immunomagnetic negative selection easysep mouse t cell isolation kit  (STEMCELL Technologies Inc)

Journal: Nature Communications

Article Title: Bacteroides fragilis polysaccharide A induces IL-10 secreting B and T cells that prevent viral encephalitis

doi: 10.1038/s41467-019-09884-6

Figure Lengend Snippet: PSA reduces CNS inflammation in HSV-infected WT but not Rag mice. a % and b total numbers (#) of CD45 high leukocytes and CD45 high Ly6C high inflammatory monocytes (IM) infiltrating the brainstem (BS) of Rag mice. c % (left y -axis) and # (right y -axis) infiltrating CD45 high leukocytes in the BS of WT mice. d % CD11b + cells within BS infiltrating CD45 high cells; e % Ly6C high and CD107 + IM within the CD11b + population; f % CD4 + and CD8 + T cells within CD45 high cells in the BS of WT mice. Data compiled from 2 to 4 experiments with n = 6–8/group at day 6 pi. All data show mean ± SEM. *** p < 0.0005, **** p < 0.0001, ns: not significant, as determined by two-tailed Student's t -test

Article Snippet: T cells and B cells were isolated using EasySep mouse CD4, CD8 or CD3 T cell and CD19 B cell immunomagnetic negative selection isolation kits by as per manufacturers recommendations (Stemcell Technologies).

Techniques: Infection, Two Tailed Test

Journal: Nature Communications

Article Title: Bacteroides fragilis polysaccharide A induces IL-10 secreting B and T cells that prevent viral encephalitis

doi: 10.1038/s41467-019-09884-6

Figure Lengend Snippet: PSA protection from HSE is independent of induced Tregs. a % FoxP3 + CD4 Tregs and b CD69 + CD4 T cells in spleen and CLN of PSA or PBS-treated WT mice at day 6 pi. c CD25 expression within FoxP3 + CD4 Tregs in WT mice at day 6 pi, % and mean fluorescence intensity (MFI) in () shown in right top quadrant. d % CD25 within FoxP3 + Tregs (left plot) and FoxP3 − CD4 + T cells (right plot), e CD103 expression within FoxP3 + Tregs in WT mice at day 6 pi; % and MFI in () shown in right top quadrant. f CD103 within FoxP3 + Tregs (left plot) and FoxP3 − CD4 + T cells (right plot) in the spleen or CLN of WT mice at day 6 pi. Data from three experiments shown. g PSA-treated Treg depleted and control WT mice were monitored for survival after HSV infection and ACV treatment as in Fig. , ns: not significant determined by log rank Mantel–Cox test ( n = 11–12 mice). After administration of three (1 week) or six doses (2 weeks) of PSA, MLN in uninfected WT mice were monitored for h cellularity, i % CD4 and CD8 T cells, and j # ICOS + , CD39 + , and CD73 + CD4 and CD8 T cells ( n = 3 mice); **** p < 0.0001, ** p < 0.01 as determined by two-way ANOVA or one-way ANOVA with Sidaks or Turkeys correction, respectively, for multiple comparisons tests. All data show mean ± SEM

Article Snippet: T cells and B cells were isolated using EasySep mouse CD4, CD8 or CD3 T cell and CD19 B cell immunomagnetic negative selection isolation kits by as per manufacturers recommendations (Stemcell Technologies).

Techniques: Expressing, Fluorescence, Control, Infection

Journal: Nature Communications

Article Title: Bacteroides fragilis polysaccharide A induces IL-10 secreting B and T cells that prevent viral encephalitis

doi: 10.1038/s41467-019-09884-6

Figure Lengend Snippet: PSA increases IL-10 and IFNγ-secreting T cells. CD4 and CD8 T cells and B cells in spleens, mesenteric lymph nodes (MLN), and cervical lymph nodes (CLN) of PSA or PBS-treated WT mice at day 6 pi were analyzed for a IL-10 and b IFNγ secretion, n = 2 experiments; * p < 0.05, ** p < 0.01, **** p < 0.0001, as determined by two-way ANOVA with Sidak’s multiple comparisons test. Survival of PSA or PBS treated c IL-10KO mice or d IFN-GKO mice ( n = 8–16 mice); ns: not significant. Bar plots show e % CD45 high leukocytes, f (left y -axis) % Ly6C high IM and (right y -axis) % Ly6G + neutrophils (PMN) within CD45 high CD11b + cells infiltrating the BS of PSA treated 129 WT, IL10KO, and GKO mice at day 6 pi, n = 3 experiments with 2–3 BS/group; * p < 0.05, **** p < 0.0001 as determined by ordinary one-way ANOVA with Turkey’s multiple comparisons tests

Article Snippet: T cells and B cells were isolated using EasySep mouse CD4, CD8 or CD3 T cell and CD19 B cell immunomagnetic negative selection isolation kits by as per manufacturers recommendations (Stemcell Technologies).

Techniques:

Journal: Nature Communications

Article Title: Bacteroides fragilis polysaccharide A induces IL-10 secreting B and T cells that prevent viral encephalitis

doi: 10.1038/s41467-019-09884-6

Figure Lengend Snippet: PSA protection against HSE requires B and T cells secreting IL-10. a Experimental design for experiments in b and c Donor WT (In black text): Naïve Rag mice were transferred with WT CD4 + or CD8 + T cells or CD19 + B cells 7 days before PSA treatment. Donor IL-10KO (magenta text) and WT (Blue text): four groups of naïve Rag mice were transferred with combinations of donor WT B and T cells, IL-10KO B and T cells, WT B and IL-10KO T cells, IL-10KO B and WT T cells 7-days before PSA treatment. All Rag recipients received six doses of PSA before HSV infection and ACV treatment. b Survival of B cell-depleted mice (BKO, n = 20 mice) and Rag recipients of WT single cell subsets ( n = 6–9 mice/group). B cell depletion in WT mice was initiated 10 days prior to PSA treatment and continued throughout infection, ns: not significant. c Survival of Rag recipients of WT and IL-10KO combination of T and B cells ( n = 10–13/group). *** p < 0.001, * p < 0.05, ns: not significant as determined by log rank (Mantel–Cox) test. FACS plots of BS CD45 high cells (left), Ly6G + PMN (left middle), CD11b + cells within CD45 high cells (right middle), and Ly6C high IM and Ly6C int CD11b + PMN within CD45 high CD11b + cells (right) were analyzed at day 6 pi in the BS of Rag recipients of d IL-10KO B + WT T cells (brown circle) and e WT B + IL-10KO T cells (green circle)

Article Snippet: T cells and B cells were isolated using EasySep mouse CD4, CD8 or CD3 T cell and CD19 B cell immunomagnetic negative selection isolation kits by as per manufacturers recommendations (Stemcell Technologies).

Techniques: Infection

Journal: Nature Communications

Article Title: Bacteroides fragilis polysaccharide A induces IL-10 secreting B and T cells that prevent viral encephalitis

doi: 10.1038/s41467-019-09884-6

Figure Lengend Snippet: Role of the bacterial symbiosis factor PSA in preventing viral encephalitis. HSV infection of susceptible 129 WT mice provokes excessive production of neutrophils (PMN) and Ly6C high inflammatory monocytes (IM) in the bone marrow that invade the brainstem in massive numbers resulting in fatal HSV encephalitis (HSE), despite antiviral treatment from day 4 pi. The bacterial symbiosis factor, PSA given orally is bound by B cells/CD138 + plasmablasts (PB) in the small intestine, which induces IL-10 and IFNγ production by regulatory CD4 and CD8 T cells resulting in the suppression of pathogenic inflammatory myeloid cells concomitant with the induction of IFNγ inducible chemokines in the BS. This novel study reveals the immunomodulatory potential of PSA in protecting from lethal viral infections of the CNS in combination with an antiviral. Cells involved in this protective mechanism are shown in the key. Inhibitory pathways indicated by red blocking arrows

Article Snippet: T cells and B cells were isolated using EasySep mouse CD4, CD8 or CD3 T cell and CD19 B cell immunomagnetic negative selection isolation kits by as per manufacturers recommendations (Stemcell Technologies).

Techniques: Infection, Blocking Assay