Simultaneous quantitative LC-MS method of ketamine, midazolam and their metabolites (dehydronorketamine, norketamine and 1hydroxymidazolam) for its application in patients on extracorporeal membrane oxygenation (ECMO) therapy.
氯胺酮、咪达唑仑及其代谢产物 (去氢氯胺酮、去甲氯胺酮和 1 羟基咪达唑仑) 的同时定量 LC-MS 方法，用于体外膜肺氧合 (ECMO) 治疗患者。
- 作者列表："Rochani A","Lam E","Tanjuakio J","Hirose H","Kraft WK","Kaushal G
:Ketamine (Ket) and midazolam (MDZ) are commonly administered drugs in the intensive care setting for analgesia and sedation. Ket and MDZ are metabolized to dehydro-norketamine (DHNK), nor-ketamine (NK) and 1-hydroxy midazolam (1HMDZ). Limited studies evaluating their pharmacokinetics exists in patients on extracorporeal membrane oxygenation (ECMO) therapy. Therefore, we developed a quantitative, high-performance liquid chromatography-mass spectrometry (with single ion monitoring) method to simultaneously detect Ket, MDZ and their (DHNK, NK and 1HMDZ) metabolites in human plasma. Considerable sensitivity was obtained for the analytes using a C18 HILIC column operated by a high-performance liquid chromatography system coupled with a Thermo Exactive Orbitrap mass spectrometer. Calibration curves were developed for analyte molecules (n = 5) in the presence of carbamazepine (CBZ) as an internal standard. The lower limits of quantitation (LLOQ) for Ket and MDZ were 20 and 10 ng/mL, respectively with the LLOQ for DHNK, NK and 1HMDZ at 470, 320 and 150 ng/ml. Moreover, the percent coefficient of variance and precision for inter- and intra-day runs were within the standards set forth by the ICH and FDA guidelines. This method is sensitive and has been successfully applied to an ongoing pharmacokinetic study in patients on ECMO therapy.
: 氯胺酮 (Ket) 和咪达唑仑 (MDZ) 是重症监护环境中常用的镇痛和镇静药物。Ket 和 MDZ 代谢为去氢去甲氯胺酮 (DHNK) 、 nor-氯胺酮 (NK) 和 1-羟基咪达唑仑 (1HMDZ)。在体外膜肺氧合 (ECMO) 治疗的患者中存在评价其药代动力学的有限研究。因此，我们开发了一种定量、高效液相色谱-质谱 (单离子监测) 方法，同时检测 Ket 、 MDZ 及其 (DHNK 、 NK 和 1HMDZ) 人血浆中的代谢物。使用 C18 HILIC 柱，通过高效液相色谱系统与热外露 Orbitrap 质谱仪联用，获得了相当高的分析物灵敏度。在卡马西平 (CBZ) 作为内标物存在下，对分析物分子 (n = 5) 建立校准曲线。Ket 和 MDZ 的定量下限 (LLOQ) 分别为 20 和 10 ng/mL，DHNK 、 NK 和 1HMDZ 的 LLOQ 分别为 470 、 320 和 150 ng/ml。此外，日间和日内运行的变异百分比系数和精密度均在 ICH 和 FDA 指南规定的标准范围内。该方法灵敏，已成功应用于 ECMO 治疗患者的药代动力学研究。
METHODS:BACKGROUND AND PURPOSE:A critical role for sphingosine kinase/sphingosine-1-phosphate (S1P) pathway in the control of airway function has been demonstrated in respiratory diseases. Here, we address S1P contribution in a mouse model of mild chronic obstructive pulmonary disease (COPD). EXPERIMENTAL APPROACH:C57BL/6J mice have been exposed to room air or cigarette smoke up to 11 months and killed at different time points. Functional and molecular studies have been performed. KEY RESULTS:Cigarette smoke caused emphysematous changes throughout the lung parenchyma coupled to a progressive collagen deposition in both peribronchiolar and peribronchial areas. The high and low airways showed an increased reactivity to cholinergic stimulation and α-smooth muscle actin overexpression. Similarly, an increase in airway reactivity and lung resistances following S1P challenge occurred in smoking mice. A high expression of S1P, Sph-K2 , and S1P receptors (S1P2 and S1P3 ) has been detected in the lung of smoking mice. Sphingosine kinases inhibition reversed the increased cholinergic response in airways of smoking mice. CONCLUSIONS AND IMPLICATIONS:S1P signalling up-regulation follows the disease progression in smoking mice and is involved in the development of airway hyperresponsiveness. Our study defines a therapeutic potential for S1P inhibitors in management of airways hyperresponsiveness associated to emphysema in smokers with both asthma and COPD.
METHODS::The interim results from this 90-day multi-dose, inhalation toxicology study with life-time post-exposure observation has shown an important fundamental difference in persistence and pathological response in the lung between brake dust derived from brake-pads manufactured with chrysotile, TiO2 or chrysotile alone in comparison to the amphiboles, crocidolite and amosite asbestos. In the brake dust exposure groups no significant pathological response was observed at any time. Slight macrophage accumulation of particles was noted. Wagner-scores, were from 1 to 2 (1 = air-control group) and were similar to the TiO2 group. Chrysotile being biodegradable, shows a weakening of its matrix and breaking into short fibers & particles that can be cleared by alveolar macrophages and continued dissolution. In the chrysotile exposure groups, particle laden macrophage accumulation was noted leading to a slight interstitial inflammatory response (Wagner-score 1-3). There was no peribronchiolar inflammation and occasional very slight interstitial fibrosis. The histopathology and the confocal analyses clearly differentiate the pathological response from amphibole asbestos, crocidolite and amosite, compared to that from the brake dust and chrysotile. Both crocidolite and amosite induced persistent inflammation, microgranulomas, and fibrosis (Wagner-scores 4), which persisted through the post exposure period. The confocal microscopy of the lung and snap-frozen chestwalls quantified the extensive inflammatory response and collagen development in the lung and on the visceral and parietal surfaces. The interim results reported here, provide a clear basis for differentiating the effects from brake dust exposure from those following amphibole asbestos exposure. The subsequent results through life-time post-exposure will follow.
METHODS::The respiratory tract is lined by a pseudo-stratified epithelium from the nose to terminal bronchioles. This first line of defense of the lung against external stress includes five main cell types: basal, suprabasal, club, goblet and multiciliated cells, as well as rare cells such as ionocytes, neuroendocrine and tuft/brush cells. At homeostasis, this epithelium self-renews at low rate but is able of fast regeneration upon damage. Airway epithelial cell lineages during regeneration have been investigated in the mouse by genetic labeling, mainly after injuring the epithelium with noxious agents. From these approaches, basal cells have been identified as progenitors of club, goblet and multiciliated cells, but also of ionocytes and neuroendocrine cells. Single-cell RNA sequencing, coupled to lineage inference algorithms, has independently allowed the establishment of comprehensive pictures of cell lineage relationships in both mouse and human. In line with genetic tracing experiments in mouse trachea, studies using single-cell RNA sequencing (RNAseq) have shown that basal cells first differentiate into club cells, which in turn mature into goblet cells or differentiate into multiciliated cells. In the human airway epithelium, single-cell RNAseq has identified novel intermediate populations such as deuterosomal cells, 'hybrid' mucous-multiciliated cells and progenitors of rare cells. Novel differentiation dynamics, such as a transition from goblet to multiciliated cells have also been discovered. The future of cell lineage relationships in the respiratory tract now resides in the combination of genetic labeling approaches with single-cell RNAseq to establish, in a definitive manner, the hallmarks of cellular lineages in normal and pathological situations.