Biomedical subjects

M Oehmichen

Publications and source records attributed to M Oehmichen.

At least 19 recordsLinked to original sources

PubMed2002

Quantification of human mitochondrial DNA in a real time PCR.

Recently, a moderately priced machine for real-time quantitative PCR has become available, the Perkin Elmer 5700. The rapid and quantitative assay of mitochondrial DNA (mtDNA) copy number is potentially useful in a variety of molecular, evolutionary and forensic fields. Using this new tool, we have evaluated the precision and reliability of the real time PCR to quantify undeleted mitochondrial genome copy number, and to determine the frequency of an age-associated deletion of 4977 base pairs in length, in 42 human iliopsoas muscle DNA samples from persons of known age. We have evaluated the accuracy with which age can be predicted, knowing only the frequency of this common 4977 bp deletion, and derived a statistical formula which describes the confidence with which the 4977 bp frequency predicts age. The results indicate that the mutation frequency could be used to distinguish between tissue from young and old individuals. However in this data set, while there was considerable agreement of 4977 bp frequency among replicates from the same individual sample, there was substantial diversity of mean mutation frequency between individuals of the same or similar ages. The simplest interpretation of these results is that there are biological modifiers of 4977 bp frequency that are age-independent, which are potentially interesting but may limit the usefulness of this deletion frequency alone as a "molecular forensic clock."

Autopsy↗

PubMed2002

Mitotic brain cells are just as prone to mitochondrial deletions as neurons: a large-scale single-cell PCR study of the human caudate nucleus.

Mitochondria are considered a key element in the process of organismic aging, because of their fundamental role in cellular energy generation. In the course of oxidative phosphorylation, harmful free radicals are continuously produced damaging the mitochondrial (mt) genome. One of the consequences is the occurrence of large-scale deletions in mtDNA molecules. The 4977 bp common deletion accumulates exponentially with age, in a mosaic pattern, especially in postmitotic tissues. In order to investigate whether certain cell characteristics underlie this pattern of distribution, and to look for possible age-related changes, two cell types in the caudate nucleus of the human brain from five young and five senescent subjects were analysed by single-cell PCR.MAP2-positive neurons and GFAP-positive astrocytes were isolated by micromanipulation. For each of the 10 cases, at least 30 cells of each type were collected and subjected to PCR individually. Screening for the presence of the common deletion yielded no significant differences in relative distribution, neither between astrocytes and neurons, nor between healthy young and old humans. Our results imply that the age-dependent increase of the common deletion cannot come about by an increase of independent deletion events in a greater proportion of cells, and that mitotic rate is not a major cellular risk factor for deletion accumulation in the caudate nucleus.

Adolescent↗

PubMed2001

Time of drug elimination in chronic drug abusers. Case study of 52 patients in a "low-step" detoxification ward.

The elimination time of illicit drugs and their metabolites is of both clinical and forensic interest. In order to determine the elimination time for various drugs and their metabolites we recruited 52 volunteers in a protected, low-step detoxification program. Blood samples were taken from each volunteer for the first 7 days, daily, urine sample for the first 3 weeks, daily. Urine was analyzed using a fluorescence-polarization immunoassay (FPIA) and gas chromatography/mass spectrometry (GC/MS), serum using GC/MS. The elimination times of the drugs and/or their metabolites in urine and serum as well as the tolerance intervals/confidence intervals were determined. Due to the sometimes extremely high initial concentrations and low cut-off values, a few of the volunteers had markedly longer elimination times than those described in the literature. The cut-off values were as follows: barbiturates II (200ng/ml), cannabinoids (20ng/ml), cocaine metabolites (300ng/ml), opiates (200ng/ml). GC/MS detected the following maximum elimination times: total morphine in urine up to 270.3h, total morphine and free morphine in serum up to 121.3h, monoacetylmorphine in urine up to 34.5h, 11-nor-9-carboxy-delta-9-tetrahydrocannabinol (THC-COOH) in urine up to 433.5h, THC-COOH in serum up to 74.3h, total codeine in urine up to 123h, free codeine in urine up to 97.5h, total codeine in serum up to 29h, free codeine in serum up to 6.3h, total dihydrocodeine (DHC) in urine up to 314.8h, free DHC in urine up to 273.3h, total and free DHC in serum up to 50.1h. Cocaine and its metabolites were largely undetectable in the present study.

Adult↗

PubMed2001

Brain injury after survived gunshot to the head: reactive alterations at sites remote from the missile track.

Gunshot wounds to the brain usually lead to acute respiratory arrest or death after a brief survival period, even in cases involving only slight direct tissue damage. It can be assumed therefore that the damage extends beyond the zone of recognizable destruction and hemorrhages. To determine the true extent of the tissue injury resulting from gunshot wounds to the brain, we carried out microscopic investigations for reactive changes (emigration of leukocytes and macrophages, axonal expression of beta-amyloid precursor protein (beta-APP) in 10 cases of gunshot wound to the narrow channel of the brain with survival times >2h. Demonstration of leukocytes expressing naphthol AS-D chloroacetate esterase activity in the brain tissue at the border of the missile track established the vitality of the gunshot effect. The presence of macrophages (CD68-epitope) allowed demarcation of a 1-2mm wide necrotic zone around the permanent cavity. Within this zone and beyond, beta-APP showed an initial increase followed by a decline in the number of injured axons. Three types of beta-APP positive staining could be differentiated. In the immediate vicinity of the missile track beta-APP positive neurons were present at a distance of 2-4mm from the margin of the permanent cavity (type 1) as a result of primary injured neuronal tissue by the gunshot itself. At longer distances from the narrow channel and the permanent cavity single beta-APP positive axons or axon fragments and two additional types were found; type 2 shows a parallel, wave-like arrangement of the damaged fibers, which suggests that the injury was produced by mechanical acceleration of the brain tissue created by the energy the projectile expended within the brain; irregular aggregation of beta-APP positive axons or axon fragments within a local edema represents type 3, which may be attributed to secondary ischemia or edema.

Adult↗

PubMed2001

Death from thermal effects and burns.

One hundred and fifteen unselected autopsy cases of death from thermal effects and/or fire between 1990 and 1999 were analyzed with regard to time of death, signs of vitality at the scene of the fire, the manner and cause of death, and the extent of soft tissue loss. The cases represented approximately 6% of all autopsy cases at the Institute of Legal Medicine responsible for a catchment area with approximately 700,000 inhabitants. In 23 victims suffering burn injuries, death occurred during initial medical care or clinical treatment. The causes of death were primary respiratory arrest due to smoke poisoning or delayed shock caused by thermal injuries to the skin. Death occurred at the scene of the fatal event in 85 cases: eight cases exhibited no thermal effects; the cause of death in one of these cases was polytrauma incurred in a leap from a height; in seven cases there was poisoning due to smoke inhalation. The remaining 77 cases were characterized by signs of intensive thermal and/or fire effects. Clear signs of vitality (carboxyhemoglobin (COHb) in blood, inhalation and/or swallowing of soot) were found in 84.7% of the victims dying at the site of the fatal event. Of the 13 victims showing no signs of vitality at the scene, a cause of death could be determined in only seven cases; death in the other six cases remains unexplained. Quantification of the soft tissue loss revealed a possible correlation with the temperature and time course of heat exposure.

Adolescent↗

PubMed2001

[The mitochondrial genome and aging].

There is a lot of evidence that age-associated alterations of the mitochondrial genome occur, especially in postmitotic tissues such as brain, heart and skeletal muscle. These alterations are supposed to be a result of an attack of free radicals generated as normal byproducts of oxidative phosphorylation and lead to damage of proteins, lipids, and DNA. The alterations of mtDNA include oxidative damage of base pairs, point mutations, large-scale deletions or duplications. The 4977 bp deletion or "common deletion" reveals an age-dependent accumulation in postmitotic tissues, but not in fast-dividing tissues such as blood cells. In addition, it is observed that a tissue-specific accumulation occurs with the highest abundance in the basal ganglia, followed by skeletal muscle, heart, and lowest in cerebellar tissue. Third, pathological alterations of specific tissue, like ischemia/reperfusion events, display a pronounced accumulation of the deletion compared to age-matched controls. Because there are many mtDNA mutations, further analysis of all alterations of mtDNA will elucidate its role in the phenomenon of aging. Despite some criticisms of this free radical theory of aging, there is a lot of experimental evidence to support the important role of mitochondria in organismal aging.

Aged↗

PubMed2001

Regional potassium distribution in the brain in forensic relevant types of intoxication preliminary morphometric evaluation using a histochemical method.

A histochemical-morphometric method was used to measure potassium (K+) levels in gray and white matter of rats following sublethal intoxication with 11 different neurotoxic compounds of high forensic significance. Six rats were each given a single substance applied intraperitoneally, the same dosage being given to two animals each. The animals were subsequently killed, the brains immediately frozen, and cryosections cut. K+ levels were evaluated morphometrically. A drop in K+ levels was used as the criterion for cytotoxic edema. Application of ethanol, atropine, carbromal, carbon monoxide, morphine or triethyltin led to a rise in K+ levels in the gray matter and a simultaneous decline in the white matter. By contrast, administration of amitriptyline, glycerin, potassium cyanide, parathion or phenobarbital initiated an increase in K+ levels in both gray and white matter. A cytotoxic edema could thus be reliably excluded in these intoxications. Although the study design allows no statistical analysis, these conclusions are supported by the marked differences in K+ levels in gray and white matter induced by the different toxicants.

Alcoholic Intoxication↗

PubMed2001

Proliferative activity of epidermal basal cells after wounding. AgNOR counts compared with bromodeoxyuridine reactivity in rats.

Quantitative changes in nucleolus organizer regions (NORs) are known markers of proliferation that can be demonstrated by a specific silver staining technique on paraffin-embedded sections. Wounding of skin induces proliferation of basal epidermal cells at the wound margin. The degree of proliferation depends on the survival time and can be measured by morphometric assessment of argyrophilic NORs (AgNORs). Following incision wounding of the pinnae, rats were allowed to survive for different intervals (7 rats per interval) up to 120 hours. Before each sacrifice, biopsies were taken and incubated in a bromodeoxyuridine (BrdU) solution, embedded in paraffin, and stained with an antibody against BrdU. At the same time morphometric analysis of AgNOR counts was performed on sections made from the same material. BrdU incorporating nuclei were assessed by simple counting, whereas morphometric analysis of AgNOR counts was computer aided. Both methods revealed an increase in the number of proliferating cells, a plateau phase being reached after about 36 hours, followed by a decline after about 70 hours. Both methods thus allowed a reliable temporal classification of the skin injury according to survival time. The molecular background of the AgNOR changes in relation to the proliferation of cellular elements is discussed in detail.

Animals↗

PubMed2001

Creutzfeldt-Jakob disease (CJD) in a case of suspected chronic heavy metal poisoning.

We describe a patient who died of suspected heavy metal poisoning after a nine-month history of rapidly worsening dementia. Autopsy at a forensic-pathological institute established the postmortem diagnosis of sporadic Creutzfeldt-Jakob disease (CJD) based on demonstration of the proteinase-resistant prion protein (PrPsSc) in Western-Blot on native brain tissue. Microscopic examination of the macroscopically largely inconspicuous brain revealed marked spongiform changes in the gray matter--mainly affecting the cerebral cortex, nucleus caudatus, and putamen--with confluent vacuoles. Patchy or perivacuolar deposits of PrPSc were found as well as granular PrPsc deposits. The cerebellum contained focal PrPsc deposits. There was an astrogliosis in the white matter and a proliferation of microglia in the gray matter with a simultaneous clear reduction in neuronal elements. The differential diagnosis is discussed, as is the potential risk to those performing autopsy on forensic cases with a clinical picture of rapidly progressing dementia, especially in cases where a prion disease is not initially suspected.

Brain↗

PubMed2000

Quantification of mitochondrial DNA in human blood cells using an automated detection system.

The 4977 bp deletion of mitochondrial DNA (mtDNA) accumulates in postmitotic tissues with advancing age. The purpose of our study was to detect and quantify these deletion even in blood cells with a high turnover activity. Whole venous blood, isolated human platelets and peripheral blood mononuclear cells (PBMCs) were collected from 10 unrelated donors aged 20-71 years and total DNA was extracted. PCR was performed for total and mutated mtDNA using two different primer pairs and two fluorogenic probes labeled with the fluorescent dyes FAM and VIC. Specific PCR products were generated, detected and quantified in a real-time PCR. The amplification products of total and deleted mtDNA could be detected in each sample and did not exhibit any differences in the amount of the deleted mtDNA in whole blood, human platelets or PBMCs. Our data did not show any accumulation of the 4977 bp deletion with increasing age as it was observed for several other tissues.

Adult↗

PubMed2000

The shaking trauma in infants - kinetic chains.

The findings in three children who died as a consequence of shaking and those in another child who survived are presented. In the three fatal cases, a combination of anatomical lesions were identified at autopsy which appear to indicate the sites where kinetic energy related to the shaking episodes had been applied thus enabling the sequence of events resulting in the fatal head injury to be elucidated. Such patterns of injuries involved the upper limb, the shoulder, the brachial nerve plexus and the muscles close to the scapula; hemorrhages were present at the insertions of the sternocleidomastoid muscles due to hyperextension trauma (the so-called periosteal sign) and in the transition zone between the cervical and thoracic spine and extradural hematomas. Characteristic lesions due to traction were also found in the legs. All three children with lethal shaking trauma died from a subdural hematoma only a few hours after the event. The surviving child had persistant hypoxic damage of the brain following on massive cerebral edema. All the children showed a discrepancy between the lack of identifiable external lesions and severe internal ones.

Arm Injuries↗

PubMed2000

AgNORs during the process of wound healing. Time dependency as evaluated in vital and postmortem biopsy.

Silver staining of paraffin sections to detect changes in nucleolar organizer regions (AgNORs) is an established method for detecting cellular proliferation. To determine whether AgNORs are helpful in assessing wound age and vitality, we examined intravital and postmortem skin biopsies from rats surviving incised wounds to both pinnae for defined intervals up to 384 h using, 7 rats per time interval. One biopsy was taken immediately before death, a second 24 h after been sacrificed and storage at 8 degrees C. Interactive computer-assisted image analysis revealed that in the first 120 h after trauma the total number of AgNORs, the mean AgNOR sum area in the nucleus, and the mean AgNOR area per nucleus were dependent on survival time. Taken as indicators of proliferative activity these morphological phenomena revealed an increase in proliferative activity after survival times ranging from 10 to a maximum of 96 h. The findings were the same in both intravital and postmortem biopsies. These findings are in accordance with those obtained using bromodeoxyuridine. The value of these experimental findings in assessing wound age of human beings is discussed.

Animals↗

PubMed2000

Histochemical characterization of cytotoxic brain edema. Potassium concentrations after cerebral ischemia and during the postmortem interval.

Cytotoxic edema is a phenomenon of the ischemically damaged brain. In the present study we tested a histochemical method that detects this phenomenon based on potassium (K+) levels in the brain. In a first series focal cerebral ischemia was induced by arterial occlusion in 23 gerbils (Meriones unguiculatus). After survival times of 30, 60 and 120 min, the animals were killed and brain section histochemically stained for potassium and quantitatively evaluated with a morphometric method. The results were compared with those using physicochemical techniques. A distinct K+ depletion could be demonstrated in the area of the focal ischemia within a survival time of 30 min, the depletion growing thereafter with increasing survival time. In a second series histochemical and chemical methods were used to study the stability of K+ levels in undamaged brains of 15 healthy rats during postmortem intervals of 2.5 and 5 h. Within these intervals K+ levels were clearly depleted, apparently as a result of cerebral spinal fluid (CSF) diffusion. Even if neuronal injury can be demonstrated histochemically after very brief survival times of about 30 min, postmortem storage of the cadavers rendered detection impossible due to electrolyte and water diffusion. In autoptic human cases, therefore, this technique is of no practical utility in detecting cytotoxic brain edema in postmortem tissue.

Animals↗

PubMed2000

Brain injury after gunshot wounding: morphometric analysis of cell destruction caused by temporary cavitation.

In addition to the primary destruction of brain tissue readily visible at autopsy (permanent track), gunshot wounding to the brain creates a pulsating temporary cavity due to radial expansion along the bullet's track. To determine the maximum extent of this temporary cavitation in brains of victims of gunshots from weapons with low muzzle velocity, we carried out morphometric studies on 20 cases of death from gunshot wounding to the head from bullets with a muzzle energy <500 J and a survival time of <90 min. The brains were fixed in formalin, examine macroscopically and microscopically, and subjected to morphometric analyses. Surrounding the permanent track, a narrow mantle-like zone of astrocyte destruction was found within an area of hemorrhagic extravasation. Axons near the permanent track had been broken into tiny fragments. The axonal damage lessened with increasing distance from the permanent track, although axons continued to be fragmented and to exhibit varicose changes and clumping until assuming their normal structure beyond 18 mm. Nerve cells were extremely shrunken close to the permanent track but gradually took on their normal shape with increasing distance. We also assessed the loss of glial fibrillary acid protein expression by astrocytes in the white matter, the extent of traumatic bleeding, and damage to axons and neurons as measured radially from the permanent track. Axonal and neuronal damage were found to extend about 18 mm radially from the permanent track, tapering gradually along the track from entry point to exit point. The destruction was probably produced by the temporary cavitation and accords with theoretical considerations and experimental observations.

Adult↗

PubMed2000

Mononuclear cells in subcutaneous haemorrhage with special consideration of myeloid percursor cells.

Various hematogenous markers were used to differentiate and quantify the types of mononuclear cells present in subcutaneous haemorrhages. Fifty samples of subcutaneous bleeding with a survival time of a few minutes to more than 48 hours were studied. The various cell types were detected using the following stains: Naphthol AS-D chloracetate esterase for myeloid cells, including mast cells; (alpha1-antichymotrypsin for monocytes/macrophages; UCHL1 for T-lymphocytes; and L26 for B lymphocytes. The percentage of monocytes/macrophages was found to increase in dependence on survival time, whereas T-lymphocytes declined. Within minutes of injury neutrophilic granulocytes had emigrated into the surrounding tissue and mast cell degranulation had occurred within the haemorrhagic zone. Esterase-positive mononuclear cells, namely metamyelocytes, were detected within minutes after injury and were still present after survival times exceeding 48 hours; however, no dependence on survival time or cause of death was found. Although the increasing number of monocytes/ macrophages and T-lymphocytes was expected, the sometimes high percentage of myeloid precursor cells within the wound were surprising. Possible explanations for this phenomenon are discussed.

Adolescent↗

PubMed2000

Petechiae of the baby's skin as differentiation symptom of infanticide versus SIDS.

The successive killing of three siblings by their biological mother at two-year intervals is described. The children were 367 days, 75 days and 3 years old. Although sudden infant death syndrome (SIDS) or interstitial pneumonia could not be ruled out as the cause of death in the two younger children, who were killed first, the third child exhibited discrete signs of violence in the mouth and throat area which were interpreted as proof of infanticide. All three children had petechiae of the skin of the face and throat, the upper thorax, the shoulders and the mucous membranes of the mouth. None of the children exhibited signs of a disease-related hemorrhagic tendency. After the mother was convicted of murdering the three-year-old boy by smothering in combination with compression of the thorax, she confessed to having killed the other two children in a similar manner. In the absence of hemostatic disease, the presence of petechiae of the skin extending over the entire drainage area of the Vena cava superior can be regarded as evidence of an increase in pressure in the thoracic cavity secondary to obstruction of the airways with simultaneous chest compression.

Asphyxia↗

PubMed2000

Sudden unexpected infant death due to fibroma of the heart.

A 7-month-old previously healthy female infant was found dead in her crib by her mother shortly after having been laid down to sleep following the noontime feeding. Because the child did not suffer from an acute illness and no other evidence pointed to a cause of death, it was initially assumed by the police that she had died of sudden infant death syndrome. At autopsy, however, the cause of death was determined to be cardiac arrhythmia secondary to fibroma of the heart.

Cause of Death↗

PubMed1999

Estimation of age at death based on quantitation of the 4977-bp deletion of human mitochondrial DNA in skeletal muscle.

The 4977-bp deletion in human mitochondrial DNA (mtDNA) is known to accumulate in various tissues with age. Since this deletion in mtDNA correlates closest with age in muscle tissue, iliopsoas muscle tissue was taken at autopsy from 50 persons aged 24-97 years to determine whether age at death can be estimated based on the amount of the 4977-bp deletion in skeletal muscle. Total DNA (nuclear and mtDNA) was extracted from 100 mg tissue and the 4977-bp deletion quantified using a kinetic polymerase chain reaction (PCR) followed by visualization of the products on silver stained polyacrylamide gels. The amount of the 4977-bp deletion of mtDNA ranged from 0.00049% to 0.14% depending on age, with a correlation coefficient of r = 0.83 (P = 0.0001). In forensic practice this method can aid in the estimation of age at death with a relatively wide confidence interval, thus enabling a discrimination between young and elderly persons in the identification of human remains based solely on skeletal muscle.

Acrylic Resins↗