Temporal Sequence of Post-Mortem Autolysis in the Mouse Retina.

In order to better differentiate ante-mortem lesions from post-mortem retinal autolysis, the temporal sequence of post-mortem changes was studied in a well-controlled mouse model. Mice were of the same strain, age and sex, and were held at a constant ambient temperature. Eyes were collected at various times up to 72 h after death and immersion-fixed in either Davidson's fixative or 10% neutral buffered formalin, paraffin-embedded and sections cut and stained with haematoxylin and eosin. The most prominent, and early, autolytic change was retinal detachment, and subsequent folding, which occurred immediately after death in formalin-fixed eyes, but not until 2 h post mortem with Davidson's fixative. Retinal separation was complete at 16 h, or almost complete by 2 h, in formalin, but in Davidson's fixative, was only partial and segmental, the latter not becoming total until much later. Retinal detachment was attended by progressively more severe disruption and dissolution of photoreceptors and, particularly in Davidson's-fixed retinas, the rod outer segment often showed marked homogenization from 30 min to 4 h after death. The other major early change was nuclear pyknosis in the inner nuclear layer. Ganglion cells initially had cytoplasmic swelling, followed by shrinkage and basophilia (at 4 h with formalin and 16 h with Davidson's), with nuclear pyknosis becoming increasingly common over time. While the three retinal neuronal layers eventually became more attenuated and depleted of cells, the thickness of these layers was augmented by severe swelling. These findings show that the post-mortem interval at which histological interpretation of retinal changes becomes potentially compromised is dependent on the duration of this interval and the fixative used.

Autólise da camada de células granulares do cerebelo em bovinos / Conglutination of the cerebellar granule cell layer in cattle

A autólise da camada de células granulares do cerebelo é um achado postmortem observado em bovinos e relacionado ao prolongado tempo de duração entre a morte e a fixação do encéfalo. Para observar a prevalência desse artefato, 228 cerebelos foram examinados histologicamente ao longo do verme cerebelar. Trinta e seis por cento desses casos apresentaram conglutinação cerebelar não relacionada a doença específica. Em 74,4 por cento dos casos em que foi observada conglutinação cerebelar, não foram observadas alterações histopatológicas. Histologicamente, a conglutinação cerebelar caracterizou-se por dissolução segmentar ou difusa da camada de células granulares. A descrição deste artefato é importante, pois já foi confundido como lesão relacionada a doença específica do sistema nervoso de bovinos.

Cerebellar conglutination of the granule cell layer is a postmortem artifact observed in cattle. It is related to increased time between death and fixation of the brain. In this study, 228 cerebellum were histologically analyzed throughout cerebellar vermis. Cerebellar conglutination was observed in 36 percent of the cases and it was not related to specific diseases. Seventy-four percent of the cases did not present any histological lesions in the brain. Microscopically, cerebellar conglutination was characterized by segmentar or diffuse dissolution of the granule cell layer. This artifact has been misinterpreted as resulting from specific diseases of the central nervous system in cattle, and it has to be differentiated from important antemortem lesions.

A comparison of rapid bovine spongiform encephalopathy testing methods on autolyzed bovine brain tissue.

In 1999, the European Union (EU) approved 3 rapid methods for the testing of bovine brain samples for the presence of bovine spongiform encephalopathy (BSE). The evaluation that led to the approval did not include an analysis of autolyzed material. Member states of the EU have active surveillance programs for BSE, which target fallen stock as well as other categories of cattle. Autolysis is a common feature of fallen stock samples because there can be a considerable delay between death and collection of samples. Therefore, it is important to know whether these tests perform optimally on autolyzed samples. The Veterinary Laboratories Agency (VLA) selected 250 positive fallen stock samples. These had been detected during routine testing using the Prionics-Check Western blot and confirmed as BSE cases by immunohistochemistry or electron microscopy. Samples were graded according to the degree of autolysis and then tested by the 3 methods: Prionics-Check Western blot, Platelia test, and Enfer test. All 3 methods correctly classified the samples as positive BSE cases, therefore alleviating doubt about their ability to do so. Subsequent EU validation exercises, such as those conducted in 2002--2003, have included the testing of autolyzed material. It is important that all new methods be evaluated on autolyzed tissue before approval for official use.

Hepatocyte vacuolation and autolytic changes in the liver of pilot whales, Globicephala melas, stranded on Cape Cod, MA, USA.

Most cetacea available for internal sampling in recent times have died through mass or single stranding events. It is important to know how the time elapsed between death and sampling affect quality of tissues. This study evaluated histological quality in the liver of long-finned pilot whales that either died or were euthanased after mass stranding events. Histological detection of significant autolysis was found in animals when 2 or more hours elapsed between death and sampling. In addition, hepatocytes often had marked idiopathic cytoplasmic vacuolation that did not stain with hematoxylin and eosin. The extent of this vacuolation did not show any correlation with time between death and sampling, but did appear more often in animals of greater total length. These observations suggest that when animals die or are euthanased at a single or mass stranding, every effort should be made to obtain samples as soon as possible, although meaningful histological observations can still be made in the presence of significant autolysis. These data also suggest that a multi-disciplinary study should be conducted to determine whether increasing autolysis is associated with changes in the organic chemical residues, molecular biology, histopathology and microbiology of those tissues.

Microscopic postmortem changes in adrenal glands of the domestic fowl.

Eighty-four male white leghorn chickens were killed by CO2 gas to determine the type, rate, and sequence of microscopic postmortem changes in the adrenal glands of dry and wet intact carcasses. They were held at 29 or 18 C with 50% relative humidity for different times postmortem. The sequence of microscopic postmortem changes was similar in all chickens except at 18 C, when karyorrhexis of cortical and medullary cells was observed. Cellular changes occurred earlier at 29 C than at 18 C and in dry chickens but not in chickens wet with detergent solution before storage, although slight quantitative and qualitative differences between wet and dry chickens were noted. Medullary cells underwent postmortem changes earlier than cortical cells. Nuclei of medullary cells decreased in size, with chromatin clumping leading to pyknosis, followed by cytoplasmic vacuolation, cellular shrinkage, and finally karyolysis and cell dissociation. Cortical cells had nuclear chromatin marginated, nuclei reduced in size initially, and some nuclear fading, followed by pyknosis and karyolysis. Karyorrhexis was not a prominent feature of cortical and medullary cells, although it occasionally occurred before pyknosis. Cytoplasm of cortical cells remained eosinophilic, granular, and vacuolated, but vacuoles became finer later. Pyknotic medullary cells with vacuolated cytoplasm were observed as early as 3 hr postmortem, regardless of the temperature. Diffuse pyknosis of medullary cells was noted at 18 hr in chickens held at 29 C and at 48 hr in chickens held at 18 C. Marked cortical pyknosis was noted only at 36 hr in wet chickens held at 29 C, when bacterial invasion started. Dry chickens held 36 hr at 29 C had diffuse cellular dissociation, karyolysis and cytoplasmic acidophilia, and marked bacterial invasion. Erythrocytes were pyknotic and had cytoplasmolysis. It was concluded that adrenal glands may still be useful for histopathological examination before 18 hr at 29 C and before 48 hr at 18 C.

Sequential sterile autolysis in the ovine fetus: macroscopic changes.

Sterile intrauterine autolysis was experimentally produced in 28 ovine fetuses in the last 3rd of gestation by umbilical artery ligation. The fetuses were retrieved by cesarean section after periods of sterile intrauterine autolysis ranging from 0.5 to 168 hours. The following autolytic changes, frequently reported as significant lesions in infectious abortions in cattle and sheep, were observed: subcutaneous blood-tinged gelatinous edema, blood-tinged fluid in the serous cavities, renal cortical softening, uniform reddish brown tissues, hepatic friability and mottling, and cloudy yellow to cloudy red abomasal contents. The appearance of these and other autolytic changes corresponded to sterile sequential autolytic changes reported in the rabbit fetus and used to determine duration of retention of stillborn infants and are of comparative biomedical significance. The sequential autolytic changes may be used as an index to duration of retention after death of a ruminant fetus and provide a basis for reappraising various infective abortifacients common to ruminants by allowing separation of specific lesions from strictly autolytic changes.