Label-Free Mass Spectrometry-Based Quantitative Proteomics to Evaluate the Effects of the Calcium-Sensing Receptor Agonist Cinacalcet on Protein Expression in Rat Brains and Livers.

BACKGROUND Cinacalcet is a calcium-sensing receptor agonist that is clinically approved for the treatment of secondary hyperparathyroidism in chronic kidney disease and hypercalcemia in patients with parathyroid carcinoma. This study aimed to use quantitative mass spectrometry-based label-free proteomics to evaluate the effects of cinacalcet on protein expression in rat brains and livers. MATERIAL AND METHODS We randomly assigned 18 Wistar rats to 2 groups: an untreated control group (n=6) and a group treated with cinacalcet at a dose corresponding to the maximum dose used in humans (2 mg/kg/body weight, 5 days/week) divided into 7-day (n=6) and 21-day (n=6) treatment subgroups. A mass-spectrometry-based label-free quantitative proteomics approach using peptides peak area calculation was used to evaluate the changes in protein expression in examined tissues. Bioinformatics analysis of quantitative proteomics data was done using MaxQuant and Perseus environment. RESULTS No changes in protein expression were revealed in the 7-day treatment subgroup. We detected 10 upregulated and 3 downregulated proteins in the liver and 1 upregulated protein in the brain in the 21-day treatment subgroup compared to the control group. Based on Gene Ontology classification, all identified differentially expressed proteins were indicated as molecular functions involved in the enzyme regulator activity (36%), binding (31%), and catalytic activity (19%). CONCLUSIONS These findings indicate that long-term cinacalcet therapy can impair phase II of enzymatic detoxication and can cause disturbances in blood hemostasis, lipid metabolism, and inflammatory mediators or contribute to the acceleration of cognitive dysfunction; therefore, appropriate patient monitoring should be considered.

Label-Free Mass Spectrometry-Based Proteomic Analysis in Lamb Tissues after Fish Oil, Carnosic Acid, and Inorganic Selenium Supplementation.

Selenium is an essential nutrient, building twenty five identified selenoproteins in humans known to perform several important biological functions. The small amount of selenium in the earth's crust in certain regions along with the risk of deficiency in organisms have resulted in increasingly popular dietary supplementation in animals, implemented via, e.g., inorganic selenium compounds. Even though selenium is included in selenoproteins in the form of selenocysteine, the dietary effect of selenium may result in the expression of other proteins or genes. Very little is known about the expression effects modulated by selenium. The present study aimed to examine the significance of protein expression in lamb tissues obtained after dietary supplementation with selenium (sodium selenate) and two other feed additives, fish oil and carnosic acid. Label-free mass spectrometry-based proteomic analysis was successfully applied to examine the animal tissues. Protein-protein interaction network analysis of forty differently-expressed proteins following inorganic selenium supplementation indicated two significant clusters which are involved in cell adhesion, heart development, actin filament-based movement, plasma membrane repair, and establishment of organelle localization.

Fluorine-Containing Drug Administration in Rats Results in Fluorination of Selected Proteins in Liver and Brain Tissue.

In many pharmaceuticals, a hydrogen atom or hydroxyl group is replaced by a fluorine to increase bioavailability and biostability. The fate of fluorine released from fluorine-containing drugs is not well investigated. The aim of this study was to examine possible fluorination of proteins in rat liver and brain after administration of the fluorinated drug cinacalcet. We assigned 18 Wistar rats to a control group (n = 6) and a group treated with cinacalcet (2 mg kg-1/body weight, 5 days/week), divided into 7 day (n = 6) and 21 day (n = 6) treatment subgroups. Fluorinated proteins were identified using a free proteomics approach; chromatographic separation and analysis by high-resolution mass spectrometry; peptide/protein identification using the Mascot search algorithm; manual verification of an experimentally generated MS/MS spectrum with the theoretical MS/MS spectrum of identified fluorinated peptides. Three fluorinated proteins (spectrin beta chain; carbamoyl-phosphate synthase [ammonia], mitochondrial; 6-phosphofructo-2-kinase/fructose-2, 6-bisphosphatase 1) were identified in the liver and four (spectrin beta chain, dihydropyrimidinase-related protein 4, prominin-2, dihydropyrimidinase-related protein 4) in the brain tissue after 21 days of cinacalcet treatment, but not in the control group. Introduction of fluorine into an organism by administration of fluorinated drugs results in tissue-specific fluorination of proteins.