INPP5D/SHIP1 regulates endo-lysosomal function and cargo-selective phagocytosis in human microglia

Two studies observed no change in plaque burden with reduced Inpp5d levels.

“While one study reported amelioration of plaque burden by Inpp5d knockdown in a mouse model of AD56, another study reported increased plaque burden78 and two studies observed no change with reduced Inpp5d levels57,61.”

• cites [57] Lin, P. B. C. , et al. INPP5D deficiency attenuates amyloid pathology in a mouse model of Alzheimer’s disease. Alzheimer’s & Dementia 19, (2022). sourceDisputed

  Section 3.1 reports that Inpp5d haplodeficiency reduced both diffuse and dense-core plaques in the cortex and hippocampus of 5xFAD mice. The Discussion likewise reports an overall reduction in plaque density, contrary to the attributed finding of no change. (LLM, conf=0.92)

  Checked by a reasoning model — may be wrong.

  Experimental — accuracy not yet measured
• cites [61] Pedicone, C. et al. Pan-SHIP1/2 inhibitors promote microglia effector functions essential for CNS homeostasis. J. Cell Sci. 133, (2020). sourceCould not verify

  The indexed passage describing the study reports increased microglial phagocytosis of Aβ after SHIP1/2 inhibition, not a measurement of plaque burden. Searches for reduced Inpp5d levels, plaque burden, and plaque load found no passage reporting that plaque burden was unchanged. (LLM, conf=0.76)

  Experimental — accuracy not yet measured

Recent GWAS have identified several genes associated with AD that are predominantly or exclusively expressed in myeloid cells, including INPP5D.

“Recent genome-wide association studies (GWAS) have identified several genes associated with AD that are predominantly or exclusively expressed in myeloid cells, including INPP5D (inositol polyphosphate 5-phosphatase 1), which encodes SHIP17–10.”

• cites [7] sourcePartly supported

  The paper identifies an Alzheimer’s disease–associated locus at INPP5D (Abstract; Table 2, Statistical analysis). It says INPP5D is expressed at low levels in the brain, but does not establish that it or the other associated genes are predominantly or exclusively expressed in myeloid cells. (LLM, conf=0.87)

  Experimental — accuracy not yet measured
• cites [8] sourcePartly supported

  The Discussion (page unavailable) identifies several AD-associated genes that are mainly expressed in microglia, supporting the general point about myeloid-cell-associated genes. The passages found for INPP5D do not identify it as one of those genes or establish that it is predominantly or exclusively expressed in myeloid cells. (LLM, conf=0.84)

  Experimental — accuracy not yet measured
• cites [9] sourcePartly supported

  The Results section (‘Candidate gene prioritization at genome-wide loci’; page not provided) identifies INPP5D as a gene at a previously identified Alzheimer’s disease GWAS locus. It also reports enrichment of associated genes in monocytes and macrophages, but does not establish that INPP5D itself is predominantly or exclusively expressed in myeloid cells. The retrieved passages do not identify INPP5D as encoding SHIP1. (LLM, conf=0.86)

  Experimental — accuracy not yet measured
• cites [10] sourcePartly supported

  The Tissue type, cell type and gene-set enrichment section (page unavailable) reports that Alzheimer’s-associated genes are enriched in microglia, supporting the broad link between GWAS findings and immune-related cells. The passages do not identify INPP5D as an associated gene or establish that it is predominantly or exclusively expressed in myeloid cells; searches for INPP5D and myeloid-cell expression found no such passage. (LLM, conf=0.82)

  Experimental — accuracy not yet measured

INPP5D loss-of-function in iMGs reduces autophagic flux.

“we previously reported a reduction in autophagic flux upon INPP5D loss-of-function in iMGs11”

• cites [11] sourceCould not verify

  The Results section, “Measuring knockout fitness effects” (page unavailable), discusses bacterial gene knockouts such as pykF, not INPP5D in iMGs. Searches for the claim and for microglia, autophagy, and autophagic flux found no passage addressing the attributed finding in our copy. (LLM, conf=0.78)

  Experimental — accuracy not yet measured

RNA expression data from human brain tissue reveal the presence of isoforms of INPP5D that encode truncated SHIP1 that lacks phosphatase domain.

“Intriguingly, RNA expression data from human brain tissue reveal the presence of isoforms of INPP5D that encode truncated SHIP1 that lacks phosphatase domain12.”

• cites [12] Zajac, D. J. et al. Expression of INPP5D Isoforms in Human Brain: Impact of Alzheimer’s Disease Neuropathology and Genetics. Genes (Basel ). 14, 763 (2023). sourceSupported

We detected several SHIP1 binding partners previously identified in other myeloid cells, such as SH3KBP1, GRB2, SHC1, DOK2, ARAP1, CD2AP.

“We detected several SHIP1 binding partners previously identified in other myeloid cells, such as SH3KBP117, GRB218, SHC118, DOK218,19, ARAP120 and CD2AP21 (Figure 1D).”

• cites [17] sourceCould not verify

  Citation 21 is attached specifically to CD2AP. The cited paper’s Results discuss protein networks in stem-cell-derived neurons and Alzheimer’s disease, but the passages available from our copy do not address CD2AP as a SHIP1 binding partner in myeloid cells. Searches for CD2AP, SHIP1/INPP5D, and the other named partners found no supporting passage; page numbers were not provided. (LLM, conf=0.76)

  Experimental — accuracy not yet measured
• cites [18] sourceNot checked — no full text of the source
• cites [19] sourceNot checked — no full text of the source

3AC robustly induces secretion of IL-1β and IL-18 upon activation of the NLRP3 inflammasome.

“3AC, which we previously showed to robustly induce the secretion of IL-1β and IL-18 upon activation of the NLRP3 inflammasome11”

• cites [11] sourceCould not verify

  The Results section discusses bacterial knockout fitness and correlations across genetic backgrounds, not inflammasome activation. Searches for 3AC, IL-1β, IL-18, and NLRP3 found no passage in our copy addressing the claimed secretion effect; no page numbers were available. (LLM, conf=0.78)

  Experimental — accuracy not yet measured

SHIP1 and SHIP2's enzymatic substrate, PI(3,4,5)P3, can induce lysosome de-acidification by activating the lysosomal potassium channel TMEM175.

“their enzymatic substrate, PI(3,4,5)P3, can induce lysosome de-acidification by activating the lysosomal potassium channel TMEM17567.”

• cites [67] sourceCould not verify

  The Discussion describes lysosomal de-acidification in Alzheimer’s disease neurons, but does not connect it to PI(3,4,5)P3 or TMEM175. Searches for TMEM175, PI(3,4,5)P3, SHIP1 and SHIP2 found no passage in our copy supporting the claimed mechanism. (LLM, conf=0.76)

  Experimental — accuracy not yet measured

Levels of full-length SHIP1 protein that include its phosphatase domain are reduced in brain tissue of individuals with AD.

“levels of full-length SHIP1 protein that include its phosphatase domain are reduced65.”

• cites [65] Chou, V. et al. INPP5D regulates inflammasome activation in human microglia. Nat. Commun. (2023). sourceSupported

Active cathepsin B can leak into the cytoplasm after lysosomal membrane permeabilization (LMP) in contexts where lysosomal membrane integrity is compromised, such as upon lysosomal deacidification.

“Second, active cathepsin B can leak into the cytoplasm after lysosomal membrane permeabilization (LMP) in contexts where lysosomal membrane integrity is compromised, such as upon lysosomal deacidification44,69.”

• cites [44] 2010, SHIP1 Inhibition Increases Immunoregulatory Capacity and Triggers Apoptosis of Hematopoietic Cancer Cells, The Journal of Immunology sourceCould not verify

  The Results section reports apoptosis after SHIP1 inhibition, and Materials and Methods describes assays for apoptosis-related proteins, but neither addresses cathepsin B release or lysosomal membrane permeabilization. Searches for cathepsin B and lysosomal deacidification found no passage in our copy supporting the attributed mechanism; page numbers were not available. (LLM, conf=0.76)

  Experimental — accuracy not yet measured
• cites [69] sourcePartly supported

  The “LysoKGF regulates lysosomal function” section reports lysosomal alkalinization in TMEM175-knockout neurons, and Extended Data Fig. 9 describes increased lysosomal permeability under a separate α-synuclein-seeding condition. Neither passage reports active cathepsin B leaking into the cytoplasm after LMP, or establishes that deacidification caused that leakage. Page numbers are unavailable in our copy. (LLM, conf=0.83)

  Experimental — accuracy not yet measured

Increased GPNMB secretion indicates a response to lysosomal stress to restore lysosomal acidification.

“which indicates a response to lysosomal stress to restore lysosomal acidification39”

• cites [39] sourceCould not verify

  The Introduction discusses lysosomal acidification in relation to cathepsin activity and NLRP3 activation, and the Methods describe measuring lysosomal integrity (page numbers unavailable). Searches for GPNMB secretion and restoration of lysosomal acidification found no passage connecting either to a response to lysosomal stress. (LLM, conf=0.75)

  Experimental — accuracy not yet measured

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

Cathepsin B leaking from destabilized lysosomes may mediate NLRP3 inflammasome activation upon SHIP1 inhibition.

“Based on the previous literature42,43,46, we reasoned that cathepsin B leaking from the destabilized lysosomes may mediate the NLRP3 inflammasome activation upon SHIP1 inhibition11.”

• cites [42] Hernandez, M. L. , Marone, M. , Gorse, K. M. & Lafrenaye, A. D . Cathepsin B Relocalization in Late Membrane Disrupted Neurons Following Diffuse Brain Injury in Rats. ASN Neuro 14, (2022). sourcePartly supported

  The Discussion (page unavailable) reports cathepsin B outside lysosomes after diffuse brain injury and suggests lysosomal membrane permeabilization could release it into the cytosol. Our copy does not link that release to NLRP3 inflammasome activation or SHIP1 inhibition, the condition next to this citation in the citing sentence. (LLM, conf=0.84)

  Experimental — accuracy not yet measured
• cites [43] sourcePartly supported

  In Results (“Lysosomal damage triggers release of cathepsin B”; page unavailable), the paper reports that amyloid-β damages microglial lysosomes, releases cathepsin B, and links cathepsin B to inflammasome activation. It does not examine SHIP1 inhibition, so it supports the proposed mechanism as a rationale, not its occurrence upon SHIP1 inhibition. (LLM, conf=0.88)

  Experimental — accuracy not yet measured
• cites [46] sourceCould not verify

  The Results section discusses microglial responses to apoptotic neurons, not cathepsin B leakage or NLRP3 activation. Searches of our copy for the proposed lysosomal mechanism and for SHIP1 inhibition found no passage addressing either point; page numbers were not available. (LLM, conf=0.72)

  Experimental — accuracy not yet measured

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.
• AssumptionRests on an assumption: Acute SHIP1 inhibition with 3AC specifically inhibits SHIP1 in iMGs.

The mRNA expression of INPP5D is increased in brain tissue of individuals with AD.

“while the mRNA expression of INPP5D is increased in brain tissue of individuals with AD”

• cites [65] Chou, V. et al. INPP5D regulates inflammasome activation in human microglia. Nat. Commun. (2023). sourceSupported

One mechanism by which SHIP1 deficiency increases the phagocytosis of lipid-rich substrates is through upregulation of TREM2 signaling.

“We confirmed that one mechanism by which SHIP1 deficiency increases the phagocytosis of lipid-rich substrates is through upregulation of TREM2 signaling (Figures 6D-G, S6E) in line with previous reports55,57.”

• cites [55] sourceNot checked — no full text of the source
• cites [57] Lin, P. B. C. , et al. INPP5D deficiency attenuates amyloid pathology in a mouse model of Alzheimer’s disease. Alzheimer’s & Dementia 19, (2022). sourcePartly supported

  The TREM2-SYK results section reports increased TREM2 expression and SYK phosphorylation in Inpp5d-haplodeficient 5xFAD mice, and the Highlights report increased microglial amyloid uptake. But the Discussion explicitly calls TREM2 activation as the mechanism a postulate requiring confirmation; the paper does not establish that it causes increased phagocytosis of lipid-rich substrates. (LLM, conf=0.87)

  Experimental — accuracy not yet measured

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.
• AssumptionRests on an assumption: Antibody-mediated inhibition of TREM2 specifically blocks TREM2 signaling in iMGs.

Loss-of-function of TREM2 did not ameliorate lysosomal deficits in other contexts.

“and while we did not test this directly in this study, loss-of-function of TREM2 did not ameliorate lysosomal deficits in other contexts76,77.”

• cites [76] 2022, Microglial INPP5D limits plaque formation and glial reactivity in the PSAPP mouse model of Alzheimer’s disease, Alzheimer’s & DementiaNot matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

• cites [77] sourceCould not verify

  The Results and Discussion passages report effects of the CSF1R inhibitor PLX3397 in 5XFAD mice, not effects of TREM2 loss-of-function on lysosomal deficits. Searches for TREM2 loss-of-function and lysosomal deficits found no passage in our copy addressing that claim; the passages have no page numbers. (LLM, conf=0.76)

  Experimental — accuracy not yet measured

Aβ can bind to lipoproteins such as clusterin (ApoJ) and ApoE in the brain, which facilitates its clearance by microglia via TREM2.

“However, Aβ can bind to lipoproteins such as clusterin (ApoJ) and ApoE in the brain82–84, which facilitates its clearance by microglia via TREM281.”

• cites [82] sourceCould not verify

  The citation is attached to the claim that lipoprotein-bound Aβ is cleared by microglia via TREM2. Our copy discusses 5xFAD plaque burden (Introduction, p. 2) and mentions Trem2 only when excluding crossbred models from a database search (Reporting of Thy1 promoter-driven transgenic AD models, p. 14). Searches for TREM2-mediated clearance and for clusterin/ApoJ and ApoE binding found no passage addressing that mechanism. (LLM, conf=0.74)

  Experimental — accuracy not yet measured
• cites [83] sourceCould not verify

  The Results section on genetic screens discusses interferon signaling in BV2 cells, not amyloid-β clearance. The only microglia-related item found is a title in the References; searches for Aβ binding to clusterin or ApoE and for TREM2-mediated clearance found no passage addressing the claim. Page numbers are unavailable in our copy. (LLM, conf=0.78)

  Experimental — accuracy not yet measured
• cites [84] sourceCould not verify

  The Methods sections on LPS challenge and respiratory burst concern immune responses in songbirds, not amyloid-β clearance in the brain. Searches for TREM2, microglia, ApoE, and clusterin found no passage in our copy addressing the claim. (LLM, conf=0.79)

  Experimental — accuracy not yet measured
• cites [81] sourcePartly supported

  In the lysosomal dysfunction section (p. 10), TREM2-deficient microglia have more APOE/Aβ-associated aggregates, supporting a role for TREM2 in limiting their accumulation. The paper does not establish that binding of Aβ to ApoE or clusterin facilitates its clearance via TREM2; its uptake experiments (pp. 12, 15) concern ApoE and do not test that full mechanism. (LLM, conf=0.82)

  Experimental — accuracy not yet measured

Reduction of SHIP1 levels via genome editing impairs lysosomal function.

“Reduction of SHIP1 levels via genome editing impairs endosome maturation and lysosomal function”

Reduction of SHIP1 levels leads to leakage of lysosomal cathepsin B into the cytosol.

“leakage of lysosomal cathepsin B into the cytosol”

Reduction of SHIP1 levels leads to lipid droplet accumulation.

“leading to lipid droplet accumulation”

Amyloid-β accumulated intracellularly due to defective lysosomal degradation in SHIP1-deficient microglia.

“amyloid-β accumulated intracellularly due to defective lysosomal degradation”

CITE-seq profiling of SHIP1-deficient microglia revealed a shift from an immune-responsive state toward a DAM-like, phagocytic state.

“CITE-seq profiling of SHIP1-deficient microglia revealed a shift from an immune-responsive state toward a DAM-like, phagocytic state”

SHIP1 localizes to both the plasma membrane and to endo-lysosomal compartments.

“We show that SHIP1 localizes to both the plasma membrane and to endo-lysosomal compartments”

Reduction of SHIP1 levels via genome editing impairs endosome maturation.

“Reduction of SHIP1 levels via genome editing impairs endosome maturation”

SHIP1-deficient microglia have enhanced phagocytosis of synaptic material and apoptotic neurons via TREM2.

“enhanced phagocytosis of synaptic material and apoptotic neurons via TREM2”

SHIP1-deficient microglia have impaired response to LPS.

“accompanied by impaired response to LPS”

Leakage of lysosomal cathepsin B into the cytosol activates the NLRP3 inflammasome.

“which in turn activates the NLRP3 inflammasome”

SHIP1 binds to the CapZ family of proteins.

“binds to the CapZ family of proteins”

Amyloid-β uptake was not affected by SHIP1 deficiency.

“While amyloid-β uptake was not affected”

BR24 HET iMGs show a pronounced reduction in LysoTracker fluorescence.

“HET BR24 iMGs showed a pronounced reduction in LysoTracker fluorescence”

In HET iMGs, a decrease in Rab11+ puncta was not observed.

“a decrease in Rab11+ puncta was not observed”

We identified 38 proteins that were significantly enriched across both genetic backgrounds more than 1.5-fold in WT microglia compared to KO microglia.

“We identified 38 proteins that were significantly (adj. p-value < 0.05) enriched across both genetic backgrounds more than 1.5-fold in WT microglia compared to KO microglia (Figures 1C-E).”

HET BR33 iMGs express SHIP1 at levels comparable to BR24 WT iMGs.

“HET BR33 iMGs expressed SHIP1 at levels comparable to BR24 WT iMGs”

Rab11 signal intensity is increased in HET iMGs.

“the intensity of Rab11 signal was increased in HET iMGs”

BR33 HET iMGs have unchanged mTFP1/mCherry ratio.

“this ratio was unchanged in BR33 HET iMGs”

The percentage of SHIP1 puncta co-localized with DQ-BSA is significantly higher in HET iMGs than WT.

“the percentage of SHIP1 puncta co-localized with DQ-BSA was significantly higher in HET iMGs”

In BR24 HET iMGs, relative area of LAMP1 puncta over IBA1+ area is lower than WT.

“In BR24 iMGs, LAMP1 protein levels measured by WB and the relative area of LAMP1 puncta over IBA1+ area measured by immunostaining were both lower in HET iMGs”

BR33 WT iMGs have significantly higher SHIP1 levels than BR24 WT iMGs.

“BR33 WT iMGs have significantly higher SHIP1 levels than BR24 WT iMGs”

BR24 HET iMGs have increased mTFP1/mCherry ratio.

“mTFP1/mCherry ratio was increased in BR24 HET iMGs”

HET iMGs have decreased Rab5 protein levels.

“found decreased Rab5 protein levels”

BR33 WT iMGs exhibit higher LysoTracker fluorescence than BR24 WT iMGs.

“BR33 WT iMGs also exhibit higher LysoTracker fluorescence”

Amyloid-β accumulation further drives lipid droplet formation.

“further driving lipid droplet formation”

BR33 WT iMGs have approximately twice as many LAMP1+ puncta compared with BR24 WT iMGs.

“approximately twice as many LAMP1⁺ puncta compared with BR24 WT iMGs”

The increase in pHrodo fluorescence is stunted in HET iMGs compared to WT.

“the increase in fluorescence was stunted in HET iMGs”

HET iMGs have reduced overall SHIP1 protein levels compared to WT.

“While there was reduced overall protein levels of SHIP1 in HET iMGs”

Treatment of iMGs with bafilomycin A1 increases SHIP1 levels.

“upon treatment of iMGs with the autophagy inhibitor bafilomycin A1 (Baf), we detected increased SHIP1 levels via WB (Figure 2D)”

HET iMGs have reduced Rab5 puncta area.

“reduced Rab5 puncta area in HET iMGs”

In WT iMGs, a subset of SHIP1 co-localizes with DQ-BSA+ puncta.

“Immunostaining of microglia after the DQ-BSA assay revealed co-localization of subset of SHIP1 with DQ-BSA+ puncta in WT iMGs.”

Reduction of INPP5D does not affect endocytic uptake.

“While reduction of INPP5D does not affect endocytic uptake (Figure 2B)”

BR33 HET iMGs show only mildly reduced LysoTracker fluorescence.

“HET BR33 iMGs showed only mildly reduced fluorescence”

In BR24 HET iMGs, LAMP1 protein levels are lower than WT.

“In BR24 iMGs, LAMP1 protein levels measured by WB and the relative area of LAMP1 puncta over IBA1+ area measured by immunostaining were both lower in HET iMGs”

HET iMGs accumulate more lipid droplets at baseline than WT.

“We found that HET iMGs accumulate more LDs at baseline (Figures 3A, S3A).”

Cluster 2 is significantly expanded in HET iMGs.

“Cluster 2 is significantly expanded in HET iMGs”

Both cell lines show a reduction in lysosomal degradation capacity upon INPP5D haploinsufficiency.

“both cell lines show a reduction in lysosomal degradation capacity upon INPP5D haploinsufficiency”

Within cluster 3, CD13 is significantly downregulated at the cell surface in HET iMGs.

“within cluster 3, CD13 (the top marker of cluster 3) was significantly downregulated at the cell surface in HET iMGs”

A significantly higher percentage of HET iMGs had taken up synaptosomes compared to WT iMGs.

“We found that a significantly higher percentage of HET iMGs had taken up synaptosomes”

In HET BR24 iMGs, cathepsin B puncta area is reduced.

“immunostaining for cathepsin B showed a reduction in cathepsin B puncta area”

Cluster 4 differentially expresses cell proliferation-related markers such as MKI67 and CDK1.

“Cluster 4, which differentially expresses cell proliferation-related markers such as MKI67 and CDK1”

Cathepsin B activity is elevated in HET BR33 iMGs.

“cathepsin B activity was elevated in HET BR33 iMGs”

The ratio of cathepsin B to LAMP1 is significantly elevated in HET BR24 iMGs.

“the ratio of cathepsin B to LAMP1 is significantly elevated in HET BR24 iMGs”

GPNMB secretion is decreased in BR24 iMGs with INPP5D reduction.

“it was decreased in BR24 iMGs with INPP5D reduction”

Cluster 3 is marked by chemokine-encoding genes CCL2, CCL3 and CCL4.

“is marked by chemokine-encoding genes CCL2, CCL3 and CCL4”

Reduced LPS-induced IL-6 transcription in HET iMGs is paralleled by diminished IL-6 secretion.

“The reduced LPS-induced IL-6 transcription in HET iMGs was paralleled by diminished IL-6 secretion (Figure S5D).”

In HET BR33 iMGs, cathepsin B intensity is trending increased.

“a trending increase in BR33 iMGs”

LPS treatment fails to increase expression of IL1B, CCL2, and IL6 in HET iMGs from both genetic backgrounds.

“it failed to do so in HET iMGs from both genetic backgrounds (Figures 5H-J, S5C).”

Extracellular SPP1 and APOE protein levels are significantly increased in HET iMG cultures from both genetic backgrounds.

“we observed a significant increase in extracellular SPP1 and APOE protein levels in HET iMG cultures from both genetic backgrounds”

• Also stated in: § INPP5D haploinsufficiency induces a shift from an immune-responsive state to a DAM-like, phagocytic state in microglia · plain

  “Notably, we observed a significant increase in extracellular SPP1 and APOE protein levels in HET iMG cultures from both genetic backgrounds (Figures S5C-D)”

CD13 is one of the top protein markers of cluster 3.

“One of the top protein markers of this cluster is CD13”

Within cluster 3, other proteins involved in immune response, such as LAIR1, CCR6 and C5aR, are significantly downregulated at the cell surface in HET iMGs.

“alongside other proteins involved in immune response, such as LAIR1, CCR6 and C5aR (Figures 5G-H, Supplementary Table 7).”

HET iMGs do not show a difference in baseline expression of IL1B, CCL2, and IL6 relative to WT iMGs.

“HET iMGs did not show a difference in the expression of either gene at baseline relative to WT iMGs.”

Cluster 4 shows minor expansion in SHIP1-deficient microglia.

“The minor expansion of this cluster in SHIP1-deficient microglia”

The top protein marker of cluster 2 is the Fc gamma receptor CD64.

“the top protein marker of cluster 2 is the Fc gamma receptor CD64.”

Cathepsin B activity is unchanged in HET BR24 iMGs.

“it was unchanged in HET BR24 iMGs”

A higher percentage of HET iMGs are positive for Gamillus compared to WT iMGs.

“We observed a higher percentage of HET iMGs positive for Gamillus”

SIRP-A is the strongest protein marker of cluster 1.

“SIRP-A is the strongest protein marker”

The increased synaptosome uptake by HET iMGs is blocked by treatment with the actin polymerization inhibitor cytochalasin D.

“which was blocked by treatment with the actin polymerization inhibitor cytochalasin D (Figures 6A, S6D).”

The percentage of HET iMGs positive for pHrodo remained higher than WT iMGs in the presence of bafilomycin A1.

“The percentage of HET iMGs positive for pHrodo remained higher than WT iMGs in the presence bafilomycin A1 (Figure 6B)”

Cluster 3 shows enrichment for inflammatory signaling-related GO terms.

“shows enrichment for inflammatory signaling-related GO terms.”

Cluster 0 is reduced in HET iMGs.

“Cluster 0, which is reduced in HET iMGs”

Cluster 2 is enriched in canonical DAM markers such as APOE, SPP1 and TREM2.

“is enriched in canonical disease-associated microglia (DAM) markers such as APOE, SPP1 and TREM2”

LPS treatment increases expression of IL1B, CCL2, and IL6 in WT iMGs.

“while LPS treatment resulted in increased expression of the three genes in WT iMGs as expected”

6-hour treatment of BR33 iMGs with 3AC does not affect cell viability.

“while not affecting cell viability”

HET iMGs have higher average Gamillus fluorescence compared to WT iMGs.

“as well as higher average Gamillus fluorescence in HET iMGs compared to WT iMGs”

6-hour treatment of BR33 iMGs with 3AC dose-dependently reduces LysoTracker intensity.

“6-hour treatment of BR33 iMGs with the SHIP1-specific inhibitor 3AC41 dose-dependently reduced LysoTracker intensity”

Cluster 1 is proportionally similar across WT and HET iMGs.

“cluster 1, which is proportionally similar across WT and HET iMGs”

Antibody-mediated blocking of TREM2 reduces the uptake of synaptosomes and apoptotic neurons by HET iMGs.

“Antibody-mediated blocking of TREM2 reduced the uptake of synaptosomes and ANs by HET iMGs”

Cluster 0 shows enrichment for MHC-related GO terms.

“shows enrichment (adjusted p-value < 0.05) for MHC-related GO terms”

CA-074-Me pre-treatment rescues 3AC-induced IL-1β and IL-18 secretion in a dose-dependent manner.

“CA-074-Me pre-treatment rescued the 3AC-induced IL-1β and IL-18 secretion in a dose-dependent manner”

Several phagocytic receptors, such as FCER1A, CD32/FCGR2, CD11b and CD33, are significantly upregulated at the cell surface in HET iMGs within cluster 2.

“Several phagocytic receptors, such as FCER1A, CD32/FCGR2, CD11b and CD33 were significantly upregulated at the cell surface in HET iMGs within cluster 2”

WT and HET iMGs cluster compositions were significantly shifted.

“the cluster compositions were significantly shifted”

HET iMGs show a similar increase in the percentage positive for pHrodo when taking up apoptotic neurons.

“we also tested the uptake of pHrodo-labeled apoptotic neurons (ANs) and found a similar increase in the percentage of iMGs positive for pHrodo (Figures 6C, S6E).”

The top protein marker for cluster 0 is CD1c.

“the top protein marker for cluster 0 is the antigen-presenting protein CD1c”

The ratio of cathepsin B to LAMP1 is trending elevated in HET BR33 iMGs.

“while it was trending in HET BR33 iMGs”

In HET BR24 iMGs, cathepsin B intensity is significantly increased.

“a significant increase in cathepsin B intensity in HET BR24 iMGs”

Cluster 3 is strongly diminished in HET iMGs.

“Cluster 3, which is strongly diminished in HET iMGs”

Clearance of extracellular Aβ is not affected by the presence of HET compared to WT iMGs.

“clearance of extracellular Aβ was not affected by the presence of HET compared to WT iMGs (Figures 7F-G).”

In INPP5D haploinsufficient iMGs, cathepsin B levels and enzymatic activity appear higher than expected.

“In INPP5D haploinsufficient iMGs, cathepsin B levels and enzymatic activity appear higher than expected, while cathepsin B± puncta, which indicate lysosomally localized cathepsin B, are reduced (Figures 4A-E).”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

INPP5D haploinsufficiency does not affect the fluorescence intensity of Aβ within iMGs.

“or the fluorescence intensity of Aβ within iMGs (Figures 7A-B, S7A).”

HET iMGs from both genetic backgrounds display higher FITC fluorescence 24 hours after washout compared to WT iMGs.

“HET iMGs from both genetic backgrounds displayed higher FITC fluorescence 24 hours after washout compared to WT iMGs”

SHIP1-deficient BR33 iMGs displayed increased GPNMB secretion.

“they displayed increased GPNMB secretion (Figures 3E,G)”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

SHIP1 deficiency leads to an overall reduction of lysosomal degradation of internalized material in microglia.

“leads to an overall reduction of lysosomal degradation of internalized material (Figures 2B, S2C, 8)”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

INPP5D haploinsufficiency does not affect the percentage of iMGs positive for Aβ.

“INPP5D haploinsufficiency did not affect the percentage of iMGs positive for Aβ”

Inhibition of cathepsin B activity with CA-074-Me rescued the NLRP3 inflammasome activation upon SHIP1 inhibition.

“Indeed, inhibition of cathepsin B activity with CA-074-Me rescued the NLRP3 inflammasome activation upon SHIP1 inhibition (Figure 4F), supporting a role for cathepsin B in mediating the NLRP3 inflammasome activation downstream of SHIP1 deficiency in microglia (Figure 8).”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.
• AssumptionRests on an assumption: CA-074-Me specifically inhibits cathepsin B activity in iMGs.

fAD iNs secrete an increased ratio of Aβ42/Aβ40 compared to paired isogenic WT iNs.

“fAD iNs secreted an increased ratio of Aβ42/Aβ40, compared to paired isogenic WT iNs, as expected (Figures 7C-D, S7B).”

Acute SHIP1 inhibition with 3AC robustly destabilizes lysosomes in BR33 iMGs.

“we tested the effect of acute SHIP1 inhibition with 3AC and found that it robustly destabilizes lysosomes in BR33 iMGs (Figure 3H).”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.
• AssumptionRests on an assumption: Acute SHIP1 inhibition with 3AC specifically inhibits SHIP1 in iMGs.

SHIP1-deficient BR33 iMGs did not show reduced lysosome acidification.

“While SHIP1-deficient BR33 iMGs did not show reduced lysosome acidification, they displayed increased GPNMB secretion (Figures 3E,G), which indicates a response to lysosomal stress to restore lysosomal acidification39.”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

SHIP1 deficiency impairs early endosome maturation in microglia.

“impairs early endosome maturation (Figures 2G, S2E)”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

SHIP1 binds to these and other adaptor proteins in human microglia.

“We confirmed SHIP1’s binding to these and other adaptor proteins in human microglia”

WT iMGs co-cultured with fAD iNs have reduced SHIP1 levels compared to those co-cultured with WT iNs.

“WT iMGs co-cultured with fAD iNs had reduced SHIP1 levels compared to those co-cultured with WT iNs”

SHIP1 deficiency reduces the number of Rab5+ early endosomes in microglia.

“it reduces the number of Rab5+ early endosomes (Figures 2E-F, S2D)”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

Aβ accumulation over 24 hours induces significant lipid droplet accumulation in HET iMGs but not WT iMGs.

“Moreover, Aβ accumulation over 24 hours induced significant lipid droplet accumulation in HET, but not WT iMGs (Figure 7L).”

In INPP5D haploinsufficient iMGs, cathepsin B± puncta, which indicate lysosomally localized cathepsin B, are reduced.

“cathepsin B± puncta, which indicate lysosomally localized cathepsin B, are reduced (Figures 4A-E)”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

SHIP1 deficiency in microglia does not affect endocytic uptake.

“While SHIP1 deficiency in microglia does not affect endocytic uptake (Figures SB, S2B)”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

SHIP1-deficient BR24 iMGs had deacidified lysosomes.

“In line with the second possibility, we found that SHIP1-deficient BR24 iMGs had deacidified lysosomes (Figure 3E).”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

DiI staining revealed a lower number of dendritic spines in iNs co-cultured with HET iMGs.

“Moreover, DiI staining revealed a lower number of dendritic spines in iNs co-cultured with HET iMGs (Figures 6K-L).”

SHIP2 is another novel binding partner of endogenous SHIP1 in microglia.

“We identified SHIP2, the main SHIP paralog expressed in non-hematopoietic cells, as another novel binding partner of endogenous SHIP1 in microglia (Figures 1B,F-G).”

Antibody-mediated inhibition of TREM2 reduced the enhanced phagocytosis in SHIP1-deficient iMGs.

“Antibody-mediated inhibition of TREM2 reduced the enhanced phagocytosis in SHIP1-deficient iMGs (Figures 6F-GA, S6E)”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.
• AssumptionRests on an assumption: Antibody-mediated inhibition of TREM2 specifically blocks TREM2 signaling in iMGs.

Excessive release of the proinflammatory cytokines IL-1β and IL-18 can be prevented by pharmacological inhibition of cathepsin B.

“Importantly, we show that the excessive release of the proinflammatory cytokines IL-1β and IL-18 can be prevented by pharmacological inhibition of cathepsin B (Figure 4F), which opens a therapeutic avenue for mitigating the consequences of microglial neuroinflammation upon SHIP1 deficiency.”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.
• AssumptionRests on an assumption: CA-074-Me specifically inhibits cathepsin B activity in iMGs.

SHIP1 deficiency causes aberrant synapse engulfment.

“These include aberrant synapse engulfment (Figures 6A-B,I-L), intracellular accumulation of Aβ due to impaired lysosomal degradation (Figures 7I-K), excessive lipid droplet formation at baseline that is exacerbated by Aβ accumulation (Figures 3A, 7M, S3A), and transcriptional downregulation of immune response programs that render microglia insensitive to inflammatory stimuli (Figures 5E,H,I-K, S5E”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

Exposure to fAD neurons reduces SHIP1 levels in microglia.

“We show that exposure to fAD neurons reduces SHIP1 levels in microglia (Figure 7E), and that SHIP1 deficiency recapitulates a number of microglial phenotypes closely associated with AD pathology.”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

The paper did not find evidence supporting a role for SHIP1 in regulating either synthetic or cell-derived Aβ uptake in human iMGs.

“We did not find evidence supporting a role for SHIP1 in regulating either synthetic or cell-derived Aβ uptake in human iMGs.”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

SHIP1 deficiency causes intracellular accumulation of Aβ due to impaired lysosomal degradation.

“intracellular accumulation of Aβ due to impaired lysosomal degradation (Figures 7I-K)”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

SHIP1 deficiency causes transcriptional downregulation of immune response programs that render microglia insensitive to inflammatory stimuli.

“transcriptional downregulation of immune response programs that render microglia insensitive to inflammatory stimuli (Figures 5E,H,I-K, S5E-F)”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

SHIP1 deficiency enhances the uptake of synaptic material and apoptotic neurons, both lipid-rich substrates, in human microglia.

“In line with these findings, we found that SHIP1 deficiency enhances the uptake of synaptic material and apoptotic neurons, both lipid-rich substrates (Figures 6A-C,I-L, S6D) in human microglia (Figure 8).”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

SHIP1 deficiency causes excessive lipid droplet formation at baseline.

“excessive lipid droplet formation at baseline that is exacerbated by Aβ accumulation (Figures 3A, 7M, S3A)”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

SHIP1-deficient microglia exhibit exaggerated immune response through NLRP3 inflammasome activation in the absence of external stimuli.

“In parallel, SHIP1-deficient microglia exhibit exaggerated immune response through NLRP3 inflammasome activation in the absence of external stimuli, a process we identify as being driven by endo-lysosomal dysfunction.”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

Aβ accumulation exacerbates excessive lipid droplet formation.

“excessive lipid droplet formation at baseline that is exacerbated by Aβ accumulation (Figures 3A, 7M, S3A)”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

SHIP1 deficiency compromises Rab5-mediated early endosomal maturation and acidification.

“These results suggest that SHIP1 deficiency compromises Rab5-mediated early endosomal maturation and acidification”

The effect of INPP5D haploinsufficiency on LAMP1 differs between BR24 and BR33 genetic backgrounds.

“indicating a difference across these two genetic backgrounds”

SHIP1 deficiency potentially leads to impaired delivery of internalized cargo for lysosomal degradation.

“potentially leading to impaired delivery of internalized cargo for lysosomal degradation”

SHIP1 is degraded via autophagy.

“This result suggests that SHIP1 is degraded via autophagy”

SHIP1 deficiency impairs degradation of both intracellular and extracellular cargo.

“These results suggest that SHIP1 deficiency impairs the degradation of both intracellular and extracellular cargo.”

Reduction of SHIP1 levels may increase accumulation of SHIP1 in lysosomes due to impaired autophagy.

“reduction of SHIP1 levels may increase the accumulation of SHIP1 in lysosomes due to an impairment in autophagy”

SHIP1 regulates endo-lysosome function via interactions with endo-lysosomal proteins.

“where it regulates endo-lysosome function via interactions with endo-lysosomal proteins (Figure 1E)”

The absolute SHIP1 protein levels or extent of SHIP1 deficiency in BR33 iMGs may not be sufficient to trigger lysosome dysfunction.

“Therefore, the absolute SHIP1 protein levels or the extent of SHIP1 deficiency in BR33 iMGs may not be sufficient to trigger lysosome dysfunction.”

Genetic background can modify the effect of SHIP1 deficiency on lysosome number and acidification.

“Therefore, genetic background can modify the effect of SHIP1 deficiency on lysosome number and acidification.”

SHIP1 may localize to lysosomes.

“Therefore, SHIP1 may localize to lysosomes”

SNPs at the INPP5D locus have been identified that are associated with AD.

“Single nucleotide polymorphisms (SNPs) at the INPP5D locus have been identified that are associated with AD.”

SHIP1 deficiency may lead to a shift towards recycling of cargo from trafficking to lysosomes.

“These results suggest that SHIP1 deficiency may lead to a shift towards recycling of cargo from trafficking to lysosomes”

INPP5D loss-of-function suggests broad lysosome dysfunction.

“suggesting broad lysosome dysfunction”

INPP5D haploinsufficiency in microglia induces a shift from an immune responsive state to a DAM-like, potentially hyper-phagocytic state.

“Taken together, these findings suggest that INPP5D haploinsufficiency in microglia induces a shift from an immune responsive state to a DAM-like, potentially hyper-phagocytic state.”

• AssumptionRests on an assumption: The observed phenotypes in HET iMGs are caused by reduced SHIP1 function rather than other genetic or off-target effects.

Cathepsin B activity was higher than expected in HET iMGs.

“cathepsin B activity was higher than expected in HET iMGs”

The reduction of cathepsin B puncta area despite elevated overall protein levels and enzymatic activity may indicate a cytosolic localization for cathepsin B in HET iMGs.

“Therefore, the reduction of cathepsin B puncta area despite elevated overall protein levels and enzymatic activity may indicate a cytosolic localization for cathepsin B in HET iMGs.”

Cluster 1 represents homeostatic microglia.

“suggesting that homeostatic microglia make up cluster 1”

INPP5D haploinsufficiency in iMGs increases synaptosome uptake even with induced lysosomal degradation impairment.

“suggesting that INPP5D haploinsufficiency in iMGs increases synaptosome uptake even with induced lysosomal degradation impairment.”

BR33 iMGs may have successfully restored lysosomal homeostasis to some degree.

“These results suggest that BR33 iMGs may have successfully restored lysosomal homeostasis to some degree.”

INPP5D haploinsufficiency enhances the uptake of multiple lipid-rich substrates.

“These results suggest that INPP5D haploinsufficiency enhances the uptake of multiple lipid-rich substrates.”

The expansion of cluster 2 has the potential to influence the entire microglial population in a non-autonomous manner.

“suggesting that the expansion of this cluster has the potential to influence the entire population in a non-autonomous manner.”

• AssumptionRests on an assumption: Secreted proteins such as SPP1 and APOE can influence other microglia in a non-autonomous manner.

Cluster 0 represents antigen-presenting microglia.

“suggesting that this subcluster represents antigen-presenting microglia”

SHIP1 activity regulates lysosome homeostasis.

“Together, these data support a role for SHIP1 activity in regulating lysosome homeostasis.”

Increased TREM2 signaling is one mechanism that mediates the enhanced phagocytosis phenotype in HET iMGs.

“suggesting that the increased TREM2 signaling is one mechanism that mediates this phenotype (Figures 6F-G).”

Cluster 4 likely represents proliferative microglia.

“likely represents proliferative microglia.”

SHIP1 deficiency impairs NF-κB-dependent response to LPS in microglia.

“Therefore, SHIP1 deficiency impairs NF-κB-dependent response to LPS in microglia.”

• AssumptionRests on an assumption: IL1B, CCL2, and IL6 are NF-κB-dependent genes.

These data do not suggest a role for SHIP1 in regulating Aβ uptake in this context.

“Taken together, these data do not suggest a role for SHIP1 in regulating Aβ uptake in this context.”

SHIP1 deficiency can trigger lysosomal deacidification.

“Therefore, SHIP1 deficiency can trigger lysosomal deacidification, which may elicit compensatory lysosomal stress responses such as GPNMB upregulation, and the degree of efficacy of this response is sensitive to genetic variation.”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

SHIP1 regulates Rab5+ early endosome numbers and maturation potentially via its interaction with CapZ.

“potentially via its interaction with CapZ.”

Exposure to fAD iNs leads to downregulation of SHIP1 in iMGs.

“suggesting that exposure to fAD iNs leads to downregulation of SHIP1 in iMGs (Figure 7E).”

Excess lipid droplet accumulation potentially exacerbates neuroinflammation.

“thereby potentially exacerbating neuroinflammation.”

SHIP1 deficiency may elicit compensatory lysosomal stress responses such as GPNMB upregulation.

“which may elicit compensatory lysosomal stress responses such as GPNMB upregulation”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

INPP5D haploinsufficiency enhances the engulfment of synaptic material from co-cultured neurons.

“indicating that INPP5D haploinsufficiency enhances the engulfment of synaptic material from co-cultured neurons (Figures 6H-J).”

There is intracellular accumulation of Aβ in HET iMGs.

“suggesting an intracellular accumulation of Aβ (Figures 7I-K, S7C).”

SHIP1-SHIP2 complexes may regulate endo-lysosomal function through a combination of interactions with other proteins (such as CapZ) and their shared phosphatase activity.

“This complex may regulate endo-lysosomal function through a combination of interactions with other proteins (such as CapZ) and their shared phosphatase activity.”

SHIP1 deficiency in microglia induces excessive engulfment of synapses.

“it also induces excessive engulfment of synapses.”

Novel binding partners of SHIP1 point to a role for SHIP1 in regulating the endo-lysosomal system.

“identified novel binding partners that pointed to a role for SHIP1 in regulating the endo-lysosomal system (Figures 1C-E).”

Lysosomal membrane permeabilization and leakage of cathepsin B into the cytosol may precede this putative compensatory response.

“Nonetheless, lysosomal membrane permeabilization and leakage of cathepsin B into the cytosol may precede this putative compensatory response.”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

SHIP1 and SHIP2 share highly conserved SH2, 5′-phosphatase, and proline-rich domains, providing a structural basis for formation of a SHIP1-SHIP2 complex.

“Although SHIP1 and SHIP2 have divergent C-terminal domains, they share highly conserved SH2, 5′-phosphatase, and proline-rich domains, providing a structural basis for formation of a SHIP1-SHIP2 complex.”

Internalized materials are increasingly recycled in SHIP1 deficient microglia due to defects in endosome maturation.

“Therefore, it is plausible that internalized materials are increasingly recycled in SHIP1 deficient microglia due to defects in endosome maturation.”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

The degree of efficacy of the compensatory lysosomal stress response is sensitive to genetic variation.

“the degree of efficacy of this response is sensitive to genetic variation”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

SHIP1 deficiency in microglia enhances the clearance of cell debris.

“Therefore, while SHIP1 deficiency in microglia enhances the clearance of cell debris”

SHIP1 deficiency recapitulates a number of microglial phenotypes closely associated with AD pathology.

“We show that exposure to fAD neurons reduces SHIP1 levels in microglia (Figure 7E), and that SHIP1 deficiency recapitulates a number of microglial phenotypes closely associated with AD pathology.”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

Cathepsin B mediates NLRP3 inflammasome activation downstream of SHIP1 deficiency in microglia.

“supporting a role for cathepsin B in mediating the NLRP3 inflammasome activation downstream of SHIP1 deficiency in microglia (Figure 8)”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.
• AssumptionRests on an assumption: CA-074-Me specifically inhibits cathepsin B activity in iMGs.

SHIP1 deficiency has been associated with increased phagocytosis of various substrates in other models.

“However, SHIP1 deficiency has been associated with increased phagocytosis of various substrates in other models73,74.”

• cites [73] sourceNot checked — no full text of the source
• cites [74] sourceNot checked — no full text of the source

Loss of SHIP1 function is a key molecular event that drives the convergence of multiple pathogenic microglial phenotypes observed in AD.

“Taken together, our findings point to loss of SHIP1 function as a key molecular event that drives the convergence of multiple pathogenic microglial phenotypes observed in AD.”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

The study identifies SHIP1 as a central regulator of convergent microglial mechanisms of AD pathogenesis.

“In this study, we identify SHIP1 as a central regulator of convergent microglial mechanisms of AD pathogenesis.”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

The exaggerated immune response through NLRP3 inflammasome activation in SHIP1-deficient microglia is driven by endo-lysosomal dysfunction.

“In parallel, SHIP1-deficient microglia exhibit exaggerated immune response through NLRP3 inflammasome activation in the absence of external stimuli, a process we identify as being driven by endo-lysosomal dysfunction.”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.

SHIP1 deficiency may promote the clearance of Aβ-lipoprotein complexes in the brain more efficiently than free, unbound Aβ.

“Therefore, SHIP1 deficiency may promote the clearance of Aβ-lipoprotein complexes in the brain more efficiently than free, unbound Aβ.”

• AssumptionRests on an assumption: iPSC-derived microglia (iMGs) are a valid model of human microglia for studying AD-related endo-lysosomal and phagocytic functions.