Journal: Acta Neuropathologica

Article Title: Alteration of gene expression and protein solubility of the PI 5-phosphatase SHIP2 are correlated with Alzheimer’s disease pathology progression

doi: 10.1007/s00401-024-02745-7

Figure Lengend Snippet: Gene expression of EGFR and INPPL1 is upregulated in AD brains. a-b A significant increase of normalized transcripts of EGFR and INPPL1 was observed in AD brains of Mayo clinic TCX cohort (Braak 0-II n = 42, Braak III-IV n = 36, Braak V-VI n = 82). Normalized gene expression of EGFR ( a ) and INPPL1 ( b ) was significantly increased in AD brains. **** p < 0.0001, *p < 0.05 for EGFR and ** p < 0.01 for INPPL1 by one-way ANOVA test with a Tukey’s multiple-comparisons post hoc test. 100% was given to the average of normalized mRNA level in the control with Braak 0-II

Article Snippet: Mouse monoclonal anti-SHIP2 (3E6) antibody was purchased from Novus (H00003636-M01), Cambridge, UK.

Techniques: Gene Expression, Control

Journal: Acta Neuropathologica

Article Title: Alteration of gene expression and protein solubility of the PI 5-phosphatase SHIP2 are correlated with Alzheimer’s disease pathology progression

doi: 10.1007/s00401-024-02745-7

Figure Lengend Snippet: SHIP2 and EGFR proteins are increased and co-enriched in the RIPA-insoluble fraction of AD brains. a In the total homogenate of human brain T1 isocortex, normalized EGFR and SHIP2 protein levels were not significantly altered in the T1 isocortex brain lysates of AD compared to those of control non-demented. b In the RIPA-soluble fraction of AD brains, SHIP2 protein level was largely reduced. In the same fraction, EGFR protein level was not significantly altered in AD brains compared to control. c Both EGFR and SHIP2 protein levels were increased in RIPA-insoluble fraction of AD brains. d-f Spearman non-parametric analyses showed significant correlations between EGFR and SHIP2 protein levels (Spearman r = 0.5657, **** p < 0.0001) ( d ), between pTau (measured using PHF-1 antibody) and EGFR (Spearman r = 0.4666, **** p < 0.0001) ( e ) and pTau and SHIP2 (Spearman r = 0.2389, * p = 0.0307) ( f ) in the insoluble fraction. T1 isocortex from control of Braak 0-II ( n = 24), Braak III-IV ( n = 7) and AD ( n = 51) including 2 FAD cases with APP or PSEN1 mutation were analysed. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001 by one-way ANOVA test with a Tukey’s multiple-comparisons post hoc test. ( a-c ) or by Spearman non-parametric correlation test ( d-f ). 100% was given to the average of the control non-demented group data normalized to GAPDH

Article Snippet: Mouse monoclonal anti-SHIP2 (3E6) antibody was purchased from Novus (H00003636-M01), Cambridge, UK.

Techniques: Control, Mutagenesis

Journal: Acta Neuropathologica

Article Title: Alteration of gene expression and protein solubility of the PI 5-phosphatase SHIP2 are correlated with Alzheimer’s disease pathology progression

doi: 10.1007/s00401-024-02745-7

Figure Lengend Snippet: SHIP2 immunoreactive structures are increased in AD brains and partially associated with amyloid plaques. a-b Representative images of immunostaining of SHIP2 using 3E6 antibody in CA1-2 hippocampal area of a control ( a ) and an AD ( b ) cases. Black arrows show 3E6-positive structures. c-d The average size ( c ) and percentage of the area occupied by SHIP2-immunolabelling ( d ) were significantly increased in the CA1-2 of AD brains compared to controls. e–f, h-i Representative images of immunostaining of anti-Aβ 6E10 ( e–f ) and anti-pTau AT8 ( h-i ) antibodies in CA1-2 hippocampal area of a control ( e, h ) and an AD ( f, i ) cases. g, j There were significant correlations between SHIP2 positive area and amyloid load ( g ) or tau load ( j ). * p < 0.05 and ** p < 0.01 by Mann–Whitney U test ( c, d ) or by Spearman non-parametric correlation analyses ( g, j ). k-n. A double immunostaining for SHIP2 ( k , green ) and Aß42 ( l , red ) suggested that SHIP2 immunoreactivity was partly associated with amyloid plaques. Mature dense-core plaque (arrowhead) showed accumulation of SHIP2-immunolabelled structure while diffuse plaque (asterisk) showed a weak SHIP2 immunoreactivity ( m merge ). Scale bars 20 μm. n Colour profile of the double staining of SHIP2 and Aß42 shows partial association of SHIP2 staining with amyloid plaque (green and red, respectively)

Article Snippet: Mouse monoclonal anti-SHIP2 (3E6) antibody was purchased from Novus (H00003636-M01), Cambridge, UK.

Techniques: Immunostaining, Control, MANN-WHITNEY, Double Immunostaining, Double Staining, Staining

Journal: Acta Neuropathologica

Article Title: Alteration of gene expression and protein solubility of the PI 5-phosphatase SHIP2 are correlated with Alzheimer’s disease pathology progression

doi: 10.1007/s00401-024-02745-7

Figure Lengend Snippet: SHIP2 immunoreactivity is increased in the presence of amyloid pathology in the human brain of related neurodegenerative diseases. a-l Representative photos of immunostaining of SHIP2 in CA1-2 area of a case of Braak III-IV, Thal 2–3 ( a ), DS/AD (Braak VI, Thal 5) ( b ), CAA (Braak II, Thal 1) ( c ), LBD/AD (Braak VI, Thal 4) ( d ), DLBD without AD lesion (Braak I, Thal 0) ( e ), FTD-U with TDP-43 accumulation due to PRG mutation ( f ), PSP ( g ), CBD ( h ) Pick disease ( i ) and FTD-tau with MAPT mutation of P301L ( j ), L266V ( k ) and G335A ( l ). There was a global correlation between the levels of amyloid load and SHIP2. Primary tau pathology such as MAPT- G335A did not lead to an increased SHIP2 immunoreactivity. CA1-2 area of hippocampus was analysed for a - f , i - l , striatum was analysed for PSP cases and temporal cortex was analysed for CBD cases. Scale bar 20 μm

Article Snippet: Mouse monoclonal anti-SHIP2 (3E6) antibody was purchased from Novus (H00003636-M01), Cambridge, UK.

Techniques: Immunostaining, Mutagenesis

Journal: Acta Neuropathologica

Article Title: Alteration of gene expression and protein solubility of the PI 5-phosphatase SHIP2 are correlated with Alzheimer’s disease pathology progression

doi: 10.1007/s00401-024-02745-7

Figure Lengend Snippet: Increased SHIP2 immunoreactivity is associated with amyloid but not with tau pathology. a-d Representative images of SHIP2 immunolabelling in the brain stem of 10-month-old male WT ( a ), Tg30 ( b ), 5xFAD ( c ) and 5xFAD X Tg30 ( d ) mice. There was a stronger SHIP2 labelling in 5xFAD and 5xFAD X Tg30 mouse brains. e–f Thresholding measurement of the average size ( e ) and area ( f ) immunostained by SHIP2 in the brain stem of WT, Tg30, 5xFAD and 5xFAD X Tg30 mice. The average size and percentage of the area occupied by 3E6 staining were increased in the presence of amyloid pathology but not of tau pathology. * p < 0.05 and ** p < 0.01 by * p < 0.05, ** p < 0.01 and *** < 0.001 by one-way ANOVA with Tukey post-hoc test. (WT: n = 5, Tg30: n = 5, 5xFAD: n = 4, 5xFAD X Tg30: n = 3). g-i A double immunostaining for SHIP2 ( g , green ) and Aß42 ( h , red ) suggested that SHIP2 immunoreactivity was detected in the brain stem of 5xFAD mouse brains. Merged image is shown in i . j Color profile of the double staining of SHIP2 and Aß42 shows partial association of SHIP2 staining with amyloid plaque (green and red, respectively). Scale bars 20 μm.

Article Snippet: Mouse monoclonal anti-SHIP2 (3E6) antibody was purchased from Novus (H00003636-M01), Cambridge, UK.

Techniques: Staining, Double Immunostaining, Double Staining

Journal: Acta Neuropathologica

Article Title: Alteration of gene expression and protein solubility of the PI 5-phosphatase SHIP2 are correlated with Alzheimer’s disease pathology progression

doi: 10.1007/s00401-024-02745-7

Figure Lengend Snippet: SHIP2 immunoreactive structures were co-stained with plaque-associated tau and astrocytes in AD brains. a-c SHIP2 ( a , green) and tau ( b , red) were partially co-stained in the AD hippocampus. d-f SHIP2 ( d ) and astrocyte marker AQP1 ( e ) were partially co-labelled. g-i SHIP2 ( g ) and microglial marker Iba1 ( h ) were not clearly co-labelled. j-l SHIP2 ( j ) and oligodendrocyte marker MBP ( k ) were not clearly co-labelled. Merged images show DAPI counterstaining in blue ( c , f , i , l ). Scale bar 20 μm

Article Snippet: Mouse monoclonal anti-SHIP2 (3E6) antibody was purchased from Novus (H00003636-M01), Cambridge, UK.

Techniques: Staining, Marker

Journal: Acta Neuropathologica

Article Title: Alteration of gene expression and protein solubility of the PI 5-phosphatase SHIP2 are correlated with Alzheimer’s disease pathology progression

doi: 10.1007/s00401-024-02745-7

Figure Lengend Snippet: SHIP2 overexpression led to increased FRET signal while siRNA-mediated SHIP2 downexpression led to decreased FRET signal in Tau RD P301S FRET biosensor cells transduced with AD-PHF. a-d . Representative image of tau RD P301S FRET biosensor cells fixed 48 h after transduction of AD-PHF and plasmid, immunostained for SHIP2 and counterstained with DAPI. Co-transduction of sarkosyl insoluble fraction from a control case with an empty vector ( a ) or a plasmid encoding SHIP2 ( b ) did not lead to a FRET positive tau inclusion. Transducing AD-PHF (sarkosyl insoluble fraction from an AD brain) led to FRET-positive tau inclusion in this model ( c , d ). The cells were co-transduced with AD-PHF and either empty vector ( c ) or SHIP2 plasmid ( d ). e–f. SHIP2 overexpression was verified by WB ( e ). There was a significant increase in FRET positive signal when AD-PHF and SHIP2 were co-transduced ( f ). g-j Tau RD P301S FRET biosensor cells were co-transduced with AD-PHF and either with a control scramble siRNA (SiC) ( g ) or siRNA targeting INPPL1 ( h ). There was a significant decrease of SHIP2 expression when siRNA-INPPL1 was transduced ( i ). INPPL1 downexpression led to a decreased FRET signal ( j ). f , j . 10,000 cells per experiment were analysed by FRET flow cytometry and the results were shown as integrated mean fluorescence intensity. Four independent experiments were carried out for overexpression and downexpression of SHIP2, respectively. ** < 0.01 and *** < 0.001 by two-way ANOVA. Scale bar 20 μm

Article Snippet: Mouse monoclonal anti-SHIP2 (3E6) antibody was purchased from Novus (H00003636-M01), Cambridge, UK.

Techniques: Over Expression, Transduction, Plasmid Preparation, Control, Expressing, Flow Cytometry, Fluorescence

Journal: Acta Neuropathologica

Article Title: Alteration of gene expression and protein solubility of the PI 5-phosphatase SHIP2 are correlated with Alzheimer’s disease pathology progression

doi: 10.1007/s00401-024-02745-7

Figure Lengend Snippet: INPPL1 locus regional plots of AD GWAS, CSF Aß42 GWAS, and CSF pTau GWAS. The association of GWAS variants within 1 Mb of INPPL1 gene coordinates (hg38 chr11:71,224,767–73,239,147) with AD risk (upper panel, from Bellenguez et al. ), with CSF Aß42 (middle panel, from Jansen et al. ) and with CSF pTau (lower panel, from Jansen et al. ). The top-associated variants for AD risk (rs35404771) and for CSF pTau (rs891322 and rs11235462) independent association signal peaks are labelled on the figure, where their proxy LD patterns are shown in yellow, blue, and green respectively. The colours with higher opacity are indicative for higher LD r 2 values. Missing variants in the LD reference panel (1 KG non-Finnish European samples, n = 404) are shown in grey. y axis, − log 10 GWAS p ; x axis, hg38 genomic position on chromosome 11

Article Snippet: Mouse monoclonal anti-SHIP2 (3E6) antibody was purchased from Novus (H00003636-M01), Cambridge, UK.

Techniques:

Journal: Acta Neuropathologica

Article Title: Alteration of gene expression and protein solubility of the PI 5-phosphatase SHIP2 are correlated with Alzheimer’s disease pathology progression

doi: 10.1007/s00401-024-02745-7

Figure Lengend Snippet: Regulation of INPPL1 gene expression by the CSF-pTau-associated variants within INPPL1 locus. a The association of GWAS and eQTL variants within 250 kb of INPPL1 gene coordinates (hg38 chr11:71,974,767–72,489,147) with CSF pTau (upper panel, from ) and with INPPL1 gene expression in brain (lower panel, ROSMAP DLPFC). The top-associated variants for CSF pTau (rs891322 and rs11235462), for INPPL1 eQTLs (rs551220731) and for both (rs11235459) independent association signal peaks are labelled on the figure, where their proxy LD patterns are shown in blue, green, yellow, and pink respectively. The colours with higher opacity are indicative for higher LD r 2 values. Missing variants in the LD reference panel (1 KG non-Finnish European samples, n = 404) are shown in grey. y axis, − log 10 GWAS p ; x axis, hg38 genomic position on chromosome 11. b GWAS-eQTL comparison plot for correlation between ROSMAP DLPFC eQTLs for INPPL1 gene expression and CSF pTau GWAS. rs11235459 is labelled on the plot, and its LD with other variants are displayed in a colour scale as shown in the legend. Missing variants in the LD reference panel (1 KG non-Finnish European samples, n = 404) are shown in grey. y axis, − log 10 eQTL p ; x axis, − log 10 GWAS p

Article Snippet: Mouse monoclonal anti-SHIP2 (3E6) antibody was purchased from Novus (H00003636-M01), Cambridge, UK.

Techniques: Gene Expression, Comparison