elisa analysis secreted cxcl5 protein levels  (R&D Systems)

Journal: The FASEB Journal

Article Title: Genome-wide effect of pulmonary airway epithelial cell–specific Bmal1 deletion

doi: 10.1096/fj.201801682R

Figure Lengend Snippet: Cell-autonomous Bmal1 function in ALI culture of primary mouse tracheal cells. ALI cultures were established using primary tracheal epithelial cells from WT and global Bmal1−/− mice, with 7 d under submerged condition and another 10 d exposed to air in the apical side of cells. A, B) Electron microscopy images for cilia in ALI primary tracheal cells from WT and global Bmal1−/− mice, respectively. C, D) Immunofluorescence staining of cilia markers (acetylated tubulin) and epithelial cell markers (E-cadherin) in ALI primary tracheal cells from WT and global Bmal1−/− mice, respectively. E, F) Cxcl5 and Cxcl15 gene expression measurement by qPCR in ALI primary tracheal cells from WT and global Bmal1−/− mice. Data shown are means ± sd; Student’s t test (n = 3/genotype). *P < 0.05, **P < 0.01. G) Expression of Bmal1 exon 8 in primary tracheal cells 2 d after infection. Data shown are means ± sd; Student’s t test (n = 3/genotype). ***P < 0.001. H) Bioluminescence traces of Per2-luc in control and Adv-Cre–transfected ALI primary tracheal cells 21 d in culture. I) Schematic description of viral Cre infection in primary tracheal cells from Bmal1flox/flox mice. J) Overlap of expression of rhythmic genes during ALI culture and LCM studies. K) KEGG pathway enrichment of DE genes in cell culture study. Pathways are ranked by significance value.

Article Snippet: CXCL5 levels were measured with a CXCL5 ELISA Kit (DY254; Bio-Techne, Minneapolis, MN, USA), according to product protocol.

Techniques: Electron Microscopy, Immunofluorescence, Staining, Expressing, Infection, Transfection, Cell Culture

Journal: The FASEB Journal

Article Title: Genome-wide effect of pulmonary airway epithelial cell–specific Bmal1 deletion

doi: 10.1096/fj.201801682R

Figure Lengend Snippet: Reverse feeding resets time of day variation in pulmonary LPS response. A) Schematic description of experiment design. The food-reversal experiment was undertaken using 2 separate cohorts. B–D) Food reversal without LPS treatment was performed in cohort 1. E–G) Aerosolized LPS exposure experiment at ZT0 vs. ZT12 was performed in cohort 2 and total BAL cells, neutrophils, and CXCL5 concentrations measured. Gene expression was determined in cohort 2. Data analyzed by 2-way ANOVA with post hoc test to examine time of day difference within genotypes (n = 6–8). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 (significant time of day difference within genotypes).

Article Snippet: CXCL5 levels were measured with a CXCL5 ELISA Kit (DY254; Bio-Techne, Minneapolis, MN, USA), according to product protocol.

Techniques: Expressing

Journal: Cell death & disease

Article Title: Cancer-derived exosomal HSPC111 promotes colorectal cancer liver metastasis by reprogramming lipid metabolism in cancer-associated fibroblasts.

doi: 10.1038/s41419-022-04506-4

Figure Lengend Snippet: Fig. 6 An increase in levels of acetyl-CoA and H3K27ac promotes CXCL5 secretion in CAFs. The effects of acetate treatment on acetyl-CoA levels in LX-2 cells incubated with ExoHCT116 (A) and ExoSW480 (B). Western blot showed the effects of acetate treatment on CXCL5 and H3K27ac levels in LX-2 cells incubated with ExoHCT116 (C) and ExoSW480 (D). E The effects of Trichostatin A (TSA) treatment on CXCL5 and H3K27ac levels in LX-2 cells incubated with ExoHCT116 KD and their relative control. F The effects of C646 treatment on CXCL5 and H3K27ac levels in LX-2 cells incubated with ExoSW480 OE and their relative control. G, H Chromatin immunoprecipitation (ChIP) assays using IgG as a control were performed with antibody against H3K23ac. Examination of H3K27 acetylation status in CXCL5 gene promoter region in LX-2 cells incubated with ExoHCT116 KD and ExoHCT116 KD Ctrl or in LX-2 cells incubated with ExoHCT116 KD and ExoHCT116 KD Ctrl and treated with TSA (G). Examination of H3K27 acetylation status in CXCL5 gene promoter region in LX-2 cells incubated with ExoSW480 OE and ExoSW480 OE Ctrl or in LX- 2 cells incubated with ExoSW480 OE and ExoSW480 OE Ctrl and treated with C646 (H). Each experiment was performed in triplicate. Data are shown as mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001.

Article Snippet: Cells were cultured in the presence of appropriate exosomes (20μg/mL) for 48 h before supernatants were collected, and the CXCL5 level was detected by CXCL5 ELISA kit (EK0728, BOSTER) according to the manufacturer’s protocols.

Techniques: Incubation, Western Blot, Control, Chromatin Immunoprecipitation

Journal: Cell death & disease

Article Title: Cancer-derived exosomal HSPC111 promotes colorectal cancer liver metastasis by reprogramming lipid metabolism in cancer-associated fibroblasts.

doi: 10.1038/s41419-022-04506-4

Figure Lengend Snippet: Fig. 8 A schematic diagram of mechanism of exosomal HSPC111 facilitates CRLM via reprogramming lipid metabolism in CAFs. Exosomal HSPC111 derived from CRC cells phosphorylates ACLY in CAFs, leading to the increased levels of acetyl-CoA and histone acetylation to secrete CXCL5, and resulting in CRC cells colonized in liver via the CXCL5-CXCR2 axis.

Article Snippet: Cells were cultured in the presence of appropriate exosomes (20μg/mL) for 48 h before supernatants were collected, and the CXCL5 level was detected by CXCL5 ELISA kit (EK0728, BOSTER) according to the manufacturer’s protocols.

Techniques: Derivative Assay

Journal: The American Journal of Pathology

Article Title: Overexpression of CXCL5 Is Associated With Poor Survival in Patients With Pancreatic Cancer

doi: 10.1016/j.ajpath.2010.11.058

Figure Lengend Snippet: Table 1

Article Snippet: Enzyme-Linked Immunosorbent Assay for Human CXCL5 Levels of CXCL5 in culture supernatant serum and tumor tissue were determined using the Quantikine Kit (R&D Systems) following the manufacturer's instructions.

Techniques: Expressing