ABSTRACT
Cytokines are released in response to brain injury and inflammation. By binding to receptors, they can cause, exacerbate or inhibit cellular injury and repair. We studied RNA expression of cytokine receptors and members of inflammatory pathways in human NT2-N neurons during 3 h of hypoxia and glucose deprivation followed by 21 h of reoxygenation, and the impact of acidosis. Right after acidotic hypoxia, RNA of IL-10RA and CXCR4 were significantly increased relative to acidotic control, while Bcl-2 and Bcl-xL were significantly decreased. After 21 h of neutral reoxygenation after neutral hypoxia, there was a significant increase in RNA of CXCR1 (relative quantification (RQ)=4.1, p<0.05), CXCR2 (3.6, p<0.05), CCR2 (3.8, p<0.05), Hsp70 (2.4, p<0.05), HIF-1alpha (1.5, p<0.001), TRAF6 (1.3, p<0.05) and TNFR1 (1.6, p<0.05). After 21 h of acidotic reoxygenation after acidotic hypoxia, we found a significant increase in RNA of IL-1R1, IL-10RA, CXCR4 and Hsp70 compared to control, and a significant decrease in FAS and TRAF6. There was a significant increase in Bax expression and a significant decrease in Bcl-2 and Bcl-xL expression in three out of four pH groups after 21 h of reoxygenation. Acidotic, relative to neutral, hypoxia and reoxygenation also influenced the expression of various genes. We conclude that inflammatory receptors and pathways are activated during hypoxia and reoxygenation in NT2-N neurons, and that this activation is pH dependent. This supports the concept that inflammatory pathways play a role in cerebral hypoxic-ischemic damage, and that they may represent important pharmacological targets.
Subject(s)
Acidosis/metabolism , Cell Hypoxia/physiology , Inflammation/metabolism , Neurons/metabolism , Receptors, Cytokine/biosynthesis , Cells, Cultured , Gene Expression , Humans , Reperfusion Injury/metabolismABSTRACT
BACKGROUND: Only a few infants develop acute bronchiolitis when exposed to respiratory syncytial virus (RSV), and host, environmental, and viral properties are probably all of importance in determining the severity of infection. METHODS: Microarray analysis was used to identify differentially expressed single genes and gene sets in cord blood from 5 infants hospitalized with RSV bronchiolitis versus cord blood from 5 control infants exposed to RSV without bronchiolitis during infancy. Quantitative real-time polymerase chain reaction (QRT-PCR) was performed on single genes in both the 5 infants selected for microarray analysis and 13 more infants hospitalized with the same disease. Gene set enrichment analysis (GSEA) was performed to identify differentially expressed gene sets within the microarray experiments. RESULTS: Microarray analysis identified 15 single genes to be significantly differentially expressed between case and control infants. Eleven of these genes were evaluated with QRT-PCR, and the genes FAM102A, TNFRSF25, and STMN3 were down-regulated in all but 1 of the 18 infants. A pathway involved in regulation of the actin cytoskeleton was found to be clearly down-regulated when analyzed with GSEA. CONCLUSIONS: FAM102A, TNFRSF25, and STMN3 and a pathway involved in regulation of the actin cytoskeleton are down-regulated in cord blood from infants hospitalized with RSV bronchiolitis.
Subject(s)
Bronchiolitis/metabolism , Bronchiolitis/virology , Fetal Blood , Gene Expression Regulation/physiology , Respiratory Syncytial Virus Infections/metabolism , Female , Gene Expression Profiling , Humans , Infant , Male , Respiratory Syncytial Viruses , Reverse Transcriptase Polymerase Chain Reaction/methodsABSTRACT
Benzalkonium chloride (BC) is a commonly used disinfectant and preservative. This study describes changes in expression level at the transcriptomic and proteomic level for Escherichia coli K-12 gradually adapted to a tolerance level to BC of 7-8 times the initial MIC. Results from DNA arrays and two-dimensional gel electrophoresis for global gene and protein expression studies were confirmed by real-time quantitative PCR. Peptide mass fingerprinting by MALDI-TOF MS was used to identify differentially expressed proteins. Changes in expression level in adapted cells were shown for porins, drug transporters, glycolytic enzymes, ribosomal subunits and several genes and proteins involved in protection against oxidative stress and antibiotics. Adapted strains showed increased tolerance to several antibiotics. In conclusion, E. coli K-12 adapted to higher tolerance to BC acquired several general resistance mechanisms, including responses normally related to the multiple antibiotic resistance (Mar) regulon and protection against oxidative stress. The results revealed that BC treatment might result in superoxide stress in E. coli.
Subject(s)
Anti-Infective Agents, Local/pharmacology , Benzalkonium Compounds/pharmacology , Escherichia coli/drug effects , Escherichia coli/genetics , Gene Expression Regulation, Bacterial/drug effects , Proteome , Transcription, Genetic , Adaptation, Physiological , Anti-Bacterial Agents/pharmacology , DNA-Binding Proteins/metabolism , Drug Tolerance , Electrophoresis, Gel, Two-Dimensional , Escherichia coli/chemistry , Escherichia coli Proteins/genetics , Escherichia coli Proteins/metabolism , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization , Trans-Activators/metabolismABSTRACT
BACKGROUND: Respiratory syncytial virus (RSV) is a major cause of viral bronchiolitis in infants worldwide, and environmental, viral and host factors are all of importance for disease susceptibility and severity. To study the systemic host response to this disease we used the microarray technology to measure mRNA gene expression levels in whole blood of five male infants hospitalised with acute RSV, subtype B, bronchiolitis versus five one year old male controls exposed to RSV during infancy without bronchiolitis. The gene expression levels were further evaluated in a new experiment using quantitative real-time polymerase chain reaction (QRT-PCR) both in the five infants selected for microarray and in 13 other infants hospitalised with the same disease. RESULTS: Among the 30 genes most differentially expressed by microarray nearly 50% were involved in immunological processes. We found the highly upregulated interferon, alpha-inducible protein 27 (IFI27) and the highly downregulated gene Charcot-Leyden crystal protein (CLC) to be the two most differentially expressed genes in the microarray study. When performing QRT-PCR on these genes IFI27 was upregulated in all but one infant, and CLC was downregulated in all 18 infants, and similar to that given by microarray. CONCLUSION: The gene IFI27 is upregulated and the gene CLC is downregulated in whole blood of infants hospitalised with RSV, subtype B, bronchiolitis and is not reported before. More studies are needed to elucidate the specificity of these gene expressions in association with host response to this virus in bronchiolitis of moderate severity.
