As2O3 oxidation by vitamin C: cell culture studies

The ability of As₂O₃ to induce apoptosis in various malignant cell lines has made it a potential treatment agent for several malignancies. In this study the chemical stability of As₂O₃ (As(III)) in cell-free growth media with various compositions was studied (MEM with different amount of amino acids and DMEM). Special attention was given to evaluate the influence of serum (FBS; fetal bovine serum) absence and vitamin C addition on the oxidation of As(III) to As(V) in cell-free growth media. FBS is an important source of antioxidants and vitamin C (ascorbic acid) is acting as a prooxidant in millimolar concentrations. Media were incubated with As(III) (0.6, 2 and 7 μmol l⁻¹) up to 72 h. Experiments were performed at 37°C in light or/and in the dark, with or without added serum (10%) or vitamin C (1.4, 0.14 mM). Metabolites were followed with high-performance liquid chromatography directly coupled to a hydride generation-atomic fluorescence spectrometry system. After 72 h up to 30% of As(III) was transformed into As(V) in MEMs and up to 35% in DMEM when exposed in dark. Light had no influence on transformations in MEMs, but changed the situation dramatically in DMEM where almost all As(III) was oxidized to As(V) after 72 h when exposed to light. Except for some faster oxidation rate the absence of FBS had little effect on the transformation rate in all media. The most visible impact on As(III) oxidation was observed by addition of vitamin C. Addition of vitamin C (1.4 mM) transformed almost all As(III) to As(V) within 72 h. In lower concentrations (0.14 mM) a pro-oxidative effect was still observed reaching approximately 60% oxidation of As(III) during 72 h. All oxidation processes could be explained by pseudo first order reaction kinetics, yielding reaction rates increasing with initial As(III) concentration and vitamin C concentration whereas the FBS content additionally increased the As(III) oxidation rate in the DMEM (light). The temporal oxidation of As(III) to As(V) in various cell-free growth media necessitates routine checking of the valence state of arsenic during cell culture experiments and the results of biological effects attributed to As(III) should be interpreted with caution. Special attention is needed particularly in cases with vitamin C which was acting pro-oxidatively in all conditions examined.     

Quantitative Proteome Analysis Reveals RNA Processing Factors As Modulators of Ionizing Radiation-Induced Apoptosis in the C. elegans Germline

The nematode Caenorhabditis elegans is an organism most recognized for forward and reverse genetic and functional genomic approaches. Proteomic analyses of DNA damage-induced apoptosis have not been shown because of a limited number of cells undergoing apoptosis. We applied mass spectrometry-based quantitative proteomics to evaluate protein changes induced by ionizing radiation (IR) in isolated C. elegans germlines. For this purpose, we used isobaric peptide termini labeling (IPTL) combined with the data analysis tool IsobariQ, which utilizes MS/MS spectra for relative quantification of peak pairs formed during fragmentation. Using stringent statistical critera, we identified 48 proteins to be significantly up- or down-regulated, most of which are part of a highly interconnected protein-protein interaction network dominated by proteins involved in translational control. RNA-mediated depletion of a selection of the IR-regulated proteins revealed that the conserved CAR-1/CGH-1/CEY-3 germline RNP complex acts as a novel negative regulator of DNA-damage induced apoptosis. Finally, a central role of nucleolar proteins in orchestrating these responses was confirmed as the H/ACA snRNP protein GAR-1 was required for IR-induced apoptosis in the C. elegans germline.     

Hemochromatosis: As a conformational disorder

Hereditary hemochroamtosis (HH) refers to a unique clinicopathologic subset of iron overload syndromes that includes the disorder related to C282Y homozygous mutation of the hemochromatosis protein (HFE), the most common form of hereditary hemochromatosis. Recent reports have highlighted analogies with the class of disorders, known as the conformational diseases whereby HFE C282Y mutant protein forms aggregates and is subsequently retained in the endoplasmic reticulum (ER). In conformational disorders, accumulation of unfolded or misfolded proteins in the ER can activate a complex cascade linked to the regulation of diverse physiologic processes, disease onset and progression. To-date, reviews on HFE C282Y HH have largely dealt with the end-stage consequence of this disorder (iron overload). However, our review focuses on upstream molecular events resulting from the mislocalization of the aggregation-prone HFE C282Y protein leading to potential advances in treatment and diagnosis.