H2S is an important signalling molecule involved in diverse biological
processes. It mediates the formation of cysteine persulfides (R-S-SH), which affect
the activity of target proteins. Like thiols, persulfides show reactivity towards
electrophiles and behave similarly to other cysteine modifications in a biotin
switch assay. In this manuscript, we report on qPerS-SID a mass spectrometry-based
method allowing the isolation of persulfide containing peptides in the mammalian
proteome. With this method, we demonstrated that H2S donors differ in
their efficacy to induce persulfides in HEK293 cells. Furthermore, data analysis
revealed that persulfide formation affects all subcellular compartments and various
cellular processes. Negatively charged amino acids appeared more frequently adjacent
to cysteines forming persulfides. We confirmed our proteomic data using pyruvate
kinase M2 as a model protein and showed that several cysteine residues are prone to
persulfide formation finally leading to its inactivation. Taken together, the
site-specific identification of persulfides on a proteome scale can help to identify
target proteins involved in H2S signalling and enlightens the biology of
H2S and its releasing agents.
Despite the efficacy of neuroprotective approaches in animal models of stroke, their translation has so far failed from bench to bedside. One reason is presumed to be a low quality of preclinical study design, leading to bias and a low a priori power. In this study, we propose that the key read-out of experimental stroke studies, the volume of the ischemic damage as commonly measured by free-handed planimetry of TTC-stained brain sections, is subject to an unrecognized low inter-rater and test-retest reliability with strong implications for statistical power and bias. As an alternative approach, we suggest a simple, open-source, software-assisted method, taking advantage of automatic-thresholding techniques. The validity and the improvement of reliability by an automated method to tMCAO infarct volumetry are demonstrated. In addition, we show the probable consequences of increased reliability for precision, p-values, effect inflation, and power calculation, exemplified by a systematic analysis of experimental stroke studies published in the year 2015. Our study reveals an underappreciated quality problem in translational stroke research and suggests that software-assisted infarct volumetry might help to improve reproducibility and therefore the robustness of bench to bedside translation.
PHD3 expression induced by cytokines is NF-κB dependent in mesangial cells. Endogenously produced NO further augments PHD3 expression via HIF-1α. PHD3 expression is induced by NO in anti-Thy-1 glomerulonephritis.
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