Erja-Leena Paukkeri

PPARα agonists inhibit nitric oxide production by enhancing iNOS degradation in LPS-treated macrophages

Nitric oxide (NO) production through the inducible nitric oxide synthase (iNOS) pathway is increased in response to pro‐inflammatory cytokines and bacterial products. In inflammation, NO has pro‐inflammatory and regulatory effects. Peroxisome proliferator‐activated receptors (PPARs), members of the nuclear steroid receptor superfamily, regulate not only metabolic but also inflammatory processes. The aim of the present study was to investigate the role of PPARα in the regulation of NO production and iNOS expression in activated macrophages.

YKL-40 as a Novel Factor Associated with Inflammation and Catabolic Mechanisms in Osteoarthritic Joints

YKL-40 is associated with tissue injury and inflammation, and consequently to diseases in which these mechanisms lead to tissue degradation, for example, asthma and rheumatoid arthritis. The purpose of the present study was to investigate if YKL-40 is also a significant factor in osteoarthritis (OA) by assessing associations of YKL-40 with mediators related to the pathogenesis of OA: cartilage destructing matrix metalloproteinases (MMPs) and proinflammatory cytokines interleukin-6 (IL-6) and interleukin-17 (IL-17). Cartilage, synovial fluid (SF), and plasma samples were obtained from 100 OA patients undergoing total knee replacement surgery. SF levels of YKL-40 (1027.9 ± 78.3 ng/mL) were considerably higher than plasma levels (67.2 ± 4.5 ng/mL) and correlated with YKL-40 released from cartilage samples obtained from the same patients (r = 0.37, P = 0.010), indicating that YKL-40 is produced by OA cartilage. Interestingly, YKL-40 concentrations in OA SF correlated positively with MMP-1 (r = 0.36, P = 0.014), MMP-3 (r = 0.46, P = 0.001), IL-6 (r = 0.57, P < 0.001), and IL-17 (r = 0.52, P = 0.010) levels. Moreover, IL-6 and IL-17 enhanced YKL-40 production in human primary chondrocyte cultures. The present study introduces YKL-40 as a cartilage-derived factor associated with mediators of inflammation and cartilage destruction involved in the pathogenesis of OA.

Compounds that increase or mimic cyclic adenosine monophosphate enhance tristetraprolin degradation in lipopolysaccharide-treated murine j774 macrophages.

Tristetraprolin (TTP) is a trans-acting factor that can regulate mRNA stability by binding to the cis-acting AU-rich element (ARE) in the 3′-untranslated region in mRNAs of certain transiently expressed genes. The best-studied target of TTP is tumor necrosis factor (TNF)-α. By binding to ARE, TTP increases the degradation of TNF-α mRNA, thereby reducing the expression of TNF-α. We examined the effects of cAMP analogs and the cAMP-elevating agents forskolin and β2-agonists on lipopolysaccharide (LPS)-induced TTP mRNA and protein expression by quantitative real-time reverse transcriptase-polymerase chain reaction and Western blotting in activated macrophages. All of these agents caused a slight increase in LPS-induced expression of TTP mRNA. However, TTP protein levels were significantly reduced when the cells were treated with the combination of LPS and cAMP-elevating agent compared with LPS alone. Proteasome inhibitors MG132 (N-[(phenylmethoxy)-carbonyl]-l-leucyl-N-[(1S)-1-formyl-3-methylbutyl]-l-leucinamide) and lactacystin increased TTP protein levels and abolished the effects of cAMP-enhancing compounds on TTP protein levels. The results suggest that mediators and drugs that enhance intracellular cAMP reduce TTP expression in macrophages exposed to inflammatory stimuli by increasing TTP degradation through the proteasome pathway.

Anti-inflammatory properties of a dual PPARgamma/alpha agonist muraglitazar in in vitro and in vivo models

IntroductionPeroxisome proliferator-activated receptor (PPAR) agonists are widely used drugs in the treatment of diabetes and dyslipidemia. In addition to their metabolic effects, PPAR isoforms PPARα and PPARγ are also involved in the regulation of immune responses and inflammation. In the present study, we investigated the effects of a dual PPARγ/α agonist muraglitazar on inflammatory gene expression in activated macrophages and on carrageenan-induced inflammation in the mouse.MethodsJ774 murine macrophages were activated by lipopolysaccharide (LPS) and treated with dual PPARγ/α agonist muraglitazar, PPARγ agonist GW1929 or PPARα agonist fenofibrate. The effects of PPAR agonists on cytokine production and the activation of inducible nitric oxide synthase (iNOS) pathway were investigated by ELISA, Griess method, Western blotting and quantitative RT-PCR. Nuclear translocation, DNA-binding activity and reporter gene assays were used to assess the activity of nuclear factor kappa B (NF-kB) transcription factor. Carrageenan-induced paw oedema was used as an in vivo model of acute inflammation.ResultsMuraglitazar as well as PPARγ agonist GW1929 and PPARα agonist fenofibrate inhibited LPS-induced iNOS expression and NO production in activated macrophages in a dose-dependent manner. Inhibition of iNOS expression by muraglitazar included both transcriptional and post-transcriptional components; the former being shared by GW1929 and the latter by fenofibrate. All tested PPAR agonists also inhibited IL-6 production, while TNFα production was reduced by muraglitazar and GW1929, but not by fenofibrate. Interestingly, the anti-inflammatory properties of muraglitazar were also translated in vivo. This was evidenced by the finding that muraglitazar inhibited carrageenan-induced paw inflammation in a dose-dependent manner in mice as did iNOS inhibitor L-NIL and anti-inflammatory steroid dexamethasone.ConclusionsThese results show that muraglitazar has anti-inflammatory properties both in vitro and in vivo and these effects reflect the agonistic action through both PPARα and PPARγ.

The Inflammatory Phenotype in Failed Metal-On-Metal Hip Arthroplasty Correlates with Blood Metal Concentrations

Introduction Hip arthroplasty is the standard treatment of a painful hip destruction. The use of modern metal-on-metal (MOM) bearing surfaces gained popularity in total hip arthroplasties during the last decade. Recently, worrisome failures due to adverse reaction to metal debris (ARMD), including pseudotumor response, have been widely reported. However, the pathogenesis of this reaction remains poorly understood. The aim of the present study was to investigate the ARMD response by flow cytometry approach. Methods Sixteen patients with a failed Articular Surface Replacement (ASR) hip prosthesis were included in the study. Samples of pseudotumor tissues collected during revision surgery were degraded by enzyme digestion and cells were typed by flow cytometry. Whole blood chromium and cobalt concentrations were analyzed with mass spectrometry before revision surgery. Results Flow cytometry analysis showed that the peri-implant pseudotumor tissue expressed two principal phenotypes, namely macrophage-dominated and T-lymphocyte-dominated response; the average portions being 54% (macrophages) and 25% (T-lymphocytes) in macrophage-dominated inflammation and 20% (macrophages) and 54% (T-lymphocytes) in T-lymphocyte-dominated response. The percentages of B-lymphocytes and granulocytes were lower in both phenotypes. Interestingly, the levels of blood chromium and cobalt were significantly higher in patients with macrophage-dominated response. Conclusions The results suggest that the adverse tissue reactions induced by MOM wear particles contain heterogeneous pathogeneses and that the metal levels are an important factor in the determination of the inflammatory phenotype. The present results support the hypothesis that higher metal levels cause cytotoxicity and tissue injury and macrophages are recruited to clear the necrotic debris. On the other hand, the adverse response developed in association with lower metal levels is T-lymphocyte-dominated and is likely to reflect hypersensitivity reaction.

Cobalt(II) Chloride Modifies the Phenotype of Macrophage Activation

Cobalt (Co) is vital for cells in trace amounts, but excessive exposure to Co is possible due to surgical devices such as artificial metal‐on‐metal joints. Cobalt(II) chloride (CoCl2) has also been shown to imitate hypoxic conditions in cells by stabilizing the transcription factor hypoxia‐inducible factor‐1α (HIF‐1α). The purpose of this study was to investigate the possible immunomodulatory action of CoCl2 by investigating its effects on the expression of inflammatory genes in macrophages. The following factors were assessed: inducible nitric oxidase synthase (iNOS), nicotinamide adenine dinucleotide phosphate‐oxidase 2 (NOX2), interleukin‐6 (IL‐6), arginase‐1 and HIF‐1α. In the absence of exogenous cytokines, CoCl2 enhanced alternative (M2) macrophage activation as demonstrated by increased arginase‐1 expression, but had no direct effect on inflammatory factors associated with classical (M1) activation. Interestingly, in lipopolysaccharide (LPS)‐stimulated macrophages, CoCl2 modified the M1‐type activation profile by increasing iNOS expression and nitric oxide production and decreasing NOX2 and IL‐6. Also, CoCl2 increased HIF‐1α levels in unstimulated and LPS‐stimulated cells as expected. In conclusion, we showed that CoCl2 enhanced alternative (M2) activation in resting macrophages. In addition, CoCl2 was found to remodel the classical M1 phenotype of macrophage activation by changing the balance of iNOS, NOX2 and IL‐6.