RETRACTED ARTICLE: Mast cells are the main interleukin 17-positive cells in anticitrullinated protein antibody-positive and -negative rheumatoid arthritis and osteoarthritis synovium

Introduction  

Mast cells have been implicated to play a functional role in arthritis, especially in autoantibody-positive disease. Among the cytokines involved in rheumatoid arthritis (RA), IL-17 is an important inflammatory mediator. Recent data suggest that the synovial mast cell is a main producer of IL-17, although T cells have also been implicated as prominent IL-17 producers as well. We aimed to identify IL-17 expression by mast cells and T cells in synovium of arthritis patients.     

Phyto (in)stabilization of elements

The effects of plants (corn, soybean, and sunflower) and fertilizer on mobility of more than 60 elements were assessed in a greenhouse experiment. Unplanted columns with the same soil served as controls. Half the columns received fertilizer and all columns were watered at the same rate. At the end of the experiment, the columns were watered to mimic a rainstorm event such that water drained from the bases of the columns, which was collected and analyzed for element content. Soil from between the roots of the plants was also collected and the water-extractable fraction determined. It was expected that (1) more mobile elements, as measured by water extraction, would be leached from the soils at a higher rate compared to less mobile elements, (2) plants would immobilize most elements, but that some would be immobilized, and (3) that this would depend on plant species. The results led to the following conclusions: plants cause metal mobility to vary over a wide range for a specific soil and do mobilize some elements (e.g., Th) while immobilizing others (e.g., U). The effects depended on plant species for some elements. Water-extractable fractions of elements do not predict mobility.     

WW domain sequence activity relationships identified using ligand recognition propensities of 42 WW domains

WW domains mediate protein-protein interactions in a number of different cellular functions by recognizing proline-containing peptide sequences. We determined peptide recognition propensities for 42 WW domains using NMR spectroscopy and peptide library screens. As potential ligands, we studied both model peptides and peptides based on naturally occurring sequences, including phosphorylated residues. Thirty-two WW domains were classified into six groups according to detected ligand recognition preferences for binding the motifs PPx(Y/poY), (p/phi)P(p,g)PPpR, (p/phi)PPRgpPp, PPLPp, (p/xi)PPPPP, and (poS/poT)P (motifs according to modified Seefeld Convention 2001). In addition to these distinct binding motifs, group-specific WW domain consensus sequences were identified. For PPxY-recognizing domains, phospho-tyrosine binding was also observed. Based on the sequences of the PPx(Y/poY)-specific group, a profile hidden Markov model was calculated and used to predict PPx(Y/poY)-recognition activity for WW domains, which were not assayed. PPx(Y/poY)-binding was found to be a common property of NEDD4-like ubiquitin ligases.     

Identification of anti-repressor elements that confer high and stable protein production in mammalian cells

The expression of transgenic proteins is often low and unstable over time, a problem that may be due to integration of the transgene in repressed chromatin. We developed a screening technology to identify genetic elements that efficiently counteract chromatin-associated repression. When these elements were used to flank a transgene, we observed a substantial increase in the number of mammalian cell colonies that expressed the transgenic protein. Expression of the shielded transgene was, in a copy number-dependent fashion, substantially higher than the expression of unprotected transgenes. Also, protein production remained stable over an extended time period. The DNA elements are small, not exceeding 2,100 base pairs (bp), and they are highly conserved between human and mouse, at both the functional and sequence levels. Our results demonstrate the existence of a class of genetic elements that can readily be applied to more efficient transgenic protein production in mammalian cells.     

No evidence for DUP25 in patients with panic disorder using a quantitative real-time PCR approach

A duplication of chromosome 15q24-q26 (DUP25) has been reported to be associated with anxiety disorders. We tested for the presence of DUP25 in a sample of 50 patients with panic disorder and 50 controls using a quantitative real-time PCR approach. Contrary to the original finding, our results were compatible with the absence of DUP25, and no significant difference could be detected between patients and controls (P=1.0). Thus, our study does not support the hypothesis of an involvement of DUP25 in panic disorder.     

Expression and distribution of the prolactin receptor in normal rat liver and in experimental liver cirrhosis.

Recent results have suggested a role for prolactin (PRL) as a regeneration factor in the liver. In order to investigate the involvement of prolactin in the pathogenesis of liver cirrhosis, we studied the expression of the prolactin receptor (PRLR) and PRL during the development of cirrhosis in an animal model. 30 male rats were exposed to CCl4 by inhalation. Phenobarbitone was added to the drinking water to accelerate the formation of toxic metabolites by enzyme induction. Two control groups of 30 animals each were treated with phenobarbitone only or received no treatment. 10 animals of each group were sacrificed 35, 55, and 70 days after initiation of treatment. Liver tissue was subjected to histological examination, which demonstrated fibrosis of different grades and cirrhosis in the CCl4-treated rats. Expression of PRLR mRNA was investigated by mRNA extraction, RT-PCR and computer-supported densitometric evaluation. Compared to control liver, PRLR mRNA was expressed at a higher level in fibrotic and cirrhotic liver specimens. In normal tissue, immunohistochemical staining showed a high concentration of PRLR around the central vein and in the epithelium of the bile ducts. This pattern of distribution was lost in fibrosis and cirrhosis. An accumulation of PRLR was demonstrated within the damaged cells. Neither PRL nor PRL mRNA was detectable in normal, fibrotic, or cirrhotic liver. We conclude that PRLR is distributed in normal rat liver in a typical pattern which is lost with increasing fibrosis. PRL is not produced by rat liver, indicating that PRL does not act through autocrine or paracrine mechanisms.     

Early elicitor-induced events in chickpea cells: functional links between oxidative burst, sequential occurrence of extracellular alkalinisation and acidification, K+/H+ exchange and defence-related gene activation

Elicitation of cultured chickpea (Cicer arietinum L.) cells stimulates a signal transduction pathway leading to several rapid responses: (1) oxidative burst, (2) extracellular alkalinisation, (3) extracellular acidification, (4) transient K+ efflux, and (5) activation of defence related genes all within 2 hours. Induced genes are encoding acidic and basic chitinases, a thaumatin-like protein and isoflavone reductase. All these elicitor-induced responses are inhibited by the Ser/Thr protein kinase inhibitor staurosporine and the anion channel blocker anthracene-9-carboxylic acid but stimulated by the Ser/Thr protein phosphatase 2A inhibitor cantharidin. The oxidative burst leads to a transient extracellular H2O2 accumulation which seems to be preceded by O2- production, indicating dismutation of O2- to H2O2. The oxidative burst is accompanied by transient alkalinisation of the culture medium which is followed by long-lasting extracellular acidification. An 80 percent inhibition of the alkalinisation after complete inhibition of the H2O2 burst with diphenylene iodonium indicates that the elicitor induced increase of extracellular pH is mainly based on a proton consumption for O2-dismutation. A simultaneous deactivation of the plasma membrane H+-ATPase during oxidative burst and extracellular alkalinisation is also suggested. The elicitor-stimulated extracellular acidification is inhibited by the plasma membrane H+-ATPase inhibitor N, N'-dicyclohexylcarbodiimide assuming a reactivation of the H+-ATPase 25 min after elicitation. Extracellular acidification seems not to be necessary for elicitor-induced activation of defence related genes. Opposite modulation of K+ and proton fluxes after elicitation and/or treatment with the H+-ATPase effectors fusicoccin or N, N'-dicyclohexylcarbodiimide indicate that the elicitor induced transient K+ efflux is regulated by a K+/H+ exchange reaction.     

Reliability of computerized image analysis for the evaluation of serial synovial biopsies in randomized controlled trials in rheumatoid arthritis

Analysis of biomarkers in synovial tissue is increasingly used in the evaluation of new targeted therapies for patients with rheumatoid arthritis (RA). This study determined the intrarater and inter-rater reliability of digital image analysis (DIA) of synovial biopsies from RA patients participating in clinical trials. Arthroscopic synovial biopsies were obtained before and after treatment from 19 RA patients participating in a randomized controlled trial with prednisolone. Immunohistochemistry was used to detect CD3⁺ T cells, CD38⁺ plasma cells and CD68⁺ macrophages. The mean change in positive cells per square millimetre for each marker was determined by different operators and at different times using DIA. Nonparametric tests were used to determine differences between observers and assessments, and to determine changes after treatment. The intraclass correlations (ICCs) were calculated to determine the intrarater and inter-rater reliability. Intrarater ICCs showed good reliability for measuring changes in T lymphocytes (R = 0.87), plasma cells (R = 0.62) and macrophages (R = 0.73). Analysis by Bland–Altman plots showed no systemic differences between measurements. The smallest detectable changes were calculated and their discriminatory power revealed good response in the prednisolone group compared with the placebo group. Similarly, inter-rater ICCs also revealed good reliability for measuring T lymphocytes (R = 0.68), plasma cells (R = 0.69) and macrophages (R = 0.72). All measurements identified the same cell types as changing significantly in the treated patients compared with the placebo group. The measurement of change in total positive cell numbers in synovial tissue can be determined reproducibly for various cell types by DIA in RA clinical trials.