Whole blood gene expression in infants with respiratory syncytial virus bronchiolitis

Abstract

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.

Figure 1

Evaluation of genes differentially expressed by microarray (blue bars) with QRT-PCR (red bars). The microarray experiment was conducted as a case-control study with whole blood gene expressions given as ratio of signal intensity in each of five male infants hospitalised with respiratory syncytial virus, subtype B, bronchiolitis versus signal intensity in a corresponding one year old male control exposed to the virus during infancy but not hospitalised and/or treated for bronchiolitis. The quantitative real-time polymerase chain reaction (QRT-PCR) study used TaqMan Low Density Array (TLDA) cards with gene expression given as mean relative quantification (RQ) from triplets of each gene and analyzed in the same five male infants as selected for microarray. A pooled sample from four of the five one year old male controls was used as exogenous control and beta-glucuronidase (GUSB) was used as endogenous control in the QRT-PCR study. The following genes (gene symbol and name) were evaluated; BPGM: 2,3-bisphosphoglycerate mutase, CLC: Charcot-Leyden crystal protein, DNAPTP6: DNA polymerase-transactivated protein 6, EPSTI1: Epithelial stromal interaction 1 (breast), ERAF: Erythroid associated factor, G1P2: Interferon, alpha-inducible protein (clone IFI-15K), HBD: Hemoglobin, delta, HBE1: Hemoglobin, epsilon 1, HP: Haptoglobin, IFI27: Interferon, alpha-inducible protein 27, IFI44: Interferon-induced protein 44, IFI44L: Interferon-induced protein 44-like, MARCO: Macrophage receptor with collagenous structure, MMP9: Matrix metalloproteinase 9, MS4A4A: Membrane-spanning 4-domains, subfamily A, member 4, NQO2: NAD(P)H dehydrogenase, quinone 2, STXBP2: Syntaxin binding protein 2

Figure 2

QRT-PCR study of six whole blood immune response genes as given by microarray. A quantitative real-time polymerase chain reaction (QRT-PCR) study using TaqMan Low Density Array (TLDA) cards was performed with gene expressions given as mean relative quantification (RQ) from triplets of each gene. Analyzed in 18 infants hospitalised with respiratory syncytial virus, subtype B, bronchiolitis versus a pooled sample from four one year old male children exposed to RSV during infancy but not treated and/or hospitalised for bronchiolitis during infancy as exogenous control and with beta-glucuronidase (GUSB) as endogenous control. The following genes (gene symbol and name) were studied; CLC: Charcot-Leyden crystal protein, G1P2: Interferon, alpha-inducible protein (clone IFI-15K), IFI27: Interferon, alpha-inducible protein 27, IFI44: Interferon-induced protein 44, IFI44L: Interferon-induced protein 44-like, MARCO:Macrophage receptor with collagenous structure