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

Article Title: Involvement of CD56brightCD11c+ cells in IL-18-mediated expansion of human γδ T cells.

doi: 10.4049/jimmunol.1001919

Figure Lengend Snippet: FIGURE 1. Effect of IL-18 on the growth of gd T cells and CD56brightCD11c+ cells. A, Effect of IL-18 (100 ng/ml) on expansion of gd T cells. PBMCs derived from a representative healthy adult individual were stimulated by ZOL (1 mM)/IL-2 (10 ng/ml) or 2M3BPP (100 nM–1 mM)/IL-2 (solid bar); ZOL/IL-2/IL-18 or 2M3BPP/IL-2/IL-18 (open bar); or ZOL/IL- 2/anti–IL-18Ra mAb (2 mg/ml; gray bar) or 2M3BPP/IL-2/anti–IL-18Ra mAb. The total number of gd T cells was quantified by trypan blue dye exclusion and flow cytometry. **p , 0.001. B, Massive cell aggregation induced by ZOL/IL-2/IL-18. Cell clusters were photo- graphed using a Nikon Digital Sight TE300- HM-2 (310) microscope (Nikon, Tokyo, Ja- pan) after 5 d of culture and analyzed by Lu- mina Vision Software (Mitani, Tokyo, Japan). C, Proportion of gd TCR-bearing cells in cul- ture on days 0 and 14. PBMCs were incubated in the presence of ZOL/IL-2/IL-18 for 14 d and analyzed for expression of CD3 and gd TCR. D, Phenotypic analysis of PBMCs in culture on days 0 and 14. PBMCs incubated with ZOL/ IL-2 or ZOL/IL-2/IL-18 were analyzed for expression of CD11c, CD3, and CD56 on days 0 and 14. CD56intCD11c2, CD56intCD11c+, CD56brightCD11c2, and CD56brightCD11c+ cells were gated, respectively. E, Time course of the expansion of gd T cells and CD56bright

Article Snippet: Depletion of CD3 +, CD56+, and CD14+ cells from PBMCs was conducted using LD columns and microbeads conjugated with mAbs to CD3, CD56, and CD14 (Miltenyi Biotec, Auburn, CA), respectively. gd T cells and CD14+ cells were purified by positive selection using MS columns (anti-TCR gd MicroBeads kit; Miltenyi Biotec).

Techniques: Derivative Assay, Cytometry, Microscopy, Software, Incubation, Expressing

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

Article Title: Involvement of CD56brightCD11c+ cells in IL-18-mediated expansion of human γδ T cells.

doi: 10.4049/jimmunol.1001919

Figure Lengend Snippet: FIGURE 2. Effect of depletion of CD56+ or CD14+ cells on the expansion of gd T cells. Whole PBMCs and PMBCs depleted of CD56+ or CD14+

Article Snippet: Depletion of CD3 +, CD56+, and CD14+ cells from PBMCs was conducted using LD columns and microbeads conjugated with mAbs to CD3, CD56, and CD14 (Miltenyi Biotec, Auburn, CA), respectively. gd T cells and CD14+ cells were purified by positive selection using MS columns (anti-TCR gd MicroBeads kit; Miltenyi Biotec).

Techniques:

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

Article Title: Involvement of CD56brightCD11c+ cells in IL-18-mediated expansion of human γδ T cells.

doi: 10.4049/jimmunol.1001919

Figure Lengend Snippet: FIGURE 3. Expansion of CD56brightCD11+ cells by IL-2/IL-18 in CD3-depleted PBMCs. A, Expan- sion of CD56brightCD11c+ cells in CD3-depleted PBMCs. CD3-depleted PBMCs were incubated in the presence of ZOL/IL-2 or ZOL/IL-2/IL-18 and then analyzed for expression of CD3, CD11c, and CD56. B, Proliferation of PBMCs depleted of CD3+ cells. CD3-depleted PBMCs were incubated in the presence of various cyto- kines in combinations, as indicated, and the number of living cells was counted at indicated time points. C, Expansion of CD56brightCD11c+ cells in CD3- depleted PBMCs. The total number of CD56bright

Article Snippet: Depletion of CD3 +, CD56+, and CD14+ cells from PBMCs was conducted using LD columns and microbeads conjugated with mAbs to CD3, CD56, and CD14 (Miltenyi Biotec, Auburn, CA), respectively. gd T cells and CD14+ cells were purified by positive selection using MS columns (anti-TCR gd MicroBeads kit; Miltenyi Biotec).

Techniques: Incubation, Expressing

Journal: bioRxiv

Article Title: Alteration of nociceptive Schwann cells in a mouse model of peripheral neuropathy in prediabetic condition

doi: 10.1101/2024.03.12.584541

Figure Lengend Snippet: (A) PGP9.5 immunofluorescence with DAPI staining in foot skin section of control (n=5) and HFD-fed mice (n=5). (B) Quantitation of IENFD is presented as the number of fibers/mm of epidermis. (C) L1CAM immunofluorescence with DAPI staining in foot skin section of control (n=5) and HFD-fed mice (n=5). Arrowhead indicate nerve fibers in the epidermis of the foot skin. Arrows indicate nociceptive Schwann cells and their cellular extensions at the border of the epidermis and the dermis. (D) Mean fluorescence intensity quantification of L1CAM immunostaining at the localization of nociceptive Schwann cells (at the border of epidermis and dermis). (E) Quantification of L1CAM-positive cells and their cellular extensions presented in number/mm of epidermis. ** P < 0.01, *** P < 0.001: control diet vs. HFD. Data are presented as means ±SEM. Scale bar: 50 μm.

Article Snippet: The same protocol was applied for L1 cell adhesion molecule (L1CAM, 1:500, #20659-1-AP, ProteinTech) antibody but the sections were incubated only overnight at 4°C.

Techniques: Immunofluorescence, Staining, Control, Quantitation Assay, Fluorescence, Immunostaining

Journal: The Journal of Biological Chemistry

Article Title: The long non-coding RNA HOTAIR enhances pancreatic cancer resistance to TNF-related apoptosis-inducing ligand

doi: 10.1074/jbc.M117.786830

Figure Lengend Snippet: Overexpression of HOTAIR attenuates TRA-8-induced apoptosis in sensitive pancreatic cancer cells. A and B, BxPC3 (A) and MiaPaCa-2 (B) cells were infected with lentiviruses carrying control vector or HOTAIR cDNA (HOTAIR), and stable clones were selected by puromycin. Panels Aa and Ba, HOTAIR expression, as determined by qRT-PCR and normalized by β-actin expression (n = 3, ***, p < 0.001). Panels Ab and Bb, TRA-8-induced apoptosis. BxPC3 and MiaPaCa-2 cells with HOTAIR overexpression and their control vector cells were seeded into 6-well plates at 2 × 105 per well. After culturing for 24 h, cells were exposed to TRA-8 (1 μg/ml) for 24 h, and apoptosis was determined by flow cytometry using Annexin PE and 7 AAD staining kit. TRA-8-induced apoptosis is shown in the hatched bars (n = 3, ***, p < 0.001). Panels Ac and Bc, Western blot analysis of the expression of caspase-8 (Casp8). The expression of β-actin was used as a loading control. Representative blots from three independent experiments are shown.

Article Snippet: Antibodies were purchased as follows: rabbit anti-DR5 antibody (ProSci, no. 2019, lot number 5355–1502), mouse anti-caspase 8 (BIOSOURCE no. AHZ0502, lot number 22363–01S), mouse anti-β-actin (Sigma, no. A5541–2MI, lot number 014M4759), rabbit anti-EZH2 (Cell Signaling Technology, no. D2C9, lot number 7), and mouse anti-trimethyl-histone H3 (lysine 27) (Active Motif, no. 61017, lot number 23115012).

Techniques: Over Expression, Infection, Plasmid Preparation, Clone Assay, Expressing, Quantitative RT-PCR, Flow Cytometry, Staining, Western Blot