The p38 MAPK Regulates IL-24 Expression by Stabilization of the 3′ UTR of IL-24 mRNA

IL-24 (melanoma differentiation-associated gene-7 (mda-7)), a member of the IL-10 cytokine family, possesses the properties of a classical cytokine as well as tumor suppressor effects. The exact role of IL-24 in the immune system has not been defined but studies have indicated a role for IL-24 in inflammatory conditions such as psoriasis. The tumor suppressor effects of IL-24 include inhibition of angiogenesis, sensitization to chemotherapy, and p38 mitogen-activated protein kinase (MAPK)-mediated apoptosis. Current knowledge on the regulation of IL-24 expression is sparse. Previous studies have suggested that mRNA stabilization is of major importance to IL-24 expression. Yet, the mechanisms responsible for the regulation of IL-24 mRNA stability remain unidentified. As p38 MAPK is known to regulate gene expression by interfering with mRNA degradation we examined the role of p38 MAPK in the regulation of IL-24 gene expression in cultured normal human keratinocytes.In the present study we show that anisomycin- and IL-1beta- induced IL-24 expression is strongly dependent on p38 MAPK activation. Studies of IL-24 mRNA stability in anisomycin-treated keratinocytes reveal that the p38 MAPK inhibitor SB 202190 accelerates IL-24 mRNA decay suggesting p38 MAPK to regulate IL-24 expression by mRNA-stabilizing mechanisms. The insertion of the 3' untranslated region (UTR) of IL-24 mRNA in a tet-off reporter construct induces degradation of the reporter mRNA. The observed mRNA degradation is markedly reduced when a constitutively active mutant of MAPK kinase 6 (MKK6), which selectively activates p38 MAPK, is co-expressed.Taken together, we here report p38 MAPK as a regulator of IL-24 expression and determine interference with destabilization mediated by the 3' UTR of IL-24 mRNA as mode of action. As discussed in the present work these findings have important implications for our understanding of IL-24 as a tumor suppressor protein as well as an immune modulating cytokine

Waiting for JAK inhibitor safety data

The US Food and Drug Administration (FDA) has recently added a new ‘black box warning’ on all currently approved Janus kinase (JAK) inhibitors indicated for the treatment of arthritis and other inflammatory conditions based on results from the ORAL Surveillance study of tofacitinib versus tumour necrosis factor alpha inhibitors in rheumatoid arthritis. This is a warning difficult to ignore because the data, being from a randomised controlled trial, are of high fidelity and hard to reproach. It is especially problematic because safety data for all the other JAK inhibitors will be pending for several years. So how might we proceed, without being bound by our stasis? The lack of absolute certainty seems to require a pragmatic approach to the routine care use of JAK inhibitors. The patients who were at greatest risk were older and had other risk factors for the corresponding adverse events, in keeping with effect modification. This highlights the need to focus on risk stratification when tailoring therapy. In this viewpoint, we propose a simple illustration to guide clinical decision-making. First, identify general risk factors for venous thromboembolic event (VTE), major adverse cardiac event (MACE) and cancer (age>65 years and smoking) and whether there is a previous history of VTE, MACE or cancer. Then, evaluate risk based on the number of other risk factors for VTE and the number of other risk factors for MACE. Ultimately, ‘treat-to-target’ will in the end always be ‘treat-to-agreement’. As we have done in the past, and will do in the future, the optimal treatment strategy will have to be tailored based on individual patient risk factors and preferences in a shared-decision process

Intermittent Hypoxic Therapy Inhibits Allogenic Bone-Graft Resorption by Inhibition of Osteoclastogenesis in a Mouse Model

Systemic Intermittent Hypoxic Therapy (IHT) relies on the adaptive response to hypoxic stress. We investigated allogenic bone-graft resorption in the lumbar spine in 48 mice. The mice were exposed to IHT for 1 week before surgery or 1 week after surgery and compared with controls after 1 and 4 weeks. Complete graft resorption was observed in 33–36% of the animals in the control group, but none in the preoperative IHT group. Increased bone-graft volume was demonstrated by micro-computed tomography in the preoperative IHT group after 1 week (p = 0.03) while a non-significant difference was observed after 4 weeks (p = 0.12). There were no significant differences in the postoperative IHT group. Increased concentration of immune cells was localized in the graft area, and more positive tartrate-resistant acid phosphatase (TRAP) staining was found in controls compared with IHT allogenic bone grafts. Systemic IHT resulted in a significant increase of the major osteoclast inhibitor osteoprotegerin as well as osteogenic and angiogenic regulators Tgfbr3, Fst3l, Wisp1, and Vegfd. Inflammatory cytokines and receptor activator of nuclear factor kappa-B ligand (RANKL) stimulators IL-6, IL-17a, IL-17f, and IL-23r increased after 1 and 4 weeks, and serum RANKL expression remained constant while Ccl3 and Ccl5 decreased. We conclude that the adaptive response to IHT activates numerous pathways leading to inhibition of osteoclastic activity and inhibition of allogenic bone-graft resorption

p38 MAPK regulates IL-24 expression at a posttranscriptional level.

<p>(A and B) NHEK were pre-stimulated with anisomycin (300 ng/ml) for 2 hours at which point medium (control), the transcription inhibitor actinomycin D, or SB 202190 were added to the cultures. Cells were then harvested at 30 minute intervals, total RNA extracted and (A) IL-24 and (B) TNFα-mRNA levels (relative to 18 s RNA) were determined. *p<0.05 compared with control (n = 5). Points indicate mean ± SD. (C) As control NHEK were pre-incubated with or without actinomycin D for one hour before treatment with medium or anisomycin (300 ng/ml) for another 2 hours. Total RNA was extracted IL-24 mRNA levels (relative to 18 s RNA) were determined. *p<0.05 compared with medium control (n = 4). Bars indicate mean ± SD.</p

Inhibition of p38 MAPK activation abrogates IL-24 expression.

<p>(A) Whole cell extracts were prepared from NHEK incubated with or without (control) anisomycin (300 ng/ml) for 30 minutes in the absence or presence of the 38 MAPK inhibitor SB 202190 (10 µM). Proteins were separated by SDS-PAGE. After electroblotting, separated proteins were probed with anti-phospho-MK2 (recognizing the phosphorylated form of MK2 isoforms 1 and 2) and anti-total p38 MAPK. A gel representative of three different experiments is shown. NHEK were cultured with or without SB 202190 (10 µM) for 30 minutes before incubation with (B) anisomycin (300 ng/ml) or medium control for 4 hours or (C) IL-1β (10 ng/ml) or medium control for 2 hours. Total RNA was extracted and IL-24 mRNA (relative to housekeeping gene 18 s RNA) levels were determined. *p<0.05 compared with medium control (n = 5 cultures). Bars indicate mean ± SD. (D) NHEK were cultured in the presence or absence of SB 202190 (10 µM) with medium control or anisomycin (300 ng/ml) for 18 hours. Using ELISA IL-24 concentration in the supernatants was determined and subsequently normalized to the total protein concentration of whole cell extracts. *p<0.05 compared with medium control (n = 5 cultures). Bars indicate mean ± SD.</p