Journal: PLoS Pathogens

Article Title: Gut microbiota from high-risk men who have sex with men drive immune activation in gnotobiotic mice and in vitro HIV infection

doi: 10.1371/journal.ppat.1007611

Figure Lengend Snippet: Ileum and colon tissues were collected from mouse recipients at 21 days post gavage and analyzed for frequencies of CD69+ and CD103+ T cells. (A) Gating strategy for CD69+ T cells. Representative plots from ileum tissues are shown. (B-C) Frequencies of (B) CD69+ CD8+ T cells, (C) CD103+ CD8+ T cells, and (D) CD103+ CD4+ T cells in the ileum. (E) Frequencies of CD69+ CD4+ T cells in the colon. Each data point represents a single mouse gavaged with an individual donor’s feces, and a representative mouse was used for stools tested in replicate mice. Lines represent medians. Statistical analyses were performed using t-tests to compare groups if data from both groups had normal distributions and Mann-Whitney tests to compare groups if data from at least one group had a non-parametric distribution. ** = p<0.01, * = p<0.05.

Article Snippet: 0.5–1 x 10 6 ileum and colon cells from each mouse were stained in FACS buffer for 30 min at 4° C with the following antibodies: CD3-PerCP-Cy5.5 (17A2, Biolegend), CD4-FITC (RM4-4, eBioscience), CD8-BV510 (53–6.7, Biolegend), CD69-PE-Cy7 (h1.2F3, eBioscience), CD103-PE-Dazzle-594 (2E7, Biolegend).

Techniques: MANN-WHITNEY

Journal: PLoS Pathogens

Article Title: Gut microbiota from high-risk men who have sex with men drive immune activation in gnotobiotic mice and in vitro HIV infection

doi: 10.1371/journal.ppat.1007611

Figure Lengend Snippet: mLN from mouse recipients were collected and analyzed for effector memory CD4+ T cell frequency (CD44+ CD62L-) using flow cytometry. (A) Comparison of frequencies of CD44+ CD62- CD4+ T cells in the mLN across recipient groups. (B) Spearman correlations of effector memory CD4+ T cell (T em ) frequencies in the mLN with frequencies of CD69+ CD4+ T cells in the colon. Each data point represents a single mouse gavaged with an individual donor’s feces, and a representative mouse was used for stools tested in replicate mice. Lines represent medians. Statistical analyses were performed using t-tests to compare groups if data from both groups had normal distributions and Mann-Whitney tests to compare groups if data from at least one group had a non-parametric distribution. * = p<0.05.

Article Snippet: 0.5–1 x 10 6 ileum and colon cells from each mouse were stained in FACS buffer for 30 min at 4° C with the following antibodies: CD3-PerCP-Cy5.5 (17A2, Biolegend), CD4-FITC (RM4-4, eBioscience), CD8-BV510 (53–6.7, Biolegend), CD69-PE-Cy7 (h1.2F3, eBioscience), CD103-PE-Dazzle-594 (2E7, Biolegend).

Techniques: Flow Cytometry, MANN-WHITNEY

Journal: PLoS Pathogens

Article Title: Gut microbiota from high-risk men who have sex with men drive immune activation in gnotobiotic mice and in vitro HIV infection

doi: 10.1371/journal.ppat.1007611

Figure Lengend Snippet: Microbial correlates with donor/recipient T cell activation and gut homing, and in vitro HIV infection.

Article Snippet: 0.5–1 x 10 6 ileum and colon cells from each mouse were stained in FACS buffer for 30 min at 4° C with the following antibodies: CD3-PerCP-Cy5.5 (17A2, Biolegend), CD4-FITC (RM4-4, eBioscience), CD8-BV510 (53–6.7, Biolegend), CD69-PE-Cy7 (h1.2F3, eBioscience), CD103-PE-Dazzle-594 (2E7, Biolegend).

Techniques: Activation Assay, In Vitro, Infection

Journal: bioRxiv

Article Title: Androgens modulate the immune profile in a mouse model of polycystic ovary syndrome

doi: 10.1101/2024.02.22.581579

Figure Lengend Snippet: ( a ) Experimental design. ( b ) Frequency of neutrophils in blood, expressed as percent of CD45 + immune cells (n = 11 Control, 9 DHT, 9 DHT + Flutamide). ( c ) Frequency of eosinophils in blood (n = 12, 11, 8). ( d ) Frequency of monocytes in blood (n = 11, 9, 9). ( e ) Neutrophil count in blood (n = 5, 10, 6). ( f ) CD45+ immune cell count in blood (n = 5, 10, 7). ( g ) Frequency of CD3 + T cells in blood (n = 8, 10, 9). ( h ) Frequency of CD4 + T helper cells in blood (n = 8, 10, 9). ( i ) Frequency of CD8 + cytotoxic T cells in blood (n = 8, 10, 9). ( j ) Thymus weight (n = 12, 11, 8). ( k ) Frequency of NK cells in blood (n = 11, 11, 8). ( l ) CD69 expression on NK cells in blood (n = 11, 11, 8). ( m ) Frequency of NK cells in spleen (n = 12, 11, 8). ( n ) CD69 expression on NK cells in spleen (n = 12, 11, 8). Data are presented as means ± SD. n indicates the number of biologically independent samples examined. Statistical analysis was assessed by one-way ANOVA with Dunnett’s multiple comparison ( b, e - j, m ) or by Kruskal-Wallis with Dunn’s multiple comparison ( c - d, k - l, n ) and significant differences were indicated with p values. Source data are provided as a Source Data File.

Article Snippet: Following incubation with Fc-block (anti-mouse CD16/32, clone 2.4G2, BD Biosciences), cell surface markers were detected using fluorochrome-conjugated antibodies: CD45 (30-F11, Invitrogen/ eBioscience), CD11b (M1/70, BD Biosciences), Siglec-F (E50-2440, BD Biosciences), Ly6G (1A8, BD Biosciences), Ly6C (AL-21, BD Biosciences), F4/80 (BM8, eBioscience), MHC II (M5/114.15.2, eBioscience), CD11c (N418, BD Biosciences), CD3e (500A2, BD Biosciences), NK-1.1 (PK136, BD Biosciences), CD8a (53-6.7, BD Biosciences), CD4 (RM4-5, BD Biosciences), CD25 (PC61, BD Biosciences), CD69 (H1.2F3, eBioscience).

Techniques: Cell Counting, Expressing, Comparison

Journal: bioRxiv

Article Title: Androgens modulate the immune profile in a mouse model of polycystic ovary syndrome

doi: 10.1101/2024.02.22.581579

Figure Lengend Snippet: ( a ) Frequency of eosinophils among immune cells in uterus (n = 11 Control, 11 DHT, 7 DHT + Flutamide). ( b ) Representative plots of CD45 + immune cells, gated on live single cells. ( c ) Frequency of NK cells in uterus (n = 12, 11, 7). ( d ) CD69 expression on NK cells in uterus (n = 12, 11, 7). ( e ) Frequency of macrophages in uterus (n = 11, 11, 7). ( f ) MHC-II expression on macrophages (n = 11, 11, 7). ( g ) CD11c expression on macrophages (n = 11, 11, 7). ( h ) Eotaxin (CCL11) levels in uterus (n = 10, 9, 7). ( i ) IL-5 levels in uterus (n = 10, 9, 7). ( j ) IFN-γ levels in uterus (n = 10, 9, 7). ( k ) TNF-α levels in uterus (n = 10, 9, 7). ( l ) CCL2 levels in uterus (n = 10, 9, 7). Data are presented as means ± SD. n indicates the number of biologically independent samples examined. Statistical analysis was assessed by Kruskal-Wallis with Dunn’s multiple comparison ( a, g ), one-way ANOVA with Dunnett’s multiple comparison ( c - f ) or mixed-effects ANOVA with Bonferroni’s multiple comparison test ( h - l ) and significant differences were indicated with p values. Source data are provided as a Source Data File.

Article Snippet: Following incubation with Fc-block (anti-mouse CD16/32, clone 2.4G2, BD Biosciences), cell surface markers were detected using fluorochrome-conjugated antibodies: CD45 (30-F11, Invitrogen/ eBioscience), CD11b (M1/70, BD Biosciences), Siglec-F (E50-2440, BD Biosciences), Ly6G (1A8, BD Biosciences), Ly6C (AL-21, BD Biosciences), F4/80 (BM8, eBioscience), MHC II (M5/114.15.2, eBioscience), CD11c (N418, BD Biosciences), CD3e (500A2, BD Biosciences), NK-1.1 (PK136, BD Biosciences), CD8a (53-6.7, BD Biosciences), CD4 (RM4-5, BD Biosciences), CD25 (PC61, BD Biosciences), CD69 (H1.2F3, eBioscience).

Techniques: Expressing, Comparison

Journal: bioRxiv

Article Title: Androgens modulate the immune profile in a mouse model of polycystic ovary syndrome

doi: 10.1101/2024.02.22.581579

Figure Lengend Snippet: ( a ) Frequency of macrophages in ovaries, expressed as percent of CD45 + immune cells (n = 9 Control, 8 DHT, 6 DHT + Flutamide). ( b ) Frequency of NK cells in ovaries (n = 8, 10, 9). ( c ) CD69 expression on NK cells in ovaries (n = 8, 10, 9). ( d ) Frequency of CD4 + T helper cells in ovaries (n = 8, 10, 9). ( e ) Frequency of CD8 + cytotoxic T cells in ovaries (n = 8, 10, 9). Data are presented as means ± SD. n indicates the number of biologically independent samples examined. Statistical analysis was assessed by one-way ANOVA with Dunnett’s multiple comparison ( a - c ), or Kruskal-Wallis with Dunn’s multiple comparison ( d - e ) and significant differences were indicated with p values. Source data are provided as a Source Data File.

Article Snippet: Following incubation with Fc-block (anti-mouse CD16/32, clone 2.4G2, BD Biosciences), cell surface markers were detected using fluorochrome-conjugated antibodies: CD45 (30-F11, Invitrogen/ eBioscience), CD11b (M1/70, BD Biosciences), Siglec-F (E50-2440, BD Biosciences), Ly6G (1A8, BD Biosciences), Ly6C (AL-21, BD Biosciences), F4/80 (BM8, eBioscience), MHC II (M5/114.15.2, eBioscience), CD11c (N418, BD Biosciences), CD3e (500A2, BD Biosciences), NK-1.1 (PK136, BD Biosciences), CD8a (53-6.7, BD Biosciences), CD4 (RM4-5, BD Biosciences), CD25 (PC61, BD Biosciences), CD69 (H1.2F3, eBioscience).

Techniques: Expressing, Comparison

Journal: bioRxiv

Article Title: Androgens modulate the immune profile in a mouse model of polycystic ovary syndrome

doi: 10.1101/2024.02.22.581579

Figure Lengend Snippet: ( a ) Frequency of macrophages among immune cells in VAT (n = 8 Control, 10 DHT, 7 DHT + Flutamide). ( b ) Representative plots of subpopulations of macrophages (CD45 + CD11b + SSC low/mid F4/80 + Ly6G mid ) based on expression of CD11b and CD11c and frequency CD11b high CD11c + . ( c ) MHC-II expression on CD11b high CD11c + macrophages (n = 8, 10, 7). ( d ) MHC-II expression on CD11b mid CD11c - macrophages (n = 8, 10, 7). ( e ) Frequency of eosinophils in VAT (n = 8, 10, 7). ( f ) CD69 expression on NK cells in VAT (n = 10, 11, 8). ( g ) Frequency of NK cells in VAT (n = 10, 11, 8). ( h ) IL-5 levels in VAT (n = 16, 14, 15). ( i ) Eotaxin (CCL11) levels in VAT (n = 10, 9, 7). ( j ) IL-4 levels in VAT (n = 16, 14, 15). ( k ) IL-13 levels in VAT (n = 10, 9, 7). ( l ) IFN-γ levels in VAT (n = 16, 16, 15). ( m ) TNF-α levels in VAT (n = 16, 16, 15). ( n ) IL-10 levels in VAT (n = 16, 14, 15). ( o ) IL-2 levels in VAT (n = 16, 16, 15). Data are presented as means ± SD. n indicates the number of biologically independent samples examined. Statistical analysis was assessed by one-way ANOVA with Dunnett’s multiple comparison ( a - f ), Kruskal-Wallis with Dunn’s multiple comparison ( g ), or mixed-effects ANOVA with Bonferroni’s multiple comparison test ( h - o ) and significant differences were indicated with p values. Source data are provided as a Source Data File.

Article Snippet: Following incubation with Fc-block (anti-mouse CD16/32, clone 2.4G2, BD Biosciences), cell surface markers were detected using fluorochrome-conjugated antibodies: CD45 (30-F11, Invitrogen/ eBioscience), CD11b (M1/70, BD Biosciences), Siglec-F (E50-2440, BD Biosciences), Ly6G (1A8, BD Biosciences), Ly6C (AL-21, BD Biosciences), F4/80 (BM8, eBioscience), MHC II (M5/114.15.2, eBioscience), CD11c (N418, BD Biosciences), CD3e (500A2, BD Biosciences), NK-1.1 (PK136, BD Biosciences), CD8a (53-6.7, BD Biosciences), CD4 (RM4-5, BD Biosciences), CD25 (PC61, BD Biosciences), CD69 (H1.2F3, eBioscience).

Techniques: Expressing, Comparison

Journal: PLoS ONE

Article Title: Bortezomib Resistance Can Be Reversed by Induced Expression of Plasma Cell Maturation Markers in a Mouse In Vitro Model of Multiple Myeloma

doi: 10.1371/journal.pone.0077608

Figure Lengend Snippet: A. Fluorescence-activated cell sorting analysis of BzS (solid black line), BzR (dark grey histogram) and I-BzR (light grey histogram) cells stained with indicated antibodies. B. Quantitative RT-PCR analysis of Cd93 and Cd69 relative mRNA expression in 595 BzS and BzR and 589 BzS, BzR, and I-BzR cell lines. Values were normalized to Gapd mRNA and error bars represent PCR triplicates. Significance was determined using a one-tailed Student’s t-test (*p<0.05; **p<0.01; ****p<0.0001). C. Fluorescence-activated cell sorting dot plot analysis of BzS and BzR cells double stained with CD93 and CD69 antibodies. Isotype controls for CD69 (FITC) and CD93 (APC) staining have been provided in .

Article Snippet: Cells were stained with the following anti-mouse antibodies: CD184/Cxcr4 PE (clone 2B11, also used for anti-human Cxcr4), CD69 FITC (clone H1.2F3), CD93 APC (clone AA4.1), CD20 PE (clone AISB12), CD22 FITC (clone 2D6), CD27 FITC (clone LG.7F9) (all from eBioscience, San Diego, CA), CD138/syndecan-1 PE (clone 281-2), CD19 PE (clone 1D3), B220 APC (clone RA3-6B2) (all from BD Biosciences, Franklin Lakes, NJ), IgM-FITC (SouthernBiotech, Birmingham, AL), CD38 (BioLegend, San Diego, CA) and analyzed using the FACSCalibur (BD Biosciences, Franklin Lakes, NJ).

Techniques: Fluorescence, FACS, Staining, Quantitative RT-PCR, Expressing, One-tailed Test

Journal: PLoS ONE

Article Title: Bortezomib Resistance Can Be Reversed by Induced Expression of Plasma Cell Maturation Markers in a Mouse In Vitro Model of Multiple Myeloma

doi: 10.1371/journal.pone.0077608

Figure Lengend Snippet: A–B. Fluorescence-activated cell sorting analysis of 589 BzS cells in the presence (+) or absence (−) of 64 nM Bz for 48 hours. Live cells were gated based on FSC and SSC and dot plots representing CD93 and CD69 double stained populations in A. In B., histograms show live untreated BzS cells (solid black line), BzS cells treated with 64 nM Bz for 48 hours (dotted black line) and untreated BzR cells (dark grey histogram) stained with CD93 and CD69 antibodies. C. Quantitative RT-PCR analysis of Cd93 and Cd69 mRNA in 589 lines corresponding to . Values were normalized to Gapd mRNA and error bars represent PCR triplicates. Significance was determined using a one-tailed Student’s t-test (****p<0.0001; NS = not significant).

Article Snippet: Cells were stained with the following anti-mouse antibodies: CD184/Cxcr4 PE (clone 2B11, also used for anti-human Cxcr4), CD69 FITC (clone H1.2F3), CD93 APC (clone AA4.1), CD20 PE (clone AISB12), CD22 FITC (clone 2D6), CD27 FITC (clone LG.7F9) (all from eBioscience, San Diego, CA), CD138/syndecan-1 PE (clone 281-2), CD19 PE (clone 1D3), B220 APC (clone RA3-6B2) (all from BD Biosciences, Franklin Lakes, NJ), IgM-FITC (SouthernBiotech, Birmingham, AL), CD38 (BioLegend, San Diego, CA) and analyzed using the FACSCalibur (BD Biosciences, Franklin Lakes, NJ).

Techniques: Fluorescence, FACS, Staining, Quantitative RT-PCR, One-tailed Test

Journal: PLoS ONE

Article Title: Bortezomib Resistance Can Be Reversed by Induced Expression of Plasma Cell Maturation Markers in a Mouse In Vitro Model of Multiple Myeloma

doi: 10.1371/journal.pone.0077608

Figure Lengend Snippet: A. Quantitative RT-PCR analysis of Irf-4 , Blimp-1 , Ddit3 , and Cxcr4 mRNA expression in 589 untreated cells and following 72 hours of LPS treatment. Values were normalized to Gapd mRNA, and error bars represent PCR triplicates. Significance was determined using a one-tailed Student’s t-test (*p<0.05; **p<0.01; ***p<0.001; ****p<0.0001; NS = not significant). B. Densitometry values representing the percentage of spliced/unspliced (s/u) Xbp-1 in 589 BzS, BzR and I-BzR cells untreated or treated with LPS for 72 hours. C. Quantitative RT-PCR analysis of Cd69 mRNA expression in 589 untreated cells and 72 hour LPS-treated cells. Values were normalized to Gapd mRNA and error bars represent PCR triplicates. Significance was determined using a one-tailed Student’s t-test (*p<0.05; ***p<0.001). D. Fluorescence-activated cell sorting analysis of untreated (solid black line) BzS (top panel), BzR (middle panel), or I-BzR (bottom panel) and LPS-treated (dotted lines) cells stained for CD69 protein expression.

Article Snippet: Cells were stained with the following anti-mouse antibodies: CD184/Cxcr4 PE (clone 2B11, also used for anti-human Cxcr4), CD69 FITC (clone H1.2F3), CD93 APC (clone AA4.1), CD20 PE (clone AISB12), CD22 FITC (clone 2D6), CD27 FITC (clone LG.7F9) (all from eBioscience, San Diego, CA), CD138/syndecan-1 PE (clone 281-2), CD19 PE (clone 1D3), B220 APC (clone RA3-6B2) (all from BD Biosciences, Franklin Lakes, NJ), IgM-FITC (SouthernBiotech, Birmingham, AL), CD38 (BioLegend, San Diego, CA) and analyzed using the FACSCalibur (BD Biosciences, Franklin Lakes, NJ).

Techniques: Quantitative RT-PCR, Expressing, One-tailed Test, Fluorescence, FACS, Staining