Psychophysiological responses to stress after stress management training in patients with rheumatoid arthritis

Background

Stress management interventions may prove useful in preventing the detrimental effects of stress on health. This study assessed the effects of a stress management intervention on the psychophysiological response to stress in patients with rheumatoid arthritis (RA).

Methods

Seventy-four patients with RA, who were randomly assigned to either a control group or a group that received short-term stress management training, performed a standardized psychosocial stress task (Trier Social Stress Test; TSST) 1 week after the stress management training and at a 9-week follow-up. Psychological and physical functioning, and the acute psychophysiological response to the stress test were assessed.

Results

Patients in the intervention group showed significantly lower psychological distress levels of anxiety after the training than did the controls. While there were no between-group differences in stress-induced tension levels, and autonomic (α-amylase) or endocrine (cortisol) responses to the stress test 1 week after the intervention, levels of stress-induced tension and cortisol were significantly lower in the intervention group at the 9-week follow-up. Overall, the response to the intervention was particularly evident in a subgroup of patients with a psychological risk profile.

Conclusion

A relatively short stress management intervention can improve psychological functioning and influences the psychophysiological response to stress in patients with RA, particularly those psychologically at risk. These findings might help understand how stress can affect health and the role of individual differences in stress responsiveness.

Trial Registration

TrialRegister.nl NTR1193     

An analysis of genetic variation across the MBL2 locus in Dutch Caucasians indicates that 3′ haplotypes could modify circulating levels of mannose-binding lectin

Mannose-binding protein (MBL) is a critical component of innate immunity and provides first-line protection against pathogens. Both circulating MBL serum levels and functional activity have been correlated with common genetic variants in the MBL2 gene. Associations between MBL deficiency and severe infections have been reported in immuno-incompetent patients and for autoimmune disorders; however, measured MBL serum levels do not fully correlate with the ‘secretor haplotypes’. Previously, the MBL2 locus was resequenced and determined that a recombination hotspot divides MBL2 into two haplotype blocks. It was sought to investigate whether additional variants, in either block structure could associate with MBL serum levels. Therefore, 31 common variants were analysed across the locus in 212 DNA samples of healthy Caucasian individuals with known MBL serum concentrations. The additional 5′ variants were in strong linkage to the elements of the ‘secretor haplotypes’; functional alleles B, C and D also lie on restricted haplotypes. Four variants in the 3′ block (Ex4-1483T>C, Ex4-1067G>A, Ex4-901G>A and Ex4-710G>A) are components of a distinct haplotype block. The results of this study suggest that additional 5′ variants as well as markers of distinct 3′ haplotype blocks in MBL2 may contribute to circulating protein levels, but further studies are required to confirm these observations. Last, there could be a selective advantage for diversification of the 3′ region of the gene.Electronic Supplementary Material Supplementary material is available for this article at     

Short-term regulation of multidrug resistance-associated protein 3 in rat and human hepatocytes

The short-term regulation of multidrug resistance-associated protein 3 (Mrp3/MRP3) by cAMP and PKC was investigated in sandwich-cultured rat and human hepatocytes and isolated perfused rat livers. The modulator glucagon (500 nM) and the phorbol ester PMA (0.1 muM) were utilized to increase intracellular cAMP and PKC levels, respectively. In glucagon-treated rat hepatocytes, efflux of the Mrp3 substrate 5-(6)-carboxy-2',7'-dichlorofluorescein (CDF) increased approximately 1.5-fold, even in hepatocytes treated with the organic anion transporter (Oatp) inhibitor sulfobromophthalein (BSP). Confocal microscopy revealed more concentrated Mrp3 fluorescence in the basolateral membrane (less diffuse staining pattern) with glucagon treatment. PMA had no effect on Mrp3 activity or localization in sandwich-cultured rat hepatocytes. Glucagon and PMA treatment in isolated perfused rat livers resulted in a threefold increase (14 +/- 4.6 mul.min(-1).g liver(-1)) and a fourfold decrease (1.3 +/- 0.3 mul.min(-1).g liver(-1)) in CDF basolateral clearance compared with control livers (4.7 +/- 2.3 mul.min(-1).g liver(-1)), whereas CDF biliary clearance was not statistically different. In sandwich-cultured human hepatocytes, glucagon treatment resulted in a 1.3-fold increase in CDF efflux and a concomitant increase in MRP3 fluorescence in the basolateral membrane. In summary, cAMP and PKC appear to be involved in the short-term regulation of Mrp3/MRP3, as demonstrated by alterations in activity and localization in rat and human hepatocytes.     

Optimisation of the fluorescein diacetate antibacterial assay

The fluorescein diacetate (FDA) antibacterial assay relies on the cleavage of fluorescein diacetate by metabolically active bacteria. The recent finding that microbiological media can lead to significant levels of cleavage has reduced the reliability of the assay. Using the nucleophilic scavengers N-ethylmaleimide and maleic anhydride, we have demonstrated that this abiotic cleavage is most likely due to nucleophiles such as cysteine and histidine commonly present in the media. To increase the reliability of the assay we have modified the original assay conditions to include use of dilute medium (peptone 0.2% w/v, yeast extract 0.1% w/v and NaCl 0.1% w/v) in a non-nucleophilic buffer and overnight incubation of the medium after addition of antibacterial agents. The optimised fluorescein diacetate assay has been used to determine the MIC of gentamicin, tetracycline and chloramphenicol for Escherichia coli, Staphyloccocus aureus and Pseudomonas aeruginosa and gave quantitative results that were reproducible and consistent with published data.     

O-acetylation of cryptococcal capsular glucuronoxylomannan is essential for interference with neutrophil migration

The capsular polysaccharide glucuronoxylomannan (GXM) of Cryptococcus neoformans has been shown to interfere with neutrophil migration. Although several receptors have been implied to mediate this process, the structural perspectives are unknown. Here, we assess the contribution of 6-O-acetylation and xylose substitution of the (1-->3)-alpha-d-mannan backbone of GXM, the variable structural features of GXM, to the interference with neutrophil migration. We compare chemically deacetylated GXM and acetyl- or xylose-deficient GXM from genetically modified strains with wild-type GXM in their ability to inhibit the different phases of neutrophil migration. Additionally, we verify the effects of de-O-acetylation on neutrophil migration in vivo. De-O-acetylation caused a dramatic reduction of the inhibitory capacity of GXM in the in vitro assays for neutrophil chemokinesis, rolling on E-selectin and firm adhesion to endothelium. Genetic removal of xylose only marginally reduced the ability of GXM to reduce firm adhesion. In vivo, chemical deacetylation of GXM significantly reduced its ability to interfere with neutrophil recruitment in a model of myocardial ischemia (65% reduction vs a nonsignificant reduction in tissue myeloperoxidase, respectively). Our findings indicate that 6-O-acetylated mannose of GXM is a crucial motive for the inhibition of neutrophil recruitment.     

ZNF674: a new kruppel-associated box-containing zinc-finger gene involved in nonsyndromic X-linked mental retardation

Array-based comparative genomic hybridization has proven to be successful in the identification of genetic defects in disorders involving mental retardation. Here, we studied a patient with learning disabilities, retinal dystrophy, and short stature. The family history was suggestive of an X-linked contiguous gene syndrome. Hybridization of full-coverage X-chromosomal bacterial artificial chromosome arrays revealed a deletion of ~1 Mb in Xp11.3, which harbors RP2, SLC9A7, CHST7, and two hypothetical zinc-finger genes, ZNF673 and ZNF674. These genes were analyzed in 28 families with nonsyndromic X-linked mental retardation (XLMR) that show linkage to Xp11.3; the analysis revealed a nonsense mutation, p.E118X, in the coding sequence of ZNF674 in one family. This mutation is predicted to result in a truncated protein containing the Kruppel-associated box domains but lacking the zinc-finger domains, which are crucial for DNA binding. We characterized the complete ZNF674 gene structure and subsequently tested an additional 306 patients with XLMR for mutations by direct sequencing. Two amino acid substitutions, p.T343M and p.P412L, were identified that were not found in unaffected individuals. The proline at position 412 is conserved between species and is predicted by molecular modeling to reduce the DNA-binding properties of ZNF674. The p.T343M transition is probably a polymorphism, because the homologous ZNF674 gene in chimpanzee has a methionine at that position. ZNF674 belongs to a cluster of seven highly related zinc-finger genes in Xp11, two of which (ZNF41 and ZNF81) were implicated previously in XLMR. Identification of ZNF674 as the third XLMR gene in this cluster may indicate a common role for these zinc-finger genes that is crucial to human cognitive functioning.     

Evidence that complement protein C1q interacts with C-reactive protein through its globular head region

C1q acts as the recognition unit of the first complement component, C1, and binds to immunoglobulins IgG and IgM, as well as to non-Ig ligands, such as C-reactive protein (CRP). IgG and IgM are recognized via the globular head regions of C1q (C1qGR), whereas CRP has been postulated to interact with the collagen-like region (C1qCLR). In the present study, we used a series of nine mAbs to C1q, five directed against C1qGR and four against C1qCLR, to inhibit the interaction of C1q with CRP. The F(ab')(2) of each of the five mAbs directed against C1qGR inhibited binding of C1q to polymerized IgG. These five mAbs also successfully inhibited the interaction of C1q with CRP. Moreover, these five mAbs inhibited C1 activation by CRP as well as by polymerized IgG in vitro. In contrast, none of the four mAbs against C1qCLR inhibited C1q interaction with CRP or IgG, or could reduce activation of complement by CRP or polymerized IgG. These results provide the first evidence that the interaction of C1q with CRP or IgG involves sites located in the C1qGR, whereas sites in the CLR do not seem to be involved in the physiological interaction of C1q with CRP.     

Multiplexed expression and screening for recombinant protein production in mammalian cells

Background  

A variety of approaches to understanding protein structure and function require production of recombinant protein. Mammalian based expression systems have advantages over bacterial systems for certain classes of protein but can be slower and more laborious. Thus the availability of a simple system for production and rapid screening of constructs or conditions for mammalian expression would be of great benefit. To this end we have coupled an efficient recombinant protein production system based on transient transfection in HEK-293 EBNA1 (HEK-293E) suspension cells with a dot blot method allowing pre-screening of proteins expressed in cells in a high throughput manner.