Journal: Frontiers in Immunology

Article Title: Mesenchymal stromal cells induced regulatory B cells are enriched in extracellular matrix genes and IL-10 independent modulators

doi: 10.3389/fimmu.2022.957797

Figure Lengend Snippet: MSC induce regulatory B cells enriched in transitional B cell phenotypes and secrete IL-10. (A) Graphical protocol for the generation of in vitro induced regulatory B cells (iBreg) and activated B cells and downstream analysis including gating strategy of flow cytometry samples. (B) Representative FACS plots showing the percentage of Transitional B (CD19 + CD24 Hi CD38 Hi ) cells from alive B cells (CD19 + 7AAD - ). (C) Summarized data for the frequency of Transitional B cells, showing a significant increase at day 7 (D) Representative FACS plots showing the percentage of naïve B cells (CD19 + CD27 - ) from alive B cells (CD19 + 7AAD - ). (E) Summarized data for the frequency of Naïve B cells showing a significant increase in the presence of MSC. (F) Supernatant cytokine quantification by ELISA of IL-10. IL-10 production increased after day 5, with a peak production observed at day 7. (G) Supernatant cytokine quantification by ELISA of TNFα. TNFα production was absent in iBreg and activated B cells. Data shows at least three independent experiments in each group. Cells from 2 MSC donors and 3 Tonsil donors were used. Error bars represent SD. Two-way ANOVA was performed to determine statistical significance. *p < 0.05, **p < 0.01, ****p < 0.0001.

Article Snippet: To asses surface marker expression, iBreg cultures were labelled for flow cytometry analysis with the following antibodies: IgD-APC-Cy7 (BioLegend, clone IA6-2, San Diego, CA, USA), CD19-BV510 (BD Horizon, clone SJ25C1), CD24-APC (Invitrogen, clone eBioSN3), CD27-PE-Cy7 (Invitrogen, clone 0323), CD38-PE (Invitrogen, clone HB7) and 7AAD (BD Pharmingen).

Techniques: In Vitro, Flow Cytometry, Enzyme-linked Immunosorbent Assay

Journal: PLoS ONE

Article Title: Presence of Rheumatoid Factor during Chronic HCV Infection Is Associated with Expansion of Mature Activated Memory B-Cells that Are Hypo-Responsive to B-Cell Receptor Stimulation and Persist during the Early Stage of IFN Free Therapy

doi: 10.1371/journal.pone.0144629

Figure Lengend Snippet: ( A), HCV RNA copies IU/mL HCV RF- (●, n = 23) and HCV RF+ (▼, n = 23). ( B) Shown an uninfected donor sample for total immature transitional B cells (CD19+CD20+CD10+) and its differentiation stages: T1 stage (CD19 + CD20 + CD10 + CD21-) and T2 stage (CD19 + CD20 + CD10 + CD21 + ). Mature activated B cells(CD19 + CD20 + CD10 - CD21 - CD27 + ), resting memory B cells (CD19 + CD20 + CD21 + CD27 + ), naïve (CD19 + CD20 + CD21 + CD27 - ),Tissue like memory B cells (CD19 + CD20 + CD10 - CD21 - CD27 - ) and Plasmablast cells (CD19 + CD20 - CD38 + )

Article Snippet: Lymphocytes were assessed by forward and side scatter, and stained for the following: anti-CD19-PECy5 (clone HIB19),anti- CD20- APC-H7 (clone L27), anti-CD10- APC (clone HI10a), anti-CD21- PE (clone B-ly4), anti- CD27- PECy7 (clone M-T271) (all from BD Biosciences, San Jose, CA) and anti-CD38- Alexa Fluor 700 (clone HIT2, Biolegend, San Diego, CA).

Techniques:

Journal: PLoS ONE

Article Title: Presence of Rheumatoid Factor during Chronic HCV Infection Is Associated with Expansion of Mature Activated Memory B-Cells that Are Hypo-Responsive to B-Cell Receptor Stimulation and Persist during the Early Stage of IFN Free Therapy

doi: 10.1371/journal.pone.0144629

Figure Lengend Snippet: ( A) Absolute count of CD19+ B cell per uL of whole blood. (B) Representative flow plot of altered B cell subset distribution in whole blood from Uninfected donor, HCV RF-, and HCV RF+ donors. (C) Summary data of Proportion of mature activated memory B cells (%) in whole blood (CD19+CD20+CD10-CD21-/loCD27+) for Uninfected donors ( ▄ n = 23) , HCV RF- (● n = 23) , and HCV RF+ donors ( ▼, n = 20). (D). Summary data of Proportion of naïve B cells (CD19+CD20+CD10-CD21+CD27-) in whole blood for Uninfected donors ( ▄ n = 23) , HCV RF- (● n = 23) , and HCV RF+ donors ( ▼, n = 20) . Kruskal- Wallis was used to analyzed differences among the three groups of donors and Mann-Whitney U test was used between two groups of donors (p≤0.05)

Article Snippet: Lymphocytes were assessed by forward and side scatter, and stained for the following: anti-CD19-PECy5 (clone HIB19),anti- CD20- APC-H7 (clone L27), anti-CD10- APC (clone HI10a), anti-CD21- PE (clone B-ly4), anti- CD27- PECy7 (clone M-T271) (all from BD Biosciences, San Jose, CA) and anti-CD38- Alexa Fluor 700 (clone HIT2, Biolegend, San Diego, CA).

Techniques: MANN-WHITNEY

Journal: PLoS ONE

Article Title: Presence of Rheumatoid Factor during Chronic HCV Infection Is Associated with Expansion of Mature Activated Memory B-Cells that Are Hypo-Responsive to B-Cell Receptor Stimulation and Persist during the Early Stage of IFN Free Therapy

doi: 10.1371/journal.pone.0144629

Figure Lengend Snippet: ( A) Plasma HCV RNA level, IU/ml in HCV RF- donors (● n = 10) and HCV RF+ donors ( ▼, n = 20) . (B) Summary data of proportion of mature activated memory B cells (%) in frozen PBMCs (CD19+CD20+CD10-CD21-/loCD27+) for Uninfected donors ( ▄ n = 10) , HCV RF- (● n = 10) , and HCV RF+ donors ( ▼, n = 10). ( C) Summary data of proportion of naïve B cells (CD19+CD20+CD10-CD21+CD27-) in frozen PBMCs for Uninfected donors ( ▄ n = 10) , HCV RF- (● n = 10) , and HCV RF+ donors ( ▼, n = 10) .(D) Summary data of CD86 expression on mature activated memory B cells (CD19+CD20+CD10-CD21-/loCD27+) for Uninfected donors (▄ n = 10) and HCV RF+ donors (▼, n = 10). (E) D) Summary data of Ki67 expression on mature activated memory B cells (CD19+CD20+CD10-CD21-/loCD27+) for Uninfected donors (▄ n = 10) and HCV RF+ donors (▼, n = 10). Kruskal- Wallis was used to analyzed differences among the three groups of donors (*) and Mann-Whitney U test was used between two groups of donors (p≤0.05)

Article Snippet: Lymphocytes were assessed by forward and side scatter, and stained for the following: anti-CD19-PECy5 (clone HIB19),anti- CD20- APC-H7 (clone L27), anti-CD10- APC (clone HI10a), anti-CD21- PE (clone B-ly4), anti- CD27- PECy7 (clone M-T271) (all from BD Biosciences, San Jose, CA) and anti-CD38- Alexa Fluor 700 (clone HIT2, Biolegend, San Diego, CA).

Techniques: Expressing, MANN-WHITNEY

Journal: PLoS ONE

Article Title: Phenotypic, Genomic and Functional Characterization Reveals No Differences between CD138 ++ and CD138 low Subpopulations in Multiple Myeloma Cell Lines

doi: 10.1371/journal.pone.0092378

Figure Lengend Snippet: (A) Dot plots showing the percentage of CD138 ++ and CD138 low cells in MM cell lines. (B) Top: summary of the expression of CD19, CD20, CD27, CD38, CD45 and CD56 in CD138 ++ and CD138 low subpopulations in MM cell lines. Code: – (negative); −/+ (heterogeneity); dim (weak positive); + (positive); ++ (high positive). Bottom: representative dot plots corresponding to non-stained RPMI-8226 cells (negative control; light grey) and stained CD138 ++ (black) and CD138 low (dark grey) RPMI-8226 cells. (C) Expression of CD138 by real-time quantitative PCR in CD138 ++ and CD138 low RPMI-8226 subpopulations. Relative values were calculated by the 2 −ΔCt method (ΔCt = Ct (Gene) −Ct (GAPDH) ). The GAPDH gene was used as a control gene. Results are expressed as the means ± SEM (n = 3). (D–K) Confocal images corresponding to the immunocytochemistry for CD138 (red) in CD138 ++ and CD138 low RPMI-8226 and NCI-H929 cells. Nuclear DNA was stained with DAPI (blue). Magnification of the lens, 63x. Specific “4× zoom” was made in E, G, I, K.

Article Snippet: The origin of the antibodies used in immunocytochemistry and flow cytometry was as follows: anti-CD138-APC (clone B-B4), used for immunocytochemistry and flow cytometry, from Miltenyi Biotec (Auburn, CA); anti-CD20-FITC (clone L27), anti-CD138-PerCP-Cy5 (clone MI15), anti-CD56-APC (clone NCAM16.2), anti-CD45-AmCyan (clone 2D1) and anti-CD38-PE (clone HB7) from BD Biosciences (San Jose, CA, USA); anti-CD19-PacificBlue (clone HIB19) and anti-CD27-PE-Cy7 (clone O323) antibodies from eBioscience (San Diego CA, USA); anti-CD38-AlexaFluor700 antibody (clone HIT2) from Exbio (Vestec, Czech Republic) and anti-CD138-FITC (clone B-A38) from Cytognos S.L. (Salamanca, Spain).

Techniques: Expressing, Staining, Negative Control, Real-time Polymerase Chain Reaction, Immunocytochemistry