Journal: Frontiers in Immunology

Article Title: GPR182 is a broadly scavenging atypical chemokine receptor influencing T-independent immunity

doi: 10.3389/fimmu.2023.1242531

Figure Lengend Snippet:

Article Snippet: Anti-mCXCL13 , 143614 , 1:1000 , RnD Systems , MAB470.

Techniques: Concentration Assay

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: CXCL13 Blockade Disrupts B Lymphocyte Organization in Tertiary Lymphoid Structures without Altering B Cell Receptor Bias or Preventing Diabetes in Nonobese Diabetic Mice

doi: 10.4049/jimmunol.0903710

Figure Lengend Snippet: CXCL13 is expressed in inflamed islets of NOD mice. A, Gel electrophoresis showing RT-PCR products from isolated pancreatic islets using cxcl13-specific primers. First three lanes are from inflamed islets of three independent female NOD mice. Last three lanes are from islets of three independent female C57BL/6 mice. Lower lanes are controls using hprt primers from the same islets. B, Real-time PCR showing cxcl13 levels, normalized to hprt, from isolated islets. At least 20 islets were isolated from three mice per group, and pooled for each mouse. Normalized cxcl13 levels are significantly higher in islets from 12-wk-old NOD mice than from any other group. **p < 0.0001, compared with noninfiltrated NOR controls or 3-wk-old NOD mice; p < 0.001 for 6-wk-old NOD mice; p = 0.004 compared with 14-wk-old NOR mice. C, Immunohistochemical staining of frozen NOD pancreatic sections shows CXCL13+ (dark brown) cells among invading lymphocytes in early insulitis. Right panel shows control section processed with secondary reagents, but without primary anti-CXCL13 Ab (original magnification ×20).

Article Snippet: CXCL13 blockade Wild-type or V H 125 transgenic NOD mice were i.p. injected with 100 μg monoclonal, rat-derived, anti-CXCL13 Ab (mAB470, R&D Systems, Minneapolis, MN) in sterile PBS three times weekly, from age 3 wk to the end of each study.

Techniques: Nucleic Acid Electrophoresis, Reverse Transcription Polymerase Chain Reaction, Isolation, Real-time Polymerase Chain Reaction, Immunohistochemical staining, Staining, Control

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: CXCL13 Blockade Disrupts B Lymphocyte Organization in Tertiary Lymphoid Structures without Altering B Cell Receptor Bias or Preventing Diabetes in Nonobese Diabetic Mice

doi: 10.4049/jimmunol.0903710

Figure Lengend Snippet: CXCL13 receptor is expressed on B cells in pancreas and spleen. Flow cytometry shows CXCR5 expression on B220+ B cells from pancreas and spleen of the same female NOD mouse. Image is representative of data from six mice.

Article Snippet: CXCL13 blockade Wild-type or V H 125 transgenic NOD mice were i.p. injected with 100 μg monoclonal, rat-derived, anti-CXCL13 Ab (mAB470, R&D Systems, Minneapolis, MN) in sterile PBS three times weekly, from age 3 wk to the end of each study.

Techniques: Flow Cytometry, Expressing

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: CXCL13 Blockade Disrupts B Lymphocyte Organization in Tertiary Lymphoid Structures without Altering B Cell Receptor Bias or Preventing Diabetes in Nonobese Diabetic Mice

doi: 10.4049/jimmunol.0903710

Figure Lengend Snippet: CXCL13 blockade does not stop B lymphocyte invasion of pancreatic islets, but scrambles tertiary lymphoid organization. A, Immunofluorescent staining of a typical, untreated, inflamed islet from a 12-wk-old female NOD mouse, with B220+ B cells (pseudocolored pink) surrounding CD3+ T cells (pseudocolored blue) (original magnification ×20). B, Immunofluorescent staining of a typical inflamed islet from a 12-wk-old female NOD mouse treated with CXCL13-blocking Ab from age 3 wk (original magnification ×20). C, Bar chart showing B lymphocytes as proportion of total lymphocytes extracted from pancreata of 12-wk-old female NOD mice treated with anti-CXCL13, isotype control, or not treated, analyzed by flow cytometry. Bar height shows average for three mice per group, with SD as indicated. D, Bar chart showing percent of inflamed islets classified as organized, disorganized, or intermediate from 12-wk-old female NOD mice treated with anti-CXCL13, isotype control Abs, or not treated. p < 0.001 for CXCL13 versus isotype-treated controls; p < 0.001 for CXCL13 versus untreated controls; p = 0.9 for isotype-treated controls versus untreated controls. Independent islets from anti-CXCL13, n = 34; isotype control, n = 27; untreated, n = 18; anti-CXCL13–treated mice, n = 3; isotype control mice, n = 2; untreated controls mice, n = 2.

Article Snippet: CXCL13 blockade Wild-type or V H 125 transgenic NOD mice were i.p. injected with 100 μg monoclonal, rat-derived, anti-CXCL13 Ab (mAB470, R&D Systems, Minneapolis, MN) in sterile PBS three times weekly, from age 3 wk to the end of each study.

Techniques: Staining, Blocking Assay, Control, Flow Cytometry

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: CXCL13 Blockade Disrupts B Lymphocyte Organization in Tertiary Lymphoid Structures without Altering B Cell Receptor Bias or Preventing Diabetes in Nonobese Diabetic Mice

doi: 10.4049/jimmunol.0903710

Figure Lengend Snippet: CXCL13 blockade fails to alter selection of the B cell repertoire found in pancreatic islets. A, LC gene families expressed by B cells from pancreata (black bars) and draining PLNs (gray bars) of 12-wk-old prediabetic female VH125/NOD mice treated with anti-CXCL13 from the age of 3 wk. p = 0.002 by Fisher exact test; mice, n = 3; independent clones, n = 53. B, The ratio of predominant B lymphocyte Vκ families found in pancreata, relative to those from PLNs, in untreated VH125/NOD (black bars) and anti-CXCL13–treated VH125/NOD mice (gray bars). p = 0.066; 95% CI 0.77–1.01 by Poisson regression; mice, n = 8; independent clones, n = 104.

Article Snippet: CXCL13 blockade Wild-type or V H 125 transgenic NOD mice were i.p. injected with 100 μg monoclonal, rat-derived, anti-CXCL13 Ab (mAB470, R&D Systems, Minneapolis, MN) in sterile PBS three times weekly, from age 3 wk to the end of each study.

Techniques: Selection, Clone Assay

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: CXCL13 Blockade Disrupts B Lymphocyte Organization in Tertiary Lymphoid Structures without Altering B Cell Receptor Bias or Preventing Diabetes in Nonobese Diabetic Mice

doi: 10.4049/jimmunol.0903710

Figure Lengend Snippet: CXCL13 blockade does not significantly decrease CDR mutations found in BCR LCs from pancreata. Gene segments encoding BCR κ LCs cloned from pancreata of VH125/NOD mice were examined for evidence of somatic hypermutation as indicated by mutations in the CDRs. A, A total of 35% of BCR LC gene segments from pancreata of untreated controls have mutations in the CDR-encoding region. B, A total of 34% of BCR LC gene segments from anti-CXCL13–treated mice have mutations in the CDR-encoding regions. Keys show shading corresponding to number of mutations. Untreated, n = 45 clones; anti-CXCL13, n = 29 clones, from at least three mice per group. p = 0.861 by negative binomial regression.

Article Snippet: CXCL13 blockade Wild-type or V H 125 transgenic NOD mice were i.p. injected with 100 μg monoclonal, rat-derived, anti-CXCL13 Ab (mAB470, R&D Systems, Minneapolis, MN) in sterile PBS three times weekly, from age 3 wk to the end of each study.

Techniques: Clone Assay

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: CXCL13 Blockade Disrupts B Lymphocyte Organization in Tertiary Lymphoid Structures without Altering B Cell Receptor Bias or Preventing Diabetes in Nonobese Diabetic Mice

doi: 10.4049/jimmunol.0903710

Figure Lengend Snippet: CXCL13 blockade does not protect against development of diabetes. Female VH125/NOD mice were administered monoclonal anti-CXCL13 (◆) or isotype control (■), or were untreated (▲). Blood glucose levels were checked weekly, and mice diagnosed with diabetes at >200 mg/dl. Study was stopped for lack of efficacy when at least 50% of anti-CXCL13–treated mice had become diabetic. Mice, n = 10 per group. Log rank p = 0.94 CXCL13 versus isotype control treated; log rank p = 0.27 anti-CXCL13 versus untreated controls.

Article Snippet: CXCL13 blockade Wild-type or V H 125 transgenic NOD mice were i.p. injected with 100 μg monoclonal, rat-derived, anti-CXCL13 Ab (mAB470, R&D Systems, Minneapolis, MN) in sterile PBS three times weekly, from age 3 wk to the end of each study.

Techniques: Control

Journal: The Journal of Clinical Investigation

Article Title: Bronchus-associated lymphoid tissue–resident Foxp3 + T lymphocytes prevent antibody-mediated lung rejection

doi: 10.1172/JCI122083

Figure Lengend Snippet: (A) Foxp3+ (green), B cells (blue), and CD4+ T cells (red) in BALB/c lungs at least 30 days after transplantation into immunosuppressed B6 Foxp3-IRES GFP recipient (n = 3). Scale bar: 10 μm. CXCR5+ B cells from secondary host (recipient) in BALB/c lungs, initially transplanted into immunosuppressed (B) WT or (C) Foxp3-DTR B6 (CD45.2+) recipient and, at least 30 days later, retransplanted into DT-treated B6 CD45.1+ hosts. Plots are gated on live CD45.2–CD45.1+ cells. (D) CD45.1+CXCR5+ B cells in (circles) control and (inverted triangles) Foxp3+ T cell–depleted lungs 7 days after retransplantation (n = 4 each). CXCL13 (brown) in (E) control and (F) Foxp3+ T cell–depleted grafts 7 days after retransplantation. Scale bars: 100 μm. CD4+ T cells (green) and B cells (blue) in BALB/c lungs, initially transplanted into immunosuppressed B6 Foxp3-DTR recipients and, at least 30 days later, retransplanted into B6 hosts, treated with (G) DT/control-Ig (arrows: CD4+ T–B cell interactions) or (H) DT/anti-CXCL13 (n = 2 each) (red, quantum dots). Scale bars: 20 μm. (I) Contact duration between CD4+ T and B cells, (J) CD4+ T, and (K) B cell mean square displacements and (L) CD4+ T and (M) B cell velocities within retransplanted Foxp3+ T cell–depleted BALB/c lungs with and without CXCL13 inhibition. (N) Gross, (O) histological appearance (H&E), staining for (P) MT (blue), (Q) CCSP (red), and AcT (green) in BALB/c lungs, transplanted into immunosuppressed B6 Foxp3-DTR mice and, at least 30 days later, retransplanted into DT- and anti-CXCL13–treated B6 hosts. Scale bars: 100 μm. (R) Donor-specific IgM titers after retransplantation of BALB/c lungs into DT-treated control (blue) or DT/anti-CXCL13 antibody–treated (red) B6 recipients after initial engraftment into immunosuppressed B6 Foxp3-DTR mice (n = 4 mice per group). Data are expressed as mean ± SEM. Mann-Whitney U test was used to compare the means.

Article Snippet: Primary antibodies used were rabbit monoclonal anti-human Foxp3 (clone SP97, 1:400, Novus), rabbit polyclonal anti-mouse CXCL13 (1:100, Bioss), and rat monoclonal anti-mouse PNAd (clone MECA-79, 1:100, BD Biosciences).

Techniques: Transplantation Assay, Inhibition, Staining, MANN-WHITNEY

Journal: The Journal of Experimental Medicine

Article Title: Follicular dendritic cells help establish follicle identity and promote B cell retention in germinal centers

doi: 10.1084/jem.20111449

Figure Lengend Snippet: DTx-mediated selective FDC ablation disrupts B cell follicle architecture despite retention of CXCL13. Unimmunized CD21-DTR chimeras were control (Ctrl) or DTx treated 2 d before analysis of spleen (spl) and pLN sections. (A) Immunofluorescence analysis for BP3 + stromal cells, CD35 + FDCs, and IgD + follicular B cells. (B) In situ hybridization for Cxcl13 mRNA with serial sections stained for B220 and CD35 or for Cxcl13 and CD35. (C) Quantitative RT-PCR analysis for Cxcl13 , Cr1 (CD35), Mfge8 , and Tnsf13B (BAFF) mRNA in pLNs, spleen, and mLNs, normalized to Hprt mRNA. Data are representative of four experiments (mean ± SEM). Statistical analysis was performed with the two-tailed unpaired Student’s t test. *, P < 0.05; n.s., not significant. (D) Immunohistochemical analysis for CXCL13 and CD3 or B220 in serial sections of spleen and pLNs. Data in A, B, and D are representative of at least three experiments (at least two mice of each type per experiment). Bars: (A [left] and B [left and middle]) 200 µm; (A and D, right) 100 µm; (B [right] and D [left]) 50 µm.

Article Snippet: Cryosections of 7 µm were fixed and stained immunohistochemically as previously described ( ) with the following first antibodies: PE-conjugated anti-IgD (11-26c.2a; BD), biotin-conjugated anti-CD35 (8C12; BD), rat anti–mouse fibroblast (ER-TR7; Novus Biologicals), rabbit anti–mouse Collagen IV (Abcam), rat anti–mouse B220 (RA-6B2; BD), biotin-conjugated anti–T/B cell activation antigen or Ly77 (GL7; BD), biotin conjugated anti–mouse CD3e (145-2C11; BD), goat anti–mouse CXCL13 (R&D Systems), goat anti–mouse CCL21 (R&D Systems), rat anti-CD169 (Ser4; P. Crocker, University of Glasgow, Glasgow, Scotland, UK), rat anti–mouse madcam-1 (MECA-367; BD), rat anti–mouse FDC (FDC-M1; BD), and FITC-conjugated rat anti–mouse CD16/32 (FcγRII/III; UCSF Hybridoma Core).

Techniques: Immunofluorescence, In Situ Hybridization, Staining, Quantitative RT-PCR, Two Tailed Test, Immunohistochemical staining