H2S biosynthesis and catabolism: new insights from molecular studies

Hydrogen sulfide (H2S) has profound biological effects within living organisms and is now increasingly being considered alongside other gaseous signalling molecules, such as nitric oxide (NO) and carbon monoxide (CO). Conventional use of pharmacological and molecular approaches has spawned a rapidly growing research field that has identified H2S as playing a functional role in cell-signalling and post-translational modifications. Recently, a number of laboratories have reported the use of siRNA methodologies and genetic mouse models to mimic the loss of function of genes involved in the biosynthesis and degradation of H2S within tissues. Studies utilising these systems are revealing new insights into the biology of H2S within the cardiovascular system, inflammatory disease, and in cell signalling. In light of this work, the current review will describe recent advances in H2S research made possible by the use of molecular approaches and genetic mouse models with perturbed capacities to generate or detoxify physiological levels of H2S gas within tissue

Aggregation induced nucleic acids recognition by homodimeric asymmetric monomethyne cyanine fluorochromes in mesenchymal stem cells

In the light of recent retrovirus pandemics, the issue of discovering new and diverse RNA-specific fluorochromes for research and diagnostics became of acute importance. The great majority of nucleic acid-specific probes either do not stain RNA or cannot distinguish between DNA and RNA. The versatility of polymethine dyes makes them suitable as stains for visualization, analysis, and detection of nucleic acids, proteins, and other biomolecules. We synthesized the asymmetric dicationic homodimeric monomethine cyanine dyes 1,1′-(1,3-phenylenebis(methylene))bis(4-((3-methylbenzo[d]thiazol-2(3H)-ylidene)methyl)pyridin-1-ium) bromide (Т1) and 1,1′-(1,3-phenylenebis(methylene))bis(4-((3-methylbenzo[d]thiazol-2(3H)-ylidene)methyl)quinolin-1-ium) bromide (M1) and tested their binding specificity, spectral characteristics, membrane penetration in living and fixed cells, cellular toxicity, and stability of fluorescent emission. Mesenchymal cells have diverse phenotypes and extensive proliferation and differentiation properties. We found dyes T1 and M1 to show high photochemical stability in living mesenchymal stem cells from apical papilla (SCAP) with a strong fluorescent signal when bound to nucleic acids. We found M1 to perform better than control fluorochrome (Hoechst 33342) for in vivo DNA visualization. T1, on the other hand, stains granular cellular structures resembling ribosomes in living cells and after permeabilization of the nuclear membrane stains the nucleoli and not the chromatin in the nucleus. This makes T1 suitable for the visualization of structures rich in RNA in living and fixed cells

Quantitative analysis of volatile organic compounds released and consumed by rat L6 skeletal muscle cells in vitro

Knowledge of the release of volatile organic compounds (VOCs) by cells provides important information on the origin of VOCs in exhaled breath. Muscle cells are particularly important, since their release of volatiles during the exertion of an effort contributes considerably to breath concentration profiles. Presently, the cultivation of human skeletal muscle cells is encountering a number of obstacles, necessitating the use of animal muscle cells in in vitro studies. Rat L6 skeletal muscle cells are therefore commonly used as a model for studying the molecular mechanisms of human skeletal muscle differentiation and functions, and facilitate the study of the origin and metabolic fate of the endogenously produced compounds observed in breath and skin emanations. Within this study the production and uptake of VOCs by rat L6 skeletal muscle cells were investigated using gas chromatography with mass spectrometric detection, combined with head-space needle trap extraction as the pre-concentration technique (HS-NTE-GC-MS). Seven compounds were found to be produced, whereas sixteen species were consumed (Wilcoxon signed-rank test, p < 0.05) by the cells being studied. The set of released volatiles included two ketones (2-pentanone and 2-nonanone), two volatile sulphur compounds (dimethyl sulfide and methyl 5-methyl-2-furyl sulphide), and three hydrocarbons (2-methyl 1-propene, n-pentane and isoprene). Of the metabolized species there were thirteen aldehydes (2-propenal, 2-methyl 2-propenal, 2-methyl propanal, 2-butenal, 2-methyl butanal, 3-methyl butanal, n-pentanal, 2-methyl 2-butenal, n-hexanal, benzaldehyde, n-octanal, n-nonanal and n-decanal), two esters (n-propyl propionate and n-butyl acetate), and one volatile sulphur compound (dimethyl disulfide). The possible metabolic pathways leading to the uptake and release of these compounds by L6 cells are proposed and discussed. An analysis of the VOCs showed them to have huge potential for the identification and monitoring of some molecular mechanism and conditions