Tyrosine phosphorylation of the N-Methyl-D-Aspartate receptor 2B subunit in spinal cord contributes to remifentanil-induced postoperative hyperalgesia: the preventive effect of ketamine

<p>Abstract</p> <p>Background</p> <p>Experimental and clinical studies showed that intraoperative infusionof remifentanil has been associated with postoperative hyperalgesia. Previous reports suggested that spinal N-methyl-D-aspartate (NMDA) receptors may contribute to the development and maintenance of opioid-induced hyperalgesia. In the present study, we used a rat model of postoperative pain to investigate the role of tyrosine phosphorylation of NMDA receptor 2B (NR2B) subunit in spinal cord in the postoperative hyperalgesia induced by remifentanil and the intervention of pretreatment with ketamine.</p> <p>Results</p> <p>Intraoperative infusion of remifentanil (0.04 mg/kg, subcutaneous) significantly enhanced mechanical allodynia and thermal hyperalgesia induced by the plantar incision during the postoperative period (each lasting between 2 h and 48 h), which was attenuated by pretreatment with ketamine (10 mg/kg, subcutaneous). Correlated with the pain behavior changes, immunocytochemical and western blotting experiments in our study revealed that there was a marked increase in NR2B phosphorylation at Tyr1472 in the superficial dorsal horn after intraoperative infusion of remifentanil, which was attenuated by pretreatment with ketamine.</p> <p>Conclusions</p> <p>This study provides direct evidence that tyrosine phosphorylation of the NR2B at Tyr1472 in spinal dosal horn contributes to postoperative hyperalgesia induced by remifentanil and supports the potential therapeutic value of ketamine for improving postoperative hyperalgesia induced by remifentanil.</p

Enhanced Enzymatic Reactivity for Electrochemically Driven Drug Metabolism by Confining Cytochrome P450 Enzyme in TiO₂ Nanotube Arrays

Understanding the enzymatic reaction kinetics that occur within a confined space or interface is a significant challenge. Herein, a nanotube array enzymatic reactor (CYP2C9/Au/TNA) was constructed by electrostatically adsorbing enzyme on the inner wall of TiO₂ nanotube arrays (TNAs). TNAs with different dimensions could be fabricated by the anodization of titanium foil through varying the anodization potential or time. The electrical conductivity of TNAs was improved by electrodepositing Au nanoparticles on the inner wall of TNAs. The cytochrome P450 2C9 enzyme (CYP2C9) was confined inside TNAs as a model. The enzymatic activity of CYP2C9 and tolbutamide metabolic yield could be effectively regulated by changing the nanotube diameter and length of TNAs. The enzymatic rate constant <i>k</i>_cat and apparent Michaelis constant <i>K</i>_m^app were determined to be 9.89 s^–1 and 4.8 μM at the tube inner diameter of about 64 nm and length of 1.08 μm. The highest metabolic yield of tolbutamide reached 14.6%. Furthermore, the designed nanotube array enzymatic reactor could be also used in situ to monitor the tolbutamide concentration with sensitivity of 28.8 μA mM^–1 and detection limit of 0.52 μM. Therefore, the proposed nanotube array enzymatic reactor was a good vessel for studying enzyme biocatalysis and drug metabolism, and has potential applications including efficient biosensors and bioreactors for chemical synthesis

Menthol in Electronic Cigarettes: A Contributor to Respiratory Disease?

Menthol is widely used in tobacco products. This study compared the effects of menthol on human bronchial epithelium using submerged cultures, a VITROCELL® cloud chamber that provides air liquid interface (ALI) exposure without solvents or heating, and a Cultex ALI system that delivers aerosol equivalent to that inhaled during vaping. In submerged culture, menthol significantly increased calcium influx and mitochondrial reactive oxygen species (ROS) via the TRPM8 receptor, responses that were inhibited by a TRPM8 antagonist. VITROCELL® cloud chamber exposure of BEAS-2B monolayers increased mitochondrial protein oxidation, expression of the antioxidant enzyme SOD2, activation of NF-κB, and secretion of inflammatory cytokines (IL-6 and IL-8). Proteomics data collected following ALI exposure of 3D EpiAirway tissue in the Cultex showed upregulation of NRF-2-mediated oxidative stress, oxidative phosphorylation, and IL-8 signaling. Across the three platforms, menthol adversely effected human bronchial epithelium in a manner that could lead to respiratory disease

Nanocomposites of Graphene and Cytochrome P450 2D6 Isozyme for Electrochemical-Driven Tramadol Metabolism

Cytochrome P450 enzymes (cyt P450s) with an active center of iron protoheme are involved in most clinical drugs metabolism process. Herein, an electrochemical platform for the investigation of drug metabolism <i>in vitro</i> was constructed by immobilizing cytochrome P450 2D6 (CYP2D6) with cyt P450 reductase (CPR) on graphene modified glass carbon electrode. Direct and reversible electron transfer of the immobilized CYP2D6 with the direct electron transfer constant of 0.47 s^–1 and midpoint potential of -0.483 V was obtained. In the presence of substrate tramadol, the electrochemical-driven CYP2D6 mediated catalytic behavior toward the conversion of tramadol to <i>o</i>-demethyl-tramadol was confirmed. The Michaelis–Menten constant (<i>K</i>_m^app) and heterogeneous reaction rate constant during the metabolism of tramadol were calculated to be 23.85 μM and 1.96 cm s^–1, respectively. The inhibition effect of quinidine on CYP2D6 catalyze-cycle was also investigated. Furthermore, this system was applied to studying the metabolism of other drugs

Label-Free Photoelectrochemical Immunosensor for Neutrophil Gelatinase-Associated Lipocalin Based on the Use of Nanobodies

Acute renal failure (ARF) represents a very important and potentially devastating disorder in clinical nephrology. Neutrophil gelatinase-associated lipocalin (NGAL) is an early biomarker for ARF in a wide range of different disease processes, which is frequently detected in clinical diagnosis. Herein, we present a label-free and sensitive photoelectrochemical (PEC) immunosensor for NGAL by utilizing a biotinylated anti-NGAL Nanobody (Nb) orientedly immobilized to streptavidin-coated cobalt 2,9,16,23-tetraaminophthalocyanine (CoPc)-sensitized TiO₂ electrode. The Nb was biotinylated at the C-terminus, which is situated at the opposite site of the antigen binding region. Using highly oriented Nb as receptor molecules, a label-free PEC immunosensor for NGAL was developed by monitoring the changes in the photocurrent signals of the electrode resulting from immunoreaction. Immobilization of Nb to streptavidin-coated CoPc-sensitized TiO₂ electrode surface provides high binding capacity to NGAL; thus, it can lead to a high sensitivity. The limit of detection (LOD) of the proposed immunosensor has been significantly lowered to 0.6 pg mL^–1. This proposed immunosensor reveals high specificity to detect NGAL, with acceptable intra-assay precision and excellent stability. In addition, the present work provides a new approach to design Nb-based PEC immunosensor and increases versatility of Nbs

Menthol in electronic cigarettes: A contributor to respiratory disease?

Menthol is widely used in tobacco products. This study compared the effects of menthol on human bronchial epithelium using submerged cultures, a VITROCELL® cloud chamber that provides air liquid interface (ALI) exposure without solvents or heating, and a Cultex ALI system that delivers aerosol equivalent to that inhaled during vaping. In submerged culture, menthol significantly increased calcium influx and mitochondrial reactive oxygen species (ROS) via the TRPM8 receptor, responses that were inhibited by a TRPM8 antagonist. VITROCELL® cloud chamber exposure of BEAS-2B monolayers increased mitochondrial protein oxidation, expression of the antioxidant enzyme SOD2, activation of NF-κB, and secretion of inflammatory cytokines (IL-6 and IL-8). Proteomics data collected following ALI exposure of 3D EpiAirway tissue in the Cultex showed upregulation of NRF-2-mediated oxidative stress, oxidative phosphorylation, and IL-8 signaling. Across the three platforms, menthol adversely effected human bronchial epithelium in a manner that could lead to respiratory disease