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Biomedical subjects

R I Carp

Publications and source records attributed to R I Carp.

At least 37 records · Page 2Linked to original sources
PubMed1999

Immunogold study of regional differences in the distribution of glucose transporter (GLUT-1) in mouse brain associated with physiological and accelerated aging and scrapie infection.

Distribution of glucose transporter (GLUT-1) in brain microvascular endothelia, representing the anatomic site of the blood-brain barrier (BBB), was studied in adult, physiologically aged, senescence-accelerated prone (SAMP8) and in scrapie-infected mice. Sections of tissue samples obtained from four brain regions (cerebral cortex, hippocampus, cerebellum, and olfactory bulb) and embedded in Lowicryl K4M were exposed to anti-GLUT-1 antiserum followed by gold-labeled secondary antibody. Labelling density was recorded over luminal and abluminal plasma membranes of the microvascular endothelial cells. We found that the density of immunosignals for GLUT-1 in the cerebral cortex showed a tendency toward insignificant diminution according to the following gradation-adult > SAMP8 > scrapie > aged mice-whereas in the hippocampus, this gradation was slightly different: adult > aged > scrapie > SAMP8 mice. In the cerebellum, immunolabelling was insignificantly diminished in aged mice, whereas it was significantly decreased in scrapie-infected and SAMP8 mice. The intensity of labelling of the vascular endothelium in the olfactory bulb was significantly lower than that in other brain regions, showing a slight decrease in the following sequence: adult > aged > scrapie > SAMP8 mice. These findings suggest that the process of aging as well as of related neurodegenerative disease affects unequally the distribution of GLUT-1 in the vasculature of different brain regions.

Aging↗
PubMed1999

Alteration of free radical metabolism in the brain of mice infected with scrapie agent.

Alteration of free radical metabolism in the mouse brain by scrapie infection was evaluated. The infection of mice with scrapie agent, 87V strain, slightly increased the activities of catalase and glutathione-S-transferase, while it had no effect on glutathione peroxidase, glutathione reductase, and Cu, Zn-superoxide dismutase. Results show that the scrapie infection decreased the activity of mitochondrial Mn-superoxide dismutase by 50% but increased that of monoamine oxidase (p < 0.05). Scrapie infection also increased the rate of mitochondrial superoxide generation (p < 0.05). Following scrapie infection, the level of free-sulfhydryl compounds in brain homogenates slightly decreased, but the content of thiobarbituric-acid-reactive substances and malondialdehyde increased significantly. Electron microscopy indicated that the ultrastructure of mitochondria was destroyed in the brain of scrapie-infected mice. These results suggest that elevated oxygen free radical generation and lowered scavenging activity in mitochondria might cause the free radical damage to the brain. Such deleterious changes in mitochondria may contribute to the development of prion disease.

Animals↗
PubMed1999

Scrapie strain-specific interactions with endogenous murine leukaemia virus.

The finding that a senescence-accelerated mouse (SAMP8) shows early brain ageing, with histopathological changes resembling those seen in scrapie, combined with the discovery of high levels of endogenous murine leukaemia virus (MuLV) in brains of SAMP8 mice prompted us to examine the effect of scrapie infection on MuLV titres in this strain and in one of its progenitors, the AKR strain. Three scrapie strains (ME7, 22L and 139A) that had a comparatively short incubation period in SAMP8 and AKR mice caused an increase in brain MuLV titres that was scrapie strain-specific: in each mouse strain, the greatest effect was with 1 39A, and the least with ME7. The 22A scrapie strain, which has a long incubation period in SAMP8 mice, did not affect MuLV titres in brains of this mouse strain. Previous analyses of scrapie incubation periods in AKR, SAMP8 and another strain derived from an AKR cross (SAMR1) showed an inverse relationship between brain MuLV titres and scrapie incubation periods. This finding, combined with the effect of scrapie on MuLV titres, suggests an interaction between the scrapie infectious process and MuLV replication.

Animals↗
PubMed1999

Abnormal periodic acid-Schiff (PAS)-positive substance in the islets of Langerhans, pituitaries and adrenal glands of 139H scrapie-infected hamsters.

Previous studies showed that the 139H strain of scrapie injected intra-cerebrally in hamsters caused obesity, and extensive histopathological changes in islets of Langerhans and pituitaries. In the current study, we report that an abnormal granular substance, which stained positively with periodic acid-Schiff (PAS-positive substance; PPS), was found in the islets of Langerhans, pituitaries, adrenal glands, in the lumens of blood vessel cores (BVCs) and in blood vessels in 139H-infected hamsters, but not in either 263K-infected or control hamsters. This substance was found in the endocrine organs, forming grape-like or plaque-like structures, which were small, round to ovoid, and homogenous measuring up to 7 microns in diameter and usually grouped in clusters. PPS was not found in the brains of control or scrapie-infected hamsters. Using immunostaining for amyloid protein (PrP, beta A4), as well as Congo red and thioflavin-S stains, no evidence was found of amyloid plaque formation in the islets of Langerhans, the adrenal glands, or the pituitaries of 139H- or 263K-infected hamsters. PPS might relate to the pathological changes in the endocrine organs in 139H-infected hamsters.

Adrenal Glands↗
PubMed1999

Immune surveillance and antigen conformation determines humoral immune response to the prion protein immunogen.

Transmissible spongiform encephalopathies (TSE) are progressive degenerative disorders of the central nervous system. PrP(Sc) is a TSE-specific marker derived from the host-encoded glycoprotein, PrPc. The generation of antibodies to PrP plays an important role in the diagnosis of these diseases. In this study the role of the PrP immunogen and the species being immunized was examined in relation to specific epitopes. Various mammals (mice, hamsters, rabbits and PrP null mice) were immunized with formic acid-treated PrP(Sc) isolated from mice, hamsters and sheep. Both the species being immunized and the source of immunogen played an important role in the antibody response. Response to a limited number of linear epitopes was seen among the various immunized animals. One region in the C-terminal portion of PrP appeared highly immunogenic in all species. Comparison of immunoreactivity and the pepscan-defined linear epitope sites suggests both linear and conformational directed responses in many of the animals. Information on the forces directing immune responses to PrP will lead to a better understanding of host-PrP interactions. It will also assist in the development of new strategies for generating additional tools for immunodiagnosis.

Amino Acid Sequence↗
PubMed1998

Astrocytosis and amyloid deposition in scrapie-infected hamsters.

In scrapie infection, prion protein (PrPSc) is localized in areas where there is neurodegeneration and astrocytosis. It is thought that PrPSc is toxic to neurons and trophic for astrocytes. In our study, paraffin sections from scrapie infected (263K and 139H) and control hamsters were examined with histological and immunocytochemical staining. We found that PrPSc was present in the ependymal cells of both 263K- and 139H-infected hamsters. In 139H-infected hamsters, PrPSc was found in the cytoplasm of neurons in cerebral cortex and in hypothalamic paraventricular (PVN) and supraoptic (SON) nuclei. In contrast, neuronal cytoplasm and nuclei, were positive for PrPSc in most areas such as cortex, hippocampus, and thalamus in 263K-infected hamsters. Many aggregations of PrPSc could be seen in the cortex, hippocampus, substantia nigra and around the Pia mater, corpus callosum, fimbria, ventricles, and blood vessels in sections from 139H- and/or 263K-positive animals. Furthermore, PrPSc was also co-localized with glial fibrillary acidic protein (GFAP) in many reactive astrocytes (approximately 90%) in certain areas such as the hippocampus in 263K-infected hamsters, but not 139H-infected hamsters. The patterns of astrocytosis and PrPSc formation were different between 139H- and 263K-infected hamsters, which may be used for a diagnosis purpose. Our results suggest a hypothesis that multiple cell-types are capable of PrPSc production. Our results also confirm that reactive astrocytes can produce and/or accumulate PrPSc during some scrapie strain infections. The findings suggest a 'snowball effect', that is: astrocytosis might play an important role in amyloidosis, while amyloidosis may induce further astrocytosis at least in 263K-infected hamsters.

Amyloidosis↗
PubMed1998

Mitochondrial dysfunction induced by oxidative stress in the brains of hamsters infected with the 263 K scrapie agent.

Scrapie, one of the prion diseases, is a transmissible neurodegenerative disease of sheep and other animals. Clinical symptoms of prion diseases are characterized by a long latent period, followed by progressive ataxia, tremor, and death. To study the induction of neurodegeneration during scrapie infection, we have analyzed the activities of various antioxidant enzymes and mitochondrial enzymes in cerebral cortex, brain stem, and cerebellum of scrapie-infected hamsters. The activity of mitochondrial Mn-superoxide dismutase (SOD) was decreased, while the activities of cytosolic Cu/Zn-SOD and catalase were not altered in infected brains. The activities of glutathione peroxidase and glutathione reductase were increased in scrapie-infected hamsters. The decreased activity of Mn-SOD might result in increasing oxidative stress in the mitochondria of infected brain; this concept is supported by our findings of a high level of lipid peroxidation, and low levels of ATPase and cytochrome c oxidase activity in the infected cerebral mitochondria. In addition, structural abnormalities of mitochondria have been observed in the neurons of hippocampus and cerebral cortex of infected brain. These results suggest that mitochondrial dysfunction caused by oxidative stress gives rise to neurodegeneration in prion disease.

Adenosine Triphosphatases↗
PubMed1998

Analysis of the incubation periods, induction of obesity and histopathological changes in senescence-prone and senescence-resistant mice infected with various scrapie strains.

The similarity in histopathological changes seen in scrapie-infected mice and in an uninfected senescence-accelerated mouse strain led to a study in which the mouse strain that is prone to senescence (SAMP8), a strain that is resistant to senescence (SAMR1) and a progenitor strain (AKR) of these two strains were infected with three different scrapie strains, ME7, 139A and 22L. For each scrapie strain, the incubation period was shortest in AKR mice and longest in SAMR1 mice. The induction of obesity was a function of scrapie strain and not mouse strain; ME7 caused obesity in all mouse strains, whereas the average weights of mice injected with 139A and 22L did not differ significantly from mice injected with homogenates of normal mouse brain. The pattern of vacuolation seen in the brain of each mouse strain was primarily dependent on the scrapie strain injected. There were, in general, similarities to the patterns induced in other inbred strains; e.g. ME7 caused extensive forebrain vacuolation, 22L caused prominent vacuolation in the cerebellum, and the 139A strain induced characteristic white matter vacuolation. Vacuolation was also seen in the medulla and midbrain of SAMP8 mice injected with normal mouse brain, which is consistent with the occurrence of accelerated ageing changes in the brain of this strain. Further analysis of the differences among these mouse strains should provide information relating to the observed differences in scrapie incubation periods.

Aging↗
PubMed1998

Astrocytosis and proliferating cell nuclear antigen expression in brains of scrapie-infected hamsters.

Scrapie is a neurodegenerative disease in sheep and goats. Neuropathological examination shows astrocytosis. One issue is whether the astrocytosis seen in scrapie is a function of an increase in reactivity of individual cells, or whether there is actual replication of astrocytes. We used double-label immunohistochemistry for proliferating cell nuclear antigen (PCNA) and for glial fibrillary acidic protein (GFAP) to determine the mitotic state of cells and to confirm their identity as astrocytes. Brain sections from hamsters (strain LVG/LAK) infected with 139H or 263K scrapie isolates were examined. GFAP immunostaining was increased in astrocytes in most regions of the brains of scrapie-infected hamsters. These qualitative observations were confirmed by computerized image analysis quantification. A proportion of the hypertrophic astrocytes (0.5-10.8%, depending on specific location) were PCNA immunoreactive. The PCNA-immunopositive astrocytes were most frequently found in cerebral cortex, corpus callosum, subependymal areas, fimbria, caudate, thalamus, hypothalamus, hippocampus, and dentate gyrus. Our results suggest that the astrocytosis seen in scrapie-infected animals is, at least in part, owing to actual replication of astrocytes in these animals. We hypothesize that the astrocytes may be an important locus for the disease process.

Animals↗
PubMed1998

Expression of inducible nitric oxide synthase in the brains of scrapie-infected mice.

The neuronal cell damage caused by inducible nitric oxide synthase (iNOS) in brain has been reported to be associated, at least in part, with many neurodegenerative diseases including Alzheimer's disease. We recently observed vacuolation and astrocytosis in the brains of ME7 scrapie strain-infected C57BL mice. To investigate if these phenomena might have a relationship to iNOS, the level of iNOS expression was measured immunohistochemically and molecular biologically in the brains of scrapie-infected C57BL mice. The number and size of astrocytes were increased and immunoreactivity of glial fibrillary acidic protein (GFAP) was significantly enhanced. iNOS immunoreactivity was observed in the astrocytes of the scrapie-infected group, but not in the control group. iNOS mRNA levels were increased in scrapie-infected mice compared to the levels in non-infected mice of the same age. Our results suggest that iNOS induction in reactive astrocytes is a part of the neurodegenerative mechanisms in scrapie infection.

Animals↗
PubMed1997

Immunocytochemical evaluation of blood-brain barrier to endogenous albumin in scrapie-infected mice.

A quantitative immunocytochemical procedure was used for evaluation of the blood-brain barrier (BBB) to endogenous albumin in plaque-forming (PF) and non-plaque-forming (NPF) groups of scrapie-infected mice at the clinical stage of disease. Ultrathin sections of brain samples (cerebral cortex, hippocampus and cerebellum) embedded in resin (Lowicryl K4M) were exposed to anti-mouse albumin antiserum followed by protein A-gold. Using morphometry, the density of immunosignals (gold particles per microns2) was recorded over four compartments: vascular lumen, endothelium, subendothelial space, and brain parenchyma (neuropil). Morphometric and statistical analyses did not reveal significant differences in the barrier function of the microvasculature of the cerebral cortex and hippocampus in either group of mice, although a slight increase in the number of leaking vessels in the PF group was noted. In contrast, in the cerebellum, the permeability of the microvessels to albumin was significantly higher in the PF than in the NPF mouse group, and this was paralleled by the infiltration of the walls of numerous vascular profiles with amyloid deposits (amyloid angiopathy). These data also indicate the existence of distinct regional differences in BBB function in the brain of scrapie-infected mice. The vascular amyloid deposits and the amyloid plaques present in the cerebral cortex of PF mice were labeled with numerous immunosignals suggesting the affinity of extravasated albumin to these deposits. In conclusion, no convincing evidence was obtained indicating that impairment of the BBB, manifested by increased permeability of vascular segments, is directly related to the deposition of amyloid in the vascular wall and in plaques. Segmental impairment of the barrier function seems to be rather the result of disturbed structural integrity of the components of the vascular wall.

Animals↗
PubMed1997

Scrapie strains retain their distinctive characteristics following passages of homogenates from different brain regions and spleen.

The molecular basis of differences among scrapie strains is unknown. The prion theory posits that there are differences in the conformation of the host protease-resistant protein (PrP) molecules and that these differences are responsible for scrapie strains. A corollary of this theory is that the origin of host PrP variation resides in different neuronal cell types. To assess this concept, preparations from three brain regions (cerebrum, cerebellum and olfactory bulb) and from spleen were passaged in C57BL mice by intracerebral injection. After three passages of three scrapie strains in this manner, homogenates of each brain region and spleen were tested for several of the characteristics that distinguish the three strains: (1) the rank order of incubation periods in C57BL mice, (2) induction of obesity in SJL mice and (3) comparative incubation periods in mice with three genotypes for the scrapie incubation period marker. Analysis revealed that virtually all of the criteria that distinguished the three strains prior to passages of the three brain regions and spleen were retained after this series of passages. This finding argues against cellular-based PrP differences providing a basis for strain specificity.

Animals↗
PubMed1997

The 139H scrapie agent produces hypothalamic neurotoxicity and pancreatic islet histopathology: electron microscopic studies.

Neuronal degeneration, along with astrocytosis, spongiform vacuolation, and amyloid (PrPSc) formation, have long been regarded as neuropathological hallmarks of transmissible spongiform encephalopathies (TSEs). In animals, these diseases include; scrapie, transmissible mink encephalopathy, chronic wasting disease, bovine and feline spongiform encephalopathies, and in humans; kuru, Creutzfeldt-Jakob disease (CJD), and Gerstmann-Sträussler-Scheinker syndrome (GSS). The abnormal amyloid protein, (PrPSc) is toxic to neurons. Our previous studies showed that hamsters treated with 139H scrapie strain developed obesity, and generalized endocrinopathy, including lesions in hypothalamus, pituitary and pancreas. Histochemical and immunocytochemical studies revealed extensive pathological changes in the islets of Langerhans in 139H-infected hamsters, but not in hamsters infected with 263K scrapie strain. Using routine electron microscopy (EM), we have observed more details of lesions in the beta cells of islets of Langerhans in these animals. Cytoplasmic vacuolation occurred, cytoplasmic organelles were found damaged and disrupted, and membranes were occasionally ruptured. The width of endoplasmic reticulum (ER) lumina were 50-150 nm in controls, whereas in 139H-infected hamsters, they wee occasionally increased up to 4000 nm in diameter. Most beta cells showed degranulation. These EM observations suggest that the cellular death seen in the islets of Langerhans in 139H-infected hamsters is due to necrosis, not apoptosis. Since there were no amyloid deposits found in the islet of Langerhans at the EM level, and there were extremely low scrapie infectivity levels and PrPSc levels in pancreas, it is suggested that the changes noted in pancreas were not a direct toxic effect of PrPSc. Instead, our study suggests that scrapie prion protein PrPSc, acting as a neurotoxicant, alters the hypothalamic neuroendocrine regulation of the pancreas.

Animals↗
PubMed1996

Margination and diapedesis of inflammatory cells in the islets of Langerhans in hamsters infected with the 139H strain of scrapie.

The islets of Langerhans in hamsters infected with the 139H strain of scrapie contain large masses of red blood cells not surrounded by the usual arterial, venous or capillary wall cells. We have referred to these structures as "blood vessel cores" (BVCs). BVCs were almost always centrally located within the islets and surrounded by pancreatic B cells. Margination and diapedesis of inflammatory cells were observed at the BVC walls in 139H-infected hamsters. The cells consisted of the following types: single or clustered lymphocytes; and mixtures of lymphocytes and macrophages or neutrophils. Interaction observed between groups of inflammatory cells and B cells at the BVC walls and inside the islets of Langerhans indicated an inflammatory process. We refer to this interaction as the "linkage-reaction", and to the inflammatory cells as "linkage-inflammatory cells". These phenomena were not observed in other organs (adrenal, uterus, ovary, spleen, liver, kidney, oesophagus, trachea, intestine or pituitary) in 139H-affected hamsters or in the islets of Langerhans of animals infected with other scrapie strains (263K-infected hamsters; 139A-, ME7- and 22L-infected SJL mice). This appears to represent the first clear evidence of an inflammatory reaction in any organ in scrapie-infected animals.

Animals↗
PubMed1996

Histopathological changes in the pituitary glands of female hamsters infected with the 139H strain of scrapie.

Previous studies in hamsters showed that the 139H strain of scrapie injected intracerebrally caused a generalized endocrinopathy and marked hypoglycaemia and hyperinsulinaemia. The low scrapie infectivity levels in the pancreas suggested that the changes noted in that organ were of neuroendocrine origin. In the current study, female weanling Syrian hamsters were inoculated intracerebrally with scrapie strain 139H or 263K, or with homogenate of normal hamster brain. Coronal sections of the pituitary gland were stained with haematoxylin and eosin, Gomori's one-step trichrome, Congo red, thioflavin-S, and antibodies specific for several pituitary hormones. Sections were examined by light microscopy. The hamsters inoculated with scrapie strain 139H showed extensive pituitary vacuolization. Most vacuoles were located in the ventral or ventrolateral parts of the pars distalis. The pituitary glands of 139H-infected hamsters also showed cellular changes, namely, hypertrophy, atrophy and cytoplasmic vesicles. Nuclear changes such as swelling, vesicle formation, chromatin increase, pyknosis, karyorrhexis and karyolysis also occurred. The cellular and nuclear changes were most pronounced in the regions with vacuolation. Hamsters infected with the 263K strain did not show these changes. Immunocytochemical examination suggested that parenchymal cell types which produce different hormones were affected in areas of vacuolation. The changes produced by 139H were not seen in hamsters infected with strain 263K. This study provides the first evidence of cytopathological changes in the pituitary glands of scrapie-infected animals and suggests a relationship between the pituitary changes and the pathological findings in the pancreas and other endocrine organs of 139H-infected hamsters.

Animals↗
PubMed1996

Histopathological changes in the islets of Langerhans in hamsters infected with the 139H strain of scrapie: semi-thin section study.

Using histopathological analysis of semi-thin sections stained with toluidine blue, we observed profound pathological changes in the islets of Langerhans of hamsters infected with the scrapie agent (strain 139H). These included cytoplasmic vesicles, nuclear swelling, and vacuolization in the islet cells. Two types of vacuolization were seen. "Localized vacuolization" (LV) has a distinct edge and is restricted or confined within the cell. "Diffuse vacuolization" (DV) has no distinct edge and is scattered within tissues either inside or outside of cells. DV may span intracellular and extracellular regions of the islet tissues. There were abnormal structures which we termed blood vessel cores (BVCs) in the islets of 139H-infected hamsters. BVC is a hollow space filled up with blood cells. Immunocytochemical staining for insulin antibody suggested that BVC was surrounded by the B cells of the islet. In the present study, we observed that many inflammatory cells passed through the blood-tissue barriers using pathways between cell-junction in the lumen of BVC. We also observed many necklace-like hollow spaces between islet cells. They are the pockets of extracellular space. A novel concept of "the accordion effect" was described to explain a function of the extracellular space. Under normal physiological conditions, as the synthesis of insulin increase in B cells, the volume of the B cells will increase while the volume of the extracellular space will decrease. After a synchronized secretory response from the stimulated B cells, the secretory product would move from the intracellular space into the extracellular space, the volume of the B cells would be decreased and the volume of the extracellular space would be increased. Most of the secretory product might be released into the blood stream immediately, causing an insulin releasing peak in the blood stream, whereas the rest would remain in the enlarged extracellular space. As the cycle repeat, the increasing volume of the B cells will squeeze the remaining insulin into the blood stream gradually. Thus, the expandable extracellular space would serve as buffer system and a reservoir to collect and store some secretory products for future use. We refer to this concept as "the accordion effect". The concept of "the accordion effect" may also be true in other endocrine organs such as pituitary gland and adrenal gland.

Animals↗
PubMed1995

The pathological changes in peripheral organs of scrapie-infected animals.

Scrapie is an unconventional neurodegenerative disease in sheep and goats that has been known in Europe for over 260 Years. The scrapie agents affect the brain and are transmissible from animal to animal. Key features of scrapie infections are abnormal behavior and deficits in motor function. These clinical findings can be related to the damage found in the central nervous system. In some scrapie strain-host model systems there are other manifestations of disease that appear to be related to pathological changes found in the peripheral organs, especially in the endocrine organs such as pituitary, adrenal glands, the islet of Langerhans and ovary. In those model systems in which extensive histopathological changes have been seen in peripheral organs, the titers of scrapie infectivity and the levels of the scrapie specific protein, PrPSc, are relatively low in the affected organs. These data suggest but do not prove that changes in peripheral organs are secondary to the scrapie-induced neurodegeneration that is occurring in the brain. In some scrapie strain-host combinations, obesity and aberrant glucose metabolism are seen in the preclinical and clinical phases of the incubation period. There appear to be two pathways that lead to these particular clinical manifestations. In SJL mice infected by the ME7 or 22L strains of mouse-adapted scrapie and in some scrapie-infected sheep, the mechanism is related to changes induced in the hypothalamic-pituitary-adrenal axis. The other pathway is exemplified by hamsters infected with two hamster-adapted scrapie strains, 139H and 22CH; it appears that lesions found in the hypothalamic-islets of Langerhans axis are critical. A number of reviews on the pathological changes in the central nervous system have been published and therefore, in this review article, we focus on the gross and histopathological changes in peripheral organs in several scrapie strain-host combinations. The changes induced in peripheral organs in a number of scrapie strain-host combinations expand the number of diseases in which the unconventional slow infections could serve as a model. Further work in this area could help us to understand the mechanisms and pathways of the pathological changes found in the peripheral organs of the scrapie-infected animals.

Animals↗