Journal: Journal of Biological Chemistry

Article Title: Tumor Microenvironment Macrophage Inhibitory Factor Directs the Accumulation of Interleukin-17-producing Tumor-infiltrating Lymphocytes and Predicts Favorable Survival in Nasopharyngeal Carcinoma Patients

doi: 10.1074/jbc.m112.367532

Figure Lengend Snippet: FIGURE 3. Characterization and cytokine expression profiles of the Th17 cell subset among PBMCs and TILs from NPC patients. A, T cell surface markers were detected in Th17 cells from NPC patients. T cells were stimulated with phorbol 12-myristate 13-acetate/ionomycin for 4 h, then stained with fluorescence- conjugated antibodies against the markers shown, and analyzed by flow cytometry. B, graph of the percentage of cytokine-secreting Th17 cells among PBMCs and TILs from NPC patients (n 5) and PBMCs from healthy donors (n 5). Error bars represent the S.E. Significance was determined by the 2 test or paired t test. (*, p 0.001).

Article Snippet: Cytokines in the culture medium were measured using the Human Cytokine Array Panel A (Proteome Profiler; R&D Systems).

Techniques: Expressing, Staining, Fluorescence, Flow Cytometry

Journal: Journal of Biological Chemistry

Article Title: Tumor Microenvironment Macrophage Inhibitory Factor Directs the Accumulation of Interleukin-17-producing Tumor-infiltrating Lymphocytes and Predicts Favorable Survival in Nasopharyngeal Carcinoma Patients

doi: 10.1074/jbc.m112.367532

Figure Lengend Snippet: FIGURE 4. Generation and migration of Th17 cells mediated by NPC tumor cell lines. A, NPC tumor cell lines induce the differentiation of naive T cells into Th17cellsinvitro.PurifiedCD4naiveTcellsfromhealthydonorswerestimulatedwithOKT3andthenco-culturedwiththeirradiatedNPCcelllineC666(EBV) or CNE2 (EBV) in IL-2-containing medium for 7 days. The NP69 and LCL lines and the cytokines IL-1 and TGF were used as controls. All cultured cells were stained for Foxp3 and IL-17 for FACS analysis after stimulation of phorbol 12-myristate 13-acetate/ionomycin. The data represent one of three independent experiments. B, mean percentage of cytokine-secreting Th17 cells induced from CD4 naive T cells from three experiments. C, migration of Th17 cells was increased in response to cultured supernatants from the NPC cell lines C666 and CNE2, relative to that from the normal NP cell line NP69 or with medium alone. The data represent one of three independent experiments. D, mean percentage of cytokine-secreting Th17 cells in the inner well and outer well after migration induced by supernatants from NP69, C666, or CNE2 cells from three experiments. Error bars represent the S.E. Significance was determined by the 2 test or paired t test. (*, p 0.05).

Article Snippet: Cytokines in the culture medium were measured using the Human Cytokine Array Panel A (Proteome Profiler; R&D Systems).

Techniques: Migration, Cell Culture, Staining

Journal: British Journal of Cancer

Article Title: Impairment of carbonic anhydrase IX ectodomain cleavage reinforces tumorigenic and metastatic phenotype of cancer cells

doi: 10.1038/s41416-020-0804-z

Figure Lengend Snippet: a Proteome profiler Human XL Cytokine Array (ARY022, R&D Systems, MN, USA) probed with serum-free-conditioned medium (SF-CM) of C33a-FL-CA IX or C33a-NS-CA IX cells incubated under normoxic (NO) and hypoxic (HY) conditions for 48 h. Spot doublets of corresponding proteins with differential intensities are framed and indicated by arrows. Medium pooled from five independent experiments was used for the array. Spot intensities representing b pro-oncogenic basigin, and c anti-angiogenic thrombospondin-1 were analysed in culture medium samples of C33a-FL-CA IX versus C33a-NS-CA IX cells. Abundance of the proteins was assessed according to spot intensity using densitometry. Data are shown as mean ± SD. d Validation of the differential levels of basigin and thrombospondin-1, respectively, by Western blotting analysis of medium precipitates (MP) and cell lysates (CL) of normoxic (NO) and hypoxic (HY) cells. (* P < 0.05, ** P < 0.01, *** P < 0.005).

Article Snippet: PPA analyses were performed using the Human XL Cytokine Array Kit and Human Soluble Receptor Array Non-Hematopoietic panel kit (R&D Systems, MN, USA).

Techniques: Incubation, Biomarker Discovery, Western Blot

Journal: British Journal of Cancer

Article Title: Impairment of carbonic anhydrase IX ectodomain cleavage reinforces tumorigenic and metastatic phenotype of cancer cells

doi: 10.1038/s41416-020-0804-z

Figure Lengend Snippet: Analysis of the differential expression of a IGFBP-2 and b IGFBP-3 in SF-CM samples from C33a-FL-CA IX and C33a-NS-CA IX incubated under normoxic (NO) and hypoxic (HY) conditions for 48 h and evaluated by Proteome Profiler Human XL Cytokine Array (ARY022, R&D Systems, MN, USA). Abundance of the proteins was assessed according to spot intensity using densitometry. Data are shown as mean ± SD of five independent samples. c Western blotting analysis of CA IX, IGFBP-2 and IGFBP-3 protein levels in medium precipitates (MP) of C33a-FL-CA IX and C33a-NS-CA IX cells. Impairment of shedding in NS-CA IX is associated with increased IGFBP-2 and reduced IGFBP-3 secretion to culture medium. Quantitative PCR analysis of relative mRNA levels of d IGFBP-2, e IGFBP-3 and f IGF1 in C33a-FL-CA IX versus C33a-NS-CA IX cells normalised to β-actin mRNA. (*** P < 0.005, ns non-significant).

Article Snippet: PPA analyses were performed using the Human XL Cytokine Array Kit and Human Soluble Receptor Array Non-Hematopoietic panel kit (R&D Systems, MN, USA).

Techniques: Quantitative Proteomics, Incubation, Western Blot, Real-time Polymerase Chain Reaction

Journal: mSphere

Article Title: Differential innate immune responses of human macrophages and bronchial epithelial cells against Talaromyces marneffei

doi: 10.1128/msphere.00258-22

Figure Lengend Snippet: Cytokine responses of human and murine macrophages cultured in the presence or absence of plasma/serum upon Talaromyces marneffei infection. Upon T. marneffei infection, the relative expression of ( A ) TNF and ( B ) CXCL8 mRNAs in hPBDMs could only be stimulated in the presence of autologous plasma; whereas no significant change in the mRNA expression of these two cytokines was observed when the culture medium was not supplemented with autologous plasma. The infection experiment was conducted in triplicate using blood cells isolated from three independent donors. The inability of T. marneffei to induce ( C ) TNF and ( D ) CXCL8 mRNAs upon infection was also observed in the murine macrophage cell lines J774A.1 and RAW264.7. Data are presented as mean ± SD and comparisons were analyzed using unpaired t -test (two-tailed). **, P < 0.01; ***, P < 0.001.

Article Snippet: Expression of cytokines were measured semi-quantitatively using the Proteome Profiler Human XL Cytokine Array Kit (R&D Systems, Minneapolis, MN, USA) according to the manufacturer’s instruction; and all samples were tested in duplicates.

Techniques: Cell Culture, Clinical Proteomics, Infection, Expressing, Isolation, Two Tailed Test

Journal: mSphere

Article Title: Differential innate immune responses of human macrophages and bronchial epithelial cells against Talaromyces marneffei

doi: 10.1128/msphere.00258-22

Figure Lengend Snippet: Changes of cytokine mRNA expression in hPBDMs upon Aspergillus fumigatus and Talaromyces marneffei infections. Relative mRNA expressions of CXCL8 and TNF in hPBDMs infected with both fungi were significantly induced mainly after 4 h post-infection and the expression of CXCL10 mRNA in hPBDMs was only significantly induced by T. marneffei infection, in which the expression level reached the peak at 8 h post-infection. The infection experiment was conducted in triplicate using blood cells isolated from three independent donors. The relative mRNA expression levels of CXCL8 , CXCL10 , and TNF were detected by qRT-PCR. Data are presented as mean ± SD and comparisons were analyzed using unpaired t -test (two-tailed). *, P < 0.05; †, P < 0.01; ‡, P < 0.001; ×, undetectable.

Article Snippet: Expression of cytokines were measured semi-quantitatively using the Proteome Profiler Human XL Cytokine Array Kit (R&D Systems, Minneapolis, MN, USA) according to the manufacturer’s instruction; and all samples were tested in duplicates.

Techniques: Expressing, Infection, Isolation, Quantitative RT-PCR, Two Tailed Test

Journal: bioRxiv

Article Title: A 50-gene high-risk profile predictive of COVID-19 and Idiopathic Pulmonary Fibrosis mortality originates from a genomic imbalance in monocyte and T-cell subsets that reverses in survivors with post-COVID-19 Interstitial Lung Disease

doi: 10.1101/2023.10.22.563156

Figure Lengend Snippet: A . Study design of the 50-gene signature and cytokine analysis in COVID-19 patients (Cohort 1). B. Heatmap of COVID-19 patients based on the 50-gene signature discriminates three risk groups (low, intermediate, and high) based on SAMS. Every column represents a patient, and every row represents a gene. Log-based two-color scale is adjacent to the heatmap. Red denotes increased expression and green denotes decreased expression. Gene expression data is represented as Log2 normalized expression values. C-D. Time to death and time to discharge by day 60 in hospitalized patients with COVID-19, respectively. E-H. Plasma cytokine concentrations (IL6, IP-10, SPP1 and TGFβ) in low, intermediate, and high-risk profile patients with COVID-19 at days 2, 6 and 13 post admission. The data is presented as an average of triplicate values ± SEM for each group. Two-way ANOVA test (GraphPad software) Tukey’s multiple comparisons were used; * p<0.05. I. Study design of 7-gene signature analysis by RT-qPCR in PBMCs from COVID-19 patients (Cohort 2). J. Heatmap of COVID-19 patients based on the 50-gene signature discriminates three risk groups (low, intermediate, and high) based on SAMS Up scores. Heatmap nomenclature is the same as in . K-L. Time to death and time to discharge by day 60 in hospitalized patients with COVID-19 respectively in cohort two. The data is presented as an average of triplicated TUs values ± SEM for each group. * p<0.05

Article Snippet: We measured cytokine concentrations of 121 plasma samples from COVID-19 patients from Cohort 1 using a customized, Bioplex 200 compatible, human cytokine panel including, IL6, IP10, SPP1 and TGFβ-1 (#FCSTM18-06, R&D Systems).

Techniques: Expressing, Gene Expression, Clinical Proteomics, Software, Quantitative RT-PCR