcription factor suppression and activation of WNT signaling³⁶. Notably, our data suggests that HSA21 trisomy enhances BACE-1 expression and activity, leading to biased increase in APP metabolites cleaved by BACE-1. Future studies are warranted to determine the effect of these metabolites on these impairments. Studies have found elevated APP and its cleavage products, higher levels of Aβ, phospho-tau, and corresponding neurodevelopmental and neurodegenerative alterations using DS iPSC-derived neurons^35,37. Experimental reduction in APP and its products using Posiphen, has been shown to partially restore AD-linked neuronal phenotypes in mouse models^37,38. Notably, evidence suggests that metabolites of APP, which is mutated in familial forms of AD, play critical roles in developmental and postnatal neurogenesis. Of particular importance is soluble APP (sAPP), which regulates neural progenitor cell proliferation¹⁵. This highlights the essential roles of APP metabolites in neurogenesis, NPC proliferation, and fate specification. In addition, future studies should examine the implications for neuronal function. In that regard, we observed upregulation of tau species in DS neurons. Given the importance of tau in neuronal development, its upregulation may compromise proper neuronal maturation. Notably, we did not observe increased tau phosphorylation relative to total tau, suggesting that other factors may play a role in tau hyperphosphorylation or that AD-linked hyperphosphorylation takes place at later stages of brain function. One possibility is that a two-hit, such as aging and APP overdose, would be required for the pathological acceleration of tau hyperphosphorylation. While we observed enhanced expression of BACE-1 and enhanced secretion of Aβ₄₂ and Aβ₄₀ in DS neurons, in contrast to previous studies¹¹, we did not observe altered Aβ₄₂/Aβ₄₀ ratio. Possible explanations to this unaltered Aβ₄₂/Aβ₄₀ ratio could be due to differences in differentiation protocols, maturation state, or APP processing enzyme expression, as well as the possibility that APP dosage modulation in this early developmental window predominantly affects amyloidogenic output rather than selectively shifting the Aβ₄₂/Aβ₄₀ balance. Future work should examine long-term cultures and manipulations of specific secretases that may clarify how APP dosage affects its processing by γ-secretase.

Besides APP, our data suggests that several other genes on chromosome 21 contribute to the complex neuropathology of DS. DYRK1A, a kinase implicated in neurodevelopmental processes, is consistently upregulated in DS neurons and has been shown to influence neuronal proliferation, differentiation, and tau phosphorylation, thereby linking it to both developmental abnormalities and Alzheimer-like pathology^39,40. The BACE2 is also overexpressed, although its exact role in DS remains to be fully elucidated. Additionally, the calcium-binding protein S100 shows differential regulation in its dimeric and monomeric forms; notably, increased S100 dimer levels in DS neurons indicate heightened stress and inflammatory responses, whereas monomeric levels remain unchanged⁴¹. Notably, our data shows that elevated protein levels of BACE-2 in DS iPSC-derived neurons were reversed by normalization of APP gene dosage, suggesting that BACE-2 dysregulation is dependent or downstream of APP-mediated pathways. However, SOD1, DYRKIA, and S100β, were not rescued by APP gene dosage normalization, indicating that their dysregulation may be independent- or upstream- of APP pathways. Evidence from the literature supports that SOD1 overexpression exacerbates oxidative damage and impacts neuronal differentiation and survival^42,43, implicating oxidative stress as a complementary mechanism alongside APP overdose in the pathogenesis of DS neurodevelopmental and neurodegenerative phenotypes⁴⁴. Together, these gene dosage effects underscore the multifaceted molecular disruptions driving neurodevelopmental and neurodegenerative phenotypes in DS. Future studies should focus on dissecting the specific mechanistic contributions of these genes to better understand this disorder.

Limitations of our study include the 2D differentiation which does not allow full assessment of corticogenesis, and the lack of mixed cellular interactions with glia and microglia. The data is obtained from 10-week- old neurons, a relatively short culture duration, which does not allow aging-related processes, as well as the accumulation of amyloid deposition and neurofibrillary tangles. Future studies should use these iPSC lines for the development of cortical organoids with prolonged maturation to model more advanced DS-AD pathology. The DS MC and DS DS APP+/+/− lines provide a platform for the examination of the effect of APP overdose on many aspects of developmental neurogenesis and AD pathology that are yet to be elucidated.

Conclusion

In summary, this study suggests that APP overdose causes impairments in neurogenesis and facilitates amyloidosis in DS. Integrating these insights builds a nuanced view of APP’s role or its downstream pathways, in DS, as a strategy to ameliorate both neurodevelopmental and neurodegenerative sequelae in DS.

Supplementary Material

NIHMS2203834-supplement-1.pdf^{(164.6KB, pdf)}

Acknowledgements and Funding Statement

Graphical abstract, Figure 1A, 1B, and 4A were created Biorender.com. This work was financially supported by National Institutes of Health (NIH), National Institute on Aging (NIA) AG079002, AG057468, AG076940, and AG033570 (OL).

Footnotes

Conflict of Interest Statement

The authors have no conflicts of interest to declare.

References:

1.Ruparelia A, Pearn ML, Mobley WC. Aging and intellectual disability: insights from mouse models of Down syndrome. Dev Disabil Res Rev. 2013;18(1):43–50. doi: 10.1002/ddrr.1127 [DOI] [PubMed] [Google Scholar]
2.Patkee PA, Baburamani AA, Kyriakopoulou V, et al. Early alterations in cortical and cerebellar regional brain growth in Down Syndrome: An in vivo fetal and neonatal MRI assessment. NeuroImage Clin. 2020;25:102139. doi: 10.1016/j.nicl.2019.102139 [DOI] [PMC free article] [PubMed] [Google Scholar]
3.Dierssen M. Down syndrome: the brain in trisomic mode. Nat Rev Neurosci. 2012;13(12):844–858. doi: 10.1038/nrn3314 [DOI] [PubMed] [Google Scholar]
4.Korbel JO, Tirosh-Wagner T, Urban AE, et al. The genetic architecture of Down syndrome phenotypes revealed by high-resolution analysis of human segmental trisomies. Proc Natl Acad Sci U S A. 2009;106(29):12031–12036. doi: 10.1073/pnas.0813248106 [DOI] [PMC free article] [PubMed] [Google Scholar]
5.Müller UC, Deller T, Korte M. Not just amyloid: physiological functions of the amyloid precursor protein family. Nat Rev Neurosci. 2017;18(5):281–298. doi: 10.1038/nrn.2017.29 [DOI] [PubMed] [Google Scholar]
6.Young-Pearse TL, Bai J, Chang R, Zheng JB, LoTurco JJ, Selkoe DJ. A critical function for beta-amyloid precursor protein in neuronal migration revealed by in utero RNA interference. J Neurosci Off J Soc Neurosci. 2007;27(52):14459–14469. doi: 10.1523/JNEUROSCI.4701-07.2007 [DOI] [PMC free article] [PubMed] [Google Scholar]
7.Coronel R, López-Alonso V, Gallego MI, Liste I. Low Levels of Amyloid Precursor Protein (APP) Promote Neurogenesis and Decrease Gliogenesis in Human Neural Stem Cells. Int J Mol Sci. 2023;24(19):14635. doi: 10.3390/ijms241914635 [DOI] [PMC free article] [PubMed] [Google Scholar]
8.Caillé I, Allinquant B, Dupont E, et al. Soluble form of amyloid precursor protein regulates proliferation of progenitors in the adult subventricular zone. Dev Camb Engl. 2004;131(9):2173–2181. doi: 10.1242/dev.01103 [DOI] [PubMed] [Google Scholar]
9.Rice HC, Townsend M, Bai J, et al. Pancortins interact with amyloid precursor protein and modulate cortical cell migration. Dev Camb Engl. 2012;139(21):3986–3996. doi: 10.1242/dev.082909 [DOI] [PMC free article] [PubMed] [Google Scholar]
10.Müller UC, Deller T, Korte M. Not just amyloid: physiological functions of the amyloid precursor protein family. Nat Rev Neurosci. 2017;18(5):281–298. doi: 10.1038/nrn.2017.29 [DOI] [PubMed] [Google Scholar]
11.Lee SH, Kang J, Ho A, Watanabe H, Bolshakov VY, Shen J. APP Family Regulates Neuronal Excitability and Synaptic Plasticity but Not Neuronal Survival. Neuron. 2020;108(4):676–690.e8. doi: 10.1016/j.neuron.2020.08.011 [DOI] [PMC free article] [PubMed] [Google Scholar]
12.Klevanski M, Herrmann U, Weyer SW, et al. The APP Intracellular Domain Is Required for Normal Synaptic Morphology, Synaptic Plasticity, and Hippocampus-Dependent Behavior. J Neurosci. 2015;35(49):16018–16033. doi: 10.1523/JNEUROSCI.2009-15.2015 [DOI] [PMC free article] [PubMed] [Google Scholar]
13.Hick M, Herrmann U, Weyer SW, et al. Acute function of secreted amyloid precursor protein fragment APPsα in synaptic plasticity. Acta Neuropathol (Berl). 2015;129(1):21–37. doi: 10.1007/s00401-014-1368-x [DOI] [PubMed] [Google Scholar]
14.Li H, Wang Z, Wang B, et al. Genetic dissection of the amyloid precursor protein in developmental function and amyloid pathogenesis. J Biol Chem. 2010;285(40):30598–30605. doi: 10.1074/jbc.M110.137729 [DOI] [PMC free article] [PubMed] [Google Scholar]
15.Lazarov O, Demars MP. All in the Family: How the APPs Regulate Neurogenesis. Front Neurosci. 2012;6:81. doi: 10.3389/fnins.2012.00081 [DOI] [PMC free article] [PubMed] [Google Scholar]
16.Zhou Z dong, Chan CH shan, Ma Q hong, Xu X hong, Xiao Z cheng, Tan EK. The roles of amyloid precursor protein (APP) in neurogenesis, implications to pathogenesis and therapy of Alzheimer disease (AD). Cell Adhes Migr. 2011;5(4):280–292. doi: 10.4161/cam.5.4.16986 [DOI] [PMC free article] [PubMed] [Google Scholar]
17.Wang B, Wang Z, Sun L, et al. The Amyloid Precursor Protein Controls Adult Hippocampal Neurogenesis through GABAergic Interneurons. J Neurosci. 2014;34(40):13314–13325. doi: 10.1523/JNEUROSCI.2848-14.2014 [DOI] [PMC free article] [PubMed] [Google Scholar]
18.Demars MP, Bartholomew A, Strakova Z, Lazarov O. Soluble amyloid precursor protein: a novel proliferation factor of adult progenitor cells of ectodermal and mesodermal origin. Stem Cell Res Ther. 2011;2(4):36. doi: 10.1186/scrt77 [DOI] [PMC free article] [PubMed] [Google Scholar]
19.Demars MP, Hollands C, Zhao KDT, Lazarov O. Soluble amyloid precursor protein-α rescues age-linked decline in neural progenitor cell proliferation. Neurobiol Aging. 2013;34(10):2431–2440. doi: 10.1016/j.neurobiolaging.2013.04.016 [DOI] [PMC free article] [PubMed] [Google Scholar]
20.Shabani K, Pigeon J, Benaissa Touil Zariouh M, et al. The temporal balance between self-renewal and differentiation of human neural stem cells requires the amyloid precursor protein. Sci Adv. 2023;9(24):eadd5002. doi: 10.1126/sciadv.add5002 [DOI] [PMC free article] [PubMed] [Google Scholar]
21.Ovchinnikov DA, Korn O, Virshup I, Wells CA, Wolvetang EJ. The Impact of APP on Alzheimer-like Pathogenesis and Gene Expression in Down Syndrome iPSC-Derived Neurons. Stem Cell Rep. 2018;11(1):32–42. doi: 10.1016/j.stemcr.2018.05.004 [DOI] [PMC free article] [PubMed] [Google Scholar]
22.Sosa LJ, Postma NL, Estrada-Bernal A, et al. Dosage of amyloid precursor protein affects axonal contact guidance in Down syndrome. FASEB J Off Publ Fed Am Soc Exp Biol. 2014;28(1):195–205. doi: 10.1096/fj.13-232686 [DOI] [PMC free article] [PubMed] [Google Scholar]
23.Selkoe DJ. Alzheimer’s disease: genes, proteins, and therapy. Physiol Rev. 2001;81(2):741–766. doi: 10.1152/physrev.2001.81.2.741 [DOI] [PubMed] [Google Scholar]
24.Maure-Blesa L, Carmona-Iragui M, Lott I, et al. The history of Down syndrome-associated Alzheimer’s disease; past, present, and future. Alzheimers Dement J Alzheimers Assoc. 2025;21(6):e70158. doi: 10.1002/alz.70158 [DOI] [PMC free article] [PubMed] [Google Scholar]
25.Zigman WB, Lott IT. Alzheimer’s disease in Down syndrome: Neurobiology and risk. Ment Retard Dev Disabil Res Rev. 2007;13(3):237–246. doi: 10.1002/mrdd.20163 [DOI] [PubMed] [Google Scholar]
26.Ovchinnikov DA, Korn O, Virshup I, Wells CA, Wolvetang EJ. The Impact of APP on Alzheimer-like Pathogenesis and Gene Expression in Down Syndrome iPSC-Derived Neurons. Stem Cell Rep. 2018;11(1):32–42. doi: 10.1016/j.stemcr.2018.05.004 [DOI] [PMC free article] [PubMed] [Google Scholar]
27.Wu CI, Vinton EA, Pearse RV, et al. APP and DYRK1A regulate axonal and synaptic vesicle protein networks and mediate Alzheimer’s pathology in trisomy 21 neurons. Mol Psychiatry. 2022;27(4):1970–1989. doi: 10.1038/s41380-022-01454-5 [DOI] [PMC free article] [PubMed] [Google Scholar]
28.Lu HE, Yang YC, Chen SM, et al. Modeling neurogenesis impairment in Down syndrome with induced pluripotent stem cells from Trisomy 21 amniotic fluid cells. Exp Cell Res. 2013;319(4):498–505. doi: 10.1016/j.yexcr.2012.09.017 [DOI] [PubMed] [Google Scholar]
29.Utagawa EC, Moreno DG, Schafernak KT, et al. Neurogenesis and neuronal differentiation in the postnatal frontal cortex in Down syndrome. Acta Neuropathol Commun. 2022;10(1):86. doi: 10.1186/s40478-022-01385-w [DOI] [PMC free article] [PubMed] [Google Scholar]
30.Hernández-Ortega K, Canul-Euan AA, Solis-Paredes JM, et al. S100B actions on glial and neuronal cells in the developing brain: an overview. Front Neurosci. 2024;18. doi: 10.3389/fnins.2024.1425525 [DOI] [PMC free article] [PubMed] [Google Scholar]
31.Shapiro LA, Bialowas-McGoey LA, Whitaker-Azmitia PM. Effects of S100B on Serotonergic Plasticity and Neuroinflammation in the Hippocampus in Down Syndrome and Alzheimer’s Disease: Studies in an S100B Overexpressing Mouse Model. Cardiovasc Psychiatry Neurol. 2010;2010:153657. doi: 10.1155/2010/153657 [DOI] [PMC free article] [PubMed] [Google Scholar]
32.Matsui Lee IS, Suzuki M, Hayashi N, et al. Copper-dependent formation of disulfide-linked dimer of S100B protein. Arch Biochem Biophys. 2000;374(2):137–141. doi: 10.1006/abbi.1999.1595 [DOI] [PubMed] [Google Scholar]
33.Thulin E, Kesvatera T, Linse S. Molecular Determinants of S100B Oligomer Formation. PLOS ONE. 2011;6(3):e14768. doi: 10.1371/journal.pone.0014768 [DOI] [PMC free article] [PubMed] [Google Scholar]
34.Langeh U, Singh S. Targeting S100B Protein as a Surrogate Biomarker and its Role in Various Neurological Disorders. Curr Neuropharmacol. 2021;19(2):265–277. doi: 10.2174/1570159X18666200729100427 [DOI] [PMC free article] [PubMed] [Google Scholar]
35.Lu J, Esposito G, Scuderi C, et al. S100B and APP Promote a Gliocentric Shift and Impaired Neurogenesis in Down Syndrome Neural Progenitors. PLOS ONE. 2011;6(7):e22126. doi: 10.1371/journal.pone.0022126 [DOI] [PMC free article] [PubMed] [Google Scholar]
36.Stagni F, Bartesaghi R. The Challenging Pathway of Treatment for Neurogenesis Impairment in Down Syndrome: Achievements and Perspectives. Front Cell Neurosci. 2022;16:903729.doi: 10.3389/fncel.2022.903729 [DOI] [PMC free article] [PubMed] [Google Scholar]
37.Shabani K, Pigeon J, Benaissa Touil Zariouh M, et al. The temporal balance between self-renewal and differentiation of human neural stem cells requires the amyloid precursor protein. Sci Adv. 9(24):eadd5002. doi: 10.1126/sciadv.add5002 [DOI] [PMC free article] [PubMed] [Google Scholar]
38.Chen XQ. A pilot exploration with Posiphen to normalize amyloid precursor protein in Down syndrome. Neural Regen Res. 2021;16(12):2420–2421. doi: 10.4103/1673-5374.313044 [DOI] [PMC free article] [PubMed] [Google Scholar]
39.Chen XQ, Salehi A, Pearn ML, et al. Targeting increased levels of APP in Down syndrome: Posiphen-mediated reductions in APP and its products reverse endosomal phenotypes in the Ts65Dn mouse model. Alzheimers Dement J Alzheimers Assoc. 2021;17(2):271–292. doi: 10.1002/alz.12185 [DOI] [PMC free article] [PubMed] [Google Scholar]
40.Ryoo SR, Jeong HK, Radnaabazar C, et al. DYRK1A-mediated Hyperphosphorylation of Tau. J Biol Chem. 2007;282(48):34850–34857. doi: 10.1074/jbc.M707358200 [DOI] [PubMed] [Google Scholar]
41.Duchon A, Herault Y. DYRK1A, a Dosage-Sensitive Gene Involved in Neurodevelopmental Disorders, Is a Target for Drug Development in Down Syndrome. Front Behav Neurosci. 2016;10:104. doi: 10.3389/fnbeh.2016.00104 [DOI] [PMC free article] [PubMed] [Google Scholar]
42.Griffin WST, Sheng JG, McKenzie JE, et al. Life-long overexpression of S100β in Down’s syndrome: implications for Alzheimer pathogenesis. Neurobiol Aging. 1998;19(5):401–405. doi: 10.1016/S0197-4580(98)00074-8 [DOI] [PMC free article] [PubMed] [Google Scholar]
43.Guan T, Guo Y, Zhou T, et al. Oxidized SOD1 accelerates cellular senescence in neural stem cells. Stem Cell Res Ther. 2024;15(1):55. doi: 10.1186/s13287-024-03669-5 [DOI] [PMC free article] [PubMed] [Google Scholar]
44.Taylor JM, Crack PJ. Impact of Oxidative Stress on Neuronal Survival. Clin Exp Pharmacol Physiol. 2004;31(7):397–406. doi: 10.1111/j.1440-1681.2004.04017.x [DOI] [PubMed] [Google Scholar]
45.Chidambaram SB, Anand N, Varma SR, et al. Superoxide dismutase and neurological disorders. IBRO Neurosci Rep. 2024;16:373–394. doi: 10.1016/j.ibneur.2023.11.007 [DOI] [PMC free article] [PubMed] [Google Scholar]

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

NIHMS2203834-supplement-1.pdf^{(164.6KB, pdf)}

[R1] 1.Ruparelia A, Pearn ML, Mobley WC. Aging and intellectual disability: insights from mouse models of Down syndrome. Dev Disabil Res Rev. 2013;18(1):43–50. doi: 10.1002/ddrr.1127 [DOI] [PubMed] [Google Scholar]

[R2] 2.Patkee PA, Baburamani AA, Kyriakopoulou V, et al. Early alterations in cortical and cerebellar regional brain growth in Down Syndrome: An in vivo fetal and neonatal MRI assessment. NeuroImage Clin. 2020;25:102139. doi: 10.1016/j.nicl.2019.102139 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R3] 3.Dierssen M. Down syndrome: the brain in trisomic mode. Nat Rev Neurosci. 2012;13(12):844–858. doi: 10.1038/nrn3314 [DOI] [PubMed] [Google Scholar]

[R4] 4.Korbel JO, Tirosh-Wagner T, Urban AE, et al. The genetic architecture of Down syndrome phenotypes revealed by high-resolution analysis of human segmental trisomies. Proc Natl Acad Sci U S A. 2009;106(29):12031–12036. doi: 10.1073/pnas.0813248106 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R5] 5.Müller UC, Deller T, Korte M. Not just amyloid: physiological functions of the amyloid precursor protein family. Nat Rev Neurosci. 2017;18(5):281–298. doi: 10.1038/nrn.2017.29 [DOI] [PubMed] [Google Scholar]

[R6] 6.Young-Pearse TL, Bai J, Chang R, Zheng JB, LoTurco JJ, Selkoe DJ. A critical function for beta-amyloid precursor protein in neuronal migration revealed by in utero RNA interference. J Neurosci Off J Soc Neurosci. 2007;27(52):14459–14469. doi: 10.1523/JNEUROSCI.4701-07.2007 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R7] 7.Coronel R, López-Alonso V, Gallego MI, Liste I. Low Levels of Amyloid Precursor Protein (APP) Promote Neurogenesis and Decrease Gliogenesis in Human Neural Stem Cells. Int J Mol Sci. 2023;24(19):14635. doi: 10.3390/ijms241914635 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R8] 8.Caillé I, Allinquant B, Dupont E, et al. Soluble form of amyloid precursor protein regulates proliferation of progenitors in the adult subventricular zone. Dev Camb Engl. 2004;131(9):2173–2181. doi: 10.1242/dev.01103 [DOI] [PubMed] [Google Scholar]

[R9] 9.Rice HC, Townsend M, Bai J, et al. Pancortins interact with amyloid precursor protein and modulate cortical cell migration. Dev Camb Engl. 2012;139(21):3986–3996. doi: 10.1242/dev.082909 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R10] 10.Müller UC, Deller T, Korte M. Not just amyloid: physiological functions of the amyloid precursor protein family. Nat Rev Neurosci. 2017;18(5):281–298. doi: 10.1038/nrn.2017.29 [DOI] [PubMed] [Google Scholar]

[R11] 11.Lee SH, Kang J, Ho A, Watanabe H, Bolshakov VY, Shen J. APP Family Regulates Neuronal Excitability and Synaptic Plasticity but Not Neuronal Survival. Neuron. 2020;108(4):676–690.e8. doi: 10.1016/j.neuron.2020.08.011 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R12] 12.Klevanski M, Herrmann U, Weyer SW, et al. The APP Intracellular Domain Is Required for Normal Synaptic Morphology, Synaptic Plasticity, and Hippocampus-Dependent Behavior. J Neurosci. 2015;35(49):16018–16033. doi: 10.1523/JNEUROSCI.2009-15.2015 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R13] 13.Hick M, Herrmann U, Weyer SW, et al. Acute function of secreted amyloid precursor protein fragment APPsα in synaptic plasticity. Acta Neuropathol (Berl). 2015;129(1):21–37. doi: 10.1007/s00401-014-1368-x [DOI] [PubMed] [Google Scholar]

[R14] 14.Li H, Wang Z, Wang B, et al. Genetic dissection of the amyloid precursor protein in developmental function and amyloid pathogenesis. J Biol Chem. 2010;285(40):30598–30605. doi: 10.1074/jbc.M110.137729 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R15] 15.Lazarov O, Demars MP. All in the Family: How the APPs Regulate Neurogenesis. Front Neurosci. 2012;6:81. doi: 10.3389/fnins.2012.00081 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R16] 16.Zhou Z dong, Chan CH shan, Ma Q hong, Xu X hong, Xiao Z cheng, Tan EK. The roles of amyloid precursor protein (APP) in neurogenesis, implications to pathogenesis and therapy of Alzheimer disease (AD). Cell Adhes Migr. 2011;5(4):280–292. doi: 10.4161/cam.5.4.16986 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R17] 17.Wang B, Wang Z, Sun L, et al. The Amyloid Precursor Protein Controls Adult Hippocampal Neurogenesis through GABAergic Interneurons. J Neurosci. 2014;34(40):13314–13325. doi: 10.1523/JNEUROSCI.2848-14.2014 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R18] 18.Demars MP, Bartholomew A, Strakova Z, Lazarov O. Soluble amyloid precursor protein: a novel proliferation factor of adult progenitor cells of ectodermal and mesodermal origin. Stem Cell Res Ther. 2011;2(4):36. doi: 10.1186/scrt77 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R19] 19.Demars MP, Hollands C, Zhao KDT, Lazarov O. Soluble amyloid precursor protein-α rescues age-linked decline in neural progenitor cell proliferation. Neurobiol Aging. 2013;34(10):2431–2440. doi: 10.1016/j.neurobiolaging.2013.04.016 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R20] 20.Shabani K, Pigeon J, Benaissa Touil Zariouh M, et al. The temporal balance between self-renewal and differentiation of human neural stem cells requires the amyloid precursor protein. Sci Adv. 2023;9(24):eadd5002. doi: 10.1126/sciadv.add5002 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R21] 21.Ovchinnikov DA, Korn O, Virshup I, Wells CA, Wolvetang EJ. The Impact of APP on Alzheimer-like Pathogenesis and Gene Expression in Down Syndrome iPSC-Derived Neurons. Stem Cell Rep. 2018;11(1):32–42. doi: 10.1016/j.stemcr.2018.05.004 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R22] 22.Sosa LJ, Postma NL, Estrada-Bernal A, et al. Dosage of amyloid precursor protein affects axonal contact guidance in Down syndrome. FASEB J Off Publ Fed Am Soc Exp Biol. 2014;28(1):195–205. doi: 10.1096/fj.13-232686 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R23] 23.Selkoe DJ. Alzheimer’s disease: genes, proteins, and therapy. Physiol Rev. 2001;81(2):741–766. doi: 10.1152/physrev.2001.81.2.741 [DOI] [PubMed] [Google Scholar]

[R24] 24.Maure-Blesa L, Carmona-Iragui M, Lott I, et al. The history of Down syndrome-associated Alzheimer’s disease; past, present, and future. Alzheimers Dement J Alzheimers Assoc. 2025;21(6):e70158. doi: 10.1002/alz.70158 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R25] 25.Zigman WB, Lott IT. Alzheimer’s disease in Down syndrome: Neurobiology and risk. Ment Retard Dev Disabil Res Rev. 2007;13(3):237–246. doi: 10.1002/mrdd.20163 [DOI] [PubMed] [Google Scholar]

[R26] 26.Ovchinnikov DA, Korn O, Virshup I, Wells CA, Wolvetang EJ. The Impact of APP on Alzheimer-like Pathogenesis and Gene Expression in Down Syndrome iPSC-Derived Neurons. Stem Cell Rep. 2018;11(1):32–42. doi: 10.1016/j.stemcr.2018.05.004 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R27] 27.Wu CI, Vinton EA, Pearse RV, et al. APP and DYRK1A regulate axonal and synaptic vesicle protein networks and mediate Alzheimer’s pathology in trisomy 21 neurons. Mol Psychiatry. 2022;27(4):1970–1989. doi: 10.1038/s41380-022-01454-5 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R28] 28.Lu HE, Yang YC, Chen SM, et al. Modeling neurogenesis impairment in Down syndrome with induced pluripotent stem cells from Trisomy 21 amniotic fluid cells. Exp Cell Res. 2013;319(4):498–505. doi: 10.1016/j.yexcr.2012.09.017 [DOI] [PubMed] [Google Scholar]

[R29] 29.Utagawa EC, Moreno DG, Schafernak KT, et al. Neurogenesis and neuronal differentiation in the postnatal frontal cortex in Down syndrome. Acta Neuropathol Commun. 2022;10(1):86. doi: 10.1186/s40478-022-01385-w [DOI] [PMC free article] [PubMed] [Google Scholar]

[R30] 30.Hernández-Ortega K, Canul-Euan AA, Solis-Paredes JM, et al. S100B actions on glial and neuronal cells in the developing brain: an overview. Front Neurosci. 2024;18. doi: 10.3389/fnins.2024.1425525 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R31] 31.Shapiro LA, Bialowas-McGoey LA, Whitaker-Azmitia PM. Effects of S100B on Serotonergic Plasticity and Neuroinflammation in the Hippocampus in Down Syndrome and Alzheimer’s Disease: Studies in an S100B Overexpressing Mouse Model. Cardiovasc Psychiatry Neurol. 2010;2010:153657. doi: 10.1155/2010/153657 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R32] 32.Matsui Lee IS, Suzuki M, Hayashi N, et al. Copper-dependent formation of disulfide-linked dimer of S100B protein. Arch Biochem Biophys. 2000;374(2):137–141. doi: 10.1006/abbi.1999.1595 [DOI] [PubMed] [Google Scholar]

[R33] 33.Thulin E, Kesvatera T, Linse S. Molecular Determinants of S100B Oligomer Formation. PLOS ONE. 2011;6(3):e14768. doi: 10.1371/journal.pone.0014768 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R34] 34.Langeh U, Singh S. Targeting S100B Protein as a Surrogate Biomarker and its Role in Various Neurological Disorders. Curr Neuropharmacol. 2021;19(2):265–277. doi: 10.2174/1570159X18666200729100427 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R35] 35.Lu J, Esposito G, Scuderi C, et al. S100B and APP Promote a Gliocentric Shift and Impaired Neurogenesis in Down Syndrome Neural Progenitors. PLOS ONE. 2011;6(7):e22126. doi: 10.1371/journal.pone.0022126 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R36] 36.Stagni F, Bartesaghi R. The Challenging Pathway of Treatment for Neurogenesis Impairment in Down Syndrome: Achievements and Perspectives. Front Cell Neurosci. 2022;16:903729.doi: 10.3389/fncel.2022.903729 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R37] 37.Shabani K, Pigeon J, Benaissa Touil Zariouh M, et al. The temporal balance between self-renewal and differentiation of human neural stem cells requires the amyloid precursor protein. Sci Adv. 9(24):eadd5002. doi: 10.1126/sciadv.add5002 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R38] 38.Chen XQ. A pilot exploration with Posiphen to normalize amyloid precursor protein in Down syndrome. Neural Regen Res. 2021;16(12):2420–2421. doi: 10.4103/1673-5374.313044 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R39] 39.Chen XQ, Salehi A, Pearn ML, et al. Targeting increased levels of APP in Down syndrome: Posiphen-mediated reductions in APP and its products reverse endosomal phenotypes in the Ts65Dn mouse model. Alzheimers Dement J Alzheimers Assoc. 2021;17(2):271–292. doi: 10.1002/alz.12185 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R40] 40.Ryoo SR, Jeong HK, Radnaabazar C, et al. DYRK1A-mediated Hyperphosphorylation of Tau. J Biol Chem. 2007;282(48):34850–34857. doi: 10.1074/jbc.M707358200 [DOI] [PubMed] [Google Scholar]

[R41] 41.Duchon A, Herault Y. DYRK1A, a Dosage-Sensitive Gene Involved in Neurodevelopmental Disorders, Is a Target for Drug Development in Down Syndrome. Front Behav Neurosci. 2016;10:104. doi: 10.3389/fnbeh.2016.00104 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R42] 42.Griffin WST, Sheng JG, McKenzie JE, et al. Life-long overexpression of S100β in Down’s syndrome: implications for Alzheimer pathogenesis. Neurobiol Aging. 1998;19(5):401–405. doi: 10.1016/S0197-4580(98)00074-8 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R43] 43.Guan T, Guo Y, Zhou T, et al. Oxidized SOD1 accelerates cellular senescence in neural stem cells. Stem Cell Res Ther. 2024;15(1):55. doi: 10.1186/s13287-024-03669-5 [DOI] [PMC free article] [PubMed] [Google Scholar]

[R44] 44.Taylor JM, Crack PJ. Impact of Oxidative Stress on Neuronal Survival. Clin Exp Pharmacol Physiol. 2004;31(7):397–406. doi: 10.1111/j.1440-1681.2004.04017.x [DOI] [PubMed] [Google Scholar]

[R45] 45.Chidambaram SB, Anand N, Varma SR, et al. Superoxide dismutase and neurological disorders. IBRO Neurosci Rep. 2024;16:373–394. doi: 10.1016/j.ibneur.2023.11.007 [DOI] [PMC free article] [PubMed] [Google Scholar]

PERMALINK

Amyloid precursor protein dosage normalization rescues neurogenesis and Alzheimer’s Disease phenotypes associated with Down Syndrome

Deepika Patel

Karen Rakowiecki

Orly Lazarov

Abstract

Introduction

Materials and Methods