Gene expression profile in monocytes (Mφ) from multiple injured patients: An experimental pilot study

Introduction: Affection of the immune system is recognized as one of the initiating mechanisms for the induction of multiple organ failure (MOF) in patients suffering from multiple injuries (polytrauma). In particular, monocytes (Mφ) play a pivotal role within the development of MOF, however, initial Mφ mRNA expression patterns which may induce this immunological reaction are unknown. Hence, the aim of this study was to perform a screening investigation of 1172 genes on mRNA expression patterns in isolated Mφ from multiple injured patients using differential gene expression analysis by the microarray technique. Methods: Within our prospective pilot study 3 patients were enrolled presenting with blunt multiple injuries (Injury Severity Score >25 points), blood samples were drawn on admission and 6 and 12 hours after trauma. Mφ were isolated using magnetic beads (CD 14) and total mRNA was isolated by the Rneasy Kit (Qiagen, Hilden. Germany) and complementary cDNA was synthesized. For differential gene expression analysis a human cDNA microarray (Atlas human 1.2. Clontech, Heidelberg, Germany) was hybridized after labeling the probes with [P32], the activated or deactivated genes were visualized by exposure to a X-ray film and computer assisted analyzed. For semi-quantitative analysis the initial sample was set as the individual baseline mRNA expression and differences of the following samples calculated in gene expression ratios compared to the baseline level. A ratio of 2 was accepted as significant change between two sample intervals. Results: The following table depicts the number of genes which showed significantly altered expression, data are given in mean±sd, n=3 Biological function groups of scanned genes 6h 12h Number of mRNA expression difference 1 Oncogenes, tumor supressors and cell cycle regulators 5.66±3.06 8±4 2 Stress response, intracellular signal transduction modulators and effectors 2±0 3±1.73 3 Apoptosis, DNA synthesis, repair and recombination 4.33+2.52 5±4 4 Transcription factors and DNA binding proteins 7.33+3.21 13.33±5.5 5 Receptors, cell surface antigenes and cell adhesion 3.66+2.08 5.33±2.52 6 Cell-cell communication. cytoskeleton and protein turnover 3.33+1.53 2.33±1.53 Conclusion: We demonstrate for the first time an experimental screening investigation of 1172 genes using the microarray technique on M0 from multiple injured patients. Although still preliminary, identical gene spots (approx. 5% of all scanned genes) could be identified in various patients as influenced by the traumatic event. This relatively low percentage of influenced genes corresponds to animal studies using the microarray technique.