Journal: International Journal of Molecular Sciences

Article Title: GM1 Oligosaccharide Modulates Microglial Activation and α-Synuclein Clearance in a Human In Vitro Model

doi: 10.3390/ijms26157634

Figure Lengend Snippet: Effects of GM1 oligosaccharide (OligoGM1) application on Human embryonic microglial (HCM3) cells. After 2 days of culture (80% of confluence), HCM3 cells were incubated or not incubated (control, CTRL) with OligoGM1 (100 µM). After 48 h, immunofluorescence staining [(against nuclei, ionized calcium-binding adaptor molecule 1 (Iba1) and triggered receptor expressed on myeloid cells 2 (TREM2)] and ELISA for quantification of released cytokines to medium were performed, as described in the : ( a ) Representative immunofluorescence images (20× magnification, scale bar 100 µm) and number of DAPI-positive cells; ( b ) Representative immunofluorescence images (20× magnification, scale bar 100 µm) of Iba1 expression, quantification of Iba1 accumulation (Iba1 area normalized by the number of cells), and area of Iba1-positive cells (area of microglia cells normalized by the µm 2 of Iba1); ( c ) Representative immunofluorescence images (20× magnification, scale bar 100 µm) of TREM2 expression, quantification of TREM2-positive cells (TREM2 area normalized by the number of cells), and area of TREM2-positive cells (area of microglia cells normalized by the µm 2 of TREM2); ( d ) Quantification of released cytokines tumor necrosis factor alpha (TNF-α) and interleukin 6 (IL-6). All values are represented as percentage versus CTRL and expressed as mean ± standard error of the mean (SEM, n = 6; ns = not significant, Mann–Whitney test).

Article Snippet: For immunofluorescence analyses, the following antibodies were used: primary mouse monoclonal anti-TREM2 antibody [Cat. 11084-MM08, research resource identifier (RRID):AB_2860315], purchased from Sino Biological Europe GmbH, Europe (Düsseldorfer, Germany); primary rabbit polyclonal anti-αSyn antibody (Cat. 2642S, RRID not available), purchased from Ozyme, Paris, France; and primary goat polyclonal anti-Iba1 antibody (Cat. Ab5076, RRID:AB_2224402), purchased from Abcam, Cambridge, UK.

Techniques: Incubation, Control, Immunofluorescence, Staining, Binding Assay, Enzyme-linked Immunosorbent Assay, Expressing, MANN-WHITNEY

Journal: International Journal of Molecular Sciences

Article Title: GM1 Oligosaccharide Modulates Microglial Activation and α-Synuclein Clearance in a Human In Vitro Model

doi: 10.3390/ijms26157634

Figure Lengend Snippet: OligoGM1 modulation of αSyn-induced activation of HMC3 cells. After 2 days of culture (80% of confluence), HCM3 cells were incubated or not incubated (CTRL) with OligoGM1 (100 µM). After 4 h, αSyn (1 µM) was added to the culture medium for 48 h. At the end of treatment, immunofluorescence staining (against nuclei, Iba1, TREM2, and αSyn) and IL-6/TNF-α quantification from medium were performed as described in the . ( a ) Representative immunofluorescence images (20× magnification, scale bar 100 µm) and number of DAPI-positive cells; ( b ) Representative immunofluorescence images (20× magnification, scale bar 50 µm) of Iba1 expression, quantification of Iba1 accumulation (Iba1 area normalized by the number of cells), and area of Iba1-positive cells (area of microglia cells normalized by the µm 2 of Iba1); ( c ) Representative immunofluorescence images (20× magnification, scale bar 25 µm) of TREM2 and αSyn expressions, quantification of TREM2-positive cells (TREM2 area normalized by the number of cells), area of TREM2-positive cells (area of microglia cells normalized by the µm 2 of TREM2), and quantification of αSyn area accumulated within TREM(+) cells (area of αSyn normalized by the µm 2 of TREM2); ( d ) Quantification of released cytokines TNF-α and IL-6. All values are represented as percentage versus CTRL and expressed as mean ± SEM ( n = 6; * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, ns = not significant by one-way ANOVA followed by Fisher’s LSD). * p < 0.05 was considered significant.

Article Snippet: For immunofluorescence analyses, the following antibodies were used: primary mouse monoclonal anti-TREM2 antibody [Cat. 11084-MM08, research resource identifier (RRID):AB_2860315], purchased from Sino Biological Europe GmbH, Europe (Düsseldorfer, Germany); primary rabbit polyclonal anti-αSyn antibody (Cat. 2642S, RRID not available), purchased from Ozyme, Paris, France; and primary goat polyclonal anti-Iba1 antibody (Cat. Ab5076, RRID:AB_2224402), purchased from Abcam, Cambridge, UK.

Techniques: Activation Assay, Incubation, Immunofluorescence, Staining, Expressing

Journal: EMBO Molecular Medicine

Article Title: Female sex is linked to a stronger association between sTREM2 and CSF p-tau in Alzheimer’s disease

doi: 10.1038/s44321-024-00190-3

Figure Lengend Snippet: Reagents and tools table

Article Snippet: Monoclonal mouse anti-human TREM2 antibody , Santa Cruz Biotechnology; B-3, sc373828.

Techniques: Recombinant, Sequencing, Software

Journal: Frontiers in Neuroscience

Article Title: Perioperative enriched environment attenuates postoperative cognitive dysfunction by upregulating microglia TREM2 via PI3K/Akt pathway in mouse model of ischemic stroke

doi: 10.3389/fnins.2024.1520710

Figure Lengend Snippet: EE upregulated the expression of TREM2 in the hippocampus after surgery in mice with ischemic stroke. (A,B) The expression of TREM2 protein in the hippocampus on day 10 after surgery ( n = 3). (C,D) The expression of DAP12 protein in the hippocampus on day 10 after surgery ( n = 3). (E) The expression of TREM2 mRNA in the hippocampus on day 10 after surgery ( n = 3). (F) The expression of DAP12 mRNA in the hippocampus on day 10 after surgery ( n = 3). (G) The immunofluorescence of the cellular localization of TREM2 in the hippocampus of mice in the Sham+EE group on day 10 after surgery ( n = 3). DG: dentate gyrus. Scale bars represented 10 μm and 50 μm. ns: no significant; * p < 0.05, *** p < 0.001, the Sham+SE group vs. the PT + SE group; # p < 0.05, ## p < 0.01, the PT + SE group vs. the PT + EE group; ^ p < 0.05, ^^^ p < 0.001, the Sham+SE group vs. the Sham+EE group. Error bars were represented as mean ± SD.

Article Snippet: After antigen retrieval with EDTA, the paraffin sections were washed with phosphate-buffered solution (PBS) and then blocked with 5% bovine serum albumin at room temperature for 1 h. Following this, the sections were incubated with a mouse anti-TREM2 primary antibody (1:1,000, 68723-1-Ig, Proteintech) overnight at 4°C.

Techniques: Expressing, Immunofluorescence

Journal: Frontiers in Neuroscience

Article Title: Perioperative enriched environment attenuates postoperative cognitive dysfunction by upregulating microglia TREM2 via PI3K/Akt pathway in mouse model of ischemic stroke

doi: 10.3389/fnins.2024.1520710

Figure Lengend Snippet: TREM2 shRNA successfully decreased the expression of TREM2 in the hippocampus on day 21 after the injection of TREM2 shRNA. (A,B) The expression of TREM2 protein in the hippocampus ( n = 3). (C,D) The expression of DAP12 protein in the hippocampus ( n = 3). (E) The expression of TREM2 mRNA in the hippocampus ( n = 3). (F) The expression of DAP12 mRNA in the hippocampus. ns: no significant; * p < 0.05, *** p < 0.001, the SE + vehicle group vs. the SE + shRNA group; ## p < 0.01, ### p < 0.001, the EE+ vehicle group vs. the EE+ shRNA group. Error bars were represented as mean ± SD.

Article Snippet: After antigen retrieval with EDTA, the paraffin sections were washed with phosphate-buffered solution (PBS) and then blocked with 5% bovine serum albumin at room temperature for 1 h. Following this, the sections were incubated with a mouse anti-TREM2 primary antibody (1:1,000, 68723-1-Ig, Proteintech) overnight at 4°C.

Techniques: shRNA, Expressing, Injection

Journal: Frontiers in Neuroscience

Article Title: Perioperative enriched environment attenuates postoperative cognitive dysfunction by upregulating microglia TREM2 via PI3K/Akt pathway in mouse model of ischemic stroke

doi: 10.3389/fnins.2024.1520710

Figure Lengend Snippet: The knockdown of TREM2 abolished the neuroprotective effects of EE and inhibited the PI3K/Akt signaling pathway in the hippocampus of mice with ischemic stroke after surgery. (A–C) The behavioral tests of each group ( n = 6). (D) Western bolt results of p-PI3K and PI3K. (E) The protein p-PI3K / PI3K ratio in the hippocampus on day 10 after surgery ( n = 3). (F) Western bolt results of p-AKT and AKT. (G) The protein p-AKT/AKT ratio in the hippocampus on day 10 after surgery ( n = 3). ns: no significant; * p < 0.05, *** p < 0.001, the SE + vehicle group vs. the SE + shRNA group; ## p < 0.01, #### p < 0.0001, the EE+ vehicle group vs. the EE+ shRNA group. Error bars were represented as mean ± SD.

Article Snippet: After antigen retrieval with EDTA, the paraffin sections were washed with phosphate-buffered solution (PBS) and then blocked with 5% bovine serum albumin at room temperature for 1 h. Following this, the sections were incubated with a mouse anti-TREM2 primary antibody (1:1,000, 68723-1-Ig, Proteintech) overnight at 4°C.

Techniques: Knockdown, Western Blot, shRNA

Journal: Cell reports

Article Title: Altered DNA methylation underlies monocyte dysregulation and immune exhaustion memory in sepsis

doi: 10.1016/j.celrep.2024.113894

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: For Panel 2, cells were stained with antibodies against Ly6C (Pacific Blue; Biolegend #128014), CD11b (BV650; BD Biosciences #563402), Ly6g (PE-Cy5.5; Elabscience #E-AB-F1108I), CXCR2 (Alexa Fluor 488; R&D Systems # FAB2164G), CD68 (APC-Fire750; Biolegend #137041), CD172a (BV510; BD Biosciences #740159), CX3CR1 (PE-Cy7; Biolegend #149016), F4/80 (BV711; BD Biosciences #565612), CD38 (BV750; BD Biosciences #747103), PD-L1 (BV421; BD Biosciences #564716), MARCO (Alexa Fluor 647; R&D Systems #FAB29561R), FAS (BV605; Biolegend #152612), MHCII (PerCP; Biolegend #107624), CD168 (PE; Novus #NBP1–76538PE), and TREM2 (Alexa Fluor 700; R&D Systems #FAB17291N).

Techniques: Blocking Assay, Negative Control, Recombinant, Methylation, SYBR Green Assay, DNA Methylation Assay, Software, Isolation, Reverse Transcription, Multiplex Assay

Journal: bioRxiv

Article Title: Drug screening targeting TREM2-TYROBP transmembrane binding

doi: 10.1101/2024.12.02.626344

Figure Lengend Snippet: A) Three independent drops of E. coli BTH101 strain harboring different plasmid configurations grown overnight at 30ºC in LB X-gal plates with Km and Ap. ZIP domains allowed a cytoplasmic reconstitution of the adenylate cyclase as the TREM2 and TYROBP TMD do in the membrane turning blue the drops. Color background obtained from the T18 and T25 control plasmids comes from fortuitous cytoplasmic binding. B) Box plot showing median, quartiles and range values of the relative intensities compared to the empty plasmids. Semiquantitative analysis of the X-gal drops under the different configurations (T18/T25 n=99; ZIP::T18/ZIP::T25 n=81; TREM2TMD::T18/TYROBP TMD::T25 n=57).

Article Snippet: Semiconfluent (50-70%) cultures were stimulated with 1-7 μg/ml anti-TREM2 (1:1 anti-TREM2 R&D systems AF1729; anti-TREM2 R&D MAB17291) for 5 minutes at 37ºC in the presence of 1% dimetilsulfoxide (DMSO) alone or in combination with the candidate drug.

Techniques: Plasmid Preparation, Membrane, Control, Binding Assay

Journal: bioRxiv

Article Title: Drug screening targeting TREM2-TYROBP transmembrane binding

doi: 10.1101/2024.12.02.626344

Figure Lengend Snippet: A) Representative analysis of the beta-galactosidase activity and bacterial growth E. coli BTH101 with the TMD plasmids after O/N stimulation with the different candidates. Candidate in well C6 was selected for further characterization. B) Western blot of HCM3 protein extracts after αTREM2 stimulation with either 1% DMSO or varenicline 150 μM. C) Pathway stimulation after quantifying the different band intensities (n=3).

Article Snippet: Semiconfluent (50-70%) cultures were stimulated with 1-7 μg/ml anti-TREM2 (1:1 anti-TREM2 R&D systems AF1729; anti-TREM2 R&D MAB17291) for 5 minutes at 37ºC in the presence of 1% dimetilsulfoxide (DMSO) alone or in combination with the candidate drug.

Techniques: Activity Assay, Western Blot

Journal: bioRxiv

Article Title: Drug screening targeting TREM2-TYROBP transmembrane binding

doi: 10.1101/2024.12.02.626344

Figure Lengend Snippet: A) Varenicline 2D representation. B) Best fitting docking configuration of varenicline with TREM2 (left) and TYROBP (right). C) NCI representation (green area) of the contacts between varenicline and ILE-6 and LEU-13 residues from TYROBP’s TMD (right) and with the LEU-6, ALA-7 and PHE-10 residues in chain TREM2’s TMD (left).

Article Snippet: Semiconfluent (50-70%) cultures were stimulated with 1-7 μg/ml anti-TREM2 (1:1 anti-TREM2 R&D systems AF1729; anti-TREM2 R&D MAB17291) for 5 minutes at 37ºC in the presence of 1% dimetilsulfoxide (DMSO) alone or in combination with the candidate drug.

Techniques: