Journal: The Journal of Experimental Medicine

Article Title: Subspecialization of Cxcr5 + T Cells

doi:

Figure Lengend Snippet: Effector machinery of CD57 +/ −CD45RA +/ − or CD57 +/ − CXCR5 +/ − CD4 T cells. (A) Intracellular cytokine analyses of TNF-α, IL-2, IL-10, IFN-γ, and IL-4. (B) ELISA of IL-10 and IL-4. Isolated T cells were activated with 50 ng/ml PMA and 1 μg/ml ionomycin for 4 h for intracellular cytokine analyses, or 24 h for ELISA of IL-10 and IL-4. Representatives of at least four independent experiments are shown.

Article Snippet: In brief, acetone-fixed sections were stained with the primary monoclonal antibody to CXCR5 (R&D Systems), followed by incubation in biotinylated goat anti–mouse IgG (Vector Laboratories).

Techniques: Enzyme-linked Immunosorbent Assay, Isolation

Journal: The Journal of Experimental Medicine

Article Title: Subspecialization of Cxcr5 + T Cells

doi:

Figure Lengend Snippet: Identification of CD57 + CXCR5 + T cells and their localization in GCs. (A) Flow analyses of CD57 and CXCR5 expression on tonsil CD4 T cells. (B) Specific localization of CD57 + CD4 T cells in GCs. (C) Localization of CD57 + CXCR5 + CD4 T cells in GCs. Antibodies to CD57 (green), CD4 (blue), and IgD (red; B) or CXCR5 (red; C) were used for in situ immunohistochemistry.

Article Snippet: In brief, acetone-fixed sections were stained with the primary monoclonal antibody to CXCR5 (R&D Systems), followed by incubation in biotinylated goat anti–mouse IgG (Vector Laboratories).

Techniques: Expressing, In Situ, Immunohistochemistry

Journal: The Journal of Experimental Medicine

Article Title: Subspecialization of Cxcr5 + T Cells

doi:

Figure Lengend Snippet: Chemotactic responses (A) and chemokine receptor expression (B) by CD57 +/2 CXCR5 +/ − CD4 T cells. Optimal concentrations of 5 μg/ml BLC, 1 μg/ml ELC, and 100 ng/ml SDF-1 were used for chemotaxis experiments. Freshly isolated tonsil cells were used for chemotaxis and flow analyses of tonsil CD4 T cell subsets. Representatives of three independent experiments are shown.

Article Snippet: In brief, acetone-fixed sections were stained with the primary monoclonal antibody to CXCR5 (R&D Systems), followed by incubation in biotinylated goat anti–mouse IgG (Vector Laboratories).

Techniques: Expressing, Chemotaxis Assay, Isolation

Journal: The Journal of Experimental Medicine

Article Title: Subspecialization of Cxcr5 + T Cells

doi:

Figure Lengend Snippet: Spontaneous B cell helper activity of CD57 +/ − CXCR5 +/ − CD4 T cells. Sorted CD57 + CXCR5 + , CD57 − CXCR5 + , and CXCR5 − CD4 T cells were cocultured with B cells from the same tonsil for 11–13 d in the absence of any stimulatory agents followed by analyses of secreted IgG, IgA, and IgM. Representatives of at least three independent experiments are shown.

Article Snippet: In brief, acetone-fixed sections were stained with the primary monoclonal antibody to CXCR5 (R&D Systems), followed by incubation in biotinylated goat anti–mouse IgG (Vector Laboratories).

Techniques: Activity Assay

Journal: The Journal of Experimental Medicine

Article Title: Subspecialization of Cxcr5 + T Cells

doi:

Figure Lengend Snippet: Phenotype and effector function of circulating CXCR5 + T cells. (A) Surface phenotype of circulating naive or CXCR5 +/ − memory T cells. **Significant differences between CXCR5 + and CXCR5 − memory cells. (B) IL-4/IFN-γ production capabilities of CXCR5 +/ − T cells during repeated T cell receptor activation (each cycle is composed of 4-d activation with anti-CD3 and anti-CD28 followed by 3-d resting in the presence of IL-2). (C) B cell help activity of circulating CXCR5 + T cells after T cell receptor activation. Naive (IgD + ) or memory (IgD − ) B cells (2 × 10 4 ) from peripheral blood were cultured for 14 d in the presence or absence of various numbers (10 3 , 5 × 10 3 , 10 4 , 2.5 × 10 4 , 5 × 10 4 , and 10 5 ) of autologous T cells (CXCR5 − CD45RA + , CXCR5 + CD45RA − , or CXCR5 − CD45RA − ). Concentrations of IgG, IgA, and IgM in the culture supernatants were measured by ELISA. T cell numbers required for peak levels of antibody production varied among donors or experiments. One peak value with the best antibody production in each T cell group is shown. (D) Loss of CXCR5 expression during T cell receptor activation with anti-CD3 and anti-CD28. Error bars indicate SD of results from at least five different experiments (A). Representatives of three independent experiments are shown (B and D), and results from seven different donors (C) are shown.

Article Snippet: In brief, acetone-fixed sections were stained with the primary monoclonal antibody to CXCR5 (R&D Systems), followed by incubation in biotinylated goat anti–mouse IgG (Vector Laboratories).

Techniques: Activation Assay, Activity Assay, Cell Culture, Enzyme-linked Immunosorbent Assay, Expressing

Journal:

Article Title: The immunosuppressive drug mycophenolate mofetil impairs the adhesion capacity of gastrointestinal tumour cells

doi: 10.1046/j.1365-2249.2003.02290.x

Figure Lengend Snippet: Fluorescence analysis (relative fluorescence units, RFU) of endothelial cell ICAM-1, VCAM-1, E-selectin and P-selectin surface expression. For maximal stimulation, HUVEC were incubated with IL-1 (100 U/ml) for 12 h to up-regulate ICAM-1, and to induce VCAM-1 and E-selectin. HUVEC were incubated with PGE2 (10−6m) for 10 min to induce P-selectin expression. FL-1H (log) channel histogram analysis; 1 × 104 cells/scan. Control values were set at 100%. Mean ± s.d. of three experiments.

Article Snippet: After blocking, they were incubated overnight with sheep-antihuman P-selectin-antibodies (sheep IgG antibodies; 1 : 500 dilution; R&D Systems).

Techniques: Fluorescence, Expressing, Incubation, Control

Journal:

Article Title: The immunosuppressive drug mycophenolate mofetil impairs the adhesion capacity of gastrointestinal tumour cells

doi: 10.1046/j.1365-2249.2003.02290.x

Figure Lengend Snippet: MMF suppresses IL-1 induced E-selectin mRNA expression. Total RNA was isolated and E-selectin mRNA levels were determined by Northern blot analysis. GAPDH was used to assess equivalent RNA loading C−= unstimulated control cells, C+= IL-1 stimulated control cells. Results of densitometry are shown in the graph below and are expressed as the ratio of E-selectin : GAPDH mRNA, relative to the control (IL-1 activation without MMF), which was assigned a value of 100%. The figure shows one representative experiment from three.

Article Snippet: After blocking, they were incubated overnight with sheep-antihuman P-selectin-antibodies (sheep IgG antibodies; 1 : 500 dilution; R&D Systems).

Techniques: Expressing, Isolation, Northern Blot, Control, Activation Assay

Journal:

Article Title: The immunosuppressive drug mycophenolate mofetil impairs the adhesion capacity of gastrointestinal tumour cells

doi: 10.1046/j.1365-2249.2003.02290.x

Figure Lengend Snippet: Western blot analysis of P-selectin in endothelial cells. HUVEC were stimulated with 10−6m PGE2 and incubated additionally with different concentrations of MMF. Throm = control analysis of platelet P-selectin protein, C-= unstimulated HUVEC without MMF, C+ = PGE2-stimulated HUVEC without MMF. The figure shows one of four separate experiments.

Article Snippet: After blocking, they were incubated overnight with sheep-antihuman P-selectin-antibodies (sheep IgG antibodies; 1 : 500 dilution; R&D Systems).

Techniques: Western Blot, Incubation, Control

Journal:

Article Title: The immunosuppressive drug mycophenolate mofetil impairs the adhesion capacity of gastrointestinal tumour cells

doi: 10.1046/j.1365-2249.2003.02290.x

Figure Lengend Snippet: Adhesion of CD4+ T (a), CD8+ T (b) or WiDr (c) cells to immobilized adhesion receptor globulin chimeras. Purified T cells or WiDr cells were added to culture dishes coated with the extracellular domain of ICAM-1, VCAM-1, E-selectin or P-selectin. The immobilized adhesion proteins were coupled to goat-antihuman IgG. Cells were pretreated with different concentrations of MMF. Untreated cells served as controls (100% value). Adhesion was measured after 30 min incubation and after removal of non-adherent cells, according to the protocol given in Materials and methods. Each point represents the mean ± s.d. of four experiments; s.d. was usually about 20%.

Article Snippet: After blocking, they were incubated overnight with sheep-antihuman P-selectin-antibodies (sheep IgG antibodies; 1 : 500 dilution; R&D Systems).

Techniques: Purification, Incubation

Journal: Radiation Oncology (London, England)

Article Title: Quantitative proteomic analysis reveals AK2 as potential biomarker for late normal tissue radiotoxicity

doi: 10.1186/s13014-019-1351-8

Figure Lengend Snippet: AK2 is differentially expressed between patients with and without grade ≥ 2 bf+. a Immunoblot analysis of AK2, ANX1, HSPA8 and IDH2 in control (0 Gy) and 8 Gy-irradiated T lymphocytes from the indicated patients. β-actin was used as loading control. b Quantification of the immunoblot presented in A using the ImageJ software. Data are the mean ± SEM; * p < 0.05 (2-tailed Mann-Whitney test). qRT-PCR analysis of AK2 and IDH2 mRNA expression in control (0 Gy) and 8 Gy-irradiated T lymphocytes from the same patients with and without grade ≥ 2 bf+. The graph shows the expression level in irradiated samples relative to non-irradiated samples

Article Snippet: Membranes were blocked with PBST (PBS plus 0.1% Tween-20) containing 5% non-fat milk at 25 °C for 60 min. Blots were then incubated (4 °C, overnight) with primary antibodies against adenylate kinase 2 (AK2) (1/100, sc-28,786; Santa Cruz Biotechnology), annexin A1 (ANXA1) (1/100, sc-11,387; Santa Cruz Biotechnology), galectin-1 (LSGAL1) (1/100, sc-19,277; Santa Cruz Biotechnology), heat shock cognate 71 kDa protein (HSPA8) (1/500, sc-7298; Santa Cruz Biotechnology) and isocitrate dehydrogenase 2 (IDH2) (1/200, sc-134,923; Santa Cruz Biotechnology).

Techniques: Western Blot, Control, Irradiation, Software, MANN-WHITNEY, Quantitative RT-PCR, Expressing

Journal: International Journal of Molecular Sciences

Article Title: The Prognostic Significance of FKBP1A and Its Related Immune Infiltration in Liver Hepatocellular Carcinoma

doi: 10.3390/ijms232112797

Figure Lengend Snippet: The FKBP1A expression levels in various types of cancer tissues indicated by mRNA detection. ( A ) The expression level of FKBP1A in different types of tumor tissues and normal tissues in the Oncomine database. Number listed under name of each tumor is sample number ( n ) recorded in each dataset. Red highlight indicates high expression and blue indicates low expression. Darker red or darker blue represents higher or lower expression, respectively. ( B ) The expression levels of FKBP1A in different types of tumor tissues queried from the TCGA Oncomine tumor database and normal tissues via TIMER2.0 database analysis. ** p < 0.01, *** p < 0.001.

Article Snippet: The slide then was blocked with 3% bovine serum albumin (BSA) in PBS for 1 h at room temperature, and then further incubated with FKBP1A antibody (cat. no. sc-28814; 1:100; Santa Cruz Biotechnology, Dallas, TX, USA) overnight at 4 °C.

Techniques: Expressing

Journal: International Journal of Molecular Sciences

Article Title: The Prognostic Significance of FKBP1A and Its Related Immune Infiltration in Liver Hepatocellular Carcinoma

doi: 10.3390/ijms232112797

Figure Lengend Snippet: FKBP1A transcription level was significantly increased in LIHC patients. ( A ) Box plots of FKBP1A mRNA expression in normal liver and tumor samples based on GEPIA database. ( B – D ) FKBP1A mRNA level in LIHC patients based on different variables by using the UALCAN database. ( B ) Normal individuals or LIHC patients with grade 1, 2, 3, or 4 tumors. ( C ) Normal individuals or in LIHC patients in stage 1, 2, 3, or 4. ( D ) Normal individuals and LIHC patients with lymph node metastasis. The unpaired Student’s t test was used to estimate the significance of difference in gene expression levels between groups. * p < 0.05, ** p < 0.01, **** p < 0.0001 vs. normal.

Article Snippet: The slide then was blocked with 3% bovine serum albumin (BSA) in PBS for 1 h at room temperature, and then further incubated with FKBP1A antibody (cat. no. sc-28814; 1:100; Santa Cruz Biotechnology, Dallas, TX, USA) overnight at 4 °C.

Techniques: Expressing, Gene Expression

Journal: International Journal of Molecular Sciences

Article Title: The Prognostic Significance of FKBP1A and Its Related Immune Infiltration in Liver Hepatocellular Carcinoma

doi: 10.3390/ijms232112797

Figure Lengend Snippet: Human tissue microarray including hepatocellular carcinoma and normal liver tissue analysis revealed that the expression of FKBP1A was significantly elevated in LIHC patients. ( A ) Representative IHC images of FKBP1A in normal liver ( a ) or LIHC patients ( b ). ( c ) Shows the enlarged image of the black box in ( a ); ( d ) is the enlarged image of the black box in ( b ). The density distribution of ( c , d ) are shown in panels ( e , f ). n (normal) = 16; n (LIHC) = 79. ( B ) The plot profiles of the intensity of FKBP1A protein in hepatocytes of normal tissues and LIHC along the straight line (as shown ( a) and ( b ). ( C ) Quantification of the mean optical density (mean IOD) of FKBP1A protein by IHC staining in healthy liver, adjacent to LIHC or LIHC tissues. The normal group includes healthy liver and adjacent to LIHC tissues. n (healthy liver) = 6; n (adjacent to LIHC) = 10; n (normal) = 16; n (LIHC) = 79. Data are presented as the means ± SD. *** p < 0.001.

Article Snippet: The slide then was blocked with 3% bovine serum albumin (BSA) in PBS for 1 h at room temperature, and then further incubated with FKBP1A antibody (cat. no. sc-28814; 1:100; Santa Cruz Biotechnology, Dallas, TX, USA) overnight at 4 °C.

Techniques: Microarray, Expressing, Immunohistochemistry

Journal: International Journal of Molecular Sciences

Article Title: The Prognostic Significance of FKBP1A and Its Related Immune Infiltration in Liver Hepatocellular Carcinoma

doi: 10.3390/ijms232112797

Figure Lengend Snippet: FKBP1A expression distribution is accompanied with ER accumulation in LIHC based on human liver microarray analysis. ( A ) Representative images of immunofluorescence staining of endoplasmic reticulum marker RyR (green), FKBP1A (red) and nucleus (blue) from normal ( a – c ) and LIHC ( d – f ). The co-localization regions of FKBP1A and RyR are shown in yellow ( c , f ). ( g , h ) The plot profiles of the intensity of co-localization regions of FKBP1A and RyR in hepatocytes of normal tissues and LIHC along the straight line (as shown c , f ). n (normal) = 16; n (LIHC) = 79. ( B ) The plot profiles of the intensity of FKBP1A and RyR in hepatocytes of normal tissues and LIHC along the straight line (as shown c , f ). ( C ) Quantification of the mean optical density (mean IOD) of FKBP1A protein in healthy liver, adjacent to LIHC or LIHC tissues. The normal group includes healthy liver and adjacent to LIHC tissues. n (healthy liver) = 6; n (adjacent to LIHC) = 10; n (normal) = 16; n (LIHC) = 79. Data are presented as the means ± SD. *** p < 0.001, **** p < 0.0001.

Article Snippet: The slide then was blocked with 3% bovine serum albumin (BSA) in PBS for 1 h at room temperature, and then further incubated with FKBP1A antibody (cat. no. sc-28814; 1:100; Santa Cruz Biotechnology, Dallas, TX, USA) overnight at 4 °C.

Techniques: Expressing, Microarray, Immunofluorescence, Staining, Marker

Journal: International Journal of Molecular Sciences

Article Title: The Prognostic Significance of FKBP1A and Its Related Immune Infiltration in Liver Hepatocellular Carcinoma

doi: 10.3390/ijms232112797

Figure Lengend Snippet: Human liver tissue microarray analysis for the correlation between FKBP1A protein expression level and LIHC patients depending upon TNM staging or histologic grade. ( A , B ) Representative images of human normal liver tissues and LIHC patients with T2N0M0, T3N0M0, or T4N0M0 tumors ( A ) and grade 1, 2, or 3 tumors ( B ) with FKBP1A staining. ( C , D ) Quantification of the mean optical density (mean IOD) of FKBP1A protein at different TNM stages (C) and different tumor grades ( D ). ( E , F ) Quantification of the mean IOD of FKBP1A protein distributed in the endoplasmic reticulum at different TNM stages ( E ) and in different tumor grades ( F ). The normal group includes healthy liver tissues and adjacent to LIHC tissues. Tumor grade is the description of a tumor based on how abnormal the tumor cells and the tumor tissue look under a microscope. Each point represents an individual tissue. Scale bar, 100 μm. Data are presented as the means ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: The slide then was blocked with 3% bovine serum albumin (BSA) in PBS for 1 h at room temperature, and then further incubated with FKBP1A antibody (cat. no. sc-28814; 1:100; Santa Cruz Biotechnology, Dallas, TX, USA) overnight at 4 °C.

Techniques: Microarray, Expressing, Staining, Microscopy

Journal: International Journal of Molecular Sciences

Article Title: The Prognostic Significance of FKBP1A and Its Related Immune Infiltration in Liver Hepatocellular Carcinoma

doi: 10.3390/ijms232112797

Figure Lengend Snippet: High FKBP1A expression predicts poor prognosis in LIHC patients. ( A – D ), Correlation analysis between FKBP1A expression and prognostic survival in LIHC patients via Kaplan–Meier plotter analysis stratified at “best cut-off”. The hazard ratio (HR) was calculated based on the Cox proportional hazards model. ( A ) Overall survival, n = 364; ( B ) relapse-free survival, n = 316; ( C ) progression-free survival, n = 370; ( D ) disease-specific survival, n = 362. ( E , F ) Overall survival ( E ) and disease-free survival ( F ) curves of FKBP1A in LIHC patients assessed in the GEPIA database. The median value was used as the cutoff for dividing the high and low groups. The log-rank method was used for the hypothesis test.

Article Snippet: The slide then was blocked with 3% bovine serum albumin (BSA) in PBS for 1 h at room temperature, and then further incubated with FKBP1A antibody (cat. no. sc-28814; 1:100; Santa Cruz Biotechnology, Dallas, TX, USA) overnight at 4 °C.

Techniques: Expressing

Journal: International Journal of Molecular Sciences

Article Title: The Prognostic Significance of FKBP1A and Its Related Immune Infiltration in Liver Hepatocellular Carcinoma

doi: 10.3390/ijms232112797

Figure Lengend Snippet: Correlation analysis of FKBP1A expression and prognostic overall survival in LIHC patients with different clinic-pathological factors by using Kaplan–Meier plotter.

Article Snippet: The slide then was blocked with 3% bovine serum albumin (BSA) in PBS for 1 h at room temperature, and then further incubated with FKBP1A antibody (cat. no. sc-28814; 1:100; Santa Cruz Biotechnology, Dallas, TX, USA) overnight at 4 °C.

Techniques: Expressing

Journal: International Journal of Molecular Sciences

Article Title: The Prognostic Significance of FKBP1A and Its Related Immune Infiltration in Liver Hepatocellular Carcinoma

doi: 10.3390/ijms232112797

Figure Lengend Snippet: Prognostic factors for overall survival in univariable and multivariable Cox proportional hazards analyses.

Article Snippet: The slide then was blocked with 3% bovine serum albumin (BSA) in PBS for 1 h at room temperature, and then further incubated with FKBP1A antibody (cat. no. sc-28814; 1:100; Santa Cruz Biotechnology, Dallas, TX, USA) overnight at 4 °C.

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: The Prognostic Significance of FKBP1A and Its Related Immune Infiltration in Liver Hepatocellular Carcinoma

doi: 10.3390/ijms232112797

Figure Lengend Snippet: The co-expression genes with FKBP1A and function of enrichment analysis concerning FKBP1A -related genes in LIHC. ( A ) All the genes significantly associated with FKBP1A distinguished by Pearson test in LIHC cohort (LinkedOmics database). Red and green dots represent positively and negatively significantly correlated genes with FKBP1A , respectively. ( B ) KEGG pathway analysis. Dark blue and orange indicate FDR ≤ 0.05. FDR, false discovery rate. ( C ) A protein–protein interaction (PPI) network of FKBP1A protein from STRING database.

Article Snippet: The slide then was blocked with 3% bovine serum albumin (BSA) in PBS for 1 h at room temperature, and then further incubated with FKBP1A antibody (cat. no. sc-28814; 1:100; Santa Cruz Biotechnology, Dallas, TX, USA) overnight at 4 °C.

Techniques: Expressing

Journal: International Journal of Molecular Sciences

Article Title: The Prognostic Significance of FKBP1A and Its Related Immune Infiltration in Liver Hepatocellular Carcinoma

doi: 10.3390/ijms232112797

Figure Lengend Snippet: Correlation analysis between FKBP1A expression and tumor-infiltrating immune cells in LIHC. ( A ) FKBP1A expression is significantly related to tumor purity and has significant positive correlations with infiltrating levels of B cells, CD8 + T cells, CD4 + T cells, macrophages, neutrophils, and dendritic cells obtained from TIMER (purity-corrected Spearman test) in LIHC. ( B ) Five-year overall survival curve of the six tumor-infiltrating immune cells and FKBP1A expression in LIHC patients produced by Kaplan–Meier estimator from TIMER. Survival differences are compared between patients with high and low (split percentage of patients is 30%) infiltration of each kind of immune cells. Log-rank p < 0.05 is considered significant. ( C ) The Kaplan–Meier plot from the TIMER2.0 shows the difference of overall survival among patients stratified by both the estimated infiltration level of different types of macrophages (M0, M1 and M2) and FKBP1A expression level in LIHC. ( D ) Scatter plots from the “Gene Module”. Correlation of FKBP1A expression with tumor purity and with the infiltration level of M2 macrophage markers CD68 , CD163 and CD209 . p < 0.05 is considered significant.

Article Snippet: The slide then was blocked with 3% bovine serum albumin (BSA) in PBS for 1 h at room temperature, and then further incubated with FKBP1A antibody (cat. no. sc-28814; 1:100; Santa Cruz Biotechnology, Dallas, TX, USA) overnight at 4 °C.

Techniques: Expressing, Produced