Evidence for respiratory viruses interactions in asymptomatic preschool-aged children

Aim: To prospectively evaluate interferences between viruses of the upper respiratory tract in asymptomatic preschool children. Methods: Nasal-pharyngeal swabs from 233 preschool aged children were prospectively collected over four consecutive time periods, during one school year. The samples were tested using a RT-PCR DNA/RNA microarray system for nine respiratory viruses. Results: Respiratory syncytial virus (RSV) was a predictor of the presence of influenza virus (INFL) (OR: 9.12, CI: 1.52-54.75, p = 0.016), and similarly, INFL predicted the presence of RSV (OR: 4.01, CI: 1.14-14.16, p = 0.030). Also, rhinovirus (RV) was a predictor of adenovirus (ADV) presence (OR: 3.66, CI: 1.10-12.14, p = 0.034), and similarly, ADV predicted the presence of RV (OR: 4.05, CI: 1.02-16.05, p = 0.046). No other significant associations between viruses were observed. Conclusion: Our results indicate that respiratory viruses found in carrier stage in asymptomatic children may interact with other viruses and even facilitate their settling in the upper respiratory tract. The pathophysiological role of these interactions is not yet clear

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Evidence for respiratory viruses interactions in asymptomatic preschool-aged children.

AIM: To prospectively evaluate interferences between viruses of the upper respiratory tract in asymptomatic preschool children. METHODS: Nasal-pharyngeal swabs from 233 preschool aged children were prospectively collected over four consecutive time periods, during one school year. The samples were tested using a RT-PCR DNA/RNA microarray system for nine respiratory viruses. RESULTS: Respiratory syncytial virus (RSV) was a predictor of the presence of influenza virus (INFL) (OR: 9.12, CI: 1.52-54.75, p=0.016), and similarly, INFL predicted the presence of RSV (OR: 4.01, CI: 1.14-14.16, p=0.030). Also, rhinovirus (RV) was a predictor of adenovirus (ADV) presence (OR: 3.66, CI: 1.10-12.14, p=0.034), and similarly, ADV predicted the presence of RV (OR: 4.05, CI: 1.02-16.05, p=0.046). No other significant associations between viruses were observed. CONCLUSION: Our results indicate that respiratory viruses found in carrier stage in asymptomatic children may interact with other viruses and even facilitate their settling in the upper respiratory tract. The pathophysiological role of these interactions is not yet clear.

Genetic screening for individuals at high risk for type 1 diabetes in the general population using HLA Class II alleles as disease markers. A comparison between three European populations with variable rates of disease incidence.

BACKGROUND: To develop screening strategies for identification of individuals at increased genetic risk for type 1 diabetes in three populations with variable disease incidence rates and distinct ethnic origin. METHODS: A stepwise HLA DQB1-DQA1-DRB1-based screening approach was evaluated. Patients with childhood-onset type 1 diabetes were recruited from Finland (n = 1739), Hungary (n = 149), and Greece (n = 119). Consecutive newborns (2568 from Finland and 1047 from Greece) or healthy schoolchildren (n = 177 from Hungary) served as controls. RESULTS: The DQB1*02/0302 genotype conferred the highest disease risk in all populations. The DQB1*02/y (y not equal DQB1*0301,*0302,*0602,*0603, *0604) genotypes were more common and conferred a higher disease risk in the Greek population (OR 4.9) compared to the Finns (OR 1.2). DQB1*0302/x (x not equal DQB1*02, *0301, *0602, *0603, *0604) genotypes were, in contrast, more prevalent among Finnish cases (32.7%) as compared to Hungarians (18.1%) or Greeks (13.5%). The protective DQB1*0602 or *0603 positive genotypes were most common in the Finns, while DQB1*0301 was more common in Hungarians and Greeks. In all groups, DQA1 and DRB1*04 typing considerably increased the sensitivity of the DQB1-based screening. The different high-risk genotype combinations present in about 10% of the background population had a diagnostic sensitivity of 60% in Finland and 80% in Hungary and Greece. CONCLUSIONS: HLA DR-DQ-based screening is a feasible tool for the identification of individuals at increased genetic risk for type 1 diabetes in populations with diverse genetic background. The risk markers should, however, be individually selected for the target population since the screening efficiency of various markers is highly dependent on the ethnic group studied.

Protooncogene c-fos Involvement in the Molecular Mechanism of Rat Brain Sexual Differentiation.

Brain sexual differentiation is mediated through testosterone, which acts during the perinatal period in the form of both 5alpha-dihydrotestosterone and estradiol. In order to gain insight into the molecular mechanisms involved, we studied induction of c-fos, an index of functional neuronal activation, in the 2-day-old female rat brain after injection of a masculinizing dose of testosterone. Administration of testosterone resulted in induction of c-fos gene expression in the hypothalamus, as determined by Northern analysis. Following immunocytochemistry, we demonstrated an increase in the number of Fos-positive nuclei in the median and medial preoptic nucleus, the medial preoptic area extending to the lateral preoptic area, and the peri- and paraventricular area. In an effort to see whether testosterone acted as 5alpha-dihydrotestosterone or as estradiol, we injected either steroid and looked at fos induction. Estradiol mimicked the effect of testosterone, while 5alpha-dihydrotestosterone was without effect. Furthermore, injection of an estrogen receptor blocker, clomiphene, together with testosterone, abolished the testosterone-induced increase in Fos-positive nuclei, thus confirming the finding that testosterone induces c-fos by acting through estrogen receptors. Electrophoretic mobility shift assays showed that nuclear extracts from 2-day-old female hypothalamus contain a protein, most probably the estrogen receptor, which binds specifically to oligodeoxynucleotides with the sequence of either vitERE, the consensus estrogen-responsive element (ERE) found in the vittelogenin gene, or fosERE, the ERE found in the 3'-untranslated region of the mouse c-fos gene. This suggests that the effect of testosterone-derived estradiol on c-fos expression is a direct one, mediated by binding of estrogen receptors to an ERE in the c-fos gene-regulatory regions.

In utero radiation-induced apoptosis and p53 gene expression in the developing rat brain.

PURPOSE: This study addressed the question of the role of the p53 gene in prenatal low-dose radiation-induced apoptosis in the neuroepithelium, in an effort to elucidate molecular mechanisms involved in the extreme radiosensitivity of the developing brain. MATERIALS AND METHODS: Pregnant Wistar rats were exposed to a single dose of 10, 20 or 40 cGy of X-rays on day 15 or 17 of gestation. Animals were sacrificed 4 or 24h after exposure. Apoptosis was studied by gel electrophoresis of isolated DNA and in situ by the TUNEL reaction. Expression of the p53 gene was studied by immunocytochemistry and Western analysis, as well as Northern analysis, for the detection of the protein and mRNA respectively. RESULTS: In utero low-dose irradiation led to apoptosis and an increase of p53 gene expression in the developing rat brain. Apoptotic as well as p53 immunopositive cells were detected among proliferating, migratory and post-mitotic neurones in the developing neuroepithelium following prenatal irradiation, even after only l0 cGy. In addition to the p53 protein, p53 mRNA brain levels were also increased following prenatal irradiation. CONCLUSIONS: Low-dose prenatal irradiation of the developing brain led to p53 induction and cell death by apoptosis.

In utero radiation-induced changes in growth factor levels in the developing rat brain.

PURPOSE: To investigate the role of growth factors in the compensatory response to radiation injury during development of the brain. Levels of gene expression in the embryonic rat brain were assessed for IGF-I, IGF-II, BDNF and NT-3. MATERIALS AND METHODS: Pregnant Wistar rats were exposed to a single dose of 10, 20 or 40 cGy X-rays on day 15 or 17 of gestation. Animals were sacrificed 4 or 24 h after exposure. IGF-I, BDNF and NT-3 proteins were detected by immunocytochemistry, while IGF-I and IGF-II mRNA by in situ hybridization, and Northern analysis respectively. RESULTS: In utero low dose X-irradiation led to a decrease in IGF-I gene expression and a compensatory increase in the expression of IGF-II, BDNF and NT-3 in the developing rat brain. IGF-I, BDNF and NT-3 immunopositive cells were detected among proliferating, migratory and post-mitotic neurones in the developing neuroepithelium. CONCLUSIONS: Low dose prenatal irradiation of the developing brain results in down-regulation of IGF-I, which could lead to cell death by apoptosis. On the other hand, IGF-II, BDNF and NT-3 gene expression is increased following irradiation, possibly as a compensatory mechanism.

NMDA receptor mediated changes in IGF-II gene expression in the rat brain after injury and the possible role of nitric oxide.

This study was undertaken in order to investigate the role of insulin-like growth factor (IGF)-II, c-fos, N-methyl-D-aspartate (NMDA) receptors, and nNOS in the cellular processes following a penetrating brain injury. IGF-II mRNA levels, as determined by Northern analysis, were decreased at 4, 8, and 24 h after brain injury, in the lesioned, compared to the contralateral intact hemisphere. Forty-eight and 72 h after the injury, there was no difference between the lesioned and the contralateral intact hemisphere in IGF-II mRNA levels. c-fos mRNA levels followed a parallel, but opposite course: They were increased at 4, 8 and 24 h after the injury, while at 48 and 72 h c-fos mRNA levels in the lesioned hemisphere did not differ from those in the intact. Administration of MK-801 reversed the injury-induced decrease in IGF-II mRNA levels. Administration of MK-801 resulted in an increase in IGF-II mRNA in both the intact and the lesioned hemispheres. Brain injury resulted in an increase in nNOS immunopositive cells in the hippocampal formation, which was detectable at 4 and 12, but not 48 h after the injury. These results suggest that IGF-II, c-fos, NMDA receptors and nNOS are involved in the cellular responses to brain injury.

p53 expression and regulation by NMDA receptors in the developing rat brain.

The p53 tumor suppressor gene, which is considered the guardian of the genome, encodes a phosphoprotein, which is a sequence-specific transcriptional activator or repressor of target genes. The role of p53 in developmental processes has not been studied extensively, although its expression appears to undergo temporal and spatial changes during prenatal and postnatal development. In the present study, we assessed the levels of p53 mRNA and protein in the developing rat brain and its relation to developmental cell death. Furthermore, we investigated the potential role of n-methyl-d-aspartate (NMDA) receptors in regulating p53 expression, since these receptors are involved in the control of cell death. We found that p53 mRNA and protein were detectable in the rat brain throughout perinatal development. In embryos, p53 immunoreactivity was mainly localized in the nuclei of neuroepithelial cells, with a maximum in staining at embryonic day (E)12. In the neuroepithelium, we also found significant numbers of TdT-mediated dUTP nick end labeling (TUNEL)-positive cells, both in dividing periventricular cells and in migrating neurons. In neonates, immediately after birth there was a reduction in the number of apoptotic cells, which then increased to reach a maximum at postnatal day (P)5. Postnatally, apoptotic as well as p53-positive cells were detected in most brain areas. P53 immunoreactivity was also highest on P5. In most cells, p53 immunoreactivity and the TUNEL signal colocalized. P53 immunoreactivity as well as the number of TUNEL- positive cells were dramatically decreased in the brains of newborns treated with MK-801, an NMDA receptor antagonist. Our results show that p53 is involved in the control of developmental cell death, and that NMDA receptors play a regulatory role in the expression of the p53 gene, and thus in apoptosis occurring in the developing rat brain.

At least three neurotransmitter systems mediate a stress-induced increase in c-fos mRNA in different rat brain areas.

1. Protooncogene c-fos mRNA levels were determined in the rat cerebral cortex, hippocampus, and cerebellum after exposure to a combined forced swimming and confinement stress. The stress resulted in an increase in c-fos mRNA levels in all three brain areas. 2. In an effort to elucidate the neurotransmitter systems involved in this stress-induced increase, animals were injected, prior to exposure to the stress, with either diazepam, MK-801, or propranolol. 3. In both the cerebral cortex and the hippocampus the stress-induced increase in c-fos mRNA was inhibited by MK-801, suggesting that it is mediated via NMDA receptors. In the hippocampus, propranolol had a similar effect, indicating that beta-adrenergic receptors are also involved in the stress-induced increase in c-fos mRNA. 4. On the other hand, the increase in c-fos mRNA produced by the stress of the injection was inhibited in the cerebral cortex by diazepam or propranolol and in the hippocampus only by diazepam. Furthermore, administration of MK-801 resulted in an increase in c-fos mRNA in the hippocampus of the nonstressed animals. In the cerebellum no one of the three drugs employed affected c-fos mRNA levels in either stressed or nonstressed animals. 5. Our results thus show that various forms of stress activate, in different brain areas, neurons with either NMDA, beta-adrenergic, and/or GABA-A receptors.

Aging-related changes in IGF-II and c-fos gene expression in the rat brain.

The protein products of growth factor genes such as IGF-II and cellular oncogenes such as c-fos are believed to be necessary for the support of normal neuronal function. Steady-state levels of c-fos and IGF-II mRNA were determined in the brain of young and old rats, using Northern analysis. Both RNAs were found to be decreased in the brain of aged rats. Age-related decrease was detected in the hippocampus, hypothalamus, striatum, cerebral cortex and cerebellum, for IGF-II mRNA, and in the cerebral cortex and cerebellum for c-fos mRNA. Furthermore, changes in the degree and pattern of DNA methylation were noted at both gene loci, in the aged rat brain. Our results could reflect changes at the genomic level possibly related to the process of aging and the accompanying decline in brain function.