Antagonistic effects of oxidized low density lipoprotein and alpha-tocopherol on CD36 scavenger receptor expression in monocytes: involvement of protein kinase B and peroxisome proliferator-activated receptor-gamma

Vitamin E deficiency increases expression of the CD36 scavenger receptor, suggesting specific molecular mechanisms and signaling pathways modulated by alpha-tocopherol. We show here that alpha-tocopherol down-regulated CD36 expression (mRNA and protein) in oxidized low density lipoprotein (oxLDL)-stimulated THP-1 monocytes, but not in unstimulated cells. Furthermore, alpha-tocopherol treatment of monocytes led to reduction of fluorescent oxLDL-3,3'-dioctadecyloxacarbocyanine perchlorate binding and uptake. Protein kinase C (PKC) appears not to be involved because neither activation of PKC by phorbol 12-myristate 13-acetate nor inhibition by PKC412 was affected by alpha-tocopherol. However, alpha-tocopherol could partially prevent CD36 induction after stimulation with a specific agonist of peroxisome proliferator-activated receptor-gamma (PPARgamma; troglitazone), indicating that this pathway is susceptible to alpha-tocopherol action. Phosphorylation of protein kinase B (PKB) at Ser473 was increased by oxLDL, and alpha-tocopherol could prevent this event. Expression of PKB stimulated the CD36 promoter as well as a PPARgamma element-driven reporter gene, whereas an inactive PKB mutant had no effect. Moreover, coexpression of PPARgamma and PKB led to additive induction of CD36 expression. Altogether, our results support the existence of PKB/PPARgamma signaling pathways that mediate CD36 expression in response to oxLDL. The activation of CD36 expression by PKB suggests that both lipid biosynthesis and fatty acid uptake are stimulated by PKB.

Expression of proinflammatory cytokines tumor necrosis factor-alpha and interleukin-1beta in the brain during experimental group B streptococcal meningitis

We performed mRNA in situ hybridization for TNF-alpha and IL-1beta from infant rats with group B streptococcal meningitis. Induction of both cytokines was seen in the ependyma and the meninges at 4 h. Both cytokines were expressed in the brain parenchyma at 12 h. Induction of IL-1beta mRNA was seen in vessels within the brain cortex. Neutrophilic infiltrate at all time points examined was minimal and could not account for the observed cytokine expression.

Tumor necrosis factor-alpha contributes to apoptosis in hippocampal neurons during experimental group B streptococcal meningitis

To evaluate the role of tumor necrosis factor-alpha (TNF-alpha) in neuronal injury in experimental group B streptococcal meningitis, infected neonatal rats were treated with a monoclonal antibody against TNF-alpha (20 mg/kg intraperitoneally) or saline given at the time of infection. Histopathology after 24 h showed necrosis in the cortex and apoptosis in the hippocampal dentate gyrus. Treated animals had significantly less hippocampal injury than did controls (P < .001) but had similar cortical injury and cerebrospinal fluid (CSF) inflammation. The antibody was then administered directly intracisternally (170 microg) to test whether higher CSF concentrations reduced inflammation or cortical injury. Again, hippocampal apoptosis was significantly reduced (P < .01), while cortical injury and inflammation were not. Thus, TNF-alpha played a critical role in neuronal apoptosis in the hippocampus, while it was not essential for the development of inflammation and cortical injury in this model.

BPAG1 isoform-b: Complex distribution pattern in striated and heart muscle and association with plectin and α-actinin

BPAG1-b is the major muscle-specific isoform encoded by the dystonin gene, which expresses various protein isoforms belonging to the plakin protein family with complex, tissue-specific expression profiles. Recent observations in mice with either engineered or spontaneous mutations in the dystonin gene indicate that BPAG1-b serves as a cytolinker important for the establishment and maintenance of the cytoarchitecture and integrity of striated muscle. Here, we studied in detail its distribution in skeletal and cardiac muscles and assessed potential binding partners. BPAG1-b was detectable in vitro and in vivo as a high molecular mass protein in striated and heart muscle cells, co-localizing with the sarcomeric Z-disc protein alpha-actinin-2 and partially with the cytolinker plectin as well as with the intermediate filament protein desmin. Ultrastructurally, like alpha-actinin-2, BPAG1-b was predominantly localized at the Z-discs, adjacent to desmin-containing structures. BPAG1-b was able to form complexes with both plectin and alpha-actinin-2, and its NH(2)-terminus, which contains an actin-binding domain, directly interacted with that of plectin and alpha-actinin. Moreover, the protein level of BPAG1-b was reduced in muscle tissues from plectin-null mutant mice versus wild-type mice. These studies provide new insights into the role of BPAG1-b in the cytoskeletal organization of striated muscle.

Hepatoprotective effect of tumour necrosis factor alpha blockade in psoriatic arthritis: a cross-sectional study

Objective To evaluate the impact of tumour necrosis factor α (TNFα) blockers on the presence of liver fibrosis in patients with rheumatoid arthritis (RA) and psoriatic arthritis (PsA) treated with methotrexate (MTX). Methods Participants were consecutive patients with RA and PsA who had undergone MTX treatment for at least 1 year ± TNF blockade for over 6 months. Liver fibrosis was assessed using non-invasive transient elastography (FibroScan). Regression models were used to compare FibroScan values of patients with RA and patients with PsA receiving TNFα blockers with those who were not. Results FibroScan assessments were performed on 51 patients with RA and 43 patients with PsA. Compared to patients with RA, those with PsA were predominantly young men, received lower cumulative dosages of MTX and exhibited a higher incidence of liver steatosis and hyperlipidaemia. An abnormal result was observed in 7.1% of the anti-TNFα-naïve and in 13% of the anti-TNFα-treated patients in the RA group and in 30% of the anti-TNFα-naïve and 4.3% of the anti-TNFα-treated patients in the PsA group (OR=0.11, 95% CI 0.02 to 0.98). Results of the PsA group were robust when adjusted for baseline characteristics. Conclusion The results suggest a protective effect of TNFα inhibitors against the development of liver fibrosis in patients with PsA.

[Swiss registry for TNF-alpha blockers in children and adolescents with rheumatological diseases]

We created a registry to evaluate long term outcome, efficacy and adverse events for children treated wit TNF-alpha inhibitors in Switzerland. 106 patients (68 female/38 male) were included. 61 patients were treated with Etanercept (Enbrel) and 45 with Infliximab (Remicade). Concomitant treatment at baseline included corticosteroids in 26% and Methotrexate in 75% of the patients. Subjective disease activity three months after initiation of TNF-alpha was better in 81%, worse in 4% and stable in 15% of the patients. In total 24 adverse events in 21 patients were reported. Treatment with TNF-alpha inhibitors seems to be safe and effective for children and adolescents with rheumatologic diseases.

Immune response to influenza vaccination in children treated with methotrexate or/and tumor necrosis factor-alpha inhibitors

In children treated with immunosuppressive medication such as methotrexate and tumor necrosis factor-alpha (TNF-α) inhibitors, additional immunizations are recommended because of increased susceptibility to infections. However, it is unclear if adequate antibody response to vaccinations can be established in children receiving methotrexate and/or TNF-α inhibitors. In a prospective open label study, we assessed seroprotection and seroconversion following influenza vaccination during 2 seasons (6 strains) in 36 children with autoimmune disease treated either with methotrexate (n=18), TNF-α inhibitors (n=10) or both (n=8) and a control group of 16 immunocompetent children. Influenza antibody titers were determined by hemagglutinin inhibition assay, before and 4-8 weeks after vaccination. Post-vaccination seroprotection (defined as a titer ≥1:40) did not significantly differ between immunosuppressed and immunocompetent subjects. Seroconversion, defined as the change from a nonprotective (< 1:40) to a protective titer (≥1:40) with at least a 4-fold titer increase, was less likely to occur in immunosuppressed patients, although no significant difference from the control group was established. Safety evaluation of vaccination showed no serious adverse events. Children receiving methotrexate and/or TNF-α inhibitors can be safely and effectively immunized against influenza, with a seroprotection after vaccination comparable to immunocompetent children.

Molecular characterization of the equine collagen, type IX, alpha 2 (COL9A2) gene on horse chromosome 2p16-->p15

The mammalian collagen, type IX, alpha 2 gene (COL9A2) encodes the alpha-2 chain of type IX collagen and is located on horse chromosome 2p16-->p14 harbouring a quantitative trait locus for osteochondrosis. We isolated a bacterial artificial chromosome (BAC) clone containing the equine COL9A2 gene and determined the complete genomic sequence of this gene. Cloning and characterization of equine COL9A2 revealed that the equine gene consists of 32 exons spanning approximately 15 kb. The COL9A2 transcript encodes a single protein of 688 amino acids. Thirty two single nucleotide polymorphisms (SNPs) equally distributed in the gene were detected in a mutation scan of eight unrelated Hanoverian warmblood stallions, including one SNP that affects the amino acid sequence of COL9A2. Comparative analyses between horse, human, mouse and rat indicate that the chromosomal location of equine COL9A2 is in agreement with known chromosomal synteny relationships. The comparison of the gene structure and transcript revealed a high degree of conservation towards the other mammalian COL9A2 genes. We chose three informative SNPs for association and linkage disequilibrium tests in three to five paternal half-sib families of Hanoverian warmblood horses consisting of 44 to 75 genotyped animals. The test statistics did not reach the significance threshold of 5% and so we could not show an association of COL9A2 with equine osteochondrosis.

HCV-related advanced fibrosis/cirrhosis: randomized controlled trial of pegylated interferon alpha-2a and ribavirin

In patients with hepatitis C virus (HCV)-related advanced fibrosis/cirrhosis, 30% of sustained HCV clearance has been reported with pegylated interferon alpha-2a (PEG-IFN) alone, but the efficacy and tolerability of the PEG-IFN/ribavirin (RBV) combination remain poorly defined. A total of 124 treatment-naïve patients with biopsy proved HCV-related advanced fibrosis/cirrhosis (Ishak score F4-F6, Child-Pugh score < or =7) were randomized to 48 weeks of PEG-IFN (180 microg sc weekly) and standard dose of RBV (1000/1200 mg po daily, STD) or PEG-IFN (180 microg sc weekly) and low-dose of RBV (600/800 mg po daily, LOW). Sustained virologic response (SVR) rates with PEG-IFN/STD RBV (52%) were higher--albeit not significantly--than that with PEG-IFN/LOW RBV (38%, P = 0.153). In multivariate analysis, genotype 2/3 and a baseline platelet count > or =150 x 10(9)/L were independently associated with SVR. The likelihood of SVR was < 7% if viraemia had not declined by > or =2 log or to undetectable levels after 12 weeks. Nine adverse events in the STD RBV and 15 in the LOW RBV group were classified as severe (including two deaths); dose reductions for intolerance were required in 78% and 57% (P = 0.013), and treatment was terminated early in 23% and 27% of patients (P = n.s.). The benefit/risk ratio of treating compensated HCV-cirrhotics with STD PEG-IFN/RBV is favourable.

Function of liver activation-regulated chemokine/CC chemokine ligand 20 is differently affected by cathepsin B and cathepsin D processing

Chemokine processing by proteases is emerging as an important regulatory mechanism of leukocyte functions and possibly also of cancer progression. We screened a large panel of chemokines for degradation by cathepsins B and D, two proteases involved in tumor progression. Among the few substrates processed by both proteases, we focused on CCL20, the unique chemokine ligand of CCR6 that is expressed on immature dendritic cells and subtypes of memory lymphocytes. Analysis of the cleavage sites demonstrate that cathepsin B specifically cleaves off four C-terminally located amino acids and generates a CCL20(1-66) isoform with full functional activity. By contrast, cathepsin D totally inactivates the chemotactic potency of CCL20 by generating CCL20(1-55), CCL20(1-52), and a 12-aa C-terminal peptide CCL20(59-70). Proteolytic cleavage of CCL20 occurs also with chemokine bound to glycosaminoglycans. In addition, we characterized human melanoma cells as a novel CCL20 source and as cathepsin producers. CCL20 production was up-regulated by IL-1alpha and TNF-alpha in all cell lines tested, and in human metastatic melanoma cells. Whereas cathepsin D is secreted in the extracellular milieu, cathepsin B activity is confined to cytosol and cellular membranes. Our studies suggest that CCL20 processing in the extracellular environment of melanoma cells is exclusively mediated by cathepsin D. Thus, we propose a model where cathepsin D inactivates CCL20 and possibly prevents the establishment of an effective antitumoral immune response in melanomas.