Relative Quantification of Interleukin-6 Gene Expression in Tissues of Mice Treated with Lipopolysaccharide Using Real Time PCR.
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This study quantified Interleukin-6 (IL-6) gene expression in liver, lung and spleen mouse tissues exposed to varying amounts of bacterial lipopolysaccharide (LPS) using Relative Quantification Real-Time Polymerase Chain Reaction (RQ RT-PCR). The exposure of macrophages to bacterial LPS initiates a signal transduction cascade that leads to production of pro-inflammatory cytokines. There were three treatment groups. Group I, the control, had no exposure to LPS. Group II had four hours of LPS exposure and group III had twenty-four hours of LPS exposure, both via intraperitoneal injection. The tissue samples were extracted, lysed and purified into RNA with various reagents and buffers in the BioRobotM48 machine. The NanoDrop ND-8000 was used to test the concentration of purified RNA and Agilent RNA 6000 Nano Kit Bioanalyzer was used to test the quality and integrity of RNA. The samples underwent reverse transcription from RNA into cDNA and were then subject to RQ RT-PCR in the 7900HT Fast Real Time PCR. Treatment group II had the greatest IL-6 gene expression within the liver, lung and spleen tissues (F2,15=50.19; p<0.01, F2,15=23.93; p<0.001; F2,15=47.83, p<0.001, respectively). When analyzing only group II between organs, the liver and spleen mouse tissues had significantly (F2,15=11.75; p<0.01) higher IL-6 gene expression than the lung mouse tissues. The results can be explained by comparing and contrasting different responses each organ has to invading pathogens like LPS. Regulation of the signaling pathway is critical for controlling initial phases of the immune response to the presence of foreign organisms in the blood and tissues.