Fig 1: MiR-223 reduced donor T cell migration to recipient mice spleens. The proportion of H2kb + CD4 and CD8 T cell in recipient mice spleens on day 7 was assessed by flow cytometry. Compared with NC and aGvHD group, H2kb + CD4 and H2kb + CD8 cell percentages in miR-223 group showed significant reduction (a). The average number of H2kb + CD4 and H2kb + CD8 cells in miR-223 group showed significant decline (b). Data were presented as mean ± SEM. Measured in three independent experiments, n = 7 per group. *P < 0.05, **P < 0.01
Fig 2: T-cell infiltration is observed in patients with PD and MPTP-treated mice. (A) CD8+ and CD4+ T cells were measured in patients with or without PD using immunofluorescence. Scale bars, 20 µm. (B) CD8+ and CD4+ T-cell levels increased in the brain tissue of patients with PD compared with the control group. n=8. ***P<0.001. (C) The abundance of CD8+ T cells in the brain tissue positively associated with disease duration. MPTP-treated mice also harbored increased levels of (D) CD8+ and (E) CD4+ T cells in the brain tissue 4 days after MPTP injection. n=9. Experiments were performed in triplicate. ***P<0.001. PD, Parkinson's disease; MPTP, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine; CTRL, control.
Fig 3: EPZ004777 and Thiostrepton improved BMDCs maturation in vivo. (A and B) Expressions of CD86, MHC‐II, CCR7, and PD‐L1 on gated CD11c+ cells in BMDCs were assessed by FACS. (C and D) The expressions of CD3, CD8, and CD4 on gated CD3+ T cells, and Foxp3 and CD25 on gated CD4+ T cells from spleen were assessed by FACS. The amounts of CD3+/CD4+ cells set at 10 000. A/C is from C57BL/6J, B/D is from BABL/c mice, each group contained at least eight mice. Data were shown as means ± SD from at least three independent experiments. # P < 0.05, ## P < 0.01 compared with the model.
Fig 4: GAS2 impairment has mild effects on normal hematopoietic cells. (A) A typical photo of genotyping analyses of +/+, flox/flox and flox/flox;Vav-iCre (conditional knockout, CKO) mice was displayed. (B) The Gas2 transcript expression of peripheral blood (PB), bone marrow (BM), and kidney cells from both CKO (n = 8) and flox/flox mice (n = 8) was analyzed by RT-qPCR. (C) The BM cells from 8 weeks old CKO mice (n = 5) and flox/flox mice (n = 5) were analyzed by flow cytometry for the expression of Mac-1, Gr-1, B220, and Ter119. (D) The absolute numbers of Mac-1+, Gr-1+, B220+, and Ter119+ cells from the bone marrow (2 femurs and 2 tibias) of CKO and flox/flox mice were compared. (E) The PB cells from CKO mice (n = 5) and flox/flox mice (n = 5) were analyzed for the expression of CD4 and CD8. The thymus cells from CKO mice (n = 5) and flox/flox mice (n = 5) were analyzed for the expression of CD4 and CD8 as well. (F) The absolute number of CD4+ and CD8+ cells in PB (100 µL) from CKO and flox/flox mice were compared (n = 5). (G) The absolute numbers of CD4+, CD8+, and CD4+CD8+ cells in the thymus from CKO and flox/flox mice were compared (n = 5). (H) A cartoon was illustrated to depict the role of GAS2 in T-cell leukemogenesis. GAS2 interacts with CXCR4 and enhances its stability. This protein complex sustains the expression of NOTCH/c-MYC and promotes the growth and invasion of T-ALL cells. GAS2 or CXCR4 silencing suppresses the expression of NOTCH/c-MYC to blockade T-cell leukemogenesis. Data were represented as the mean ± SEM, and the statistical significance was estimated with Student's t-test (*P < 0.05, ***P < 0.001. n.s., not significant).
Fig 5: IDO1-shRNA treatment decreases the expression of inhibitory receptors of CD4+/CD8+ T cells in vivo. Lymphocytes from lymph nodes and spleen of LLC-bearing tumor mice were collected on day 21 injection of LLC cells. CD4+ and CD8+ T cells were stained with anti-PD-1 and anti-BTLA and analyzed by flow cytometry. Expression of PD-1 and BTLA on CD4+ T cells in the (A) lymph nodes and (B) spleen of tumor-bearing mice without treatment, or treated with scrambled-shRNA or IDO1-shRNA. (C) Expression of PD-1 and BTLA on CD8+ T cells from the lymph nodes of tumor-bearing mice without treatment or treated with scrambled-shRNA or IDO1-shRNA. Statistical analysis of the expression of PD1 and BTLA on CD4+ (right panel in A and B) and CD8+ (right panels in C) was performed from three independent experiments. *P<0.05 and **P<0.01. BTLA, B and T lymphocyte attenuator; IDO1, indoleamine 2,3-dioxygenase 1; LN, lymph node; PD-1, programmed death-1; sh, short hairpin; SP, spleen.
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