Oxidative stress-induced modifications of histidyl-tRNA synthetase affect its tRNA aminoacylation activity but not its immunoreactivity

The aminoacyl-tRNA synthetases are ubiquitously expressed enzymes that catalyze the esterification of amino acids to their cognate tRNAs. Autoantibodies against several aminoacyl-tRNA synthetases are found in autoimmune polymyositis and dermatomyositis patients. Because necrosis is often found in skeletal muscle biopsies of these patients, we hypothesized that cell-death-induced protein modifications may help in breaking immunological tolerance. Since cell death is associated with oxidative stress, the effect of oxidative stress on the main myositis-specific autoantibody target Jo-1 (histidyl-tRNA synthetase; HisRS) was studied in detail. The exposure of Jurkat cells to hydrogen peroxide resulted in the detection of several oxidized methionines and one oxidized tryptophan residue in the HisRS protein, as demonstrated by mass spectrometry. Unexpectedly, the tRNA aminoacylation activity of HisRS appeared to be increased upon oxidative modification. The analysis of myositis patient sera did not lead to the detection of autoantibodies that are specifically reactive with the modified HisRS protein. The results of this study demonstrate that the Jo-1/HisRS autoantigen is modified under oxidative stress conditions. The consequences of these modifications for the function of HisRS and its autoantigenicity are discussed.     

Experimental assessment of the probabilistic maturation reaction norm: condition matters

The probabilistic maturation reaction norm (PMRN) describes an individual''s probability of maturing at a given age as a function of size and other relevant phenotypic traits. Population-level shifts in the PMRN are often interpreted to indicate genetic as opposed to phenotypic changes in maturation in fish. Inferences derived from trends in the PMRN have been challenged, warranting an experimental assessment of the method. This was accomplished in a laboratory experiment using zebrafish (Danio rerio). Fish were reared under different food levels to induce variation in growth and maturation. Plasticity in maturation was not entirely captured by the demographic age- and length-based PMRN. Adding condition to the PMRN captured a greater amount of environmental variation in maturation probability. Nevertheless, significant differences in the PMRNs among the food levels remained after accounting for the influences of age, size and condition on maturation probability indicating plasticity of the PMRN. This was particularly pronounced between fish held on low food levels as compared with fish experiencing abundant resources, with the latter experiencing higher size-specific maturation probabilities. Our analysis emphasizes the need for incorporating salient physiological traits influencing maturation, such as condition, to make accurate inferences about documented shifts observed in the position of PMRNs on maturation trends in wild fish stocks.     

Chromosomal regions in prostatic carcinomas studied by comparative genomic hybridization, hierarchical cluster analysis and self-organizing feature maps.

Comparative genomic hybridization (CGH) is an established genetic method which enables a genome-wide survey of chromosomal imbalances. For each chromosome region, one obtains the information whether there is a loss or gain of genetic material, or whether there is no change at that place. Therefore, large amounts of data quickly accumulate which must be put into a logical order. Cluster analysis can be used to assign individual cases (samples) to different clusters of cases, which are similar and where each cluster may be related to a different tumour biology. Another approach consists in a clustering of chromosomal regions by rewriting the original data matrix, where the cases are written as rows and the chromosomal regions as columns, in a transposed form. In this paper we applied hierarchical cluster analysis as well as two implementations of self-organizing feature maps as classical and neuronal tools for cluster analysis of CGH data from prostatic carcinomas to such transposed data sets. Self-organizing maps are artificial neural networks with the capability to form clusters on the basis of an unsupervised learning rule. We studied a group of 48 cases of incidental carcinomas, a tumour category which has not been evaluated by CGH before. In addition we studied a group of 50 cases of pT2N0-tumours and a group of 20 pT3N0-carcinomas. The results show in all case groups three clusters of chromosomal regions, which are (i) normal or minimally affected by losses and gains, (ii) regions with many losses and few gains and (iii) regions with many gains and few losses. Moreover, for the pT2N0- and pT3N0-groups, it could be shown that the regions 6q, 8p and 13q lay all on the same cluster (associated with losses), and that the regions 9q and 20q belonged to the same cluster (associated with gains). For the incidental cancers such clear correlations could not be demonstrated.     

Prevention of human PC-346C prostate cancer growth in mice by a xenogeneic tissue vaccine

Vaccination, as an approach to prostate cancer, has largely focused on immunotherapy utilizing specific molecules or allogeneic cells. Such methods are limited by the focused antigenic menu presented to the immune system and by immunotolerance to antigens recognized as “self”. To examine if a xenogeneic tissue vaccine could stimulate protective immunity in a human prostate cancer cell line, a vaccine was produced by glutaraldehyde fixation of harvested PAIII prostate cancer cells tumors (GFT cell vaccine) from Lobund-Wistar rats. Immunocompetent Ncr-Foxn1<nu> mice were vaccinated with the GFT cell vaccine four times, 7 days apart. The control animals were either not vaccinated or vaccinated with media or glutaraldehyde-fixed PC346C human prostate cancer cells and adjuvant. About 8 days after the final boost, serum and spleens were harvested. The splenocytes were co-incubated with PC346C cells and then transplanted orthotopically into sygneneic immunodeficient nude mice. About 10 weeks later, the prostates were weighed and sampled for histolologic examination. The spleens were harvested from additional mice, and the splenocytes were cultured, either with or without pulsing by GFT cells, and the supernatants harvested 72 h later for cytokine analysis. Results showed that vaccination with GFT cells resulted in increased serum antibody to a PAIII cell lysate; reduced weight of the prostate/seminal vesicle complex and reduced incidence of prostate cancer in nude mice; increased splenocyte supernatant levels of TNF-α, IL-2, IFN-γ and IL-12, cytokines associated with Th1 immunity; and increased splenocyte supernatant levels of IL-4 and IL-10, cytokines associated with Th2 immunity. In summary, the results suggest that use of a xenogeneic tissue vaccine can stimulate protective immunity against human prostate cancer cells.     

Vaccination against prostate cancer using a live tissue factor deficient cell line in Lobund–Wistar rats

Reducing expression of the tissue factor gene in prostate adenocarcinoma cells (PAIII) results in a cell line that, in vivo, mimics the growth of wildtype (wt) PAIII. However, instead of continuing to grow and metastasize as wt PAIII tumors do, tissue factor deficient PAIII (TFD PAIII) masses spontaneously regress after several weeks. Although whole cell vaccines are typically inactivated prior to administration to prevent proliferation within the host, numerous studies have suggested that exposure to live, attenuated, whole tumor cells, and the extracellular microenvironment they recruit, increases immunotherapeutic potential. Here, we provide support for this notion, and a strategy through which to implement it, by demonstrating that subcutaneous vaccinations with the TFD PAIII protect the Lobund–Wistar rat against subsequent wt PAIII cell challenge. TFD PAIII immunized rats suffered significantly less metastasis of wt PAIII challenge tumors compared to unvaccinated naïve controls rats. These results offer the intriguing possibility that the TFD PAIII vaccine is an effective system for the prevention and, possibly, the treatment of prostate cancer.     

Human and Extracellular DNA Depletion for Metagenomic Analysis of Complex Clinical Infection Samples Yields Optimized Viable Microbiome Profiles

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Female,but not male,nematodes evolve under experimental sexual coevolution

Coevolution between the sexes is often considered to be male-driven: the male genome is constantly scanned by selection for traits that increase relative male fertilization success. Whenever these traits are harmful to females, the female genome is scanned for resistance traits. The resulting antagonistic coevolution between the sexes is analogous to Red Queen dynamics, where adaptation and counteradaptation keep each other in check. However, the underlying assumption that male trait evolution precedes female trait counteradaptation has received few empirical tests. Using the gonochoristic nematode Caenorhabditis remanei, we now show that 20 generations of relaxed versus increased sexual selection pressure lead to female, but not to male, trait evolution, questioning the generality of a male-driven process.