Synovial tissues concentrate secreted APRIL

Introduction

TNF-like weak inducer of apoptosis (TWEAK) has been implicated as a mediator of chronic inflammatory processes via prolonged activation of the NF-κB pathway in several tissues, including the kidney. Evidence for the importance of TWEAK in the pathogenesis of lupus nephritis (LN) has been recently introduced. Thus, TWEAK levels may serve as an indication of LN presence and activity.

Methods

Multicenter cohorts of systemic lupus erythematosus (SLE) patients and controls were recruited for cross-sectional and longitudinal analysis of urinary TWEAK (uTWEAK) and/or serum TWEAK (sTWEAK) levels as potential biomarkers of LN. The performance of TWEAK as a biomarker for nephritis was compared with routinely used laboratory tests in lupus patients, including anti-double stranded DNA antibodies and levels of C3 and C4.

Results

uTWEAK levels were significantly higher in LN patients than in non-LN SLE patients and other disease control groups (P = 0.039). Furthermore, uTWEAK was better at distinguishing between LN and non-LN SLE patients than anti-DNA antibodies and complement levels, while high uTWEAK levels predicted LN in SLE patients with an odds ratio of 7.36 (95% confidence interval = 2.25 to 24.07; P = 0.001). uTWEAK levels peaked during LN flares, and were significantly higher during the flare than at 4 and 6 months prior to or following the flare event. A linear mixed-effects model showed a significant association between uTWEAK levels in SLE patients and their disease activity over time (P = 0.008). sTWEAK levels, however, were not found to correlate with the presence of LN or the degree of nephritis activity.

Conclusions

High uTWEAK levels are indicative of LN, as opposed to non-LN SLE and other healthy and disease control populations, and reflect renal disease activity in longitudinal follow-up. Thus, our study further supports a role for TWEAK in the pathogenesis of LN, and provides strong evidence for uTWEAK as a candidate clinical biomarker for LN.     

The biological and clinical importance of the 'new generation' cytokines in rheumatic diseases

A better understanding of cytokine biology over the last two decades has allowed the successful development of cytokine inhibitors against tumour necrosis factor and interleukin (IL)-1 and IL-6. The introduction of these therapies should be considered a breakthrough in the management of several rheumatic diseases. However, many patients will exhibit no or only partial response to these therapies, thus emphasising the importance of exploring other therapeutic strategies. In this article, we review the most recent information on novel cytokines that are often members of previously described cytokine families such as the IL-1 superfamily (IL-18 and IL-33), the IL-12 superfamily (IL-27 and IL-35), the IL-2 superfamily (IL-15 and IL-21), and IL-17. Several data derived from experimental models and clinical samples indicate that some of these cytokines contribute to the pathophysiology of arthritis and other inflammatory diseases. Targeting of some of these cytokines has already been tested in clinical trials with interesting results.     

Indirect effects of leptin receptor deficiency on lymphocyte populations and immune response in db/db mice

Leptin-deficient ob/ob and leptin receptor (Ob-rb)-deficient db/db mice display a marked thymic atrophy and exhibit defective immune responses. Lymphocytes express leptin receptors and leptin exerts direct effects on T cells in vitro. In addition, ob/ob and db/db mice display multiple neuroendocrine and metabolic defects, through which leptin deficiency may indirectly affect the immune system in vivo. To study the relative contributions of direct and indirect effects of leptin on the immune system in a normal environment, we generated bone marrow chimeras (BMCs) by transplantation of leptin receptor-deficient db/db, or control db/+, bone marrow cells into wild-type (WT) recipients. The size and cellularity of the thymus, as well as cellular and humoral immune responses, were similar in db/db to WT and db/+ to WT BMCs. The immune phenotype of db/db mice is thus not explained by a cell autonomous defect of db/db lymphocytes. Conversely, thymus weight and cell number were decreased in the reverse graft setting in WT to db/db BMCs, indicating that expression of the leptin receptor in the environment is important for T cell development. Finally, normal thymocyte development occurred in fetal db/db thymi transplanted into WT hosts, indicating that direct effects of leptin are not required locally in the thymic microenvironment. In conclusion, direct effects of leptin on bone marrow-derived cells and on thymic stromal cells are not necessary for T lymphocyte maturation in normal mice. In contrast, leptin receptor deficiency affects the immune system indirectly via changes in the systemic environment.     

Cell-free co-production of an orthogonal transfer RNA activates efficient site-specific non-natural amino acid incorporation

We describe a new cell-free protein synthesis (CFPS) method for site-specific incorporation of non-natural amino acids (nnAAs) into proteins in which the orthogonal tRNA (o-tRNA) and the modified protein (i.e. the protein containing the nnAA) are produced simultaneously. Using this method, 0.9–1.7 mg/ml of modified soluble super-folder green fluorescent protein (sfGFP) containing either p-azido-l-phenylalanine (pAzF) or p-propargyloxy-l-phenylalanine (pPaF) accumulated in the CFPS solutions; these yields correspond to 50–88% suppression efficiency. The o-tRNA can be transcribed either from a linearized plasmid or from a crude PCR product. Comparison of two different o-tRNAs suggests that the new platform is not limited by Ef-Tu recognition of the acylated o-tRNA at sufficiently high o-tRNA template concentrations. Analysis of nnAA incorporation across 12 different sites in sfGFP suggests that modified protein yields and suppression efficiencies (i.e. the position effect) do not correlate with any of the reported trends. Sites that were ineffectively suppressed with the original o-tRNA were better suppressed with an optimized o-tRNA (o-tRNA^opt) that was evolved to be better recognized by Ef-Tu. This new platform can also be used to screen scissile ribozymes for improved catalysis.     

Cyclooxygenase-2 gene and epithelial ovarian carcinoma risk

In this study, we aimed to investigate a possible association of the COX-2 polymorphisms (−765G→C and −1195A→G) and with the risk of developing epithelial ovarian carcinoma (EOC). COX-2 gene polymorphisms was investigated in 111 healthy women and 57 patients with EOC. Individuals who had −765 CG, −1195 AA genotype, and −765 C allele had increased risk for ovarian carcinoma (P < 0.01) and individuals with −765 GG, −1195 AG genotypes and −1195 G allele seem to be protected from ovarian carcinoma (P < 0.01). Haplotype analysis confirmed the association of COX-2 gene variants with ovarian carcinoma and revealed that the frequencies of −765C: −1195A haplotype frequencies was significantly higher in patients as compared with those of controls (P = 0.048). We state that there appears to be a modulating role for the COX-2 −1195A→G and −765G→C polymorphisms in the development of EOC. To the best of our knowledge, this is the first study to show such an association.     

Enzymes of mannitol metabolism in the human pathogenic fungus Aspergillus fumigatus--kinetic properties of mannitol-1-phosphate 5-dehydrogenase and mannitol 2-dehydrogenase, and their physiological implications

The human pathogenic fungus Aspergillus fumigatus accumulates large amounts of intracellular mannitol to enhance its resistance against defense strategies of the infected host. To explore their currently unknown roles in mannitol metabolism, we studied A. fumigatus mannitol-1-phosphate 5-dehydrogenase (AfM1PDH) and mannitol 2-dehydrogenase (AfM2DH), each recombinantly produced in Escherichia coli, and performed a detailed steady-state kinetic characterization of the two enzymes at 25 °C and pH 7.1. Primary kinetic isotope effects resulting from deuteration of alcohol substrate or NADH showed that, for AfM1PDH, binding of D-mannitol 1-phosphate and NAD(+) is random, whereas D-fructose 6-phosphate binds only after NADH has bound to the enzyme. Binding of substrate and NAD(H) by AfM2DH is random for both D-mannitol oxidation and D-fructose reduction. Hydride transfer is rate-determining for D-mannitol 1-phosphate oxidation by AfM1PDH (k(cat) = 10.6 s(-1)) as well as D-fructose reduction by AfM2DH (k(cat) = 94 s(-1)). Product release steps control the maximum rates in the other direction of the two enzymatic reactions. Free energy profiles for the enzymatic reaction under physiological boundary conditions suggest that AfM1PDH primarily functions as a D-fructose-6-phosphate reductase, whereas AfM2DH acts in D-mannitol oxidation, thus establishing distinct routes for production and mobilization of mannitol in A. fumigatus. ATP, ADP and AMP do not affect the activity of AfM1PDH, suggesting the absence of flux control by cellular energy charge at the level of D-fructose 6-phosphate reduction. AfM1PDH is remarkably resistant to inactivation by heat (half-life at 40 °C of 20 h), consistent with the idea that formation of mannitol is an essential component of the temperature stress response of A. fumigatus. Inhibition of AfM1PDH might be a useful target for therapy of A. fumigatus infections.