Mesalazine Suppresses Proinflammatory Cytokines in Patients with Acute Anterior Uveitis Independently of HLA-B27.

PURPOSE: The aim is to unravel the mechanism of mesalazine (5-ASA) on proinflammatory cytokines in PBMCs of patients with HLA-B27 +and HLA-B27 -acute anterior uveitis (AAU), and whether this may explain the different effects of 5-ASA in both disoders. METHODS: PBMCs from 12 HLA-B27+ and 4 HLA-B27- AAU patients were preincubated with 5-ASA and stimulated with LPS. As mesalazine (5-ASA) could be involved in ER stress, proinflammatory and ER stress-associated cytokines and markers were measured. RESULTS: Mesalazine (5-ASA) suppressed IL-6 mRNA in healthy donors and in HLA-B27+ and HLA-B27- patients but did not lead to induction and secretion of IL-1ß. In HLA-B27 + or - patients the ER stress-associated markers CHOP (DDIT3) and ATF6 were suppressed. CONCLUSIONS: Here we show that mesalazine (5-ASA) inhibits the transcription of proinflammatory and (ER) stress associated cytokines and markers, independently of the HLA-B27 status. Results show the similarities of both AAU types but do not decipher the mechanism why the HLA-B27 status determines the therapeutic response to mesalazine in AAU.

ASC Speck Formation after Inflammasome Activation in Primary Human Keratinocytes.

Chronic UV irradiation results in many changes in the skin, including hyperplasia, changes in dermal structures, and alteration of pigmentation. Exposure to UVB leads to cutaneous damage, which results in inflammation characterized by increased NF-κB activation and the induction of inflammatory cytokines, such as tumor necrosis factor (TNF), interleukin- (IL-) 1, or IL-8. IL-1 secretion is the result of inflammasome activation which is besides apoptosis, a result of acute UVB treatment. Inflammasomes are cytosolic protein complexes whose formation results in the activation of proinflammatory caspase-1. Key substrates of caspase-1 are IL-1ß and IL-18, and the cytosolic protein gasdermin D (GSDMD), which is involved in inflammatory cell death. Here, we demonstrate that UVB-induced inflammasome activation leads to the formation of ASC specks. Our findings show that UVB provokes ASC speck formation in human primary keratinocytes prior to cell death, and that specks are, opposed to the perinuclear cytosolic localization in myeloid cells, formed in the nucleus. Additionally, we showed by RNAi that NLRP1 and not NLRP3 is the major inflammasome responsible for UVB sensing in primary human keratinocytes. Formation of ASC specks indicates inflammasome assembly and activation as their formation in hPKs depends on the presence of NLRP1 and partially on NLRP3. Nuclear ASC specks are not specific for NLRP1/NLRP3 inflammasome activation, as the activation of the AIM2 inflammasome by cytosolic DNA results in ASC specks too. These nuclear ASC specks putatively link cell death to inflammasome activation, possibly by binding of IFI16 (gamma-interferon-inducible protein) to ASC. ASC can interact upon UVB sensing via IFI16 with p53, linking cell death to ASC speck formation.

Differential induction of ATF3 and HO-1 in myeloid cells and keratinocytes via Dimethylfumarate or Cyclosporine A.

BACKGROUND: Chronic inflammatory skin diseases are characterized by controlled proliferation of keratinocytes. Here, activating transcription factor 3 (ATF3) might play a fundamental role. In these inflammatory diseases, proliferation is controlled and only rarely leads to cancer development which can be supported by an inflammatory microenvironment. ATF3 is a dual function protein as it suppresses pro-inflammatory IL-6 and IL-8, but also acts as a pro-oncogenic factor by the suppression of p53. We therefore analyzed ATF3 expression comparing myeloid cells with keratinocytes. OBJECTIVE: To dissect the bi-modal role of ATF3 we pharmacologically induced ATF3 and analyzed its influence on cytokine expression and secretion in a cell type specific manner. METHODS: Since inflammatory skin diseases can be treated systemically with Cyclosporin A or Dimethylfumarate we stimulated myeloid cells and primary human keratinocytes with these drugs and analyzed gene expression by quantitative real-time PCR. Cytokine secretion was measured by ELISA. RESULTS: In the present study, we could show that ATF3 is induced in PBMCs by DMF and weakly by Ebselen, while CsA is the most prominent inducer of ATF3 in keratinocytes without enhancing HO-1 transcription. Further we could show that induction of stress by LPS treatment elevates IL-1ß and IL-6 and weakly ATF3 transcription in PBMCs. While transcription of both cytokines is elevated, LPS treatment mediates IL-6 secretion with only little IL-1ß secretion. Treatment with DMF dampens LPS-induced transcription. CONCLUSIONS: Taken together, our results shed light into the different carcinogenic potential of CsA and DMF, which both target ATF3. Collectively our data demonstrate that CsA strongly induces pro-carcinogenic ATF3 in keratinocytes, whereas ATF3 induction by DMF in myeloid cells acts anti-inflammatory.