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Bone Regeneration Effect of Cassia occidentalis Linn. Extract and Its Isolated Compounds
Published in Brijesh Kumar, Vikas Bajpai, Vikaskumar Gond, Subhashis Pal, Naibedya Chattopadhyay, Phytochemistry of Plants of Genus Cassia, 2021
Brijesh Kumar, Vikas Bajpai, Vikaskumar Gond, Subhashis Pal, Naibedya Chattopadhyay
Caryophyllenes are natural sesquiterpenes present in the essential oils of many medicinal plants including CO extract. In MSCs derived from mouse bone marrow, β-caryophyllene between 0.1 and 100 μM range promotes osteoblastic differentiation but inhibits adipocytic and osteoclastic differentiation (Yamaguchi and Levy, 2016). In MC3T3-E1 cells, trans-caryophyllene promotes differentiation assessed by ALP activity, collagen production and osteocalcin release, and these effects were mediated by CB2R, a cannabinoid receptor isoform. Trans-caryophyllene also protected cells against antimycin-induced loss viability of MC3T3-E1 cells likely due to the reduction of ROS production due to the activation of anti-oxidant machinery including increased GSH levels and catalase activity. Furthermore, the endogenous antioxidant stress protectant proteins including Nrf2 and heme oxygenase 1 (HO-1) that were downregulated by antimycin A were maintained in MC3T3-E1 cells with trans-caryophyllene co-treatment (Shan et al., 2017).
Di-Calciphor-Dependent Protection Against Cell Death Due to Mitochondrial Failure
Published in John J. Lemasters, Constance Oliver, Cell Biology of Trauma, 2020
Measurement of mitochondrial membrane potential and ΔpH showed that antimycin A caused a loss in these values and that di-Calciphor protected against their loss. di-Calciphor also prevented mitochondrial phosphate loading and mitochondrial swelling. Thus, the effects of di-Calciphor on antimycin A-induced mitochondrial failure and cell death suggest that di-Calciphor is a specific mitochondrial protective agent.
High-Performance Liquid Chromatography
Published in Adorjan Aszalos, Modern Analysis of Antibiotics, 2020
Joel J. Kirschbaum, Adorjan Aszalos
Antimycin A in tissues was chromatographed using a 5-µm octadecylsilane column and precolumn (50 x 4 mm) and a mobile phase of 0.25 M acetate buffer, pH 5-methanol (25:75). Detection of this antifungal agent was either electrochemical (using a glass carbon electrode at +1.00 V versus Ag/AgCl), fluorescence at 365/418 (after derivatization with dimethylaminonaphthalene-5-sulfonyl chloride, or UV at 254 nm. The respective minimum detectable quantities were approximately 0.5, 0.5, and 5 ng [424].
Pre-imaginal exposure to mancozeb induces morphological and behavioral deficits and oxidative damage in Drosophila melanogaster
Published in Drug and Chemical Toxicology, 2023
Cynthia Camila Ziech, Nathane Rosa Rodrigues, Giulianna Echeverria Macedo, Karen Kich Gomes, Illana Kemmerich Martins, Jeferson Luis Franco, Thaís Posser
After signal stabilization, the basal respiration supported by endogenous substrates was evaluated. The Complex I (CI)-mediated leak (LEAK) respiration was determined using 5 mM pyruvate, 5 mM L-proline, and 1 mM malate. CI-mediated OXPHOS (OXPHOS) was determined using ADP (2.5 mM). The convergent electron flow during the maximal OXPHOS respiration (CIc + CIIOXPHOS) was evaluated with substrates of CIc and CII (10 mM succinate). The ETS respiration represents the not coupled respiration using FCCP (optimum concentration reached between 0.5 and 1.5 µM); CIc + CII-mediated ETS respiration (CIc + CIIets) was determined using FCCP (optimum concentration reached between 0.5 and 1.5 µM). CII-mediated E.T.S. respiration (CIIETS) was determined with 0.5 µM rotenone. The addition of 2.5 µM antimycin A inhibited Complex III, resulting in non-mitochondrial respiration, the residual oxygen consumption (ROX) with minor contributions from electron leak in the uncoupled state. Respiration of CIV was determined by adding ascorbate 0.5 mM and TMPD 1 mM. For HRR, three independent mitochondrial preparations from each experimental group were used (n = 3). All HRR experiments were performed in duplicates.
scRNA-seq reveals ATPIF1 activity in control of T cell antitumor activity
Published in OncoImmunology, 2022
Genshen Zhong, Qi Wang, Ying Wang, Ying Guo, Meiqi Xu, Yaya Guan, Xiaoying Zhang, Minna Wu, Zhishan Xu, Weidong Zhao, Hongkai Lian, Hui Wang, Jianping Ye
CD8+ T cells were isolated from the mouse spleen and activated with CD3/CD28 antibody stimulation in the culture medium. The cells (or the CAR-T cells) were loaded at 1 × 106/well into XF24 plate, which was coated with Cell-Tak (company name, 22.4 μg/mL, in sterile water) for 20 minutes to increase the adhesion of T cells. The plate was centrifuged at 200 × g (zero braking) for 1 minute to let the T cells to adhere to the culture surface. After incubation for 30 minutes at 37°C without CO2 supplementation, Oxygen Consumption Rate (OCR) and Extracellular Acidification Rate (ECAR) were determined with the Seahorse XF Cell Mito Stress Test Kit and Seahorse XF Glycolysis Stress Test kit with the Agilent Technologies equipment. The final concentrations of inhibitors were 1 μM oligomycin, 2 μM FCCP, 0.5 μM rotenone, and antimycin A. In the ECAR assay, the final concentrations of compounds were 10 mM glucose, 1 μM oligomycin, and 50 mM 2-Deoxy-D-glucose. The readings were taken after each sequential injection of corresponding chemicals.
Effects of different standard and special diets on cognition and brain mitochondrial function in mice
Published in Nutritional Neuroscience, 2022
Martina Reutzel, Rekha Grewal, Carsten Esselun, Sebastian Friedrich Petry, Thomas Linn, Annette Brandt, Ina Bergheim, Gunter P. Eckert
Half a brain hemisphere (the frontal part) was used to isolate brain mitochondria. The protocol is described in Hagl et al. [16]. The pellet obtained from the last centrifugation step was dissolved in 250 µl MIRO5. A volume of 80 µl of the resulting cell suspension was injected into the Oxygraph 2k-chamber. The capacity of the oxidative phosphorylation (OXPHOS) was determined using complex-I related substrates pyruvate (5 mM) and malate (2 mM) and ADP (2 mM) followed by the addition of succinate (10 mM). Mitochondrial integrity was measured by the addition of cyctochrom c (10 µM). Oligomycin (2 µg/ml) was added to determine leak respiration (leak (omy)) and afterwards uncoupling was achieved by carbonyl cyanide p-(trifluoromethoxy) phenyl-hydrazone (FCCP, injected stepwise up to 1–1.5 µM). Complex II respiration was measured after the addition of rotenone (0.5 µM). Complex III inhibition was achieved by the addition of antimycin A (2.5 µM) and was subtracted from all respiratory parameters. Cytochrome c oxidase (COX) activity was determined after residual oxygen consumption (ROX) determination by applying 0.5 mM tetramethylphenylenediamine (TMPD) as an artificial substrate of complex IV and 2 mM ascorbate to keep TMPD in the reduced state. Autoxidation rate was determined after the addition of sodium azide (>100 mM), and COX respiration was additionally corrected for autoxidation.