We have performed several lines of experiments to examine the interactions between the endocrine and immune systems. We showed that two small molecules an IL-6 antagonist and a PPAR-delta agonist have strong anti-inflammatory activities via STAT3 inhibition. The former compound, TB-2-081, interacted with the gp130 subunit of the IL-6-type receptors and blocked the activity not only of IL-6 but also of the leukemia inhibitory factor, oncostatin M and IL-11. A major change in the cell was the inhibition of the tyrosine phosphorylation of the STAT3 transcription factor. The latter compound, GW50516, inhibited the production of several acute phase reactants by hepatic cells showing thus major anti-inflammatory activity. This effect was also mediated by inhibition of STAT3 activity at the promoter of responsive genes. Viruses, including the human immunodeficiency syndrome type-1 (HIV-1), cytomegalovirus (CMV), and Newcastle disease (NDV) viruses, are potent activators and modulators of the host immune and endocrine systems, influencing hormonal actions in host tissues, such as leukocytes, adipose tissue and skeletal muscles. These effects further contribute to viral expansion and pathogenesis. Indeed, we demonstrated that HIV-1 accessory protein Vpr suppresses PPAR-gamma activity playing a potentially important role in the development of the characteristic AIDS-associated lipodystrophy and insulin-resistance syndrome. In dendritic cells (DCs), which play a central role in the recognition and presentation of viral antigens, infection of CMV or NDV, and perhaps HIV-1, causes dramatic changes in the expression of a group of nuclear hormone receptors, including the glucocorticoid and estrogen receptors, as well as of several transcriptional co-regulators, such as p300 and p160-type histone acetyltransferase coactivators, possibly altering secretion/production of interferons and other cytokines by these cells. We have published one manuscript based on the results obtained in the mRNA expression profiling of nuclear hormone receptors and coregulators. Since NOR1, one member of NR4A group nuclear receptors, was the most highly regulated NR upon viral infection in DCs, we obtained NOR1 knockout mice from Dr. Conneely, the Baylor collage of medicine, and have examined impact of pathogen infection to the action of DCs purified from NOR1 knockout mice. DCs from these mice showed a significant defect in the response of interleukin (IL)-12 to viral infection compared to those from wild type mice, while NOR-1 potently stimulated the IL-12 p40 promoter activity in reporter assays. NOR-1 knockout mice demonstrated significant reduction of IL-12 production against infection of Toxoplasma gondii, a protozoa against which IL-12 acts as an essential component for host defense. We are currently examining details of molecular regulation of NOR-1 on IL-12 production both in the cellular and animal systems. In the same line of experiments, we found that CMV and NDV stimulated IL-10 expression in DCs, and glucocorticoids further potentiated such virus-induced expression of this cytokine. Since IL-10 inhibits synthesis of pro-inflammatory cytokines and has ability to suppress antigen presentation by DCs and monocytes, synergistic activation of IL-10 by glucocorticoids may explain why exposure to stress and subsequent activation of the HPA axis increases susceptibility to viral infection, and possibly, subsequent development of viral-associated disorders, such as asthma, atherosclerosis and caners. We found that viral infection activated the extracellular signal-regulated kinase 1 (ERK1), a member of mitogen-activated protein kinase family, which in turn phosphorylated the human GR at serine located at amino acid position 211 and modulated the transcriptional activity of GR on the IL-10 promoter. Extracellular hyperosmolarity or osmotic stress is a major threat for land organisms, and thus strongly stimulates HPA axis through secretion of ariginine vasopression from the hypothalamus/posterior lobe of the pituitary gland. In addition to this systemic response, osmotic stress also activates a cellular signaling cascade called adaptive response to extracellular hyperosmolarity: Extracellular hyperosmolarity induces and activates a Rel-homology domain-containing transcription factor, the nuclear factor of activated T-cells 5 (NFAT5), which subsequently stimulates transcription of osmotic stress-responsive genes and causes intracellular accumulation of small organic osmolytes to maintain isotonicity between the inside and outside of the cells. We previously reported an intracellular signaling cascade responsive to osmotic stress in lymphocytes: The Rho-type guanine nucleotide exchange factor (GEF) Brx or AKAP13 is essential for the osmotic stress-stimulated expression of NFAT5, and is a key component of the intracellular signaling cascade transmitting the extracellular hyperosmolarity signal to the nucleus. Osmotic stress-mediated induction of NFAT5 requires the Brx GEF domain and p38 mitogen-activated kinase (MAPK), while Brx in response to osmotic stress attracts through its C-terminal domain the cJun kinase (JNK)-interacting protein (JIP) 4, a scaffold specific to activation of the p38 MAPK cascade and NFAT5, coupling activated Rho-type small G proteins to components of the p38 MAPK signaling pathway. Importantly, this signaling system plays a critical role in the differentiation of lymphocytes in the spleen and stimulates production of TNF-alpha and other cytokines. Osmotic stress-mediated activation of this cellular signaling system may also be an important component in immune dysregulation observed in some pathologic conditions accompanied by plasma hyperosmolarity, such as diabetes mellitus, uremia, dehydration and severe burn all of which are known to develop altered cytokine production and immune-associated symptoms/signs. To further elucidate roles of Brx/NFAT5-mediated osmotic stress on the regulation of the immune system, we are now developing mice carrying specific deletion of the brx gene in DCs/monocytes/macrophages using the Cre/LoxP system. We hypothesized that Brx/NFAT5 plays a significant roles in the high-salt diet-induced decrease of nitric oxide synthase and subsequent increase of blood pressure, as recent publication indicated that NFAT5 expressed in residential macrophages participate in this BP regulatory system. Using the conditional Brx knockout mice in DCs/macrophages, we will examine contribution of Brx on high salt diet-induced hypertension, as well as on the cardiac fibrosis, which is in part mediated by inflammation.

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Agarwal, Neeti; Iyer, Dinakar; Patel, Sanjeet G et al. (2013) HIV-1 Vpr induces adipose dysfunction in vivo through reciprocal effects on PPAR/GR co-regulation. Sci Transl Med 5:213ra164
Shrivastav, Shashi; Zhang, Liyan; Okamoto, Koji et al. (2013) HIV-1 Vpr enhances PPAR?/?-mediated transcription, increases PDK4 expression, and reduces PDC activity. Mol Endocrinol 27:1564-76
Ng, Sinnie Sin Man; Li, Andrew; Pavlakis, George N et al. (2013) Viral infection increases glucocorticoid-induced interleukin-10 production through ERK-mediated phosphorylation of the glucocorticoid receptor in dendritic cells: potential clinical implications. PLoS One 8:e63587
Kino, Tomoshige (2012) Circadian rhythms of glucocorticoid hormone actions in target tissues: potential clinical implications. Sci Signal 5:pt4
Kino, T; Charmandari, E; Chrousos, G P (2011) Glucocorticoid receptor: implications for rheumatic diseases. Clin Exp Rheumatol 29:S32-41
Kino, Tomoshige; Chrousos, George P (2011) Circadian CLOCK-mediated regulation of target-tissue sensitivity to glucocorticoids: implications for cardiometabolic diseases. Endocr Dev 20:116-26
Ng, Sinnie Sin Man; Chang, Tsung-Hsien; Tailor, Prafullakumar et al. (2011) Virus-induced differential expression of nuclear receptors and coregulators in dendritic cells: implication to interferon production. FEBS Lett 585:1331-7
Kino, Tomoshige; Segars, James H; Chrousos, George P (2010) The Guanine Nucleotide Exchange Factor Brx: A Link between Osmotic Stress, Inflammation and Organ Physiology and Pathophysiology. Expert Rev Endocrinol Metab 5:603-614
Kino, Tomoshige; Chrousos, George P (2009) Tumor-associated, estrogen receptor-related antigen EBAG9: linking intracellular vesicle trafficking, immune homeostasis, and malignancy. Mol Interv 9:294-8
Kino, Tomoshige; Takatori, Hiroaki; Manoli, Irini et al. (2009) Brx mediates the response of lymphocytes to osmotic stress through the activation of NFAT5. Sci Signal 2:ra5

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