Lipopolysaccharide-Stimulated Responses in Rat Aortic Endothelial Cells by a Systems Biology Approach

• Main Authors: Hsiang-Wen Tseng, 曾湘文
• Other Authors: Chieh-Fu Chen
• Format: Others
• Language: zh-TW
• Published: 2006
• Online Access: http://ndltd.ncl.edu.tw/handle/01907460566729474813

博士 === 國立陽明大學 === 藥理學研究所 === 94 === Systems biology, a new discipline of biology, provides a system-level understanding of biological responses by integrating three subjects, genomics, proteomics, and bioinformatics. The vascular endothelium plays an important role in regulating the immune and inflammatory responses against pathogens infection. It is well-known that lipopolysaccharide (LPS) is a critical inducer of sepsis or endotoxemia, however, the systematic responses of LPS-stimulated endothelial cells (ECs) are still unclear. The present study aims to analyze the late-phase responses of LPS-induced rat aortic ECs by using systematic biology approaches, including cDNA microarray, two-dimensional electrophoresis, matrix-assisted laser desorption/ionization time-of-fly mass spectrometer, and cytokine protein array. Furthermore, to improve the efficiency of analysis of the bulk systematic data, a set of bioinformatic tools based on BioCarta, KEGG, and Gene Ontology databases was designed to convert and integrate rat data into the corresponding human genes or proteins. Rat cDNA microarray was used to evaluate the LPS-stimulated transcriptomic change at the mRNA level. Significant changes of mRNA levels were showed that 318 genes were found to be up-regulated by more than 2-fold and 158 genes were down-regulated at least 0.5-fold by LPS-treatment as compared with control experiments. After removing of the Incyte EST genes and homologous human Unigene ID conversion, 100 genes showed up-regulation and 52 genes were found to be down-regulated. In proteomics study, 20 protein spots were identified which showed significant changes due to LPS stimulation, all of which contained 14 different proteins. The up-regulated proteins included hypoxanthine-guanine phosphoribosyltransferase, peroxiredoxin 1, ferritin heavy chain, ferritin light chain, proteasome subunit alpha type 6, Mn-superoxide dismutase, voltage-dependent anion-selective channel protein 1, cathepsin B and guanine deaminase. Transgelin, vimentin, tropomyosin beta chain, tropomyosin 1 alpha chain, and PDZ and LIM domain protein 1 were down-regulated by LPS-treatment. In the secreted protein from ECs, VEGF were increased and 7 cytokines (IL-6, MCP-1, MIP-3 alpha, CINC-2, CINC-3, LIX, and fractalkine) were inducible by LPS. Using the bioinformatics tools (BGSSJ and ArrayXPath) to annotate transcriptomics and proteomics changes, it was shown that LPS could promote some signaling or metabolic pathways as well as pathophysiologic phenomena of proliferation, atherogenesis, inflammation, and apoptosis through activated nuclear factor-kappaB pathway in ECs. Interestingly, ECs also activated the mediators of anti-inflammation, anti-apoptosis, and anti-oxidation to protect themselves. Moreover, the expressions of altered genes, proteins, and their involvement in the hypothetical signaling pathway can provide further understanding of inflammation associated responses in ECs. In summary, the systems changes in ECs primed by LPS, and many of these altered expressions of genes, proteins and biological pathways can provide valuable information for understanding the mechanism of inflammation on ECs.