Journal: Cells
Article Title: Interference of LPS H. pylori with IL-33-Driven Regeneration of Caviae porcellus Primary Gastric Epithelial Cells and Fibroblasts
doi: 10.3390/cells10061385
Figure Lengend Snippet: Assessment of apoptosis. Cell apoptosis was estimated by terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) assay ( i ), assessment of expression of anti-apoptotic Bcl-xL protein by using anti-Bcl-xL primary antibodies and secondary antibodies conjugated with isothiocyanate fluorescein (FITC) ( ii ), and 4′,6-diamino-2-phenylindole (DAPI) nuclear staining ( iii ). The results for the guinea pig primary gastric epithelial cells ( A ) or fibroblasts ( B ), transfected with siRNA IL-33, which were not treated or treated with H. pylori lipopolysaccharide (LPS), 25 ng/mL, in the culture medium without or with IL-33 (3.0 ng/mL), are presented. The cell fluorescence was measured using a fluorescence reader (Victor2, Wallac, Oy Turku, Finland) at appropriate wavelengths: 495 excitation and 519 emission for FITC, 550 nm excitation and 590 nm emission for TUNEL, and 345 nm excitation and 455 nm emission for DAPI. Results are shown as median with the range of five experiments performed in triplicate for each experimental variant. Statistical analysis was performed using the nonparametric Mann-Whitney U test with significance at p < 0.05. *cells in medium without IL-33 vs. cells in medium with IL-33, not treated with LPS H. pylori ; ● cells treated with LPS H. pylori , in medium without IL-33 vs. cells treated with LPS in medium with IL-33; ▪ cells not treated with LPS H. pylori vs. cells treated with LPS H. pylori , in medium without or with IL-33.
Article Snippet: Cells fixed with 4% formaldehyde from each experimental variant were incubated overnight at 4 °C with primary antibodies towards anti-apoptotic protein Bcl-xL (Cell Signaling Technology, Danvers, MA, USA), and then stained with FITC-conjugated secondary antibodies (Invitrogen Carlsbad, CA, USA), as previously described [ , ].
Techniques: End Labeling, TUNEL Assay, Expressing, Staining, Transfection, Fluorescence, Variant Assay, MANN-WHITNEY
