人心肌梗死心肌肌钙蛋白T脱失的免疫组化观察

目的 观察人梗死心肌中心肌肌钙蛋白T的脱失情况，探讨其在人心肌梗死死后诊断中的应用价值。方法 应用免疫组化LSAB法，对人心肌梗死心肌肌钙蛋白T的脱失情况观察。结果 人梗死心肌肌钙蛋白T有明显脱失，通过计算机图像分析显示，人梗死心肌肌钙蛋白T的脱失面积与正常人心肌之间存在显著性差异(P＜0.01)。结论 心肌肌钙蛋白T(CTnT)可能成为人心肌梗死的诊断指标之一。     

大鼠早期心肌缺血心肌肌钙蛋白T脱失的组化研究

目的研究早期心肌缺血大鼠模型上心肌肌钙蛋白T的脱失情况。方法应用免疫组化LSAB法显色观察。结果缺血15分钟,大鼠心肌肌钙蛋白T即有明显脱失,随缺血时间的延长,脱失面积呈扩大趋势。计算机图像分析显示缺血大鼠心肌肌钙蛋白T的脱失面积与正常大鼠心肌之间存在显著性差异。结论心肌肌钙蛋白T是诊断早期心肌缺血的较为敏感的指标之一。     

A study of various morphologic variables and troponin I in pericardial fluid as possible discriminators of sudden cardiac death

Pathologists frequently examine victims of sudden cardiac death. In some cases, a firm diagnosis of cardiac-related death can be made based on conclusive gross and histologic findings. In many other cases, we find evidence supportive of, but not diagnostic of, cardiac death (e.g., atherosclerotic coronary artery disease, cardiomegaly, myocardial scarring). A final cohort consists of cases of sudden death with minimal to mild cardiac disease, no other significant pathology, and negative toxicologic studies. This prospective study compared 38 cardiac-related deaths with 52 control cases with respect to concentrations of pericardial cardiac troponin I (cTnI), heart weight, evidence of old and/or recent myocardial injury, and presence of significant coronary artery disease. The influence of documented chest trauma and/or perimortem cardiopulmonary resuscitation (CPR) on levels of cTnI was also analyzed. Even though median cTnI levels were significantly higher in cardiac deaths than in controls (p = .003), cTnI was not found to be a significant predictor of cardiac deaths, as determined by discriminant analysis (p = .52). Heart weight >500 g, evidence of old and recent myocardial injury, and significant coronary artery disease were seen statistically more often in cardiac deaths than in controls (p < or = .005 in each case), and median age was significantly higher in cardiac deaths than in controls (p = .001). Based on a stepwise logistic regression model, significant coronary artery disease, old and recent myocardial injury, and heart weight >500 g were found to contribute significantly to the prediction of cardiac death. Finally, neither chest injury nor CPR significantly affected concentrations of cTnI in pericardial fluid. These data confirm that the presence of acute and remote myocardial injury, significant coronary artery disease, and cardiomegaly (heart weight >500 g) strongly supports the diagnosis of a cardiac-related death. In contrast to a recently published report, we do not find that elevated concentrations of cTnI in pericardial fluid are strong indicators of cardiac-related deaths using our methodology.     

肢体挤压伤大鼠早期心电图及血清cTnI的变化

目的观察挤压伤大鼠早期心电图的改变,并检测血清心肌肌钙蛋白I(cTnI)的变化。方法复制挤压伤大鼠动物模型,标准Ⅱ导联记录心电图变化,化学发光法检测血清cTnI水平。结果挤压伤后心电图ST段明显抬高,并持续24h,伤后6h血清cTnI则显著升高,并持续24h以上。结论肢体挤压伤早期存在心肌细胞的损伤。     

心脏肌钙蛋白T、I与心肌损伤及其法医学应用前景

心脏肌钙蛋白T、I具有很高的心脏特异性,敏感性高,能检测多种不同的心肌损伤,区别心肌损伤和骨骼肌损伤,并能用作心肌损害的危险程度分级和判断预后的指标,在临床上得到了广泛的应用。在法医学上也有一定的应用前景。     

Evaluation of anti-muscle actin monoclonal antibody HHF35 in the diagnosis of early myocardial infarction

In order to explore the specificity of anti-muscle actin monoclonal antibody HHF35 in the postmortem diagnosis of early myocardial infarction, the changes of HHF35 staining in the infarcted myocardia and other non-infarcted myocardia with direct or indirect myocardial injury were studied with immunohistochemical S-P method. The results showed that the loss of HHF35 staining of different degrees was found in the infarcted myocardia, but also in the other non-infarcted myocardia with direct or indirect myocardial injury, such as cardiac contusion, myocarditis, hemorrhagic shock, electrocution, mechanical asphyxia. So it should be cautious in diagnosing early myocardial infarction with HHF35 immunohistochemistry.     

Fluorescence microscopy in the detection of early myocardial infarction

Fluorescent microscopy is amongst the many techniques devised for the post-mortem detection of early myocardial infarction. The method has the advantage of sensitivity, speed and simplicity, compared with more complex techniques such as enzyme histochemistry and electron microscopy. Disadvantages include the difficulty of permanent preservation of the sections and lack of sharp differentiation between normal and very early infarction--though the latter problem is common to most other methods, except enzyme histochemistry. There are two main techniques in fluorescent microscopy: The examination of sections specifically stained by fluorochrome dyes; The re-examination of haematoxylin-eosin sections for 'autofluorescence'.     

早期心肌缺血猝死心肌Ⅰ型胶原的表达

目的探讨Ⅰ型胶原蛋白在心肌早期缺血死后诊断中的法医学价值。方法将收集的案例心肌蜡块分为3组：正常对照组、早期心肌缺血组、心肌梗死组,用免疫组织化学技术（S-P法）,观察猝死心肌内Ⅰ型胶原的表达情况,对其结果进行半定量分析。结果早期心肌缺血组和心肌梗死组心肌细胞质以及细胞核染色阳性,且早期心肌缺血组与心肌梗死组表达强度相近,说明早期心肌缺血组在胶原网结构方面已经发生与心肌梗死相同的改变。结论Ⅰ型胶原可以作为早期心肌缺血的一个诊断指标。     

白细胞介素-18在早期心肌缺血和心肌梗死心肌的表达

目的探讨白细胞介素-18(IL-18)对早期心肌缺血(EMI)和心肌梗塞(MI)死后诊断的法医学意义。方法筛选出34例心脏标本,分EMI(11例)、MI(10例)和对照(13例)组,检测心肌IL-18免疫组化染色(SP法)反应,并经全自动图像分析系统分析得到平均光密度值(AOD),所得数据进行统计学分析。结果对照组心肌IL-18免疫组化染色呈阴性,EMI组心肌呈阳性,MI组心肌呈强阳性;其AOD值分别是0.1193±0.0649、0.3827±0.0868、0.4792±0.0884,3组之间差异均具有显著性意义(P<0.05)。结论作为判断EMI和MI的指标之一,IL-18对EMI和MI的死后诊断具有法医学意义。     

Heart fatty acid binding protein as a marker for postmortem detection of early myocardial damage

Depletion of heart fatty acid binding protein (H-FABP) from cardiomyocytes with varying post-ischemia intervals was studied in acute myocardial infarction (AMI) model of rat, and 22 human autopsy cases were studied with streptavidin-peroxidase conjugated method (S-P). It was observed that as early as 15 min after ischemia, the depletion of H-FABP could be detected in model rats. With the ischemic time prolonged, the depletion of H-FABP was more and more evident. In all human cases with myocardial infarction, absent H-FABP staining could be found in infarcted area. And in some suspected early myocardial infarction cases, depletion of H-FABP staining could be demonstrated in areas that showed normal hematoxylin-eosin (HE) staining. The blood samples from model rats before ligation, at varying post-ischemia intervals and various postmortem time were measured for plasma concentration of H-FABP with enzyme-linked immuno-sorbent assay (ELISA) method. At 15 min after myocardial ischemia, the concentration of H-FABP was 4 times higher (546.0+/-85.3 microg/l) than that of the baseline level (103.7+/-94.1 microg/l). With the continuation of ischemic time, the concentration of H-FABP increased and peaked at 4 h (1953.5+/-405.3 microg/l), then decreased. The plasma concentration of H-FABP decreased slightly with postmortem time, but was still significant higher at any postmortem intervals than that of baseline level within 48 h after death. The results suggest that H-FABP staining can detect very early ischemic damages in human myocardium and the elevated plasma concentration of H-FABP in rat was an indicator of AMI, which was not affected by autolysis.     

Cardiac rupture in acute myocardial infarction: a reassessment

Cardiac rupture as a complication of acute myocardial infarction (AMI) has been described as occurring infrequently. Because of the recent dramatic decrease in autopsy rates, the authors believe that current studies do not accurately represent the frequency of this catastrophic complication. Autopsy protocols and archived histologic slides of patients with AMI were retrospectively reviewed to determine whether the frequency of cardiac rupture, as a complication of AMI, is altered when a non-hospital-based patient cohort after autopsy is evaluated. This review yielded 153 cases of 41 women and 112 men, whose postmortem examinations revealed gross and histologic evidence of AMI. Cardiac rupture was present in 30.7% of these cases. Of the 47 patients with rupture, 35 had no relevant medical history. The remaining 12 patients had various medical conditions. None of the patients in the rupture group had previously treated symptoms related to coronary artery conditions. Whereas women constituted 26.8% of the total AMI group, they had a cardiac rupture rate of 61%. By contrast, men with AMI had a cardiac rupture rate of 19.6%. All patients in the cardiac rupture group had heart weights over the predicted expected weight as a function of body weight. Age, gender, and heart weight were significant factors associated with cardiac rupture, whereas body mass index was not significantly related. When these factors were evaluated jointly, age was a significant explanatory factor for rupture among both men and women, whereas body mass index and heart weight were significant for men but not for women. When the rupture sites occurred on the left ventricular myocardium, the anterior wall was affected in 21 cases (45%), the posterior wall in 18 (38%), the lateral wall in 4 (9%), and the apex in 3 (6%). The right ventricular myocardium ruptured in 1 case (2%). Most of the patients had severe multivessel coronary artery disease. Histologic study of the specimens showed that the majority of ruptures occurred between 24 and 72 hours after myocardial infarction. This study showed a frequency of cardiac rupture of 30.7% in patients with AMI and sudden death according to medical examiner's records. These findings confirm and reinforce the importance of postmortem examination and autopsy as an adjunct to clinical medical practice.     

早期心肌梗死死后诊断的法医病理学研究进展

心源性猝死在成人猝死的死因中占首位,心肌梗死是冠心病猝死的主要机制之一,由于早期心肌梗死发生突然,且可在短时间内死亡,肉眼和镜下都缺乏典型的形态学改变,一直都是法医实践中研究的热点,本文回顾多年来心肌梗死的研究成果在法医学中的应用及局限,展望新的临床指标在法医工作中的可行性,期望提高死后心肌梗死诊断的准确率。     

Estimation of postmortem interval using the protein marker cardiac Troponin I

The importance of determining the time since death is crucial to criminal, civil and forensic cases. A technique exploiting the degradation of a protein, cardiac Troponin I (cTnI), to estimate the postmortem interval (PMI) was investigated. Cardiac Troponin I is a basic regulatory protein found as part of a ternary complex responsible for calcium dependent muscle contraction. An efficient extraction protocol to analyze the banding pattern of cTnI in postmortem tissue was developed. The analysis involves extraction of the protein, separation by denaturing gel electrophoresis (SDS-PAGE) and visualization by Western blot using cTnI specific monoclonal antibodies. A bovine model was used to develop and optimize the protocol. The homology of cTnI amongst mammalian species allows for the cross-reaction of human anti-cTnI antibodies with bovine cTnI. The results indicate a characteristic banding pattern amongst human cadavers (n=6), a pseudo-linear relationship between percent cTnI degraded and the log of the time since death (r>0.95), and a qualitative degradation band pattern that in a simple comparative analysis with a standard human heart (known time since death) can be used to estimate the postmortem interval. The degradation-banding pattern of tissue cTnI is useful in the determination of the early postmortem (pm) interval (0-5 days). Overall, this technique offers advantages such a wide postmortem interval, measurable degradation pattern, a temporal semiquantitative relationship and manageable temperature dependence over direct temperature methods.     

Immunostaining by complement C9: a tool for early diagnosis of myocardial infarction and application in forensic medicine

Before the first 12 hours, diagnosis of early myocardial infarctions is always difficult for forensic pathologists. We tested complement C9 expression in 121 formalin-fixed and paraffin-embedded heart samples by an immunohistochemical procedure. The heart specimens were separated into four groups: 33 cases in group 1 with typical ischemic damages histologically located, 20 cases in group 2 with death related to myocardial infarction on the basis of ischemic presentation on electrocardiogram but no obvious histological ischemic damage, 35 cases in group 3 with severe coronary disease without cause of death found at the autopsy, and 33 cases in group 4 without sign of myocardial infarction and without coronary disease. In the first group, all 33 heart samples showed a well-defined C9 expression in the necrotic areas. The second group in 17 of 20 cases showed positive areas for C9 expression. In the other three heart specimens, only few stained cells were observed whereas the painful symptoms had begun less than 1 h before death. The third group showed C9 immunopositive areas in six of 35 cases, few stained cells in 8 cases, and no C9 deposition in the 21 other cases. The last group showed no staining area. To avoid nonspecific C9 staining due to tissue autolysis, we studied C9 expression during a controlled putrefactive process in four cases included in group 1; staining was found only in infarcted myocardial areas, and was observed up to ten days. Specificity of C9 expression was evaluated to be 100% [89.4 to 100%] and sensitivity to be 85% [62.11 to 96.79%]. In conclusion, evaluation of immunohistochemical expression of C9 appears to be a highly sensitive and specific marker of early myocardial infarction, useful in forensic medicine if survival is more than 1 h after the beginning of myocyte damage.     

Immunohistochemical study of fibronectin for postmortem diagnosis of early myocardial infarction

The postmortem diagnosis of early myocardial infarction has been a puzzling problem in forensic practice. In the present study, an immunohistochemical study of fibronectin (FN) was performed for the first time on 34 autopsy hearts to determine early myocardial infarction with streptavidin/biotin/peroxidase technique. Five cases of definite myocardial infarction showed positive FN staining of cardiomyocytes; of 18 cases where early myocardial infarction was suspected, positive FN staining of cardiomyocytes was found in 15 cases, but no such staining was seen in 11 non-cardiac death controls. The results led to the conclusion that positive FN staining in cardiomyocytes is a reliable marker of acute myocardial infarction and could be used as a new, sensitive method for the postmortem diagnosis of early myocardial infarction. It is worth noting that all cases in this study were autopsied between 8 h and 4 days after death and 5 cases had been fixed in 10% formalin for over 10 years. FN immunohistochemistry still gave satisfactory results in those cases. It seemed that FN was not affected by postmortem autolysis and formalin-fixation and could be used in routine forensic practice, especially for retrospective analysis of cases.     

早期心肌梗死bFGF的免疫组化研究

采用免疫组化和图像分析与统计学处理方法,对人体早期心肌梗死猝死者心肌中碱性成纤维细胞生长因子 (basic fibroblast growth factor,bFGF)的表达进行了研究。结果发现 :心肌梗死组 24例的心肌细胞核以及血管内皮细胞核均为强阳性染色,有些细胞浆也有阳性染色,尤以心肌梗死灶周围区域心肌细胞为甚。可疑梗死组 21例中 17例见心肌细胞核以及血管内皮细胞核为强阳性染色, 4例为弱阳性染色,未见阴性染色。而正常心脏对照组 16例中 3例呈弱阳性染色,其余均为阴性。图像分析与统计学处理结果显示对照组 bFGF阳性指数明显低于心肌梗死组和可疑心肌梗死组（ P< 0.001）,而后两组无显著性差异（ P >0.05）。结果表明 bFGF免疫组化染色可望为早期心肌缺血致猝死的死后诊断提供客观的形态学依据。     

早期心肌梗死与冠状动脉痉挛致心肌肌红蛋白缺失的实验研究

本实验按Selye法结扎Wistar大鼠冠状动脉左前降支复制早期心肌梗死模型；又经颈外静脉注射垂体后叶素复制冠状动脉痉挛模型。按心肌不同缺血时间（25min、1h、3h）及对照组将以上各模型均分成四组．每组均取心关部及其相邻组织块制成石蜡切片，进行HE染色、链霉菌素-生物素-肌红蛋白免疫组化染色（LSAN－Mb染色）。结果表明：心肌梗死25min即可见左心室前壁心内膜下心肌条片状Mb缺失，随着梗死时间的延长，条片状缺染区向外层心肌扩展；冠状动脉痉挛组呈多发性、小灶性Mb缺失，右心室Mb缺染灶多于左心室，且Mb缺染灶多围绕冠状动脉周围，或沿一支冠状动脉之各分枝是葡萄串样分布。可见大鼠早期心肌梗死与冠状动脉痉挛所致心肌Mb缺失具有不同的形态学特点．故LSAB－Mb法可望为冠状动脉痉挛所致心肌缺血提供客观形态学证据．     

A rapid postmortem cardiac troponin T assay: laboratory evidence of sudden cardiac death

Postmortem examination may be useful in establishing the cause of sudden unexpected death. In many instances, however, limitations of staffing, budget, and time may force the pathologist to triage cases to external examination rather than autopsy. A rapid assay for cardiac troponin T (cTnT) to document suspected cardiac-related deaths may optimize the use of the time and resources of the autopsy pathologist. Peripheral blood was sampled percutaneously before each of 40 autopsies and placed in the well of the Cardiac T Rapid Assay unit in accordance with the included instructions, and the results were read after 15 minutes. The assay result, decedent age, postmortem interval, and evidence of cardiopulmonary resuscitation were tabulated and subsequently correlated with the cause of death. On final sign-out of each of the autopsies, the cause of death was determined to be cardiac-related (n = 20) versus the cause in non-cardiac control subjects (n = 20). This determination was made while the investigators were blinded to the cTnT assay result. Of the 20 cardiac deaths, 17 (85%) showed positive results for cTnT compared with 6 (30%) false-positive results among the 20 control cases; this result was statistically significant according to the chi-square test. In the over-50 age group, the sensitivity of this assay in detecting cardiac-related death was 91%, with a specificity of 86%. Perimortem cardiopulmonary resuscitation did not appear to result in false-positive results. In the appropriate setting, this rapid assay for cTnT can provide valuable data supportive of a cardiac-related death. This inexpensive test may best be used in triaging sudden deaths in persons over 50 to external examination versus complete autopsy.     

VEGF在早期心肌梗死死后诊断中的应用

运用免疫组化方法和图像分析与统计学处理系统 ,对人体心脏标本VEGF的表达进行定量研究。结果显示 :心肌梗死组 2 4例心脏标本心肌细胞膜及血管内皮细胞膜均可见棕黄色VEGF强阳性染色 ,有些胞浆也呈阳性 ,尤以心肌梗死灶周围区域心肌细胞为甚 ;在可疑梗死组 2 1例中 ,19例心肌细胞膜及血管内皮细胞膜VEGF强阳性 ,2例为弱阳性 ,未见阴性 ;在正常心脏对照组 16例中 ,仅 1例见心肌细胞膜及血管内皮细胞膜散在棕黄色VEGF弱阳性 ,其余均为阴性。图像分析与统计学处理结果表明 ,对照组VEGF阳性指数 ( 0 3 0± 0 10 )明显低于心肌梗死组 ( 12 46± 3 2 6,P<0 0 1)和可疑梗死组 ( 11 70± 3 5 6,P <0 0 1) ,后两组之间阳性指数无明显差异 (P >0 0 5 )。提示运用免疫组化染色方法结合图像分析处理技术定量检测心肌局部VEGF的表达可望作为因早期心肌缺血导致心脏性猝死死后诊断较为客观的病理形态学指标之一。     

Serum concentrations of cardiac troponin T in sudden death

Ischemic heart disease is the most common cause of sudden death of natural causes in most western countries. By autopsy, there may be no gross or histologic evidence of acute myocardial damage unless the patient survived for several hours following the event. Cardiac troponin in serum has become the recommended biochemical marker for myocardial injury in the clinical setting. We performed a prospective study on 102 autopsied subjects at the Central Hospital of Rogaland, Stavanger, Norway. Femoral blood was sampled for subsequent analysis of cardiac troponin T (cTnT). In the subjects with morphologic evidence of recent myocardial injury (n = 34), the mean serum cTnT level was 1.95 microg/L compared with 0.16 microg/L in the subjects with a noncardiac cause of death (n = 35) and 0.61 microg/L in the group with probable sudden cardiac death without morphologic signs of acute myocardial injury (n = 33). The observed differences in mean serum cTnT levels between the groups were statistically significant (P < 0.0001). These data suggest that elevated postmortem serum concentration of cTnT reflects ongoing myocardial damage and may support a diagnosis of cardiac-related death in cases associated with sparse or inconclusive morphologic findings postmortem.