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Macrophages are professional phagocytes that display remarkable plasticity, with a range of phenotypes that can be broadly characterized by the M1/M2 dichotomy. Glucocorticoid (GC)-induced leucine zipper (GILZ) is a protein known to mediate anti-inflammatory and some pro-resolving actions, including as neutrophil apoptosis. However, the role of GILZ in key macrophage function is not well understood. Here, we investigated the role of GILZ on macrophage reprogramming and efferocytosis. Using murine bone-marrow-derived macrophages (BMDMs), we found that GILZ was expressed in naive BMDMs and exhibited increased expression in M2-like macrophages (IL4-differentiated). M1-like macrophages (IFN/LPS-differentiated) from GILZ -/- mice showed higher expression of the M1 markers CD86, MHC class II, iNOS, IL-6 and TNF-α, associated with increased levels of phosphorylated STAT1 and lower IL-10 levels, compared to M1-differentiated cells from WT mice. There were no changes in the M2 markers CD206 and arginase-1 in macrophages from GILZ -/- mice differentiated with IL-4, compared to cells from WT animals. Treatment of M1-like macrophages with TAT-GILZ, a cell-permeable GILZ fusion protein, decreased the levels of CD86 and MHC class II in M1-like macrophages without modifying CD206 levels in M2-like macrophages. In line with the in vitro data, increased numbers of M1-like macrophages were found into the pleural cavity of GILZ -/- mice after LPS-injection, compared to WT mice. Moreover, efferocytosis was defective in the context of GILZ deficiency, both in vitro and in vivo. Conversely, treatment of LPS-injected mice with TAT-GILZ promoted inflammation resolution, associated with lower numbers of M1-like macrophages and increased efferocytosis. Collectively, these data indicate that GILZ is a regulator of important macrophage functions, contributing to macrophage reprogramming and efferocytosis, both key steps for the resolution of inflammation., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Glucocorticoids are hormones that regulate several functions in living organisms and synthetic glucocorticoids are the most powerful anti-inflammatory pharmacological tool that is currently available. Although glucocorticoids have an immunosuppressive effect on immune cells, they exert multiple and sometimes contradictory effects on neutrophils. From being extremely sensitive to the anti-inflammatory effects of glucocorticoids to resisting glucocorticoid-induced apoptosis, neutrophils are proving to be more complex than they were earlier thought to be. The aim of this review is to explain these complex pathways by which neutrophils respond to endogenous or to exogenous glucocorticoids, both under physiological and pathological conditions.

Bursera microphylla (BM), one of the common elephant trees, is widely distributed in the Sonoran Desert in Mexico. The Seri ethnic group in the Sonoran Desert uses BM as an anti-inflammatory and painkiller drug for the treatment of sore throat, herpes labialis, abscessed tooth, and wound healing. Dried stems and leaves of BM are used in a tea to relieve painful urination and to stimulate bronchial secretion. Furthermore, BM is used for fighting venereal diseases. To investigate the effects of the hexane fraction of resin methanol extract (BM-H) on cell growth, the acute myeloid cell line (OCI-AML3) was treated with 250, 25, or 2.5 µg/mL of BM-H. The first 2 concentrations were able to significantly decrease OCI-AML3 cell number. This reduced cell number was associated with decreased S-phase, blockade of the G 2 /M phase of the cell cycle, and increased cell death. Similar results were obtained on all tested tumor cell lines of different origins. We found that blockade of the cell cycle was due to upregulation of p21 protein in a p53-independent way. Increase of p21 was possibly due to upstream upregulation of p-ERK (which stabilizes p21 protein) and downregulation of p-38 (which promotes its degradation). Regarding cell death, activation of caspase-3, but not of caspase-8 or -9, was detectable after BM-H treatment. In conclusion, these data suggest that the BM's hexane fraction inhibited proliferation of cell lines mainly by a p21-dependent, p53-independent mechanism and promoted apoptosis through activation of caspase-3, but not caspase-8 or -9.

Bursera microphylla (BM), one of the common elephant trees, is widely distributed in the Sonoran desert in Mexico. The Seri ethnic group in the Sonoran desert uses BM as an anti-inflammatory and painkiller drug for the treatment of sore throat, herpes labialis, abscessed tooth, and wound healing. Dried stems and leaves of BM are used in a tea to relieve painful urination and to stimulate bronchial secretion. Furthermore, BM is used for fighting venereal diseases. To investigate the effects of the hexane fraction of resin methanol extract (BM-H) on cell growth, the acute myeloid cell line (OCI-AML3) was treated with 250, 25, or 2.5 µg/mL of BM-H. The first 2 concentrations were able to significantly decrease OCI-AML3 cell number. This reduced cell number was associated with decreased S-phase, blockade of G 2 /M phase of the cell cycle, and increased cell death. Similar results were obtained on all tested tumor cell lines of different origins. We found that blockade of the cell cycle was a result of upregulation of p21 protein in a p53-independent way. Increase of p21 was possibly a result of upstream upregulation of p-ERK (which stabilizes p21 protein) and downregulation of p-38 (which promotes its degradation). Regarding cell death, activation of caspase-3, but not of caspase-8 or -9, was detectable after BM-H treatment. In conclusion, these data suggest that BM-H inhibited proliferation of cell lines mainly by a p21-dependent, p53-independent mechanism and promoted apoptosis through activation of caspase-3 but not caspase-8 or -9.

Background: Cytokines belonging to the TNF superfamily play a relevant role in neurodegenerative processes. Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL), released during neuronal injury, has proven to potently mediate and sustain neurotoxic processes leading to neuronal death. Similarly to TRAIL, the cytokine Glucocorticoid-induced TNF receptor ligand (GITRL) is able to transduce proapoptotic signals. In spite of the array of reports suggesting relationships between TRAIL and other cytokines, scanty data are, so far, available about a GITRL/TRAIL crosstalk., Methods: Here, we investigated possible interactions between TRAIL and the GITRL system in an in vitro model of neurodegeneration, using the human cortical neuronal cell line HCN-2. Cultured HCN-2 neurons were incubated at different times with GITRL and/or TRAIL, and thereafter nucleic acid and protein expression were measured. Real-time PCR analysis showed that the human cortical neuronal cell line HCN-2 does not express GITRL mRNA, but the latter is induced after treatment with TRAIL. In addition, HCN-2 cells did not express the GITRL receptor GITR mRNA, neither in control cultures, nor after treatment with TRAIL. All mRNA data were confirmed by western blot analysis of proteins. Cell viability assay showed that TRAIL, when associated to GITRL, was able to exert additive toxic effects. A counterproof was provided in experiments performed blocking GITRL, in which TRAIL-mediated toxicity appeared significantly reduced. Results suggest that GITRL/TRAIL redundancy during neurodegenerative processes implies extended potentiation of detrimental effects of both cytokines on neurons, eventually leading to larger cell damage and death., Conclusion: Finally, characterization of novel molecular targets within the TRAIL/GITRL interplay may represent a platform for innovative therapy of neurodegenerative disorders., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.)

Glucocorticoids (GC) are widely used as antiinflammatory/immunosuppressive drugs and antitumor agents in several types of lymphoma and leukemia. Therapeutic doses of GC induce growth-suppressive and cytotoxic effects on various leukocytes including B cells. Molecular mechanisms of GC action include induction of GC target genes. Glucocorticoid-induced leucine zipper (GILZ) is a rapidly, potently, and invariably GC-induced gene. It mediates a number of GC effects, such as control of cell proliferation, differentiation, and apoptosis. Here we show that deletion of GILZ in mice leads to an accumulation of B lymphocytes in the bone marrow, blood, and lymphoid tissues. Gilz knockout (KO) mice develop a progressive nonlethal B lymphocytosis, with expansion of B220(+) cells in the bone marrow and in the periphery, dependent on increased B-cell survival. Decreased B-cell apoptosis in mice lacking GILZ correlates with increased NF-κB transcriptional activity and Bcl-2 expression. B cell-specific gilz KO mice confirmed that the effect of GILZ deletion is B-cell self-intrinsic. These results establish GILZ as an important regulator of B-cell survival and suggest that the deregulation of GILZ expression could be implicated in the pathogenesis of B-cell disorders., (© 2015 by The American Society of Hematology.)

Rhabdomyosarcoma represents about 50% of soft-tissue sarcomas and 10% of malignant solid tumors in childhood. Embryonal rhabdomyosarcoma (ERMS) is the most frequent subtype, suggested to have an origin in muscle precursor cells that fail to exit the cell cycle and terminally differentiate mainly because of overexpression of the transcription factor, PAX7, which sustains proliferation, migration and invasiveness in ERMS cells. Artesunate (ARS) is a semi-synthetic derivative of artemisinin (ART), a natural compound well known as an antimalarial drug. However, ART and its derivatives have been found efficacious even as anticancer drugs that induce cell cycle arrest and/or apoptosis in several kinds of cancer. Here, we show that ARS dose-dependently induces DNA damage and apoptosis in ERMS cell lines. Production of reactive oxygen species (ROS) and activation of p38 MAPK have a central role in triggering ARS-mediated apoptosis in ERMS cells; indeed either the antioxidant, N-acetylcysteine or the p38 MAPK inhibitor, SB203580, protects ERMS cells from ARS-induced apoptosis. Moreover, ARS treatment in ERMS cells ROS-dependently induces the expression of the myo-miRs, miR-133a and miR-206, which are down-regulated in RMS, and reduces PAX7 protein levels. Finally, ARS upregulates the expression of the adhesion molecules, NCAM and integrin β1, and reduces migration and invasiveness of ERMS cells in vitro, and ARS treatment reduces of about 50% the growth of ERMS xenografts in vivo. Our results are the first evidence of efficacy of ART derivatives in restraining ERMS growth in vivo, and suggest ARS as a potential candidate for therapeutic treatment of ERMS., (© The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Glucocorticoids play a role in regulation of T lymphocytes homeostasis and development. In particular, glucocorticoid treatment induces massive apoptosis of CD4(+)CD8(+) double-positive (DP) thymocytes. This effect is due to many mechanisms, mainly driven by modulation of gene transcription. To find out which genes are modulated, we analyzed DP thymocytes treated for 3 h with dexamethasone (a synthetic glucocorticoid) by global gene expression profiling. Results indicate modulation of 163 genes, also confirmed by either RNase protection assay or real-time polymerase chain reaction. In particular, dexamethasone caused down-regulation of genes promoting DP thymocyte survival (e.g., Notch1, suppressor of cytokine signaling 1, and inhibitor of DNA binding 3) or modulation of genes activating cell death through the ceramide pathway (UDP-glucose ceramide glucosyltransferase, sphingosine 1-phosphate phosphatase, dihydroceramide desaturase, isoform 1, and G protein-coupled receptor 65) or through the mitochondrial machinery. Among the latter, there are Bcl-2 family members (Bim, Bfl-1, Bcl-xL, and Bcl-xbeta), genes involved in the control of redox status (thioredoxin reductase, thioredoxin reductase inhibitor, and NADP(+)-dependent isocitrate dehydrogenase) and genes belonging to Tis11 family that are involved in mRNA stability. Our study suggests that dexamethasone treatment of DP thymocytes modulates several genes belonging to apoptosis-related systems that can contribute to their apoptosis.

Glucocorticoids promote thymocyte apoptosis and modulate transcription of several genes including GILZ, which is strongly up-regulated in the thymus. We used transgenic mice overexpressing GILZ in the T-cell lineage to investigate TCR-triggered functions of GILZ-overexpressing thymocytes. TCR-triggered apoptosis, but not glucocorticoid-induced apoptosis, was inhibited in transgenic mice compared to their controls. In vivo anti-CD3 administration did not reduce CD4(+)CD8(+) thymocyte number. Analysis of TCR-triggered molecular changes indicated that p65 NF-kappaB nuclear translocation and DNA binding activity was inhibited in transgenic mice, which might be linked with apoptosis inhibition. IL-10 release increased whereas release of IL-2, IFN-gamma, IL-13 and IL-4 remained unchanged. These results support the hypothesis that GILZ regulates, at least in part, T-cell development by influencing thymus function at cellular and molecular levels.

Glucocorticoids, widely used therapeutic agents for several pathologies, act upon diverse cells and tissues, including the lympho-haemopoietic system. Glucocorticoid-mediated apoptosis has been described as one of the mechanisms underlying their pharmacological and physiological effects. Glucocorticoids induce apoptosis in thymocytes through genomic and non-genomic signals. We tested thymocyte apoptosis rates as induced by a panel of glucocorticoids. Using four glucocorticoids that are widely adopted in clinical practice we compared their induction of thymocyte apoptosis and activation of non-genomic and genomic signals, including phosphatidylinositol-specific phospholipase C (PI-PLC), caspase-8, -9 and -3, and Glucocorticoid-Induced Leucine Zipper (GILZ). GILZ is a protein that is rapidly induced by glucocorticoids treatment and involved in apoptosis modulation. Results indicate different glucocorticoids have different apoptotic activity which is related to their ability to induce both genomic, evaluated as caspases activation and GILZ expression, and non-genomic effects, evaluated as PI-PLC phosphorylation.

Since its introduction, the propidium iodide (PI) flow cytometric assay has been widely used for the evaluation of apoptosis in different experimental models. It is based on the principle that apoptotic cells, among other typical features, are characterized by DNA fragmentation and, consequently, loss of nuclear DNA content. Use of a fluorochrome, such as PI, that is capable of binding and labeling DNA makes it possible to obtain a rapid (the protocol can be completed in about 2 h) and precise evaluation of cellular DNA content by flow cytometric analysis, and subsequent identification of hypodiploid cells. The original protocol enhanced the capacity for a rapid, quantitative measure of cell apoptosis. For this reason, since its publication, the PI assay has been widely used, as demonstrated by the large number of citations of the original paper and/or the continuous use of the method in many laboratories.

Glucocorticoid-induced tumor necrosis factor receptor (GITR) is a member of the tumor necrosis factor receptor superfamily, is expressed in T lymphocytes, and exerts an anti-apoptotic function in these cells. We reported that GITR is also highly expressed in the skin, specifically in keratinocytes, and that it is under negative transcriptional control of p21(Cip1/WAF1), independently from the cell cycle. Although GITR expression is higher in p21-deficient keratinocytes and skin, it is down-modulated with differentiation and in response to UVB. The combined analysis of keratinocytes with increased GITR expression versus normal keratinocytes and skin of mice with a disruption of the GITR gene indicates that this protein protects keratinocytes from UVB-induced apoptosis both in vitro and in vivo.

Two series of derivatives were synthesised. In one series (R)-4-hydroxy-2-pyrrolidinone was used as a mimic of cyclic L-carnitine analogue and in the second series 3-amino-2-piperidinone was used as a cyclic ornithine analogue. N-Benzyloxycarbonyl derivatives of some amino acids were also prepared. The newly synthesised compounds were tested for their ability to inhibit Fas-activated apoptosis of human Jurkatt T-cell line. The results confirm the previously described anti-apoptotic activity of carnitine and indicate new carnitine and amino acid analogues (1, 3, 6, 7, 20) that inhibit Fas-induced apoptosis.

We have previously shown 1,4-benzothiazine (1,4-B) derivatives induce thymocyte apoptosis in vitro and thymus cell loss in vivo. Apoptosis is mediated through a complex of biochemical events including phosphatidylcholine specific-phospholipase C (PC-PLC) activation, acidic sphingomyelinase (aSMase) activation and ceramide generation, caspase-8 and caspase-3 activation. As preliminary analysis of the structure-activity relationship (SAR) suggested some structural features were responsible for apoptosis, we synthesised several derivatives and tested for apoptosis activity at equimolar concentrations. In particular, we synthesised analogues that differed in the nature of skeleton (1,4-benzothiazine, 1,4-benzoxazine and 1,2,3,4-tetrahydroquinoline) and in the nature of side chain (imidazole, benzimidazole or piperazine as azole substituent; presence, absence or transformation of alcoholic group). Results of apoptosis induction indicate that transforming the 1,4-benzothiazine skeleton into 1,2,3,4-tetrahydroquinoline does not result in significant change. Transformation into 1,4-benzoxazine decreased activity. Replacing imidazole at the side chain with different piperazines also decreased activity while replacing it with benzimidazole does not change apoptotic activity. Finally, removal of the alcoholic group by dehydration to olefin, or by transforming it into ether, increased activity. Moreover, in an attempt to analyse further the SAR characteristics that are responsible for 1,4-B-activated apoptosis we tested the effect on caspase-8,-9 and-3 activation. 1,4-B analogues activate caspases and the structural requirements correlate with those responsible for apoptosis induction.

Dexamethasone (DEX), a glucocorticoid hormone (GCH) with specificity for the glucocorticoid receptor (GR) induces T lymphocyte and thymocyte apoptosis. DEX-activated thymocyte apoptosis requires a sequence of biochemical events including mRNA and protein synthesis. In particular, GCH treatment induces non-genomic mechanisms, such as for example Ca(2+) mobilization and PI-PLC activation, and genomic mechanisms. Most of these events, including protein synthesis, are required and precede caspase activation. As protein synthesis is required for caspases and apoptosis activation, DEX-induced GR nuclear translocation is necessary for apoptosis. Cell treatment with geldanamycin (GA) inhibits the GR nuclear translocation and consequently, caspases activation and apoptosis. Although DEX treatment induces loss of mitochondrial membrane potential (deltapsim) and cytochrome c release, deltapsim induction does not correlate with thymocyte apoptosis. In fact, while Cyclosporin-A and the caspase-9 inhibitor, Z-LEHD-FMK, inhibit DEX-induced deltapsim, do not influence apoptosis. These data indicate many biochemical events and are activated by DEX treatment of thymocytes and some, but not all, are required for apoptosis.

Glucocorticoid hormones (GCHs) regulate normal and neoplastic lymphocyte development by exerting antiproliferative and/or apoptotic effects. We have previously shown that dexamethasone (DEX)-activated thymocyte apoptosis requires a sequence of events including interaction with the glucocorticoid receptor (GR), phosphatidylinositol-specific phospholipase C (PI-PLC), and acidic sphingomyelinase (aSMase) activation. We analyzed the mechanisms of GCH-activated apoptosis by focusing on GR-associated Src kinase, cytochrome c release, and caspase-8, -9, and -3 activation. We show here that PI-PLC binds to GR-associated Src kinase, as indicated by coimmunoprecipitation experiments. Moreover, DEX treatment induces PI-PLC phosphorylation and activation. DEX-induced PI-PLC phosphorylation, activation, and apoptosis are inhibited by PP1, a Src kinase inhibitor, thus suggesting that Src-mediated PI-PLC activation is involved in DEX-induced apoptosis. Caspase-9, -8, and -3 activation and cytochrome c release can be detected 1 to 2 hours after DEX treatment. Caspase-9 inhibition does not counter cytochrome c release, caspase-8 and caspase-3 activation, and apoptosis. Caspase-8 inhibition counters cytochrome c release, caspase-9 and caspase-3 activation, and apoptosis, thus suggesting that caspase-8 inhibitor can directly inhibit caspase-9 and/or that DEX-induced caspase-8 activation is upstream to mitochondria and can regulate caspase-3 directly or through cytochrome c release and the consequent caspase-9/caspase-3 activation. DEX-induced caspase-8 activation, like ceramide-induced caspase-8 activation, correlates with the formation of Fas-associated death domain protein (FADD)/caspase-8 complex. Caspase-8 activation is countered by the inhibition of macromolecular synthesis and of Src kinase, PI-PLC, and aSMase activation, suggesting it is downstream in the DEX-activated apoptotic pathway of thymocytes.

Glucocorticoid hormones regulate essential body functions in mammals, control cell metabolism, growth, differentiation, and apoptosis. Importantly, they are potent suppressors of inflammation, and multiple immune-modulatory mechanisms involving leukocyte apoptosis, differentiation, and cytokine production have been described. Due to their potent anti-inflammatory and immune-suppressive activity, synthetic glucocorticoids (GCs) are the most prescribed drugs used for treatment of autoimmune and inflammatory diseases. It is long been noted that males and females exhibit differences in the prevalence in several autoimmune diseases (AD). This can be due to the role of sexual hormones in regulation of the immune responses, acting through their endogenous nuclear receptors to mediate gene expression and generate unique gender-specific cellular environments. Given the fact that GCs are the primary physiological anti-inflammatory hormones, and that sex hormones may also exert immune-modulatory functions, the link between GCs and sex hormones may exist. Understanding the nature of this possible crosstalk is important to unravel the reason of sexual disparity in AD and to carefully prescribe these drugs for the treatment of inflammatory diseases. In this review, we discuss similarities and differences between the effects of sex hormones and GCs on the immune system, to highlight possible axes of functional interaction. [ABSTRACT FROM AUTHOR]

Background: Glucocorticoids affect peripheral immune responses, including modulation of T-cell activation, differentiation, and apoptosis. The quantity and quality of T-cell receptor (TCR)-triggered intracellular signals modulate T-cell function. Thus, glucocorticoids may affect T cells by interfering with the TCR signaling cascade. The purpose of the study was to search for glucocorticoid-modulated kinases downstream of the TCR. Methods: Gene modulation in lymphoid cells either treated with glucocorticoids or from glucocorticoid-treated mice was studied using a RNase protection assay, real-time PCR, and western blotting. The sensitivity of genetically modified thymocytes to glucocorticoid-induced apoptosis was studied by performing hypotonic propidium iodide staining and flow cytometry. The Student’s t-test was employed for statistical evaluation. Results: We found that transcription of Itk, a non-receptor tyrosine kinase of the Tec family, was up-regulated in a mouse T-cell hybridoma by the synthetic glucocorticoid dexamethasone. In contrast, dexamethasone down-regulated the expression of Txk, a Tec kinase that functions redundantly with Itk, and Lck, the Src kinase immediately downstream of the TCR. We investigated the expression of Itk, Txk, and Lck in thymocytes and mature lymphocytes following in vitro and in vivo dexamethasone treatment at different time points and doses. Kinase expression was differentially modulated and followed distinct kinetics. Itk was up-regulated in all cell types and conditions tested. Txk was strongly up-regulated in mature lymphocytes but only weakly up-regulated or non-modulated in thymocytes in vitro or in vivo, respectively. Conversely, Lck was down-regulated in thymocytes, but not modulated or up-regulated in mature lymphocytes in the different experimental conditions. This complex behaviour correlates with the presence of both positive and negative glucocorticoid responsive elements (GRE and nGRE, respectively) in the Itk, Txk and Lck genes. To investigate the function associated with Itk up-regulation, dexamethasone-induced apoptosis of thymocytes from Itk-deficient mice was evaluated. Our results demonstrated that Itk deficiency causes increased sensitivity to dexamethasone but not to other pro-apoptotic stimuli. Conclusions: Modulation of Itk, Txk, and Lck in thymocytes and mature lymphocytes is another mechanism by which glucocorticoids modulate T-cell activation and differentiation. Itk up-regulation plays a protective role in dexamethasone-treated thymocytes. [ABSTRACT FROM AUTHOR]

Spinal cord injury (SCI) is a major cause of disability, its clinical outcome depending mostly on the extent of damage in which proapoptotic cytokines have a crucial function. In particular, the inducers of apoptosis belonging to TNF receptor superfamily and their respective ligands are upregulated after SCI. In this study, the function of the proapoptotic cytokine tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) in SCI-induced damage was investigated in the mouse. SCI resulted in severe trauma, characterized by prominent inflammation-related damage and apoptosis. Immunostaining for TRAIL and its receptor DR5 was found in the white and gray matter of the perilesional area, as also confirmed by western blotting experiments. Immunoneutralization of TRAIL resulted in improved functional recovery, reduced apoptotic cell number, modulation of molecules involved in the inflammatory response (FasL, TNF-α, IL-1β, and MPO), and the corresponding signaling (caspase-8 and -3 activation, JNK phosphorylation, Bax, and Bcl-2 expression). As glucocorticoid-induced TNF receptor superfamily-related protein (GITR) activated by its ligand (GITRL) contributes to SCI-related inflammation, interactions between TRAIL and GITRL were investigated. SCI was associated with upregulated GITR and GITRL expression, a phenomenon prevented by anti-TRAIL treatment. Moreover, the expression of both TRAIL and DR5 was reduced in tissues from mice lacking the GITR gene (GITR−/−) in comparison with wild-type mice suggesting that TRAIL- and GITRL-activated pathways synergise in the development of SCI-related inflammatory damage. Characterization of new targets within such molecular systems may constitute a platform for innovative treatment of SCI. [ABSTRACT FROM AUTHOR]

Glucocorticoids (GCs) represent the mainstay of current anti-inflammatory and immunosuppressive strategies, mediating effects that mostly result in transcriptional regulation of glucocorticoid receptor target genes. A variety of actions are tied together in the response to GC treatment. Dissecting the beneficial from the detrimental actions in GC therapy is a major challenge in basic research, raising the critical issue of whether a single target gene or gene family might eventually be linked to a specific GC function. Glucocorticoid-induced leucine zipper (GILZ) was originally discovered in studies aimed at characterizing genes targeted by dexamethasone. The first suggestion that GILZ plays an important role in GC immunomodulation came from observations of GILZ up-regulation by GCs, mainly in lymphoid organs, and inhibition of anti-CD3-induced activation and apoptosis. The identification of GILZ interaction with and inhibition of NF-κB provided a first molecular mechanistic basis for explaining GILZ effects on T cells. Subsequently, other GILZ targets have been identified, including AP-1, Raf-1, and Ras, all involved in GC effects. The finding that GILZ silencing abrogates the antiproliferative activity of dexamethasone and reduces GC inhibition of cytokine-induced COX-2 expression dearly gained GILZ a distinguished reputation within the critical mediators of GC effects. The multiple functions of Gill and their potential biological relevance are here reviewed. [ABSTRACT FROM AUTHOR]

Correct function of spermatogonia is critical for the maintenance of spermatogenesis throughout life, but the cellular pathways regulating undifferentiated spermatogonia proliferation, differentiation, and survival are only partially known. We show here that long glucocorticoid-induced leucine zipper (L-GILZ) is highly expressed in spermatogonia and primary spermatocytes and controls spermatogenesis. Gilz deficiency in knockout (gilz KO) mice leads to a complete loss of germ cell lineage within first cycles of spermatogenesis, resulting in male sterility. Spermatogenesis failure is intrinsic to germ cells and is associated with increased proliferation and aberrant differentiation of undifferentiated spermatogonia and with hyperactivity of Ras signaling pathway as indicated by an increase of ERK and Akt phosphorylation. Spermatogonia differentiation does not proceed beyond the prophase of the first meiotic division due to massive apoptosis associated with accumulation of unrepaired chromosomal damage. These results identify L-GILZ as a novel important factor for undifferentiated spermatogonia function and spermatogenesis. [ABSTRACT FROM AUTHOR]

Tsc22d3 coding for glucocorticoid-induced leucine zipper (GILZ) was initially identified as a dexamethasone-responsive gene involved in the control of T lymphocyte activation and apoptosis. However, the physiological role of this molecule and its function in the biological activity of glucocorticoids (GCs) has not been clarified. Here, we demonstrate that GILZ interacts directly with Ras in vitro and in vivo as shown by GILZ and Ras coimmunoprecipitation and colocalization upon PMA activation in primary mouse spleen T lymphocytes and thymus cells. The analysis of GILZ mutants showed that they bound Ras through the tuberous sclerosis complex box (TSC) and, depending on the Ras activation level, formed a trimeric complex with Ras and Raf, which we previously identified as a GILZ binder. As a consequence of these interactions, GILZ diminished the activation of Ras and Raf downstream targets including ERK1/2, AKT/PKB serine/threonine kinase, and retinoblastoma (Rb) phosphorylation and cyclin D1 expression, leading to inhibition of Ras- and Raf-dependent cell proliferation and Ras-induced NIH-3T3 transformation. GILZ silencing resulted in an increase in concanavalin A–induced T cell proliferation and, most notably, inhibition of dexamethasone antiproliferative effects. Together, these findings indicate that GILZ serves as a negative regulator of Ras- and Raf-induced proliferation and is an important mediator of the antiproliferative effect of GCs. [ABSTRACT FROM AUTHOR]

Glucocorticoid-induced tumor necrosis factor receptor (GITR) is a member of the tumor necrosis factor receptor superfamily, is expressed in T lymphocytes, and exerts an anti-apoptotic function in these cells. We reported that GITR is also highly expressed in the skin, specifically in keratinocytes, and that it is under negative transcriptional control of p21Cip1/WAF1, independently from the cell cycle. Although GITR expression is higher in p21-deficient keratinocytes and skin, it is down-modulated with differentiation and in response to UVB. The combined analysis of keratinocytes with increased GITR expression versus normal keratinocytes and skin of mice with a disruption of the GITR gene indicates that this protein protects keratinocytes from UVB-induced apoptosis both in vitro and in vivo. [ABSTRACT FROM AUTHOR]