Journal: Frontiers in Microbiology
Article Title: PGE2 Augments Inflammasome Activation and M1 Polarization in Macrophages Infected With Salmonella Typhimurium and Yersinia enterocolitica
Figure Lengend Snippet: Transcript analysis of PGE2 receptors in THP-1 macrophages during S. Typhimurium and Y. enterocolitica infection. THP-1 macrophages (2.5 × 10 5 ) were infected for 2 h with (A) S. Typhimurium or (B) Y. enterocolitica at an MOI of 50:1. Total RNA was extracted using a PureLink RNA mini kit, and cDNA was generated using the iScript system (Bio-Rad). RT-PCR was performed using SYBR green on a CFX96 Real-Time System (Bio-Rad) targeting prostanoid receptors EP1, EP2, EP3, EP4, and GAPDH as a reference gene. Resulting data were then normalized to GAPDH mRNA levels, and uninfected samples served as a baseline for fold change determination using the ΔΔCt method. Resulting data is representative of three biological replicates and three technical replicates and represents the mean fold change ± SD. One-way ANOVA test with Tukey’s multiple testing correction was used to establish statistical significance. p -values were indicated as follows: ∗ p ≤ 0.05; ∗∗ p ≤ 0.01; ∗∗∗ p ≤ 0.001; ∗∗∗∗ p ≤ 0.0001.
Article Snippet: The cDNA was synthesized using a Bio-Rad iScript cDNA Synthesis Kit and expression of genes encoding EP1, EP2, EP3, and EP4 was measured as stated above on a Bio-Rad CFX96 Real-Time System.
Techniques: Infection, Generated, Reverse Transcription Polymerase Chain Reaction, SYBR Green Assay