The respiratory syncytial virus (RSV) nonstructural proteins mediate RSV suppression of glucocorticoid receptor transactivation.

Respiratory syncytial virus (RSV)-induced bronchiolitis in infants is not responsive to glucocorticoids. We have shown that RSV infection impairs glucocorticoid receptor (GR) function. In this study, we have investigated the mechanism by which RSV impairs GR function. We have shown that RSV repression of GR-induced transactivation is not mediated through a soluble autocrine factor. Knock-down of mitochondrial antiviral signaling protein (MAVS), but not retinoic acid-inducible gene 1 (RIG-I) or myeloid differentiation primary response gene 88 (MyD88), impairs GR-mediated gene activation even in mock-infected cells. Over-expression of the RSV nonstructural protein NS1, but not NS2, impairs glucocorticoid-induced transactivation and viruses deleted in NS1 and/or NS2 are unable to repress glucocorticoid-induction of the known GR regulated gene glucocorticoid-inducible leucine zipper (GILZ). These data suggest that the RSV nonstructural proteins mediate RSV repression of GR-induced transactivation and that inhibition of the nonstructural proteins may be a viable target for therapy against RSV-related disease.

Respiratory syncytial virus (RSV) suppression of glucocorticoid receptor phosphorylation does not account for repression of transactivation.

Respiratory syncytial virus (RSV)-induced bronchiolitis in infants, although inflammatory in nature, is not responsive to glucocorticoids. We have recently shown that RSV-infected lung epithelial cells have impaired glucocorticoid receptor (GR)-mediated transactivation. In this study, we show that the N-terminal region of GR is required for RSV repression of GR transactivation and that RSV infection of lung epithelial cells reduces ligand-dependent GR phosphorylation at serine 211 and serine 226. However, we also show that these changes in GR phosphorylation do not account for the RSV repression of GR transactivation suggesting other regions of the GR N-terminus must also be involved.

Poly I:C and respiratory syncytial virus (RSV) inhibit glucocorticoid receptor (GR)-mediated transactivation in lung epithelial, but not monocytic, cell lines.

Respiratory syncytial virus (RSV)-induced bronchiolitis in infants is not responsive to glucocorticoids. We have recently shown that RSV infection of lung epithelial cells impairs glucocorticoid receptor (GR) function. In this current study, we have shown that the viral mimic poly I:C also represses GR-mediated gene activation in lung epithelial cells, suggesting that this might be a common phenomenon of other viral infections. However, we also show that neither RSV infection nor poly I:C affect GR-mediated gene activation in the monocytic cell line THP-1, suggesting that these effects on GR function may be cell-type specific.

Stressor-induced increase in microbicidal activity of splenic macrophages is dependent upon peroxynitrite production.

Exposing mice to a social stressor called social disruption (SDR) that involves repeated social defeat during intermale aggression results in increased circulating cytokines, such as interleukin-1α (IL-1α) and IL-1ß, and increased reactivity of splenic CD11b(+) macrophages to inflammatory stimuli. For example, upon lipopolysaccharide stimulation, macrophages from stressor-exposed mice produce higher levels of cytokines than do cells from nonstressed controls. Moreover, the SDR stressor enhances the ability of these macrophages to kill Escherichia coli both in vitro and in vivo, through a Toll-like receptor 4-dependent mechanism. The present study tested the hypothesis that stressor-enhanced bacterial killing is due to increases in the production of peroxynitrite. Male mice were exposed to the SDR stressor or were left undisturbed. Upon stimulation with E. coli, splenic macrophages from SDR-exposed mice expressed significantly increased levels of inducible nitric oxide synthase mRNA and produced higher levels of peroxynitrite. Blocking the production of peroxynitrite abrogated the SDR-induced increase in microbicidal activity. Studies in IL-1 receptor type 1 knockout mice indicated that the increased microbicidal activity and peroxynitrite production was dependent upon IL-1 signaling. These data confirm and extend the importance of IL-1 signaling for stressor-induced immunopotentiation; the finding that inhibiting superoxide or nitric oxide production inhibits both peroxynitrite production and killing of E. coli demonstrates that peroxynitrite mediates the stressor-induced increase in bacterial killing.

Respiratory syncytial virus represses glucocorticoid receptor-mediated gene activation.

Respiratory syncytial virus (RSV) is a common cause of bronchiolitis in infants. Although antiinflammatory in nature, glucocorticoids have been shown to be ineffective in the treatment of RSV-induced bronchiolitis and wheezing. In addition, the effectiveness of glucocorticoids at inhibiting RSV-induced proinflammatory cytokine production in cell culture has been questioned. In this study, we have investigated the effect of RSV infection on glucocorticoid-induced gene activation in lung epithelium-derived cells. We show that RSV infection inhibits dexamethasone induction of three glucocorticoid receptor (GR)-regulated genes (glucocorticoid-inducible leucine zipper, FK506 binding protein, and MAPK phosphatase 1) in A549, BEAS-2B cells, and primary small airway epithelial cells. UV irradiation of the virus prevents this repression, suggesting that viral replication is required. RSV is known to activate the nuclear factor κB (NFκB) pathway, which is mutually antagonistic towards the GR pathway. However, specific inhibition of NFκB had no effect on the repression of GR-induced genes by RSV infection, indicating that RSV repression of GR is independent of NFκB. RSV infection of A549 cells does not alter GR protein levels or GR nuclear translocation but does reduce GR binding to the promoters of the glucocorticoid responsive genes analyzed in this study. Repression of GR by RSV infection may account for the apparent clinical ineffectiveness of glucocorticoids in RSV bronchiolitis therapy. In addition, this data adds to our previously published data suggesting that GR may be a general target for infectious agents. Identifying the mechanisms through which this suppression occurs may lead to the development of novel therapeutics.

Glucocorticoids activate Epstein Barr virus lytic replication through the upregulation of immediate early BZLF1 gene expression.

Psychological stress-associated immune dysregulation has been shown to disrupt the steady-state expression and reactivate latent herpes viruses. One such virus is the Epstein Barr virus (EBV), which is associated with several human malignancies. EBV infects >90% of people living in North America and persists for life in latently infected cells. Although several studies have shown that glucocorticoids (GCs) can directly induce reactivation of the latent virus, the mechanism of stress hormone involvement in the control of EBV gene expression is not well understood. In this study, we tested the hypothesis that GCs can induce the latent EBV genome to lytically replicate through the induction of the EBV immediate early gene BZLF1 which encodes the lytic transactivator protein ZEBRA. We show a dose-dependent upregulation of BZLF1 mRNA expression by hydrocortisone (HC) and dexamethasone (Dex) in Daudi cells, an EBV genome positive Burkitt's lymphoma cell line, and Dex-induction of the early gene products BLLF3 (encoding for the EBV dUTPase) and BALF5 (encoding for the EBV DNA polymerase). We show that Daudi cells express glucocorticoid receptors (GR) that mediate Dex-dependent upregulation of BZLF1 mRNA levels. This effect was inhibited by both the glucocorticoid receptor antagonist RU486 and by cycloheximide. The results suggest that GCs, in addition to inducing stress-related immune dysregulation, can mediate latent EBV reactivation through the induction of the BZLF1 gene.

Bacillus anthracis lethal toxin represses MMTV promoter activity through transcription factors.

We have recently shown that the anthrax lethal toxin (LeTx) selectively represses nuclear hormone receptors. In this study, we found that LeTx repressed the activation of the mouse mammary tumor virus promoter related to overexpression of the transcription factors hepatocyte nuclear factor 3, octamer-binding protein 1, and c-Jun. LeTx transcriptional repression was associated with a decrease in the protein levels of these transcription factors in a lethal factor protease activity-dependent manner. Early administration of LeTx antagonists partially or completely abolished the repressive effects of LeTx. In contrast to the rapid cleavage of mitogen-activated protein kinase kinases by LeTx, the degradation of these transcription factors occurred at a relatively late stage after LeTx treatment. In addition, LeTx repressed phorbol-12-myristate-13-acetate-induced mouse mammary tumor virus promoter activity and phorbol 12-myristate 13-acetate induction of endogenous c-Jun protein. Collectively, these findings suggest that transcription factors are intracellular targets of LeTx and expand our understanding of the molecular action of LeTx at a later stage of low-dose exposure.

Norepinephrine upregulates VEGF, IL-8, and IL-6 expression in human melanoma tumor cell lines: implications for stress-related enhancement of tumor progression.

Studies suggest that stress can be a co-factor for the initiation and progression of cancer. The catecholamine stress hormone, norepinephrine (NE), may influence tumor progression by modulating the expression of factors implicated in angiogenesis and metastasis. The goal of this study was to examine the influence of NE on the expression of VEGF, IL-8, and IL-6 by the human melanoma cell lines, C8161, 1174MEL, and Me18105. Cells were treated with NE and levels of VEGF, IL-8, and IL-6 were measured using ELISA and real-time PCR. The expression of beta-adrenergic receptors (beta-ARs) mRNA and protein were also assessed. Finally, immunohistochemistry was utilized to examine the presence of beta1- and beta2-AR in primary and metastatic human melanoma biopsies. We show that NE treatment upregulated production of VEGF, IL-8, and IL-6 in C8161 cells and to a lesser extent 1174MEL and Me18105 cells. The upregulation was associated with induced gene expression. The effect on C8161 cells was mediated by both beta1- and beta2-ARs. Furthermore, 18 of 20 melanoma biopsies examined expressed beta2-AR while 14 of 20 melanoma biopsies expressed beta1-AR. Our data support the hypothesis that NE can stimulate the aggressive potential of melanoma tumor cells, in part, by inducing the production VEGF, IL-8, and IL-6. This line of research further suggests that interventions targeting components of the activated sympathetic-adrenal medullary (SAM) axis, or the utilization of beta-AR blocking agents, may represent new strategies for slowing down the progression of malignant disease and improving cancer patients' quality of life.

Stress hormones and immune function.

Over the past 20 years we have demonstrated both in animal models and in human studies that stress increases neuroendocrine hormones, particularly glucocorticoids and catecholamines but to some extent also prolactin, growth hormone and nerve growth factor. We have also shown that stress, through the action of these stress hormones, has detrimental effects on immune function, including reduced NK cell activity, lymphocyte populations, lymphocyte proliferation, antibody production and reactivation of latent viral infections. Such effects on the immune system have severe consequences on health which include, but are not limited to, delayed wound healing, impaired responses to vaccination and development and progression of cancer. These data provide scientific evidence of the effects of stress on immune function and implications for health.