RESUMEN
PURPOSE: Resuscitation of newborns is one of the most frequent procedures in neonatal medicine. The use of supplementary oxygen during resuscitation of the asphyxiated newborn has been shown to be detrimental to vulnerable tissues. We wanted to assess transcriptional changes in ocular tissue after the acute use of oxygen in the delivery room in a hypoxia-reoxygenation model of the newborn mouse. METHODS: C57BL/6 mice (n = 57), postnatal day 7, were randomized to receive either 120 minutes of hypoxia, at 8% O2, followed by 30 minutes of reoxygenation with 21, 40, 60, or 100% O2 or to normoxia followed by 30 minutes of 21% or 100% O2. Whole ocular homogenates were analyzed by Affymetrix 750k expression array, and RT-PCR was performed for validation. Bayesian analysis of variance for microarray data (BAMarray) was used to identify single significant genes, and Gene Set Enrichment Analysis (GSEA) was applied to reveal significant pathway systems. RESULTS: In total, â¼ 92% of the gene expression changes were altered in response to reoxygenation with 60% or 100% O2 compared to expression at the lower percentages of 21% and 40%. After 100% O2 treatment, genes involved in inflammation (Ccl12), angiogenesis (Igfr1, Stat3), and metabolism (Hk2) were upregulated. Pathway analyses after hypoxia-reoxygenation revealed significant alterations of six pathways which included apoptosis, TGF-beta signaling, oxidative phosphorylation, voltage-gated calcium channel complex, mitochondrion, and regulation of RAS protein signal transduction. CONCLUSIONS: Hypoxia-reoxygenation can induce immediate transcriptional responses in ocular tissue involving inflammation, angiogenesis, energy failure, and Ras signaling.
Asunto(s)
Regulación del Desarrollo de la Expresión Génica , Hiperoxia/genética , Hipoxia/genética , Proteínas Quimioatrayentes de Monocitos/genética , Estrés Oxidativo/genética , ARN/genética , Factor de Transcripción STAT3/genética , Animales , Animales Recién Nacidos , Modelos Animales de Enfermedad , Hiperoxia/metabolismo , Hipoxia/metabolismo , Ratones , Ratones Endogámicos C57BL , Análisis por Micromatrices , Proteínas Quimioatrayentes de Monocitos/biosíntesis , Reacción en Cadena en Tiempo Real de la Polimerasa , Factor de Transcripción STAT3/biosíntesis , Transducción de SeñalRESUMEN
BACKGROUND: The intestinal mitosis-inhibiting peptide pyroglu-His-GlyOH (pEHG), inhibits normal intestinal epithelial cells and the human colon adenocarcinoma cell line HT-29 and increases the expression of c-fos (1). In this study, we investigated the mechanisms of the growth-inhibiting effects of pEHG. MATERIALS AND METHODS: cDNA expression array was hybridized with cDNA from HT-29 cells exposed to pEHG or control. The results were confirmed with Northern blot or real-time PCR. RESULTS: pEHG(1 nM) provoked a significant increase in the expression of the early growth response protein 1 (egr-1) after an incubation of 20 minutes, while c-fos was confirmed up-regulated by the same treatment. We further studied the expression of fosB, c-jun and junB, in the AP-1 complex. fosB was up-regulated 20-fold, but only minor effects on jun variants were observed. CONCLUSION: pEHG stimulates the gene expression of some immediate-early transcription factors involved in cell proliferation.