Induction of Glucocorticoid-induced Leucine Zipper (GILZ) Contributes to Anti-inflammatory Effects of the Natural Product Curcumin in Macrophages

Abstract

GILZ (glucocorticoid-induced leucine zipper) is inducible by glucocorticoids and plays a key role in their mode of action. GILZ attenuates inflammation mainly by inhibition of NF-B and mitogen-activated protein kinase activation but does not seem to be involved in the severe side effects observed after glucocorticoid treatment. Therefore, GILZ might be a promising target for new therapeutic approaches. The present work focuses on the natural product curcumin, which has previously been reported to inhibit NF-B. GILZ was inducible by curcumin in macrophage cell lines, primary human monocyte-derived macrophages, and murine bone marrow-derived macrophages. The up-regulation of GILZ was neither associated with glucocorticoid receptor activation nor with transcriptional induction or mRNA or protein stabilization but was a result of enhanced translation. Because the GILZ 3-UTR contains AU-rich elements (AREs), we analyzed the role of the mRNA-binding protein HuR, which has been shown to promote the translation of ARE-containing mRNAs. Our results suggest that curcumin treatment induces HuR expression. An RNA immunoprecipitation assay confirmed that HuR can bind GILZ mRNA. In accordance, HuR overexpression led to increased GILZ protein levels but had no effect on GILZ mRNA expression. Our data employing siRNA in LPS-activated RAW264.7 macrophages show that curcumin facilitates its anti-inflammatory action by induction of GILZ in macrophages. Experiments with LPS-activated bone marrow-derived macrophages from wild-type and GILZ knock-out mice demonstrated that curcumin inhibits the activity of inflammatory regulators, such as NF-B or ERK, and subsequent TNF-␣ production via GILZ. In summary, our data indicate that HuR-dependent GILZ induction contributes to the anti-inflammatory properties of curcumin.The polyphenol curcumin (diferuloylmethane) is the principal bioactive compound of turmeric (Curcuma longa) preparations, which are commonly used as traditional remedies or spices. Recently, extensive research has shown that curcumin has a broad range of therapeutic effects, including anti-inflammatory, anti-oxidant, anti-proliferative, hypoglycemic, lipidlowering, anti-thrombotic, and anti-coagulant activity. At the same time, no dose-limiting toxicity was observed in clinical trials, indicating pharmacological safety. Thus, curcumin administration has been suggested as a potential therapeutic approach for the treatment of several pathologic conditions, especially inflammatory diseases such as cardiovascular pathologies, arthritis, asthma, ulcerative colitis, inflammatory bowel disease, and type II diabetes (1-5). Several in vivo studies suggested a profound effect of curcumin on cells of the mononuclear phagocyte system. In a mouse model of abdominal aortic aneurysms, oral administration of curcumin efficiently suppressed mononuclear inflammation in the aortic walls, i.e. proinflammatory cytokine expression and adverse connective tissue remodeling (6). In addition, curcumin decreased the number of proinflammatory M1 macro...

The glucocorticoid‐induced leucine zipper mediates statin‐induced muscle damage

“…A CFX96 touch Real‐Time PCR detection system, and the CFX Manager 2.1 software (Bio‐Rad, Hercules, CA, USA) were used for qPCR analysis. Data were analyzed as previously described . Housekeeping genes were chosen based on the literature, or after evaluating the expression stability of at least three candidate genes under the experimental conditions, using the geNorm, NormFinder, and BestKeeper software tools …”

“…Reporter gene assays were performed as previously described. 25 The proximal Gilz promoter (a fragment located between −1938 bp upstream and +206 bp downstream of the transcription start site) was cloned into the pGL3 luciferase reporter vector (Promega, Madison, WI, USA) using KpnI and SmaI according to the manufacturer's instructions. The FOXO3 reporter vector (FHRE-Luc) was a gift from Michael Greenberg (Addgene plasmid #1789).…”

Altered glucocorticoid metabolism represents a feature of macroph‐aging

“…Quantitative RT–PCR (qPCR) was performed as described previously (Dembek et al., 2017; Hoppstädter, Dembek, et al., 2019; Hoppstädter, Diesel, et al., 2019; Hoppstädter et al., 2015, 2016; Kessler et al., 2017). Total RNA from cells was isolated using the High Pure RNA Isolation Kit (#11828665001; Roche), following the manufacturer's instructions.…”

“…SDS–polyacrylamide gel electrophoresis, blotting, and staining were performed as described previously. Signals were detected using an Odyssey ® Near‐Infrared Imaging System and software (LI‐COR Biosciences) (Dembek et al., 2017; Hoppstädter, Dembek, et al., 2019; Hoppstädter, Diesel, et al., 2019; Hoppstädter et al., 2016).…”

“…Subsequently, samples were washed in saponin buffer and resuspended in eBioscience™ IC Fixation Buffer. The specificity of the anti‐GILZ antibody was validated by negative staining of GILZ knockout cells (Hoppstädter et al., 2016).…”