Elevated morphine concentrations determined during infant death investigations: artifacts of withdrawal of care

Abstract:  Three cases are reported of elevated postmortem blood morphine concentrations (189–3036 ng/mL) that were observed during the course of death investigations involving three children ranging in age from 1 week to 2 years, all of whom underwent withdrawal of life support. In all three cases, the presence of opiates in postmortem blood was indicated by immunoassay (ELISA) and quantitative confirmatory analysis of free morphine concentrations in postmortem blood was performed by solid-phase extraction followed by gas chromatography/mass spectrometry (GC/MS) in the selected ion monitoring mode. While the practice of withdrawing life support from terminally ill patients, with the accompanying administration of narcotics/analgesics has been reported in the medical literature, it has not been adequately described in the forensic literature. The implications of this practice on the forensic toxicological interpretation of morphine findings are discussed. To our knowledge, this is the first report of postmortem morphine concentrations arising directly from administration in conjunction with withdrawal of care in pediatric patients.     

Dominance of pre-analytical over analytical variation for measurement of methadone and its main metabolite in postmortem femoral blood

On the basis of simultaneously sampled postmortem blood specimens from the left and right femoral veins the pre-analytical variation of methadone measurements was evaluated and compared to the analytical variation. The material consisted of a series of 27 duplicate samples from routine autopsy cases comprising mainly drug addicts. A chiral LC-MS/MS method was used for measurement of the R- and S-enantiomers of methadone and its main metabolite 2-ethyl-1,5-dimethyl-3,3-diphenylpyrrolinium (EDDP). The analytical CV% was determined to be in the range 3-4% for methadone enantiomers and 4-6% for EDDP enantiomers. The total measurement uncertainty (CV(T)) was estimated from the pre-analytical variation (CV(PA)), analytical variation proper (CV(A)), and variation related to calibration (traceability) (CV(Cal)) according to the relationship CV(T) = [CV(2)(PA) + CV(2)(A) + CV(2)(cal)](0.5). Uncertainty related to calibration concerned a component related to the purity of drug reference compound and a contribution from the production of calibrator solutions (CV(Cal)<1%). Pre-analytical sampling variation was estimated from the duplicate measurements of blood samples after subtraction of the analytical component. The pre-analytical variation amounted to a CV% of 19-21% for R- and S-methadone and 30-38% for R- and S-EDDP, i.e. considerably larger than the other components. Due to the squared addition principle, the resulting total uncertainty (CV(T)) became largely identical to the CV(PA), i.e. 19-21% for R- and S-methadone and 31-38% for R- and S-EDDP enantiomers. Accordingly, CV(T) exceeded CV(A) by a factor 5 or more. Dominance of the pre-analytical component of variation may also be likely for other compounds measured in postmortem blood samples. Thus, the width of the 95%-uncertainty interval (+/-2CV(T)) for a postmortem measurement is largely determined by the pre-analytical component of variation. This should be kept in mind when judging on the uncertainty of postmortem measurement results.     

Hair analysis for cocaine: factors in laboratory contamination studies and their relevance to proficiency sample preparation and hair testing practices

Since the introduction in 2001 of a urine-based detection method for recombinant erythropoietin (rHuEPO), transfusion-doping practices have regained interest. To address this problem, an efficient antidoping test designed to obtain direct proof of allogeneic blood transfusion was developed and validated. This test, based on flow cytometry analysis of red blood cell (RBCs) phenotypes, was used to determine the absence or the presence of numerous RBCs populations in a blood sample. A such, it may constitute a direct proof of an abnormal blood population resulting from homologous transfusion. Single-blind and single-site studies were carried out to validate this method as a forensic quality standard analysis and to allow objective interpretation of real cases. The analysis of 140 blood samples containing different percentages (0-5%) of a minor RBCs population were carried on by four independent analysts. Robustness, sensitivity, specificity, precision and stability were assessed. ISO-accredited controls samples were used to demonstrate that the method was robust, stable and precise. No false positive results were observed, resulting in a 100% specificity of the method. Most samples containing a 1.5% minor RBCs population were unambiguously detected, yielding a 78.1% sensitivity. These samples mimicked blood collected from an athlete 3 months after a homologous blood transfusion event where 10% of the total RBCs present in the recipient originated in the donor. The observed false negative results could be explained by differences in antigen expression between the donor and the recipient. False negatives were more numerous with smaller minor RBCs populations. The method described here fulfils the ISO-17025 accreditation and validation requirements. The controls and the methodology are solid enough to determine with certainty whether a sample contains one or more RBCs populations. This variable is currently the best indicator for homologous blood transfusion doping.     

基于PTEN研究桃红四物汤治疗产后血瘀证的机制

目的 观察桃红四物汤对产后血瘀证大鼠子宫第10染色体缺失的碱性磷酸和张力蛋白同源物（phosphatase and tensin homolog deleted in chromosome 10，PTEN）mRNA及其蛋白表达的影响，探讨其祛瘀生新的作用机制。方法 采用米非司酮和米索前列醇灌胃法复制产后血瘀证大鼠模型，将模型大鼠分为模型组，阳性对照组（益母草颗粒4.3 g/kg），桃红四物汤高剂量（18.0 g/kg）、中剂量（9.0 g/kg）、低剂量（4.5 g/kg）组，另设正常对照组。连续灌胃给药7 d后，麻醉大鼠并分离子宫，光镜下观察子宫血瘀状态，分别采用实时PCR和Western blot法检测子宫组织PTEN mRNA及其蛋白表达水平。结果 桃红四物汤能改善产后血瘀证大鼠子宫间质的充血、水肿。与正常对照组比较，模型组PTEN mRNA及其蛋白表达水平显著升高（P<0.05）；与模型组比较，桃红四物汤高剂量组PTEN mRNA及其蛋白表达水平均显著降低（P<0.05），桃红四物汤中剂量组仅PTEN蛋白表达水平显著降低（P<0.05）。结论 桃红四物汤的祛瘀生新作用可能与其下调PTEN的表达有关。     

从气虚络瘀论特发性肺纤维化自噬机制

中医理论认为，特发性肺纤维化（idiopathic pulmonary fibrosis, IPF）的关键病机在于“气虚络瘀”。IPF患者自噬水平不足，不能及时清除异常病理物质，造成细胞外基质过度沉积而致肺纤维化。这与气虚无力推动新陈代谢，形成病理产物堆积，进而壅滞肺络，最终导致肺叶痿弱而不用相对应。因此，自噬不足导致的病理过程可视为“气虚络瘀”病机的微观机制，运用气虚络瘀理论阐释IPF自噬机制并通过调控自噬以防治本病可能成为新的重要靶点。     

旋毛虫IFN-γ诱导的溶酶体硫醇还原酶对大鼠外周血单个核细胞功能的影响

为了研究旋毛虫(Trichinella spiralis)IFN-γ诱导的溶酶体硫醇还原酶(GILT)对体外大鼠外周血单个核细胞(PBMCs)免疫功能的影响,将不同质量浓度的重组蛋白TsGILT(rTsGILT)与大鼠PBMCs体外共孵育后,用间接免疫荧光技术检测该蛋白与细胞之间的结合情况,并测定其对细胞增殖、迁移、凋亡、吞噬和NO分泌的影响。结果表明:rTsGILT可与大鼠PBMCs结合,可显著促进大鼠PBMCs的增殖、迁移、吞噬和NO的分泌(P<0.01),并呈现一定剂量依赖性;10、20μg/m L的rTsGILT对大鼠PBMCs的凋亡没有影响,40和80μg/m L的rTsGILT均能显著促进大鼠PBMCs的凋亡(P<0.001)。综上所述,旋毛虫GILT能通过多种途径影响宿主PBMCs发挥免疫作用,对宿主的免疫应答具有一定的促进作用。     

恒河猴ABO血型的研究

本文应用A型、B型、AB型血清，及抗－A与抗－B单克隆抗体对113只猕猴的ABO血型进行了测试。结果表明：在恒河猴（Macacamulatta）红细胞表面及血清中均未证明有类人凝集原和凝集素存在。但可根据其唾液中类人ABH型物质存在的情况判定该动物的类人血型。本实验检出的血型表型频率分别为：A型17．70％；B型52.21％；AB型20.36％；O型9.73％。该结果与国外报道的血型分布频率有差异。     

益气活血方对气虚血瘀证脑缺血再灌注损伤大鼠Fas、FasL蛋白表达的影响

目的：探讨益气活血方对气虚血瘀证局灶性脑缺血再灌注损伤大鼠Fas、FasL蛋白表达的影响。方法：采用气虚血瘀证局灶性脑缺血再灌注损伤大鼠模型，用线栓法阻塞大鼠大脑中动脉复制局部脑缺血再灌注模型，缺血2h后再灌注l，3，7d。用免疫组化染色法分别检测缺血皮质Fas、FasL蛋白表达。结果：与模型组比较。益气活血方组Fas、FasL蛋白表达显降低。结论：益气活血方可能通过抑制Fas、FasL蛋白表达，减轻气虚血瘀证局灶性脑缺血再灌注损伤。     

多虑平SPE-HPLC分析方法的建立及其应用

目的　建立尿样和全血中多虑平的固相萃取 高效液相色谱 (SPE HPLC)分析方法。方法　以多沙普仑为内标 ,1ml尿样或 0 5ml全血用Oasis小柱固相萃取后进Lichrospher 10 0RP 18e ( 2 5 0mm× 4mm ,5 μm)分析柱进行分析 ,2 3 0、 2 5 0nm同时进行检测。结果　尿样和全血中的检测限均 2ng/ml,线性相关系数r≥ 0 9992 ,天内和天间精密度均小于 6 75 % ,绝对回收率大于 85 % ,内源性物质不干扰测定。结论　本法快速、简便、准确 ,可用于实际案例的检测。     

补肝益肾活血法治疗中重度斑秃临床观察

目的 评价补肝益肾活血法治疗中重度斑秃的临床疗效并探究其对患者外周血CD4+CD25+调节性T细胞亚群的影响。方法 选择中重度斑秃患者50例,根据就诊先后顺序将其随机分为治疗组和对照组各25例,对照组服用复方甘草酸苷片,治疗组给予补肝益肾活血之剂治疗,观察两组治疗前后斑秃严重程度评分（severity of alopecia tool,SALT）的变化,评价临床疗效,并利用流式细胞仪分析两组患者治疗前后外周血清CD4+CD25+调节性T细胞亚群的变化情况。结果 两组患者治疗12周后,SALT评分均显著低于治疗前（P<0.05）;两组患者治疗前后SALT评分差值比较,差异有统计学意义（P<0.05）。两组临床疗效的分布比较,差异有统计学意义（P<0.05）。两组患者血清CD4+CD25+调节性T细胞表达水平均较治疗前升高（P<0.05）,但均未达到正常组水平。结论 补肝益肾活血法能够有效治疗中重度斑秃,并可以提高患者外周血CD4+CD25+调节性T细胞水平,效果优于复方甘草酸苷。