Masahiro Mukaida

Histochemical demonstration of methamphetamine by immunocytochemistry.

A method for the demonstration of methamphetamine (MA) by immunocytochemistry was established. The tissues of intoxicated mice, administered various amounts of MA in single doses of from 0.01 to 1 mg of MA-HCl, were fixed in glutaraldehyde-containing fixatives. Cryostat and paraffin slices gave a positive reaction of MA localization by staining the brain, liver, kidney, lung, stomach, spleen, and so forth, with the aid of the indirect immunoperoxidase technique. Those of animals administered a single dose of 0.1 mg or more (over 3 to 4 mg/kg--the usual dose of MA in acute intoxication death in forensic medicine), in particular, gave a strong strong reaction, so that the diagnosis of MA intoxication can be performed by macroscopic observation of stained slices. The histochemical diagnosis of MA intoxication in clinical toxicology and pathology might be regarded as a useful tool, especially in forensic pathology. The following cells gave a strong positive reaction: nerve cells and myelin sheaths, hepatocytes, epithelial cells of the distal part of the renal tubule and of the collecting tubule, alveolar and bronchial epithelial cells of the lung, chief and parietal cells of the gastric gland, capillaries of the renal glomerulus, macrophages in the blood and tissues, and striated muscle cells including cardiocytes. The morphological evidence of the pharmacodynamics and pharmacokinetics of MA can be determined at the cellular level by immunocytochemistry.

Histochemical demonstration of phenobarbital by immunocytochemistry.

A method for the demonstration of the topographical distribution of phenobarbital at the cellular level in various tissues was established. Mice that had been exposed to various doses of phenobarbital by intraperitoneal injection were killed, and their tissues were fixed with 0.1 M phosphate buffer solution (pH 7.4) containing paraformaldehyde and glutaraldehyde. Thereafter, paraffin and frozen sections were made and stained by the indirect immunoperoxidase method using antisera obtained from commercial sources and used for the immunochemical assay of the blood level of phenobarbital in clinical medicine. A specific positive reaction was observed solely in testing the intoxicated tissues, and this reaction was inhibited when phenobarbital was added to the antisera. The minimal sensitivity of the positive reaction, which can be discerned by observing the stained slides macroscopically, was in the range of 10 mg/kg. Thus, the diagnosis of phenobarbital intoxication in the forensic autopsy can be made by immunohistochemistry. A positive reaction was found in various tissue cells, including nerve cells, myelin sheaths, glia cells, hepatocytes, cells of the alveolar and bronchial wall, epithelial cells of the distal part of the renal tubules, and so forth. Endothelial cells of the capillaries in all tissues gave a strong positive reaction. The immunocytochemical electron microscopy of the hepatocytes revealed that the positive reaction in the cytoplasm was located solely in the intraluminal space of the smooth endoplasmic reticulum. These results indicate some interesting aspects of the pharmacokinetics of phenobarbital in vivo. It is expected that the antisera, which are used widely for the assay of the blood concentration of various drugs (phenobarbital, amphetamines, morphine, and so forth), may be regarded as excellent reagents for immunocytochemistry. This clearly indicates that morphological evidence in toxicology, which had so far remained obscure, can be easily obtained by applying these antisera against various drugs.

Penetrating cardiac injuries. A pathological analysis of 20 autopsy cases.

A total of 20 autopsy cases involving deaths due to penetrating cardiac injuries were analyzed pathologically: the causes of death of the victims who died before cardiorrhaphy were exsanguination or hemorrhagic shock in nine cases, pericardial tamponade in three, and a combination of the two in another three. Three of the remaining five cases of early postoperative death resulted from hemorrhagic shock, one resulted from pulmonary air embolism, and one resulted from a combination of hemorrhagic shock and coronary platelet embolism. We should pay more attention to air embolism and platelet embolism as causes of death of victims who have had penetrating cardiac injuries, whose exsanguinating hemorrhage and/or pericardial tamponade may be controlled by emergency-room thoracotomy and cardiorrhaphy.

Posttraumatic acute cholecystitis. Relationship to the initial trauma.

Posttraumatic acute cholecystitis is an often unrecognized and potentially fatal complication seen among patients hospitalized for trauma, and differs in etiology from cholecystitis which develops de novo. The cause, although not yet clearly defined, is believed to be related to bile stasis, ischemia, bacterial infection, sepsis, the activation of factor XII, and the Shwarzman reaction. A case is described in which a 53-year-old man with pelvic fractures developed acute acalculous cholecystitis and died of multiple organ failure 3 weeks following cholecystectomy. The histopathological findings are also reported; these are most likely attributed to the Shwarzman reaction or the activation of the factor XII pathways. There has been a tendency to regard posttraumatic acute acalculous cholecystitis as induced by trauma, and calculous as mere coincidence. We believe, however, that it is not calculous but histopathological findings that determine whether acute cholecystitis following trauma was more than coincidence or just mere coincidence. Although progress in clinical care has improved the chances of survival of severely traumatized patients, posttraumatic acute cholecystitis has been increasing in frequency. We cannot be careful enough in judging the relationship of this fatal complication to the initial trauma.

Purification and Characterization of Anti‐A Agglutinin from Euhadra callizoma amaliae

Abstract. Snail agglutinin (anti‐AEc, anti‐A1 of cold agglutinin type) from the albumin gland of Euhadra callizona amaliae was purified by affinity chromatography (meconium‐A aminoethyl cellulose). Its physicochemicl properties are as follows: S20, w: 5.3S. Isoelectric point: pH 3.6. Molecular weight: 8.9 × 104. Hexose content: 5.1%. The amino acid composition is that of an acidic protein.

DNA fingerprints: The importance in forensic medicine

SummaryInvestigation of genomic polymorphisms detected by a minisatellite named tentatively “Myo”, which is expected to correspond to the minisatellite in human myoglobin gene of Jeffreys et al., gives distinct and different aspects of chorionic villus and the decidual membrane in the same placenta. The chorionic villus, which is regarded as the extraembryonal tissue, represents the essential embryonal DNA fingerprint pattern, while the decidual membrane reveals the maternal one. A comparison between the DNA fingerprints from the chorion villus and from the blood sample of the suspected father provides the possibility of setting a paternity determination which can be achieved during the first trimester of a pregnancy.ZusammenfassungDie genomischen Eigenschaften der Plazenta wurden mit Hilfe von einer Sonde “Myo” gegenüber dem Minisatellit, der im Bereich vom humanen Myoglobinen vorhanden ist, analysiert. Der DNS Fingerabdruck von Chorionzotten stimmt völlig mit dem des Fötus überein, während der Fingerabdruck der Dezidua die mütterliche Herkunft darstellt. Die Analyse von verschiedenen DNS Fingerabdrücken aus Fötus, Chorionzotten, Mutter und Vater bei einer künstlichen Schwangerschaftsunterbrechung hat gezeigt, daß der DNS Fingerabdruck von Chorionzotten, die gewöhnlich bei der klinischen Chorionzottenbiopsie entnommen werden können, die entscheidende Auskunft darüber gibt, das Bestehen der Vaterschaft vollständig zu beweisen.

Immunohistochemical investigation of topographic localization of phenobarbital and methamphetamine in the rat retina

The topographic localization of methamphetamine and phenobarbital in the rat retina was studied. It is interesting that the secondary and primary ganglionic cells were stained predominantly only in the case of methamphetamine while in phenobarbital these were not stained. The staining patterns of the inner plexiform layer (strong reaction in phenobarbital and weak reaction in methamphetamine), outer nuclear layer (weak reaction in phenobarbital and strong reaction in methamphetamine) and choroid (very strong reaction in phenobarbital and no reaction in methamphetamine) also showed some differences.

Genetic Polymorphism of Red Cell Glutamate-Pyruvate Transaminase in Japanese

Six hundred and eight red cell hemolysates were screened for glutamate-pyruvate transaminase (GPT) by means of isoelectric focusing. Two new variant phenotypes were detected, neither of which could be distinguished from GPT2-1 and GPT2 by conventional starch gel electrophoresis. The two types were considered to correspond to GPT2B-1 and GPT2A-2C reported previously in samples of European origin.

Heterogeneity in the myoglobin content of chicken heart Purkinje fibers

SummaryThe myoglobin content of chicken myocardial cells was studied using indirect-immunoperoxidase histochemistry. While ordinary myocardial cells exhibited a homogeneous reaction pattern, the reactions for ventricular Purkinje fibers were remarkably heterogeneous. On the basis of the degree of staining, three types of cells, i.e., dark, intermediate, and clear, were distinguishable. In addition, the cytological heterogeneity of Purkinje cells was confirmed using conventional and immunological electron microscopy. The dark cells contained more myofibrils, mitochondria, and other organelles (e.g., ribosomes) than the clear cells.