| Objective:The aim of the study was to measure the level of mast cell carboxypeptidase A3, mast cell tryptase and mast cell chymase in postmortem serum, pericardial fluid, peritoneal fluid, pleural fluid of drug-related fatal anaphylaxis decedent; and to investigate the expression of mast cell tryptase, mast cell tryptase in heart, lungs, throat, stomach and jejunum of drug-related fatal anaphylaxis decedent. This study tries to to provide objective laboratory indicator and detection methods for forensic medicine diagnosis of drug-related fatal anaphylaxis deaths. Methods:The experiment is consist of drug-related fatal anaphylaxis death group, sudden death of coronary heart disease group and normal control group.The drug-related fatal anaphylaxis death group are 20 autopsy cases, which are diagnosed with drug related anaphylactic shock death, and excluded the coronary heart disease, mastocytosis, craniocerebral injury and drug user; and 2 cases were caused by drip intravenous infusion of penicillin, 6 cases were caused by drip intravenous infusion of ceftriaxone sodium, 3 cases were caused by drip intravenous infusion of levofloxacin, 1 cases was caused by drip intravenous infusion of lincomycin hydrochloride, 1 case was caused by drip intravenous infusion of lai ammonia aspirin in, 1 case was caused by drip intravenous infusion of lomefloxacin, 1 case was caused by drip intravenous infusion of Qingkailing injection, 1 case was caused by drip intravenous infusion of Shuang Huang Lian 1 case was caused by oral ibuprofen, 1 case was caused by clysis of chloral hydrate enema. 5 cases were postmortem examined within 48 h after death, 14 cases were postmortem examined within 7 days after death, 1 case was postmortem examined within 15 days after death of the postmortem anatomy. Sudden death of coronary heart disease group are 20 autopsy cases, excluded allergies, mastocytosis and drug users. 10 cases were postmortem examined Vwithin 48 h after death,and 10 cases were postmortem examined in 7 days after death. The normal group are 20 autopsy cases, excluded allergies, the coronary heart disease, mastocytosis, craniocerebral injury and drug user. 20 cases were postmortem examined within 48 h after death.Three groups cases were taken the postmortem serum, pericardial fluid, peritoneal fluid, pleural fluid, took supernatant after centrifugation, and placed supernatant in-80℃ refrigerator to prepare for the fluorescence enzyme immunoassay(FEIA) and the enzymes linked immunosorbent assay(ELISA). Three groups cases were extracted the heart, lungs, throat, stomach and jejunum, to prepare for immunofluorescence staining and western blot detection.(1)We measure the level of mast cell carboxypeptidase A3 and mast cell chymase in the postmortem serum, pericardial fluid, peritoneal fluid, pleural fluid by the ELISA double antibody sandwich assay.(2)We determinate the level of the mast cell tryptase in the postmortem serum, pericardial fluid, peritoneal fluid, pleural fluid by the FEIA.(3) We stain mast cell tryptase and mast cell chymase of the heart, lungs, throat, stomach and jejunum by immunofluorescence, and observe ande collect images by Olympus BX61 fluorescence microscopy, and analyse the image by Image-pro plus 6.2 software.(4) We determine mast cell tryptase content of the heart, lungs, throat, stomach and jejunum by western blot.(5) To observe mast cell degranulation in the drug-related anaphylactic shock group by the three different methods to dye mast cells with toluidine blue.(6) We do statistical analysis by SPSS17.0 statistical software for the experimental data, to analyse differences between three groups using single factor analysis of variance(ANOVA), to analyse differences between two groups using LSD method. Results:The mast cell carboxypeptidase A3 content in the postmortem serum, pericardial fluid, peritoneal fluid, pleural fluid of the drug-related fatal anaphylaxis death group was respectively 10.13±2.11ng/ml, 4.34±1.61ng/ml, 4.60±1.81ng/ml, 4.49±1.21ng/ml; mast cell tryptase content was respectively 41.57±5.48ug/L, 28.96±4.82ug/L, 29.71±4.90ug/L, 22.62±4.10ug/L; mast cell chymase content was respectively 7.85±1.46ng/ml, 7.31±1.37ng/ml, 7.32±1.79ng/ml, 7.29±1.53ng/ml. The mast cell carboxypeptidase A3 content in the postmortem serum, pericardial fluid, peritoneal fluid, pleural fluid of the coronary death group was respectively 4.51±0.91ng/ml, 2.07±0.41ng/ml, 1.85±0.61ng/ml, 1.89±0.66ng/ml; mast cell tryptase content were respectively 16.42±2.91ug/l, 11.55±2.41ug/l, 12.74±2.84ug/l, 10.66±2.76ug/l; mast cell chymase content was respectively 3.02±0.86ng/ml, 2.80±0.67ng/ml, 2.82±0.76ng/ml, 2.79±0.76ng/ml. The mast cell carboxypeptidase A3 content in the postmortem serum, pericardial fluid, peritoneal fluid, pleural fluid of the normal control group was 3.18±1.30ng/ml, 1.36±0.48ng/ml, 1.26±0.36ng/ml, 1.59±0.51ng/ml; mast cell tryptase content was 4.86±0.86ug/L, 4.64±0.95ug/L, 3.30±0.84ug/L, 3.11±0.96ug/L; mast cell chymase content was respectively 1.22±0.55ng/ml, 1.13± 0.48ng/ml, 1.15±0.58ng/ml, 1.11± 0.49ng/ml.Compared with the sudden coronary death group and the normal group,mast cell carboxypeptidase A3, mast cell tryptase, mast cell chymase content in the postmortem serum, pericardial fluid, peritoneal fluidc and pleural fluid of the drug-related fatal anaphylaxis death group was significantly higher(P<0.05), especially the level of the postmortem serum. Mast cell tryptase and mast cell chymase’ expression in the heart, lung, larynx, stomach and jejunum tissues of the drug-related fatal anaphylaxis death group were significantly higher than of the sudden coronary death group and the normal group(P<0.05).Mast cell carboxypeptidase A3, mast cell tryptase, mast cell chymase content in the postmortem serum and pericardial fluid of the sudden death of coronary heart disease group was increased significantly compared with normal control group(P<0.05), and was lower than he drug-related fatal anaphylaxis death group(P<0.05). Mast cell carboxypeptidase A3, mast cell tryptase, mast cell chymase content in peritoneal fluidc and pleural fluid of the sudden death of coronary heart disease group has no significant difference with normal control group(P>0.05). Mast cell tryptase and mast cell chymase expression in the heart of the sudden death of coronary heart disease group is significantly higher than normal control group(P<0.05), mast cell tryptase and mast cell chymase expression in the lung, throat, stomach and jejunum of the sudden death of coronary heart disease group has no significant difference with normal control group(P>0.05). Conclusion:(1) This study indicate that death of drug-related anaphylactic shock can be diagnosed by determining mast cell carboxypeptidase A3, mast cell tryptase, mast cell chymase content in the postmortem serum, pericardial fluid, peritoneal fluidc and pleural fluid with FEIA and ELISA.(2)We can observe mast cell tryptase and mast cell chymase expression in heart, lung, throat, stomach and jejunum of the cases which have no the postmortem serum, pericardial fluid, peritoneal fluid, pleural fluid by Western blot and immunofluorescence staining(3)Mast cell carboxypeptidase A3, mast cell tryptase, mast cell chymase content in the postmortem serum, pericardial fluid, peritoneal fluidc and pleural fluid of the drug-related fatal anaphylaxis death can be impacted of time of drug sensitive, survival time after anaphylaxis and autopsy time after death.(4) There isn’t different expression of mast cell tryptase and mast cell chymase in different organs. Mast cell tryptase and mast cell chymase content in heart, lung, throat, stomach and jejunum has fewer changes contrast with that in the postmortem serum, pericardial fluid, peritoneal fluidc and pleural fluid(5) Mechanism ofdrug anaphylactic shock death is complex. So to legal medical expert it is very difficult to diagnose with drug-related fatal anaphylaxis death because the accuracy and sensitivity of laboratory diagnosis index may be influenced autopepsia, cruor, hemolysis. we should diagnose with drug-related fatal anaphylaxis death by determining different indicators of different organs(or body fluid) with different laboratory method. |
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