Autophagy sustains mitochondrial respiration and determines resistance to BRAFV600E inhibition in thyroid carcinoma cells.

BRAFV600E is the most prevalent mutation in thyroid cancer and correlates with poor prognosis and therapy resistance. Although selective inhibitors of BRAFV600E have been developed, more advanced tumors such as anaplastic thyroid carcinomas show a poor response in clinical trials. Therefore, the study of alternative survival mechanisms is needed. Since metabolic changes have been related to malignant progression, in this work we explore metabolic dependencies of thyroid tumor cells to exploit them therapeutically. Our results show that respiration of thyroid carcinoma cells is highly dependent on fatty acid oxidation and, in turn, fatty acid mitochondrial availability is regulated through macroautophagy/autophagy. Furthermore, we show that both lysosomal inhibition and the knockout of the essential autophagy gene, ATG7, lead to enhanced lipolysis; although this effect is not essential for survival of thyroid carcinoma cells. We also demonstrate that following inhibition of either autophagy or fatty acid oxidation, thyroid tumor cells compensate oxidative phosphorylation deficiency with an increase in glycolysis. In contrast to lipolysis induction, upon autophagy inhibition, glycolytic boost in autophagy-deficient cells is essential for survival and, importantly, correlates with a higher sensitivity to the BRAFV600E selective inhibitor, vemurafenib. In agreement, downregulation of the glycolytic pathway results in enhanced mitochondrial respiration and vemurafenib resistance. Our work provides new insights into the role of autophagy in thyroid cancer metabolism and supports mitochondrial targeting in combination with vemurafenib to eliminate BRAFV600E-positive thyroid carcinoma cells.Abbreviations: AMP: adenosine monophosphate; ATC: anaplastic thyroid carcinoma; ATG: autophagy related; ATP: adenosine triphosphate; BRAF: B-Raf proto-oncogene, serine/threonine kinase; Cas9: CRISPR-associated protein; CREB: cAMP responsive element binding protein; CRISPR: clustered regularly interspaced short palindromic repeats; 2DG: 2-deoxyglucose; FA: fatty acid; FAO: fatty acid oxidation; FASN: fatty acid synthase; FCCP: trifluoromethoxy carbonyl cyanide phenylhydrazone; LAMP1: lysosomal associated membrane protein 1; LIPE/HSL: lipase E, hormone sensitive type; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; OCR: oxygen consumption rate; OXPHOS: oxidative phosphorylation; PRKA/PKA: protein kinase cAMP-activated; PTC: papillary thyroid carcinoma; SREBF1/SREBP1: sterol regulatory element binding transcription factor 1.

Thyroid hormones act as modulators of inflammation through their nuclear receptors.

Reciprocal crosstalk between endocrine and immune systems has been well-documented both in physiological and pathological conditions, although the connection between the immune system and thyroid hormones (THs) remains largely unclear. Inflammation and infection are two important processes modulated by the immune system, which have profound effects on both central and peripheral THs metabolism. Conversely, optimal levels of THs are necessary for the maintenance of immune function and response. Although some effects of THs are mediated by their binding to cell membrane integrin receptors, triggering a non-genomic response, most of the actions of these hormones involve their binding to specific nuclear thyroid receptors (TRs), which generate a genomic response by modulating the activity of a great variety of transcription factors. In this special review on THs role in health and disease, we highlight the relevance of these hormones in the molecular mechanisms linked to inflammation upon their binding to specific nuclear receptors. In particular, we focus on THs effects on different signaling pathways involved in the inflammation associated with various infectious and/or pathological processes, emphasizing those mediated by NF-kB, p38MAPK and JAK/STAT. The findings showed in this review suggest new opportunities to improve current therapeutic strategies for the treatment of inflammation associated with several infections and/or diseases, such as cancer, sepsis or Covid-19 infection.

V600EBRAF Inhibition Induces Cytoprotective Autophagy through AMPK in Thyroid Cancer Cells.

The dysregulation of autophagy is important in the development of many cancers, including thyroid cancer, where V600EBRAF is a main oncogene. Here, we analyse the effect of V600EBRAF inhibition on autophagy, the mechanisms involved in this regulation and the role of autophagy in cell survival of thyroid cancer cells. We reveal that the inhibition of V600EBRAF activity with its specific inhibitor PLX4720 or the depletion of its expression by siRNA induces autophagy in thyroid tumour cells. We show that V600EBRAF downregulation increases LKB1-AMPK signalling and decreases mTOR activity through a MEK/ERK-dependent mechanism. Moreover, we demonstrate that PLX4720 activates ULK1 and increases autophagy through the activation of the AMPK-ULK1 pathway, but not by the inhibition of mTOR. In addition, we find that autophagy blockade decreases cell viability and sensitize thyroid cancer cells to V600EBRAF inhibition by PLX4720 treatment. Finally, we generate a thyroid xenograft model to demonstrate that autophagy inhibition synergistically enhances the anti-proliferative and pro-apoptotic effects of V600EBRAF inhibition in vivo. Collectively, we uncover a new role of AMPK in mediating the induction of cytoprotective autophagy by V600EBRAF inhibition. In addition, these data establish a rationale for designing an integrated therapy targeting V600EBRAF and the LKB1-AMPK-ULK1-autophagy axis for the treatment of V600EBRAF-positive thyroid tumours.

Downregulation of Snail by DUSP1 Impairs Cell Migration and Invasion through the Inactivation of JNK and ERK and Is Useful as a Predictive Factor in the Prognosis of Prostate Cancer.

Dual specificity phosphatase 1 (DUSP1) is crucial in prostate cancer (PC), since its expression is downregulated in advanced carcinomas. Here, we investigated DUSP1 effects on the expression of mesenchymal marker Snail, cell migration and invasion, analyzing the underlying mechanisms mediated by mitogen-activated protein kinases (MAPKs) inhibition. To this purpose, we used different PC cells overexpressing or lacking DUSP1 or incubated with MAPKs inhibitors. Moreover, we addressed the correlation of DUSP1 expression with Snail and activated MAPKs levels in samples from patients diagnosed with benign hyperplasia or prostate carcinoma, studying its implication in tumor prognosis and survival. We found that DUSP1 downregulates Snail expression and impairs migration and invasion in PC cells. Similar results were obtained following the inhibition of c-Jun N-terminal kinase (JNK) and extracellular-signal-regulated kinase (ERK). In clinical samples, we evidenced an inverse correlation between DUSP1 expression and Snail levels, which are further associated with JNK and ERK activation. Consequently, the pattern DUSP1high/activated JNKlow/activated ERKlow/Snaillow is associated with an overall extended survival of PC patients. In summary, the ratio between DUSP1 and Snail expression, with additional JNK and ERK activity measurement, may serve as a potential biomarker to predict the clinical outcome of PC patients. Furthermore, DUSP1 induction or inhibition of JNK and ERK pathways could be useful to treat PC.

Resveratrol promotes apoptosis through the induction of dual specificity phosphatase 1 and sensitizes prostate cancer cells to cisplatin.

Resveratrol is a polyphenol with chemopreventive properties against prostate cancer; however, the mechanisms underlying its actions are not completely understood. Previously, we demonstrated that DUSP1 induces apoptosis in prostate cancer cells; therefore in the present study we investigated the role of this phosphatase on resveratrol effects. Moreover, we analysed the efficiency of combined treatment of resveratrol and the chemotherapeutic drug cisplatin on cellular viability and apoptosis and its relation with DUSP1 in prostate cancer cells. We found that resveratrol up-regulates DUSP1 expression in androgen-independent prostate cancer cells, which in turn, is involved in the inhibition of the NF-κB pathway and Cox-2 expression. This phosphatase is required for the induction of apoptosis achieved by resveratrol, but does not regulate the effects of this compound on cell cycle. Furthermore, we show that resveratrol cooperates with cisplatin both in the up-regulation of DUSP1 levels and in the promotion of apoptosis, suggesting that DUSP1 is a major determinant of cisplatin sensitivity to apoptosis. These results reveal a novel molecular mechanism by which resveratrol induces apoptosis in prostate cancer cells, and highlight the importance of DUSP1 in future therapeutic approaches based in the use of this polyphenol and cisplatin.

Author Correction: Excitotoxic inactivation of constitutive oxidative stress detoxification pathway in neurons can be rescued by PKD1.

The original version of this Article contained an error in the spelling of the author Álvaro Sebastián-Serrano, which was incorrectly given as Álvaro Sebastián Serrano. This has now been corrected in both the PDF and HTML versions of the Article.

Excitotoxic inactivation of constitutive oxidative stress detoxification pathway in neurons can be rescued by PKD1.

Excitotoxicity, a critical process in neurodegeneration, induces oxidative stress and neuronal death through mechanisms largely unknown. Since oxidative stress activates protein kinase D1 (PKD1) in tumor cells, we investigated the effect of excitotoxicity on neuronal PKD1 activity. Unexpectedly, we find that excitotoxicity provokes an early inactivation of PKD1 through a dephosphorylation-dependent mechanism mediated by protein phosphatase-1 (PP1) and dual specificity phosphatase-1 (DUSP1). This step turns off the IKK/NF-κB/SOD2 antioxidant pathway. Neuronal PKD1 inactivation by pharmacological inhibition or lentiviral silencing in vitro, or by genetic inactivation in neurons in vivo, strongly enhances excitotoxic neuronal death. In contrast, expression of an active dephosphorylation-resistant PKD1 mutant potentiates the IKK/NF-κB/SOD2 oxidative stress detoxification pathway and confers neuroprotection from in vitro and in vivo excitotoxicity. Our results indicate that PKD1 inactivation underlies excitotoxicity-induced neuronal death and suggest that PKD1 inactivation may be critical for the accumulation of oxidation-induced neuronal damage during aging and in neurodegenerative disorders.

VHL promotes immune response against renal cell carcinoma via NF-κB-dependent regulation of VCAM-1.

Vascular cell adhesion molecule 1 (VCAM-1) is an adhesion molecule assigned to the activated endothelium mediating immune cells adhesion and extravasation. However, its expression in renal carcinomas inversely correlates with tumor malignancy. Our experiments in clear cell renal cell carcinoma (ccRCC) cell lines demonstrated that von Hippel Lindau (VHL) loss, hypoxia, or PHD (for prolyl hydroxylase domain-containing proteins) inactivation decreased VCAM-1 levels through a transcriptional mechanism that was independent of the hypoxia-inducible factor and dependent on the nuclear factor κB signaling pathway. Conversely, VHL expression leads to high VCAM-1 levels in ccRCC, which in turn leads to better outcomes, possibly by favoring antitumor immunity through VCAM-1 interaction with the α4ß1 integrin expressed in immune cells. Remarkably, in ccRCC human samples with VHL nonmissense mutations, we observed a negative correlation between VCAM-1 levels and ccRCC stage, microvascular invasion, and symptom presentation, pointing out the clinical value of VCAM-1 levels as a marker of ccRCC progression.

TGFß induces epithelial-mesenchymal transition of thyroid cancer cells by both the BRAF/MEK/ERK and Src/FAK pathways.

The epithelial-mesenchymal transition (EMT) is a crucial process in tumour progression, by which epithelial cells acquire a mesenchymal phenotype, increasing its motility and the ability to invade distant sites. Here, we describe the molecular mechanisms by which V600E BRAF, TGFß and the Src/FAK complex cooperatively regulate EMT induction and cell motility of anaplastic thyroid cancer cells. Analysis of EMT marker levels reveals a positive correlation between TGFß and Snail expression, with a concomitant downregulation of E-cadherin, accompanied by an increase of cell migration and invasion. Furthermore, we show that V600E BRAF depletion by siRNA or inhibition of its activity by treatment with its inhibitor PLX4720 reverses the TGFß-mediated effects on Snail, E-cadherin, migration and invasion. Moreover, V600E BRAF induces TGFß secretion through a MEK/ERK-dependent mechanism. In addition, TGFß activates the Src/FAK complex, which in turn regulates the expression of Snail and E-cadherin as well as cell migration. The inhibition of Src with the inhibitor SU6656 or abrogation of FAK expression with a specific siRNA reverses the TGFß-induced effects. Interestingly, we demonstrate that activation of the Src/FAK complex by TGFß is independent of V600E BRAF signalling, since inhibition of this oncogene does not affect its phosphorylation. Our data strongly suggest that TGFß induces EMT and aggressiveness of thyroid cancer cells by parallel mechanisms involving both the V600E BRAF/MEK/ERK and Src/FAK pathways independently. Thus, we describe novel functions for Src/FAK in mediating the EMT program and aggressiveness regulated by TGFß, establishing the inhibition of these proteins as a possible effective approach in preventing tumour progression of V600E BRAF-expressing thyroid tumours. © 2015 Wiley Periodicals, Inc.

Hepatitis C virus-mediated Aurora B kinase inhibition modulates inflammatory pathway and viral infectivity.

BACKGROUND & AIMS: Chronic hepatitis C is a leading cause of chronic liver disease, cirrhosis and hepatocellular carcinoma. DNA methylation and histone covalent modifications constitute crucial mechanisms of genomic instability in human disease, including liver fibrosis and hepatocellular carcinoma. The present work studies the consequences of HCV-induced histone modifications in early stages of infection. METHODS: Human primary hepatocytes and HuH7.5 cells were transiently transfected with the core protein of hepatitis C virus (HCV) genotypes 1a, 1b, and 2a. Infectious genotype 2a HCV in culture was also used. RESULTS: We show that HCV and core protein inhibit the phosphorylation of Serine 10 in histone 3. The inhibition is due to the direct interaction between HCV core and Aurora B kinase (AURKB) that results in a decrease of AURKB activity. HCV and core significantly downregulate NF-κB and COX-2 transcription, two proteins with anti-apoptotic and proliferative effects implicated in the control of the inflammatory response. AURKB depletion reduced HCV and core repression of NF-κB and COX-2 gene transcription and AURKB overexpression reversed the viral effect. AURKB abrogation increased HCV specific infectivity which was decreased when AURKB was overexpressed. CONCLUSIONS: The core-mediated decrease of AURKB activity may play a role in the inflammatory pathway during the initial steps of viral infection, while ensuring HCV infectivity.

Dual specificity phosphatase 1 expression inversely correlates with NF-κB activity and expression in prostate cancer and promotes apoptosis through a p38 MAPK dependent mechanism.

Dual specificity phosphatase 1 (DUSP1) and the transcription factor NF-κB are implicated in prostate cancer since their expression levels are altered along this disease, although there are no evidences up to date demonstrating a crosstalk between them. In this report, we show for the first time that DUSP1 over-expression in DU145 cells promotes apoptosis and decreases NF-κB activity by blocking p65/NF-κB nuclear translocation. Moreover, although DUSP1 impairs TNF-α-induced p38 MAPK and JNK activation, only the specific inhibition of p38 MAPK exerts the same effects than DUSP1 over-expression on both apoptosis and NF-κB activity. Consistently, DUSP1 promotes apoptosis and decreases NF-κB activity in cells in which p38 MAPK is induced by TNF-α treatment. These results demonstrate that p38 MAPK is specifically involved in DUSP1-mediated effects on both apoptosis and NF-κB activity. Interestingly, we show an inverse correlation between DUSP1 expression and activation of both p65/NF-κB and p38 MAPK in human prostate tissue specimens. Thus, most of apparently normal glands, benign prostatic hyperplasia and low-grade prostatic intraepithelial neoplasia samples show high DUSP1 expression and low levels of both nuclear p65/NF-κB and activated p38 MAPK. By contrast, DUSP1 expression levels are low or even absent in high-grade prostatic intraepithelial neoplasia and prostatic adenocarcinoma samples, whereas nuclear p65/NF-κB and activated p38 MAPK are highly expressed in the same samples. Overall, our results provide evidence for a role of DUSP1 in the apoptosis of prostate cancer cells, through a mechanism involving the inhibition of p38 MAPK and NF-κB. Furthermore, our findings suggest that the ratio between DUSP1 and p65/NF-κB expression levels, rather than the individual expression of both molecules, is a better marker for diagnostic purposes in prostate cancer.

The use of an active learning approach to teach metabolism to students of nutrition and dietetics.

This article describes the transition from a traditional instructor-centered course, based on lectures, to a student-centered course based on active learning methodologies as part of the reform of the Spanish higher education system within the European Higher Education Area (EHEA). Specifically, we describe the use of active learning methodologies to teach metabolism to students of nutrition and dietetics during the first year of their professional training in a 4-year undergraduate degree (Bachelor of Human Nutrition and Dietetics). In the new course design, the number of didactic lectures was largely reduced and complemented with a series of activities (problems/case studies, discussion workshops, self-assessment quizzes) aimed to get students actively engaged, to encourage self-learning, and to promote sustained work throughout the length of the course. The article presents quantitative data demonstrating a clear and significant improvement in students' performance when an active approach was implemented. Importantly, the improved performance was achieved without work overload. Finally, students' responses to this new teaching methodology have been very positive and overall satisfaction high. In summary, our results strongly argue in favor of the teaching model described herein.

Balance between apoptosis or survival induced by changes in extracellular-matrix composition in human mesangial cells: a key role for ILK-NFκB pathway.

Renal fibrosis is the final outcome of many clinical conditions that lead to chronic renal failure, characterized by a progressive substitution of cellular elements by extracellular-matrix proteins, in particular collagen type I. The aim of this study was to identify the mechanisms responsible for human mesangial cell survival, conditioned by changes in extracellular-matrix composition. Our results indicate that collagen I induces apoptosis in cells but only after inactivation of the pro-survival factor NFκB by either the super-repressor IκBα or the PDTC inhibitor. Collagen I activates a death pathway, through ILK/GSK-3ß-dependent Bim expression. Moreover, collagen I significantly increases NFκB-dependent transcription, IκBα degradation and p65/NFκB translocation to the nucleus; it activates ß1 integrin and this is accompanied by increased activity of ILK which leads to AKT activation. Knockdown of ILK or AKT with small interfering RNA suppresses the increase in NFκB activity. NFκB mediates cell survival through the antiapoptotic protein Bcl-xL. Our data suggest that human mesangial cells exposed to abnormal collagen I are protected against apoptosis by a complex mechanism involving integrin ß1/ILK/AKT-dependent NFκB activation with consequent Bcl-xL overexpression, that opposes a simultaneously activated ILK/GSK-3ß-dependent Bim expression and this dual mechanism may play a role in the progression of glomerular dysfunction.

Thyroid hormone antagonizes tumor necrosis factor-alpha signaling in pituitary cells through the induction of dual specificity phosphatase 1.

Pituitary function has been shown to be regulated by an increasing number of factors, including cytokines and hormones, such as TNFalpha and T(3). Both the proinflammatory cytokine TNFalpha and T(3) have been suggested to be involved in the maintenance of tissue homeostasis in the anterior pituitary gland. In this report we show that T(3) negatively interferes with MAPK p38 and nuclear factor-kappaB (NF-kappaB) activation by TNFalpha in GH4C1 cells. Our data demonstrate that MAPK p38 is specifically activated upon exposure to TNFalpha and that T(3) abolishes this activation in a time-dependent manner by a mechanism that involves the induction of the MAPK phosphatase, DUSP1. Our data show that the pool of up-regulated DUSP1 by T(3) is mainly localized to the cytosol, and that TNFalpha does not affect this localization. On the other hand, we show that T(3) impairs the activation of the NF-kappaB pathway induced by TNFalpha, producing a significant decrease in NF-kappaB-dependent transcription, phosphorylation of IkappaBalpha, translocation of p65/NF-kappaB to the nucleus, and p65/NF-kappaB transactivation potential. Interestingly, the overexpression of DUSP1 inhibits the NF-kappaB activation achieved by either TNFalpha or ectopic expression of the upstream inducer of MAPK p38. Conversely, DUSP1 depletion abrogates the inhibitory effect of T(3) on the induction of NF-kappaB-dependent transcription by TNFalpha. Overall, our results indicate that T(3) antagonizes TNFalpha signaling in rat pituitary tumor cells through the induction of DUSP1.

Thyroid hormone-mediated activation of the ERK/dual specificity phosphatase 1 pathway augments the apoptosis of GH4C1 cells by down-regulating nuclear factor-kappaB activity.

Thyroid hormone (T3) plays a crucial role in processes such as cell proliferation and differentiation, whereas its implication on cellular apoptosis has not been well documented. Here we examined the effect of T3 on the apoptosis of GH4C1 pituitary cells and the mechanisms underlying this effect. We show that T3 produced a significant increase in apoptosis in serum-depleted conditions. This effect was accompanied by a decrease in nuclear factor-kappaB (NF-kappaB)-dependent transcription, IkappaBalpha phosphorylation, translocation of p65/NF-kappaB to the nucleus, phosphorylation, and transactivation. Moreover, these effects were correlated with a T3-induced decrease in the expression of antiapoptotic gene products, such as members of the inhibitor of apoptosis protein and Bcl-2 families. On the other hand, ERK but not c-Jun N-terminal kinase or MAPK p38, was activated upon exposure to T3, and inhibition of ERK alone abrogated T3-mediated apoptosis. In addition, T3 increased the expression of the MAPK phosphatase, dual specificity phosphatase 1 (DUSP1), in an ERK-dependent manner. Interestingly, the suppression of DUSP1 expression abrogated T3-induced inhibition of NF-kappaB-dependent transcription and p65/NF-kappaB translocation to the nucleus, as well as T3-mediated apoptosis. Overall, our results indicate that T3 induces apoptosis in rat pituitary tumor cells by down-regulating NF-kappaB activity through a mechanism dependent on the ERK/DUSP1 pathway.

Low cell cholesterol levels increase NFkappaB activity through a p38 MAPK-dependent mechanism.

Cholesterol, p38 MAPK and NFkappaB have been shown to participate in inflammation and cellular differentiation. Here, we examined the effect of cholesterol on NFkappaB-dependent transcription and the mechanisms underlying this effect in NIH3T3 cells. We show that chronic cholesterol depletion achieved with lipoprotein-deficient serum (LPDS) and 25-hydroxycholesterol (25-HC) treatment resulted in a significant increase in NFkappaB-dependent transcription, NFkappaB-DNA binding, IkappaBalpha degradation and p65/NFkappaB translocation to the nucleus, and the addition of exogenous cholesterol reversed these effects. Previously, we have shown that low cell cholesterol levels activate p38 MAPK. Here, we found that inhibition of p38 MAPK with the specific inhibitor SB203580 blocked the increase in NFkappaB activity, IkappaBalpha degradation and p65/NFkappaB translocation to the nucleus induced by cholesterol depletion. Moreover, the inhibition of the p38 MAPK downstream effector MSK1 with the specific inhibitor H89, or the overexpression of a kinase defective MSK1 abrogated the NFkappaB-dependent transcription induced by cholesterol depletion. On the other hand, the transactivation potential of p65/NFkappaB depends on phosphorylation of S276 by MSK1. We observed that cholesterol depletion increased the p65/NFkappaB transactivation capacity. This effect was reversed by cell cholesterol repletion or incubation with the SB203580 inhibitor. Moreover, the expression of a p65/NFkappaB S276A mutant was insensitive to cholesterol depletion. Together, our results demonstrate that cholesterol depletion induces NFkappaB transcriptional activity, not only by affecting the IkappaBalpha degradation and the translocation of p65/NFkappaB to the nucleus, but also regulating the p65/NFkappaB transactivating potential through a p38 MAPK/MSK1 mediated pathway.

RhoA and p38 MAPK mediate apoptosis induced by cellular cholesterol depletion.

Cholesterol is essential for cell viability, and homeostasis of cellular cholesterol is crucial to various cell functions. Here we examined the effect of cholesterol depletion on apoptosis and the mechanisms underlying this effect in NIH3T3 cells. We show that chronic cholesterol depletion achieved with lipoprotein-deficient serum (LPDS) and 25-hydroxycholesterol (25-HC) treatment resulted in a significant increase in cellular apoptosis and caspase-3 activation. This effect is not due to a deficiency of nonsterol isoprenoids, intermediate metabolites of the cholesterol biosynthetic pathway, but rather to low cholesterol levels, since addition of cholesterol together with LPDS and 25-HC nearly abolished apoptosis, whereas addition of farnesyl pyrophosphate or geranylgeranyl-pyrophosphate did not reverse the cell viability loss induced by LPDS plus 25-HC treatment. These effects were accompanied by an increase in ERK, JNK and p38 MAPK activity. However, only the inhibition of p38 MAPK with the specific inhibitor SB203580 or the overexpression of a kinase defective MKK6 resulted in a significant decrease in apoptosis and caspase-3 cleavage induced by cholesterol depletion. Furthermore, LPDS plus 25-HC increased RhoA activity, and this effect was reversed by addition of exogenous cholesterol. Finally, overexpression of the dominant negative N19RhoA inhibited p38 MAPK phosphorylation and apoptosis induced by low cholesterol levels. Together, our results demonstrate that cholesterol depletion induces apoptosis through a RhoA- and p38 MAPK-dependent mechanism.

Crosstalk between glucocorticoids and mitogen-activated protein kinase signalling pathways.

Synthetic glucocorticoids are potent modulators of the immune system, and are clinically invaluable in the treatment of a wide variety of conditions. However, their use can be limited by harmful side effects, and their mechanisms of action remain poorly understood and controversial. It has recently been reported that glucocorticoids induce the expression of two genes that participate in cross-talk with the mitogen-activated protein kinase signalling pathways. It is possible that both therapeutic and harmful effects of glucocorticoids may be mediated by these genes.

Dexamethasone causes sustained expression of mitogen-activated protein kinase (MAPK) phosphatase 1 and phosphatase-mediated inhibition of MAPK p38.

The stress-activated protein kinase p38 stabilizes a number of mRNAs encoding inflammatory mediators, such as cyclooxygenase 2 (Cox-2). In HeLa cells the anti-inflammatory glucocorticoid dexamethasone destabilizes Cox-2 mRNA by inhibiting p38 function. Here we demonstrate that this effect is phosphatase dependent. Furthermore, in HeLa cells dexamethasone induced the sustained expression of mitogen-activated protein kinase phosphatase 1 (MKP-1), a potent inhibitor of p38 function. The inhibition of p38 and the induction of MKP-1 by dexamethasone occurred with similar dose dependence and kinetics. No other known p38 phosphatases were induced by dexamethasone, and other cell types which failed to express MKP-1 also failed to inhibit p38 in response to dexamethasone. The proinflammatory cytokine interleukin 1 (IL-1) induced MKP-1 expression in a p38-dependent manner and acted synergistically with dexamethasone to induce MKP-1 expression. In HeLa cells treated with IL-1 or IL-1 and dexamethasone, the dynamics of p38 activation mirrored the expression of MKP-1. These observations suggest that MKP-1 participates in a negative-feedback loop which regulates p38 function and that dexamethasone may inhibit proinflammatory gene expression in part by inducing MKP-1 expression.