Journal: Journal of Neuroinflammation

Article Title: LPS- induced inflammation exacerbates phospho-tau pathology in rTg4510 mice

doi: 10.1186/1742-2094-7-56

Figure Lengend Snippet: Age-related, Tau Induced CD45 activation . Immunohistochemistry was performed for CD45 in rTg4510 and nontransgenic (nTg) littermates aged 1 (A, B, H, I), 5 (C, D, J, K), and 9 (E, F, L, M) months in the anterior cortex (A-F) and hippocampus (H-M). Panels G and N present mean ± S.E.M of % Area for immunostaining of CD45+ microglia in the anterior cortex and hippocampus, respectively. Statistical analysis was performed using 2-way ANOVA followed by Fisher's PLSD multiple comparison test. Area stained for CD45 significantly increased in rTg4510 mice at 9 months of age in the anterior cortex and hippocampus compared with nontransgenic littermates or with younger mice. Lines above relevant bars display significant differences between groups (*p < 0.05), n = 4-5. Sections were digitized and representative images were taken at 40× magnification. Scale bar represents 50 μm.

Article Snippet: Sections were incubated with primary antibodies rat anti-mouse CD45 (1:3000) (Serotec, Raleigh, NC), rat anti-major histocompatibility complex -II (1:5000) (MHCII; BD Pharmigen), rabbit anti-mouse chitinase 3-like-3 (1:3000) (YM1; StemCell Technologies, Vancouver, Canada), chicken anti-arginase 1 (1:50,000)(generous gift from Dr. S.M.

Techniques: Activation Assay, Immunohistochemistry, Immunostaining, Comparison, Staining

Journal: Journal of Neuroinflammation

Article Title: LPS- induced inflammation exacerbates phospho-tau pathology in rTg4510 mice

doi: 10.1186/1742-2094-7-56

Figure Lengend Snippet: LPS-Induced CD45 Activation in rTg4510 and nontransgenic mice . The microglia marker CD45 was examined by immunohistochemical staining in anterior cortex, hippocampus (CA1), and entorhinal cortex (ECX) after LPS administration. rTg4510 mice or nontransgenic (nTg) littermates were injected unilaterally with LPS or vehicle into the anterior cortex and hippocampus, followed by 1 week survival. Representative images from anterior cortex of nontransgenic (A, C) or rTg4510 mice (B, D) from vehicle- (A, B) or LPS-injected (C, D) mice are shown. Images (F-I) were collected from the CA1 stratum radiatum (CA1) of non-transgenic (F, H) or rTg4510 mice (G, I) after vehicle (F, G) or LPS (H, I) injection. Similar images (K-N) were collected from the entorhinal cortex (ECX) of nontransgenic (K, M) or rTg4510 mice (L, N) after vehicle (K, L) or LPS (M, N) injection. Quantitation of % area containing positive immunostaing is presented for each brain region in E, J, and O (mean ± S.E.M, n = 6-8). CD45+ staining increased in both rTg4510 and nontransgenic littermates in anterior cortex, CA1, and ECX of mice treated with LPS compared to vehicle-treated mice. Statistical analysis was performed using 2-way ANOVA followed by Fisher's PLSD multiple comparison test. The asterisk indicates *p < 0.05), n = 6 = 8. Scale bar represents 20 μm.

Article Snippet: Sections were incubated with primary antibodies rat anti-mouse CD45 (1:3000) (Serotec, Raleigh, NC), rat anti-major histocompatibility complex -II (1:5000) (MHCII; BD Pharmigen), rabbit anti-mouse chitinase 3-like-3 (1:3000) (YM1; StemCell Technologies, Vancouver, Canada), chicken anti-arginase 1 (1:50,000)(generous gift from Dr. S.M.

Techniques: Activation Assay, Marker, Immunohistochemical staining, Staining, Injection, Transgenic Assay, Quantitation Assay, Comparison

Journal: Journal of Neuroinflammation

Article Title: LPS- induced inflammation exacerbates phospho-tau pathology in rTg4510 mice

doi: 10.1186/1742-2094-7-56

Figure Lengend Snippet: LPS-Induced Microglia Activation on Contralateral Hemisphere

Article Snippet: Sections were incubated with primary antibodies rat anti-mouse CD45 (1:3000) (Serotec, Raleigh, NC), rat anti-major histocompatibility complex -II (1:5000) (MHCII; BD Pharmigen), rabbit anti-mouse chitinase 3-like-3 (1:3000) (YM1; StemCell Technologies, Vancouver, Canada), chicken anti-arginase 1 (1:50,000)(generous gift from Dr. S.M.

Techniques: Activation Assay

Journal: Journal of Neuroinflammation

Article Title: LPS- induced inflammation exacerbates phospho-tau pathology in rTg4510 mice

doi: 10.1186/1742-2094-7-56

Figure Lengend Snippet: Immunofluorescent labeling of microglial activation and phospho-tau in rTg4510 mice treated with LPS . Panels A-C shows immunofluorescent staining of arginase-1 positive microglia (green; A, C) and phospho-tau ser396 (red; B, C) in the anterior cortex. Panels D-F shows YM1 positive microglia (green; D, F) and AT8 (phospho-tauSer202/Thr204) (red; E, F) in the anterior cortex. Panels G-L shows increased CD45 positive microglia (Green; G, I, J, L), phospho-tauSer199/202 (red; H, I), and full-length tau (red; K, L) in the hippocampus CA region. Activated microglia failed to co-label with phospho-tau markers. Images were taken at 20x objective. Scale bar represents 20 μm.

Article Snippet: Sections were incubated with primary antibodies rat anti-mouse CD45 (1:3000) (Serotec, Raleigh, NC), rat anti-major histocompatibility complex -II (1:5000) (MHCII; BD Pharmigen), rabbit anti-mouse chitinase 3-like-3 (1:3000) (YM1; StemCell Technologies, Vancouver, Canada), chicken anti-arginase 1 (1:50,000)(generous gift from Dr. S.M.

Techniques: Labeling, Activation Assay, Staining

Journal: Cell Proliferation

Article Title: p75NTR −/− mice exhibit an alveolar bone loss phenotype and inhibited PI3K/Akt/β‐catenin pathway

doi: 10.1111/cpr.12800

Figure Lengend Snippet: P75NTR −/− EMSCs exhibit decreased osteogenic differentiation capacity compared to WT EMSCs. (A) The p75NTR and mesenchymal stem cell surface markers (CD14, CD90, CD146 and CD166) and hematopoietic markers (CD45) were detected on WT and p75NTR −/− EMSCs by the Flow cytometry. (B) The third passage (P3) cells of E12.5d WT and p75NTR −/− EMSCs. Scale bar represents 50 μm. (C) Representative images of colonies formed by E12.5d WT and p75NTR −/− EMSCs and the analysis of colony formation. (D) The proliferation ratio of E12.5d WT and p75NTR −/− EMSCs was assessed by CCK‐8. WT and p75NTR −/− EMSCs were induced with osteogenic induction medium. On days 7, 14 and 21, the (E) protein levels of Runx2, Col1 and β‐catenin were detected by Western blot and (F) grayscale analysis was performed and the levels of the indicated proteins are expressed relative to the levels of GAPDH. (G) mRNA levels of Runx2, Col1 and β‐catenin were detected by real‐time PCR, respectively, using GAPDH as control. (H) On day 7, ALP staining was used to detect their potential of differential mineralization. On day 21, Alizarin Red staining was used to detect their mineralized nodules. Scale bar represents 50 μm. KO represent p75NTR −/− . Data are shown as mean ± SD from three independent experiments. * P < .05, ** P < .01

Article Snippet: The primary antibodies including anti‐mouse CD14, anti‐mouse CD90, anti‐mouse CD146, anti‐mouse CD166, anti‐mouse CD45 (1:100; Santa Cruz, USA) and anti‐mouse p75NTR‐FITC (1:100; Abcam, UK) were used to identify EMSCs.

Techniques: Flow Cytometry, CCK-8 Assay, Western Blot, Real-time Polymerase Chain Reaction, Control, Staining

Journal: Cell reports

Article Title: Activation of GPR44 decreases severity of myeloid leukemia via specific targeting of leukemia initiating stem cells

doi: 10.1016/j.celrep.2023.112794

Figure Lengend Snippet:

Article Snippet: Mouse anti-mouse CD45.1 (FITC) , Miltenyi Biotec , Cat# 130–124-211; RRID: AB_2857674.

Techniques: Virus, Recombinant, Enzyme-linked Immunosorbent Assay, Binding Assay, Staining, Modification, Saline, Concentration Assay, Over Expression, Protein Extraction, Membrane, SYBR Green Assay, Bicinchoninic Acid Protein Assay, Protease Inhibitor, CCK-8 Assay, Reverse Transcription, Selection, Plasmid Preparation, Knock-Out, Software, Real-time Polymerase Chain Reaction, Flow Cytometry

Journal: Frontiers in Immunology

Article Title: Multifocal Cerebral Microinfarcts Modulate Early Alzheimer’s Disease Pathology in a Sex-Dependent Manner

doi: 10.3389/fimmu.2021.813536

Figure Lengend Snippet: Dynamics of circulating monocytes are modulated upon multifocal microinfarction. (A) Gating strategy used to discriminate the circulating monocytes (CD11b + LyC6 + ) in leukocytes (CD45 + ) and the subsequent distribution of the inflammatory monocytes (Ly6C high ), patrolling monocytes (Ly6C low ) and neutrophils (Ly6G high ). (B) Flow cytometry analysis shows that the frequency of total monocytes in the blood circulation increases in MO-operated male WT mice while it remains unchanged in MO-operated male APP/PS1 mice compared to sham-operated male mice and decreases in MO-operated male APP/PS1 mice compared to MO-operated male WT mice. (C) Flow cytometry analysis shows that the frequency of Ly6C low monocytes decreases in the blood circulation of MO-operated male WT mice compared to sham-operated WT mice. (D) Flow cytometry analysis shows that the frequency of Ly6C int monocytes increases in the blood circulation of MO-operated male WT mice as well as MO-operated male APP/PS1 mice compared to sham-operated WT mice. (E) Flow cytometry analysis shows that the frequency of inflammatory Ly6C high monocytes decreases in the blood circulation of MO-operated male WT mice as well as MO-operated male APP/PS1 mice compared to sham-operated male mice. (F) Flow cytometry analysis shows that similarly to males, the frequency of total monocytes in the blood circulation increases in MO-operated female WT mice while it remains unchanged in MO-operated female APP/PS1 mice compared to sham-operated female mice and decreases in MO-operated female APP/PS1 mice compared to MO-operated female WT mice. (G) Flow cytometry analysis indicates that the frequency of Ly6C low monocytes increases in MO-operated female WT mice, while it decreases in the MO-operated female APP/PS1 mice compared to sham-operated female mice. (H) Flow cytometry analysis indicates that the frequency of Ly6C int monocytes increases in the blood circulation of MO-operated female WT mice and to a lesser extent MO-operated female APP/PS1 mice. (I) Flow cytometry analysis indicates that the frequency of Ly6C high monocytes is decreased in the blood circulation of MO-operated female WT mice as well as MO-operated female APP/PS1 mice compared to sham-operated female mice. (J) Flow cytometry analysis shows that the frequency of Ly6G neutrophils increases in MO-operated male WT mice compared to sham-operated male WT mice but to a lesser extent in MO-operated male APP/PS1 mice compared to sham-operated APP/PS1 male mice. (K) Flow cytometry analysis shows that the frequency of Ly6G neutrophils similarly increases in the blood circulation of MO-operated female WT mice compared to sham-operated female WT mice, whereas it remains unchanged in MO-operated female APP/PS1 mice compared to sham-operated female APP/PS1 mice. Data are mean ± SEM (n = 5-10 animals/group). *P < 0.05/**P < 0.01/****P < 0.01 compared with sham- and MO-operated male and female WT and APP/PS1 mice (two-tailed unpaired t-test).

Article Snippet: The following primary antibodies were used; anti-Aβ (6E10) (1:1000; mouse anti-human, Biolegend, 8030001), anti-ionized calcium binding adaptor molecule (IBA)-1 (1:1000, rabbit anti-mouse, WAKO, 019-19741), anti-cluster of differentiation (CD)-68 (1:1000, rat anti-mouse, Bio-Rad, Hercules, CA, USA, MCA1957) and anti-CD45 (1:500, rat anti-mouse, BD bioscience, 55376) and anti-nuclear receptor subfamily 4 group A member 1 (Nr4A1 or Nurr77) (1:250, rabbit, Abcam, ab13851).

Techniques: Flow Cytometry, Two Tailed Test

Journal: Frontiers in Immunology

Article Title: Multifocal Cerebral Microinfarcts Modulate Early Alzheimer’s Disease Pathology in a Sex-Dependent Manner

doi: 10.3389/fimmu.2021.813536

Figure Lengend Snippet: Infiltrated phagocytic monocytes are recruited to the lesion sites and Aβ plaques. (A) A tile representing a coronal brain section of MO-operated male APP/PS1/CX3CR1 GFP/+ mouse immunolabeled with CD45 (i.e. infiltrating cells, blue) and IBA1 (i.e. microglia; red) showing (A) the presence of CX3CR1 GFP/+ expressing Nurr77 + , highlighting their monocytic origin, at lesion site 1 week post-microinfarct induction. (B) A tile representing a coronal brain section of MO-operated male APP/PS1/CX3CR1 GFP/+ mouse immunolabeled with CD45 (blue) and 6E10 (Aβ plaques; red) showing (B’) the presence of CD45 + /CX3CR1 GFP/+ cells at the lesion site and (B’’) CD45 + /IBA1 + /CX3CR1 GFP/+ cells recruited to Aβ plaques. Scale bar = 20 µm (A) , 1000 µm (B) , 50 µm (B’) and 25 µm (B”) .

Article Snippet: The following primary antibodies were used; anti-Aβ (6E10) (1:1000; mouse anti-human, Biolegend, 8030001), anti-ionized calcium binding adaptor molecule (IBA)-1 (1:1000, rabbit anti-mouse, WAKO, 019-19741), anti-cluster of differentiation (CD)-68 (1:1000, rat anti-mouse, Bio-Rad, Hercules, CA, USA, MCA1957) and anti-CD45 (1:500, rat anti-mouse, BD bioscience, 55376) and anti-nuclear receptor subfamily 4 group A member 1 (Nr4A1 or Nurr77) (1:250, rabbit, Abcam, ab13851).

Techniques: Immunolabeling, Expressing

Journal: Journal of neuroinflammation

Article Title: Treatment with gelsolin reduces brain inflammation and apoptotic signaling in mice following thermal injury.

doi: 10.1186/1742-2094-8-118

Figure Lengend Snippet: Figure 8 Gelsolin affects migration of myeloid-derived cells into brain. CD45+ cells from gelsolin-treated mice 8 h postburn (A) and quantification of infiltrating CD45+ (B) cells in 10 high power fields (HPF) of the periventricle region following gelsolin treatment. Gelsolin positive cells were seen in medial habenular nucleus (MHb), stria medullaris (sm), hippocampal CA field (CA2) and blood vessel (BV). Magnifications are × 400. *P < 0.05, **P < 0.01, and ***P < 0.001 vs. sham-injured mice; #p < 0.05, ##p < 0.01, ###p < 0.001 vs. placebo mice; + +p < 0.01, and +++p < 0.001 vs. Gsn-L mice by ANOVA, Newman-Keuls post-hoc test. Data are means ± SD for n = 6-8.

Article Snippet: Sections used for immunocytochemistry were incubated in 0.3% hydrogen peroxide (H2O2) for 10 min, and incubated free-floating in antibodies (Abs) of polyclonal anti-mouse ionized calcium-binding adapter molecule 1 (Iba-1, 1:1000; Wako, Osaka, Japan), monoclonal anti-mouse CD11b (Mac-1, 1:1000; EuroBioScience, Lund, Sweden), monoclonal anti-mouse CD45 (1:1000; EuroBioScience), or rabbit anti-cleaved caspase-3 (1:50; Cell Signaling, Danvers, MA, USA) with 3% normal goat serum, 0.05%Triton-X in PBS, for 24-48 h rotating at 4°C.

Techniques: Migration, Derivative Assay

Journal: NPJ Regenerative Medicine

Article Title: Matrix-bound nanovesicle-associated IL-33 supports functional recovery after skeletal muscle injury by initiating a pro-regenerative macrophage phenotypic transition

doi: 10.1038/s41536-024-00346-2

Figure Lengend Snippet: Cardiotoxin muscle injuries were performed on 8 week old B6- il33 +/+ arg1 GFP (WT) or B6- il33 −/− arg1 GFP (KO) mice. Functional analysis was performed on POD14 male mice. Flow cytometric analysis or in situ immunolabeling was performed on male or female mice sacrificed at POD3, POD7, and POD14. a In situ contractile testing comparing il33 −/− vs. il33 + /+ mice reveals significantly different force-frequency responses ( p < 0.05, data shown as mean ± SEM) and reduced peak specific force in il33 −/− mice compared to wildtype counterparts ( p < 0.01). b Representative dot plots and frequency for inflammatory macrophages in the CD45 + CD3 - B220 - CD11b + Ly6G - gate (data shown as mean ± SEM). c Dot plots and frequency for ST2 + Treg in the CD45 + CD3 + B220 - CD4 + gate (data shown as mean ± SEM). d Representative in situ immunolabeling images showing CD11b+ macrophages (arrowheads), CD11b+Fizz1+ (arrows, left panel), or CD11b+iNOS+ (arrows, right panel). e Quantification of immunolabeling shows significantly fewer Fizz1+ macrophages in il33 −/− mice at all timepoints ( p < 0.05), and increased iNOS+ macrophages at POD3 ( p < 0.05). f Representative in situ immunolabeling image shows nuclear FoxP3 staining (left panels). Quantification of FoxP3+ T REG immunolabeling shows reduced FoxP3+ T REG accumulation in il33 −/− animals compared to wildtype counterparts. (Scale bars = 5 µm, N = 5 biological replicates, n ≥ 3 technical replicates. Results shown as Min-Max unless otherwise specified, p -values were calculated using repeated measures ANOVA (force-frequency), two-tail t-test (peak specific force and FoxP3 immunolabeling), one-way ANOVA (flow cytometry), or multiple t-tests with two stage step-up method of Benjamini, Krieger and Yekutieli false discovery rate p-value correction (immunolabeling), * p < 0.05, ** p < 0.01, **** p < 0.0001).

Article Snippet: Antibodies: PECF594 Rat Anti-Mouse CD45 (1:400 BD Horizon Cat: 562420; Clone: 30-F11), Pacific Blue Anti-Mouse F4/80 (1:200 BioLegend Cat: 123124; Clone: BM8), BV605 Rat Anti-Mouse CD38 (1:200 BD OptiBuild Cat: 740361; Clone: 90/CD38), BV650 Rat Anti-Mouse CD4 (1:200 BD Horizon Cat: 563232; Clone: GK1.5), PE Rat Anti-Mouse Ly6C (1:200 BD Pharmigen Cat: 560592; Clone: AL-21), PE-Cy5 Rat Anti-Mouse CD45R/B220 (1:200 BD Pharmigen Cat: 553091; Clone: RA3-6B2), Alexa Fluor 700 Rat Anti-Mouse CD3 Molecular Complex (1:200 BD Pharmigen Cat: 561388; Clone: 17A2), APC/Fire 750 Anti-Mouse/Human CD11b (1:200 BioLegend Cat: 101262; Clone: M1/70), BUV395 Anti-Mouse I-A/I-E (1:200 BD OptiBuild Cat: 743876; Clone: 2G9), BUV805 Rat Anti-Mouse Ly6G (1:200 BD OptiBuild Cat: 741994; Clone: 1A8), Brilliant Violet 510 Anti-Mouse CD11c (1:200 BioLegend Cat: 117338; Clone:N418), BUV737 Rat Anti-Mouse IL33R (ST2) (1:100 BD OptiBuild Cat: 749323; Clone: U29-93), PerCP-Cyanine5.5 FoxP3 Monoclonal Antibody (1:200 Thermo Fisher Cat: 45-5773-82; Clone: FJK-16s), PE-Cyanine7 iNOS Monoclonal Antibody (1:400 Thermo Fisher Cat: 25-5920-82; Clone: CXNFT), Alexa Fluor 647 Anti-Mouse CD206 (1:200 BioLegend Cat: 141712; Clone: 068C2), eBioscience Fixable Viability Dye eFluor 506 (1:500 Thermo Fisher Cat: 65-0866-14).

Techniques: Functional Assay, In Situ, Immunolabeling, Staining, Flow Cytometry

Journal: NPJ Regenerative Medicine

Article Title: Matrix-bound nanovesicle-associated IL-33 supports functional recovery after skeletal muscle injury by initiating a pro-regenerative macrophage phenotypic transition

doi: 10.1038/s41536-024-00346-2

Figure Lengend Snippet: A Absolute macrophages per gram of muscle 14 days post LysMcre-ST2 f/f injury with cardiotoxin and co-administration with vehicle or MBV. B Representative flow plots of F4_80 + macrophages expressing iNOS, Ly6C and CD206 14 days post LysMcre-ST2 f/f injury with cardiotoxin and co-administration with vehicle or MBVs. C Frequency quantification of F4_80 + macrophages expressing iNOS, Ly6C or CD206 out of CD45 live cells. D Absolute macrophages per gram of muscle expressing iNOS, Ly6C or CD206. E Absence of macrophage ST2 reduces the beneficial impact of MBV treatment on force-producing capacity of mice. Force-frequency curves for TA stimulation in mice from four different experimental groups. ( N = 5-6/group, two-way ANOVA with repeated measures, * denotes p < 0.05, ** denotes p < 0.005, *** denotes p < 0.001). N = 6 per group from 2 independent experiments. Error bars are SD. Student’s t-test.

Article Snippet: Antibodies: PECF594 Rat Anti-Mouse CD45 (1:400 BD Horizon Cat: 562420; Clone: 30-F11), Pacific Blue Anti-Mouse F4/80 (1:200 BioLegend Cat: 123124; Clone: BM8), BV605 Rat Anti-Mouse CD38 (1:200 BD OptiBuild Cat: 740361; Clone: 90/CD38), BV650 Rat Anti-Mouse CD4 (1:200 BD Horizon Cat: 563232; Clone: GK1.5), PE Rat Anti-Mouse Ly6C (1:200 BD Pharmigen Cat: 560592; Clone: AL-21), PE-Cy5 Rat Anti-Mouse CD45R/B220 (1:200 BD Pharmigen Cat: 553091; Clone: RA3-6B2), Alexa Fluor 700 Rat Anti-Mouse CD3 Molecular Complex (1:200 BD Pharmigen Cat: 561388; Clone: 17A2), APC/Fire 750 Anti-Mouse/Human CD11b (1:200 BioLegend Cat: 101262; Clone: M1/70), BUV395 Anti-Mouse I-A/I-E (1:200 BD OptiBuild Cat: 743876; Clone: 2G9), BUV805 Rat Anti-Mouse Ly6G (1:200 BD OptiBuild Cat: 741994; Clone: 1A8), Brilliant Violet 510 Anti-Mouse CD11c (1:200 BioLegend Cat: 117338; Clone:N418), BUV737 Rat Anti-Mouse IL33R (ST2) (1:100 BD OptiBuild Cat: 749323; Clone: U29-93), PerCP-Cyanine5.5 FoxP3 Monoclonal Antibody (1:200 Thermo Fisher Cat: 45-5773-82; Clone: FJK-16s), PE-Cyanine7 iNOS Monoclonal Antibody (1:400 Thermo Fisher Cat: 25-5920-82; Clone: CXNFT), Alexa Fluor 647 Anti-Mouse CD206 (1:200 BioLegend Cat: 141712; Clone: 068C2), eBioscience Fixable Viability Dye eFluor 506 (1:500 Thermo Fisher Cat: 65-0866-14).

Techniques: Expressing