Induction of the glucocorticoid-induced leucine zipper (GILZ) by glucocorticoids plays a role in their antiinflammatory action, whereas GILZ expression is reduced under inflammatory conditions. The mechanisms regulating GILZ expression during inflammation, however, have not yet been characterized. Here, we investigated GILZ expression in human alveolar macrophages (AMs) following Toll-like receptor (TLR) activation. Macrophages were shown to predominantly express GILZ transcript variant 2. Lipopolysaccharide-treated AMs, THP-1 cells, and lungs of lipopolysaccharide-exposed mice displayed decreased GILZ protein and mRNA levels. The effect was strictly dependent on the adapter molecule MyD88, as shown by using specific ligands or a knockdown strategy. Investigations on the functional significance of GILZ downregulation performed by GILZ knockdown revealed a proinflammatory response, as indicated by increased cytokine expression and NF-κB activity. We found that TLR activation reduced GILZ mRNA stability, which was mediated via the GILZ 3 -untranslated region. Finally, involvement of the mRNA-binding protein tristetraprolin (TTP) is suggested, since TTP overexpression or knockdown modulated GILZ expression and TTP was induced in a MyD88-dependent fashion. Taken together, our data show a MyD88-and TTP-dependent GILZ downregulation in human macrophages upon TLR activation. Suppression of GILZ is mediated by mRNA destabilization, which might represent a regulatory mechanism in macrophage activation.Keywords: GILZ transcript variants · Inflammation · Innate immunity · mRNA stability · Pulmonary macrophages IntroductionInflammatory processes in the lung play an important role in different diseases, such as viral and bacterial infections. Moreover, noninfectious diseases such as chronic obstructive pulmonary disCorrespondence: Prof. Alexandra K. Kiemer e-mail: pharm.bio.kiemer@mx.uni-saarland.de ease (COPD) are characterized by chronic inflammatory processes. Asthma and allergic diseases are other examples of chronic inflammatory lung diseases with major clinical relevance [1,2].Toll-like receptors (TLRs) represent an important family of innate immune receptors, which act as a first line of defense of host immunity against various pathogens. Presently, ten human TLRs are known, which recognize pathogen-associated molecular patterns including bacterial cell wall components, such as lipoproteins (TLR1/2 or TLR1/6 dimers) or lipopolysaccharide (LPS, TLR4), C 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.eji-journal.eu Eur. J. Immunol. 2012. 42: 1282-1293 Innate immunity 1283 bacterial flagellin (TLR5), viral RNA (TLR3, 7 and 8), as well as bacterial DNA (TLR9) [3]. Except for TLR3, all TLRs signal via the adapter molecule MyD88. The MyD88-independent pathway used by TLR3 can also be utilized by TLR4. The different signaling pathways employ different protein kinase complexes and show distinct cytokine profiles, but all activate NF-κB [4]. In addition to their role in pathogen defense, TLRs also play a dominant role in ...

The tumor suppressor programmed cell death 4 (Pdcd4) is lost in various tumor tissues. Loss of Pdcd4 has been associated with increased tumorigenic potential and tumor progression. While various mechanisms of Pdcd4 regulation have been described, the effect of an inflammatory tumor microenvironment on Pdcd4 protein expression has not been characterized so far. In the present study, we aimed to elucidate the molecular mechanisms of Pdcd4 protein regulation in tumor cells under inflammatory conditions. 12-O-tetradecanoylphorbol 13-acetate-induced differentiation of human U937 monocytes increased the expression and secretion of inflammatory cytokines such as tumor necrosis factor α, interleukin (IL)-6 and IL-8. Exposure to conditioned medium (CM) of these activated macrophages markedly decreased Pdcd4 protein expression in various tumor cells. Similarly, indirect coculture with such activated U937 monocyte-derived macrophages resulted in the loss of Pdcd4 protein in tumor cells. Decreased Pdcd4 protein levels were attributable to enhanced proteasomal degradation, diminishing Pdcd4 protein half-life. Proteasomal degradation required activation of phosphatidylinositol-3-kinase (PI3K)-mammalian target of rapamycin (mTOR) signaling. Since macrophage-CM sufficed to induce Pdcd4 degradation, Pdcd4 downregulation was determined to be an indirect unidirectional effect of the macrophages on the tumor cells. Pdcd4 protein expression was also attenuated in vivo in mouse colon tissue in response to dextran sodium sulfate-induced colitis. In summary, we characterized PI3K-mTOR-dependent proteasome-mediated Pdcd4 degradation in tumor cells in the inflammatory tumor microenvironment. Consequently, stabilization of Pdcd4 protein could provide a promising novel avenue for therapeutics targeting inflammation-associated tumors.

Type 1 diabetes mellitus (T1DM) is associated with impaired spermatogenesis, lower testosterone levels and epididymal weight. However, the underlying processes in the testis are unknown and remain to be elucidated. Therefore, the present study focused on the effects of T1DM on testicular function in a spontaneously diabetic rat model.BB/OKL rats after diabetes manifestation were divided into three groups: without insulin treatment, insulin treatment for a duration of 2 and of 6 weeks. Anthropometrical data, circulating levels of gonadotrophins, testosterone and inhibin B were measured. Intratesticular testosterone, oxidative stress, inflammation, and apoptosis were analyzed. Key enzymes of steroidogenesis were evaluated in the testis.Untreated diabetic rats had significantly lower serum FSH and LH levels. Serum and intratesticular testosterone levels significantly decreased in the untreated diabetic rat compared to healthy controls. Key markers of Leydig cell function were significantly downregulated on RNA level: Insl3 by 53% (p=0.006), Star by 51% (p=0.004), Cyp11A1 by 80% (p=0.003), 3Beta-Hsd2 by 61% (p=0.005) and Pbr by 52% (p=0.002). In the insulin-treated group only Cyp11A1 and 3Beta-Hsd2 transcripts were significantly lower. Interestingly the long-term insulin treated group showed significant upregulation of most steroidogenic enzymes without affecting testosterone levels. TNF-alpha and apoptosis were significantly increased in the long-term insulin treated rats. In conclusion T1DM, with a severe lack of insulin, has an adverse action on Leydig cell function. This is partially reversible with well compensated blood glucose control. Long-term T1DM adversely affects Leydig cell function due to the process of inflammation and apoptosis.

Female LD animals show impaired fertility which is restored by leptin. Future studies should assess leptin as a subfertility treatment in human leptin-deficiency disorders.

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