Role of SIRPα in regulation of mucosal immunity in the intestine

Mononuclear phagocytes such as dendritic cells (DCs) and macrophages in the lamina propria (LP) are thought to be important for both induction of inflammatory responses and maintenance of immunologic tolerance in the mammalian intestine. The molecular mechanisms by which these cells regulate intestinal immunity have remained poorly understood, however. Signal regulatory protein α (SIRPα) is a transmembrane protein that is specifically expressed in DCs, macrophages and neutrophils. Here, we show that SIRPα is abundant in CD11c(+) CD11b(+) LP cells of the mouse intestine. Whereas SIRPα did not appear to be important for the steady-state homeostasis of mucosal immunity in the intestine, the flagellin-stimulated production of IL-17 or interferon (IFN)-γ by LP cells of SIRPα mutant (MT) mice that lack the cytoplasmic region of the protein was markedly decreased compared with that observed with wild-type cells. Moreover, the flagellin-induced production of IL-6 by LP cells from SIRPα MT mice was also greatly reduced. SIRPα MT mice were also resistant to the development of colitis induced by IL-10 deficiency. Our data thus suggest that SIRPα expressed on CD11c(+) LP cells is important for the production of IL-17 or IFN-γ in the LP as well as for the development of colitis induced by IL-10 deficiency.     

Optimal Use of Epoetin-α in the Anaemia of Chronic Renal Failure

Anaemia, a major feature of chronic renal failure, is responsible for a significant proportion of the multisystemic disabling symptoms in patients with chronic renal failure (CRF). Anaemia correction with recombinant human erythropoietin (epoetin-alpha) often leads to significant amelioration of these symptoms. Starting dose should be 50 U/kg of bodyweight given subcutaneously or intravenously 1, 2 or 3 times a week. The subcutaneous route is more effective, resulting in use of less epoetin-alpha and subsequent cost-saving. Adverse effects of epoetin-alpha tend to be more severe and more likely following intravenous administration. Once the target haematocrit is achieved, the epoetin-alpha dose should be reduced by 50 to 80%, but should not be stopped, since an abrupt fall in haematocrit may ensue if epoetin-alpha is completely discontinued. There is an emerging consensus that it is deleterious, and not necessary, to allow patients to become severely anaemic before starting epoetin-alpha. Thus, the current trend is to start epoetin-alpha once the haematocrit is 30 to 35%. On the other hand, what the target haematocrit should be is controversial. The threshold haematocrit beyond which the risks of anaemia correction outweigh the benefits in patients with CRF is not established. Studies that assessed quality of life outcomes as well as clinical experience in Europe clearly demonstrate better outcomes without adverse effects in patients with a haematocrit between 36 to 40%, compared to those with haematocrit of 30%. Prior to starting epoetin-alpha in any patient with CRF, a methodological evalution should be performed to exclude other major causes of anaemia and to identify any comorbid conditions that may impair response to epoetin-alpha. Response to epoetin-alpha may be impaired by loss of residual renal function in patients with progressive CRF or kidney transplant recipients. Other major factors that impair response to epoetin-alpha include impaired utilisation of iron, absolute iron deficiency, inadequate dialysis, infection and bone marrow fibrosis.     

Recent advances in the use of Anti-TNFα therapy for the treatment of juvenile idiopathic arthritis

Juvenile Idiopathic Arthritis (JIA) encompasses a group of diseases of unknown etiology having in common arthritis in at least 1 joint that persists for 6 weeks and begins before 16 years of age, with other conditions excluded. With a prevalence of 1 per 1,000 children in the USA, JIA is the most common pediatric rheumatic illness and a major cause of acquired childhood disability. During the last 20 years, the advent of host immune response modifiers known as biologic agents, in particular the anti-TNFα agents (etanercept, infliximab, adalimumab), which directly inhibit the action of pro-inflammatory mediators, has revolutionized the treatment and the expected outcome of JIA. This article highlights treatment indications of anti-TNFα drugs and their more frequent side effects in JIA patients.     

Attenuation of shock-induced inflammation in the rat liver depends on the time of TNF-α inhibition

The relevance of tumor necrosis factor- (TNF-) inducing early inflammatory reactions in the liver after hemorrhagic shock, for example, leukocyte adhesion, has been well described. This study evaluated the anti-inflammatory effects of a monoclonal antibody against TNF- (TN3.19.12) in terms of the time of application, namely, prior to shock induction, at the time of resuscitation, and after resuscitation. The hepatic micro-circulation was investigated by intravital fluorescence microscopy in female Sprague-Dawley rats undergoing severe hemorrhagic shock for 60 min and subsequent resuscitation. TN3.19.12 or placebo was given in a randomized and blinded manner either 60 min prior to shock induction, l min prior to resuscitation, or 15 min after the onset of resuscitation. The number of firmly adherent leukocytes in the livers of treated animals depended on the time of application of TN3.19.12. Leukocyte adhesion was significantly reduced when TN3.19.12 was given prior to shock induction or at the time of resuscitation and was less effective when administered after the onset of resuscitation. The results further confirm that TNF- initiates very early pathological leukocyte adhesion in the liver 5 h following shock. Inhibition of leukocyte adhesion after shock, however, depends strongly on the time of TNF- blocking. While TN3.19.12 prior to shock induction resulted in most effective attenuation, only very early treatment allowed limitation of posttraumatically increased leukocyte adhesion.Abbreviations CO Cardiac output - HR Heart rate - IL Interleukin - MAP Mean arterial pressure - WBC White blood cells     

Isolation of the Bα and Bβ mating-type loci of Schizophyllum commune

The B mating type of the basidiomycete fungus, Schizophyllum commune is determined by two, tightly linked, multi-specificity (also called multi-allelic) loci: B and B. A plasmid library was used in DNA-mediated transformation to obtain transformants that displayed B-directed development. Plasmids that conferred B1 and B1 mating-type specificities were rescued from the transformants. Fragments of DNA from each plasmid hybridized to genomic DNA from the strain used to make the plasmid library; however, they did not hybridize, or hybridized only weakly, to genomic DNA from strains with mating-type specificities different from B1 or B1. The cloned fragments are presumed to correspond to active regions of each B mating-type locus.     

Safety of anti-TNFα agents in the treatment of psoriasis and psoriatic arthritis

Context: The efficacy and favorable safety profile of anti-tumor necrosis factor (TNF) agents in the treatment of psoriasis and psoriatic arthritis (PsA) are supported by several randomized controlled studies and meta-analyses. However, some concerns on the long-term safety of these drugs still exist, as these studies generally included small patient numbers and were performed in selected patient populations.

Objective: This review presents and discusses current evidence on the safety of anti-TNFα agents in patients with psoriasis and PsA, with a focus on European registry studies and case reports of particular importance.

Methods: Key studies on the safety of anti-TNFα agents in the treatment of adult patients with psoriasis or PsA were identified by a MEDLINE search (last updated 10 November 2011) based on several interrelated queries, with a focus on European registries. Other studies and case reports were included if deemed relevant. Studies concerning other conditions, such as rheumatoid arthritis (RA), were included as appropriate when data in psoriatic disease were unavailable or insufficient.

Results: Available data on the safety of anti-TNFα agents such as etanercept in psoriasis and PsA appear reassuring, even if some concerns still exist. Most notably, data suggest a higher incidence of infection and lymphoma amongst patients treated with the anti-TNFα monoclonal antibodies infliximab and adalimumab compared with etanercept.

Conclusion: The overall safety profile of monoclonal antibodies in patients with psoriasis, PsA and RA seems less favorable than that of etanercept, particularly in terms of risk of infection and hepatotoxicity.     

Evidence of the TNF-α System in the Human Achilles Tendon: Expression of TNF-α and TNF Receptor at both Protein and mRNA Levels in the Tenocytes

Understanding adaption to load is essential for prevention and treatment of tendinopathy/tendinosis. Cytokine release in response to load is one mechanism involved in mechanotransduction. The cytokine tumor necrosis factor alpha (TNF-α) is implicated in tendinosis and can induce apoptotic effects via tumor necrosis factor receptor 1 (TNFR1). The complete absence of information concerning the TNF-α system in Achilles tendon is a limitation as mid-portion Achilles tendinosis is very frequent. Purpose: To examine expression patterns of TNF-α and its two receptors (TNFR1 and TNFR2) in human Achilles tendinosis and control tissue and to biochemically confirm the presence of TNF-α in tendinosis tissue. Methods: TNF-α and TNFR1 mRNA were detected via in situ hybridization. TNF-α, TNFR1, and TNFR2 were demonstrated immunohistochemically. Apoptosis markers were utilized. ELISA was used to detect TNF-α. Results: TNF-α and TNFR1 mRNA was detected in tenocytes of both tendinosis and control tendons. Tenocytes from both groups displayed specific immunoreactions for TNF-α, TNFR1, and TNFR2. The widened/rounded tenocytes of tendinosis samples exhibited the most intense immunoreactions. Apoptosis was detected in only a subpopulation of the tenocytes in tendinosis tissue. TNF-α was measurable in tendinosis tissue. Inflammatory cells were not seen. Conclusion: This is the first evidence of the existence of the TNF-α system in the human Achilles tendon. Findings are confirmed at mRNA and protein levels as well as biochemically. The TNF-α system was in principle confined to the tenocytes. The connection between tenocyte morphology and the expression pattern of TNF-α, TNFR1, and TNFR2 suggests that the TNF-α system may be involved in tenocyte activation in Achilles tendinosis.     

Oxygen tension modulates the effects of TNFα in compressed chondrocytes

Objective and design

Oxygen tension and biomechanical signals are factors that regulate inflammatory mechanisms in chondrocytes. We examined whether low oxygen tension influenced the cells response to TNFα and dynamic compression.

Materials and methods

Chondrocyte/agarose constructs were treated with varying concentrations of TNFα (0.1–100 ng/ml) and cultured at 5 and 21 % oxygen tension for 48 h. In separate experiments, constructs were subjected to dynamic compression (15 %) and treated with TNFα (10 ng/ml) and/or L-NIO (1 mM) at 5 and 21 % oxygen tension using an ex vivo bioreactor for 48 h. Markers for catabolic activity (NO, PGE₂) and tissue remodelling (GAG, MMPs) were quantified by biochemical assay. ADAMTS-5 and MMP-13 expression were examined by real-time qPCR. 2-way ANOVA and a post hoc Bonferroni-corrected t test were used to analyse data.

Results

TNFα dose-dependently increased NO, PGE₂ and MMP activity (all p < 0.001) and induced MMP-13 (p < 0.05) and ADAMTS-5 gene expression (pp < 0.01) with values greater at 5 % oxygen tension than 21 %. The induction of catabolic mediators by TNFα was reduced by dynamic compression and/or L-NIO (all p < 0.001), with a greater inhibition observed at 5% than 21 %. The stimulation of GAG synthesis by dynamic compression was greater at 21 % than 5 % oxygen tension and this response was reduced with TNFα or reversed with L-NIO.

Conclusions

The present findings revealed that TNFα increased production of NO, PGE₂ and MMP activity at 5 % oxygen tension. The effects induced by TNFα were reduced by dynamic compression and/or the NOS inhibitor, linking both types of stimuli to reparative activities. Future therapeutics should develop oxygen-sensitive antagonists which are directed to interfering with the TNFα-induced pathways.

     

TNFα Inhibitors in the Treatment of Psoriasis and Psoriatic Arthritis

Psoriasis is a chronic inflammatory skin disease that can lead to significant physical and psychologic distress for patients. Psoriatic arthritis (PsA), originally thought to be quite a mild disorder, is now recognized as a progressive and destructive arthritis. To date, therapies for both these conditions have been non-specific and unable to maintain long-lasting remission. In addition, many of the current therapies have significant adverse effects, limiting their usefulness. However, elucidation of the pathogenesis of psoriasis and PsA at a molecular level and the development of selective biologic agents have led to an enormous expansion of the armamentarium available to psoriasis patients. Two agents (infliximab and etanercept) selectively block the role of the cytokine tumor necrosis factor (TNF)-alpha and have proved effective in clinical trials in the treatment of both the skin and the joint manifestations of psoriasis. A third anti-TNF alpha agent (adalimumab Humira) is licensed for the treatment of rheumatoid arthritis; however, no studies have been published to date on its use in PsA or psoriasis. It is known that TNF alpha is elevated in both the skin and synovium of psoriatic patients and the effectiveness of its blockade by these two agents in psoriasis and PsA confirms its role in their pathogenesis. Randomized, double-blind, placebo-controlled trials have been performed with both agents in the treatment of psoriasis and PsA; in the case of etanercept these have been to support US FDA approval for use in psoriatic arthropathy. These studies are supported by smaller cohorts in open-label studies and anecdotal reports in the literature. Anti-TNF alpha therapy has proved to have disease-reducing activity in PsA and psoriasis and appears to be well tolerated. These studies have generally featured small numbers of patients and, until a larger cohort of treated patients is available, vigilance must be exercised. A considerable body of post-marketing safety data exists on the use of infliximab in rheumatoid arthritis and Crohn disease and for etanercept in rheumatoid arthritis and PsA. Certain issues, particularly the risk of infection, have emerged as features of the use of these agents. It remains to be seen whether effects seen in other disease entities may be extrapolated to psoriatic patients. More long-term data and experience are needed to define the role of anti-TNF alpha agents in the management of psoriasis and PsA. In particular, more studies are required to elucidate the finer points of co-medication; in some studies both agents have been used with other medications but there have been no formal trials of various possible combinations.     

Comparison of the eIF-2α Homologous Proteins of Seven Ranaviruses (Iridoviridae)

The -subunit of the eukaryotic initiation factor 2 (eIF-2) is a key component of the translation machinery of the cell. In response to cellular stress such as viral infections, eIF-2 is phosphorylated by double-stranded RNA-dependent protein kinase (PKR) leading to the inhibition of cellular protein synthesis. The importance of eIF-2 as a regulatory mechanism for protein synthesis is illustrated by the wide variety of strategies employed by viruses to down-regulate PKR. Thus, Vaccinia virus encodes K3L protein, which resembles eIF-2 and acts as a pseudo-substrate inhibitor of PKR. Nucleotide sequencing of the genome of epizootic haematopoietic necrosis virus (EHNV), a member of the genus ranavirus of Iridoviridae, has revealed an eIF-2 equivalent gene. We have cloned and sequenced eIF-2 genes of several iridoviruses of fishes and frogs. The eIF-2 open reading frames and deduced proteins of the iridoviruses investigated exhibit a high degree of homology of both nucleotide and amino acid sequences. At the N-terminus, the iridoviral eIF-2 shows significant homology to the N-termini of cellular initiation factor 2- of various species, to full-length poxviral eIF-2 proteins, and to the S1 domain of ribosomal proteins. Comparison of amino acid sequences of corresponding iridoviral proteins with eIF-2 homologous proteins of poxviruses and eukaryotes has revealed a high conservation of motifs. A phylogenetic analysis of eukaryotic eIF-2 and poxvirus and iridovirus eIF-2 sequences has demonstrated the relationship of these iridoviruses. In order to investigate the role of the eIF-2 equivalent, respective genes have been expressed in prokaryotic and eukaryotic (insect, fish and chicken cell) systems. The iridoviral eIF-2 protein has a molecular weight of 31 kDa and is cytoplasmic. The cellular and viral protein synthesis of iridoviruses is probably regulated by a mechanism similar to that of Vaccinia virus. Frog-virus 3, the type species of the genus ranavirus of Iridoviridae, has a unique translational efficiency and, moreover, down-regulates the cellular protein synthesis of infected cells.     

Tumor necrosis factor-α mediates the early pathology in Salmonella infection of the gastrointestinal tract

Salmonella infection of the intestinal tract results in damage to the gut epithelium. While it is generally believed that bacteria and/or bacterial products account for this pathology, the role of host factors has not been explored. Using a ligated intestinal loop model, we investigated whether tumor necrosis factor-α (TNF-α) could contribute to the tissue pathology associated with Salmonella infection. Intestinal segments infected with Salmonella typhimurium had high levels of fluid secretion as early as 6 h post bacterial infection. At this time point, low levels of TNF activity were also present in the fluid obtained from infected segments. At 20 h post-infection, high levels of TNF activity were present in fluids obtained from infected intestinal segments and was characterized as TNF-α by neutralization experiments using rabbit antisera to TNF-α. TNF-α production was further verified by Northern blot analysis using RNA obtained from cells eluted from the infected intestinal segments. In contrast, no TNF activity was found in fluid obtained from intestinal segments challenged with cholera toxin, which induces fluid secretion with little to no inflammatory response. Double labeling by in situ hybridization and immunocytochemistry revealed that macrophages in the lamina propria were producing the TNF-α mRNA. To investigate what role TNF-α might play in Salmonella-induced inflammation, intestinal segments were injected with recombinant mouse TNF-α (rTNF-α) or mice were pretreated with antibody to TNF-α or a control antibody prior to Salmonella infection. The histological profile of intestinal segments injected with rTNF-α appeared identical to segments infected with S. typhimurium. Further, pathology was completely eliminated in infected mice pretreated with antibody to TNF-α. These results document the production of TNF-α in the intestinal tract following S. typhimurium infection and show that the early pathology induced by Salmonella infection of the gastrointestinal tract is mediated by immune mechanisms.     

HIF-1α Upregulation due to Depletion of the Free Ubiquitin Pool

Hypoxia-inducible factor 1alpha (HIF-1α), which transactivates a variety of hypoxia-induced genes, is rapidly degraded under nomoxia through the hydroxylation-ubiquitination-proteasome pathway. In this study, we addressed how HIF-1α is stabilized by proteasome inhibitors. The ubiquitin pool was rapidly reduced after proteasome inhibition, followed by the accumulation of non-ubiquitinated HIF-1α. The poly-ubiquitination of HIF-1α was resumed by restoration of free ubiquitin, which suggests that the HIF-1α stabilization under proteasome inhibition is attributed to depletion of the free ubiquitin pool. Ni²⁺ and Zn²⁺ also stabilized HIF-1α with depletion of the free ubiquitin pool and these effects of metal ions were attenuated by restoration of free ubiquitin. Ni²⁺ and Zn²⁺ may disturb the recycling of free ubiquitin, as MG132 does. Based on these results, the state of the ubiquitin pool seems to be another critical factor determining the cellular level of HIF-1α.

Graphical Abstract

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Pharmacokinetics and Antitumor Effects of the Drug Containing TNF-α in Nanoparticles

Antitumor activity of TNF-α incorporated in nanoparticles (VLP-TNF-α) and dynamics of its accumulation and elimination from the blood and tumor tissue were studied in ICR mice. The VLP-TNF-α preparation exhibited higher antitumor activity compared to free TNF-α, presumably due to longer circulation of the cytokine in the blood and its more intensive accumulation by tumor tissue.     

The effect of simvastatin on chemotactic capability of SDF-1α and the promotion of bone regeneration

The purpose of this study was to investigate the cooperative effects of simvastatin (SIM) and stromal cell-derived factor-1α (SDF-1α) on the osteogenic and migration capabilities of mesenchymal stem cells (MSCs), and construct a cell-free bone tissue engineering system comprising SIM, SDF-1α and scaffold. We found that 0.2 μm SIM significantly increased alkaline phosphatase activity (P < 0.05) of mouse bone marrow MSCs with no inhibition of cell proliferation, and enhanced the chemotactic capability of SDF-1α (P < 0.05). Next, we constructed a novel cell-free bone tissue engineering system using PLGA loaded with SIM and SDF-1α, and applied it in critical-sized calvarial defects in mice. New bone formation in the defect was evaluated by micro-CT, HE staining and immunohistochemistry. The results showed that PLGA loaded with SIM and SDF-1α promoted bone regeneration significantly more than controls. We investigated possible mechanisms, and showed that SDF-1α combined with SIM increased MSC migration and homing in vivo, promoted angiogenesis and enhanced the expression of BMP-2 in newly-formed bone tissue. In conclusion, SIM enhanced the chemotactic capability of SDF-1α and the cell-free bone tissue engineering system composed of SIM, SDF-1α and scaffold promoted bone regeneration in mouse critical-sized calvarial defects.     

The role of the silicatein-α interactor silintaphin-1 in biomimetic biomineralization

Biosilicification in sponges is initiated by formation of proteinaceous filaments, predominantly consisting of silicateins. Silicateins enzymatically catalyze condensation of silica nanospheres, resulting in symmetric skeletal elements (spicules). In order to create tailored biosilica structures in biomimetic approaches it is mandatory to elucidate proteins that are fundamental for the assembly of filaments. Silintaphin-1 is a core component of modularized filaments and also part of a spicule-enfolding layer. It bears no resemblance to other proteins, except for the presence of an interaction domain that is fundamental for its function as scaffold/template. In the presence of silicatein silintaphin-1 facilitates the formation of in vitro filaments. Also, it directs the assembly of γ-Fe₂O₃ nanoparticles and surface-immobilized silicatein to rod-like biocomposites, synthetic spicules. Thus, silintaphin-1 will contribute to biomimetic approaches that pursue a controlled formation of patterned biosilica-based materials. Its combination with γ-Fe₂O₃ nanoparticles and immobilized silicatein will furthermore inspire future biomedical applications and clinical diagnostics.     

Decreased expression of microRNA-130a correlates with TNF-α in the development of osteoarthritis

Objective: Increased expression of tumor necrosis factor a (TNF-α) has emerged as an important inflammatory factor in osteoarthritis (OA) and other joint diseases. The study was performed to investigate whether the expression of TNF-α in human chondrocytes was regulated by miRNAs. Methods: MiRNA-130a and TNF-α expression in cartilage specimens was examined in patients with knee osteoarthritis, chondrocytes and osteoarthritis rat model. Chondrocytes were transfected with siRNAs as a gene silencing methods. Expression of genes and proteins were analyzed by real-time PCR and western blotting respectively. Results: Increased TNF-α and decreased miRNA-130a were observed in tissues from osteoarthritis patients. Moreover, we found a highly negitive correlation between miRNA-130a and TNF-α. Next, miRNA-130a loss-of-function increased the expression of TNF-α and promoted inflammation in chondrocytes. It was reasonable that miRNA-130a regulated a distinct underlying molecular and pathogenic mechanism of OA by forming a negative feedback loop with TNF-α. Furthermore, there were the abnormalities of bone metabolism in OA rat, which showed the miRNA-130a and TNF-α dysfunction that was one of important factors for the occurrence and development of OA. Conclusions: Our results indicated that miR-130a played an important role in regulating the expression of TNF-α in human chondrocytes and identified miR-130a as a novel therapeutic target in OA.