Neuropathological analysis of pathological forms of astroglia in Wilson's disease.

A neuropathological study of Alzheimer type I (Alz I) and Alzheimer type II (Alz II) as well as Opalski (Opl) cells was performed serially on brain tissue from nine autopsied Wilson's disease (WD) cases. Conventional staining methods (Kluver-Barrera, HE, PAS) and immunocytochemical techniques (anti-GFAP and anti-Metallothionein-MT) were used. On conventional staining, each of the studied abnormal cell types retained common morphological characteristics of astroglia, and concurrently demonstrated its own distinctive features, specific only for a given cell type. Anti-GFAP staining revealed positive immunoreactivity of Alz I and Opl cells, and its absence in Alz II cells. On anti-MT staining both the cytoplasm and nucleus of Alz I and Opl cells showed positivity whereas in Alz II cells the cytoplasm was positive in contrast to the negative nucleus. The results of our study confirm the hypothesis of the astroglial origin of all three types of cells. The lack of immunoreactivity for GFAP and similar immunocytochemical staining patterns for MT in Alz II cells and protoplasmic astrocytes may suggest that Alz II cells originate from the protoplasmic type of astroglia. The fact that Alz I and Opl cells resemble fibrous astrocytes in their immunoreactive positivity for GFAP may lead to a supposition that they originate from the fibrous type of astroglia. MT-positive expression by the three abnormal cell types suggests that they may be involved in the process of copper detoxification in WD.

Neurones and microglia in central nervous system immune response to degenerative processes. Part 1: Alzheimer's disease and Lewy body variant of Alzheimer's disease. Quantitative study.

The quantitative correlation between neurone loss and brain immune response, assessed by intensity of microglia inflammatory reaction in cortical association area and limbic cortex, was investigated and compared in previously immunohistochemistry (IHC) and ultrastructural confirmed 11 cases of Alzheimer's disease (AD), 7 cases mixed form of Dementia with AD findings and Lewy bodies (AD/DLB) reported, in accordance with Consortium on Dementia, as Lewy body variant of AD (LBV) and 6 non-demented autopsy control cases from 63 to 86 years old. In the present work we investigated association and limbic cortical areas linked with memory mechanisms; there are regions characterised by early distribution of IHC confirmed AD and DLB/AD (LBV) markers, as well as a substantial physiological stability of neurone pool regardless of age. The results indicated that AD and LBV differ in their neurone loss intensity and inflammatory reaction, with much higher intensity in AD. In Alzheimer's disease, neurone loss in association temporal cortex made up 51% of control values with simultaneous 8-fold increase in the density of MHC II-positive activated microglia, whereas in LBV, both the loss of neurone density and the increase in activated microglia density, was not so high (up to 41% and 4-5-fold, respectively). Changes in the limbic cortex were less pronounced. A strong correlation in the clinical material between neurone loss and microglia activation in both processes, especially in AD (r = 0.73), speaks in favour of the hypothesis on the neuronal immune surveillance and arousal of immune brain response in conditions of declining control, due to significant neurone loss in the neurodegenerative process. The inflammatory reaction of MHC II-immunoreactive microglia, concomitant with neurodegenerative process, seems to be a consequence of increased immune response due to loss of neurones and weakening of their control upon immunosurveillance in central nervous system.

The fractal analysis of subdural haematoma.

The fractal analysis of vessel structure in the capsule of subdural haematoma has been performed. Examined material comprised 100 normal cases and 30 with long-lasting subdural haematoma. Pickworth's method and computer image processing under lmtron, Scion for Windows 98 were applied. The parallel vessels with fork-like branching were found in the subdural haematoma capsule. The essential result of the fractal analysis was that there was no difference in one-sided dimensions of subdural haematomas in contrast to differentiated one-sided dimensions of normal vessels. This finding may explain the continuous growth of subdural haematoma. The applied method allows observation of the microangioarchitecture in the capsule of subdural haematoma.

Hallervorden-Spatz disease in an adult patient.

Hallervorden-Spatz disease (HSD) is an extremely rare degenerative process. The familial studies point to inherited, autosomal recessive neurodegenerative disorder. Quite recently this disease gene has been identified to chromosome 20p12.3-p13. Clinical manifestations of HSD leading to death after several years of illness are most frequently observed in childhood. HSD in adults is very scarce. The case reported concerns a woman who at the age of 26 years began to suffer from slowly progressing psycho-organic syndrome with muscular rigidity, involuntary movements and dysarthria. The patient was hospitalized several times with successive diagnoses of multiple sclerosis, amyotrophic lateral sclerosis and Huntington's disease. Shortly before death magnetic resonance imaging (MRI) scan showed a decreased signal in both basal ganglia. The patient died at the age of 34 years after an eight-year illness. In the brain autopsy symmetric hyperpigmentation of globus pallidus (GP) and reticular part of substantia nigra (SN) was found. The microscopic observation revealed abundant deposits of brown pigment mostly in GP and SN. In addition, numerous spheroids disseminated in the basal ganglia, mesencephalon and medulla oblongata, as well as Lewy bodies in SN were noted. Pigment deposits expressed intensive iron positive reaction by Perls' Prussian-blue method. Based on the described neuropathological changes occurring mostly in GP and SN, Hallervorden-Spatz disease was diagnosed.

Border zone neovascularization in cerebral ischemic infarct.

Immunocytochemical and quantitative studies on vascular reaction (angiogenesis) in cortical border zone of infarct were undertaken. Intensity and temporal profile of angiogenesis was assessed in 60 patients aged between 48 and 69 (younger group), and between 70 and 92 (older group), with cerebral infarct in the area of middle cerebral artery vascularization, who died during the first six weeks following the stroke. We have found that angiogenesis was a multistage process in which four stages were distinguished: phase of primary activation of endothelial cells, two consecutive phases of active angiogenesis and final phase of only sporadic proliferation of vessels. The distinction of phases in a multiphase angiogenic cascade helped us to evaluate the correlation with survival time and the age of patients. The most pronounced intensification of angiogenesis and increased density of CD 31 positive capillaries in penumbra were observed in the second phase, especially in younger patients. The duration of the penumbral neovascularization decreased in the older age patient. Our results indicate that sprouting angiogenesis is a quantitatively significant source of vessels in the cerebral infarct border zone. However, non-therapeutically stimulated angiogenesis developed only 3-4 days after the stroke, that is beyond the period of reversible changes in ischemic penumbra recognized as a "therapeutical window" in the human brain. The angiogenic therapy opens a new way towards the revascularization of ischemic brain infarct.

[Modern views on the pathogenesis of Wilson's disease]. / Aktualne poglady na patogeneze choroby Wilsona.

Wilson's disease is a rare, multiorganic genetically coded disorder, induced by impaired copper transport. The recent identification of the disease gene and the discovery of gene product--copper transporting P-type ATPase that is integrate membranous protein has contributed greatly to better understanding of the pathogenesis. This protein is probably essential for incorporation of copper into ceruloplasmin and for its biliary excretion. Multiple mutations of Wilson's disease gene are responsible for the excess of so called "free" copper which is toxic to tissues. Copper toxicity involves first of all, functional disorders of many enzymatic systems, particularly those of respiratory chain enzymes. In the central nervous system, a special kind of copper toxicity is medicated by astroglia, so that a direct, harmful effect of both copper and ammonia on the brain is observed. The authors present a current review on biochemical mechanisms of copper toxicity and physiopathological significance of the CNS astroglia in Wilson's disease.

[Current views on the mechanisms of dopaminergic neuron death of the nigrostriatal system in Parkinson's disease]. / Obecne poglady na mechanizmy smierci neuronów dopaminergicznych ukladu nigrostriatalnego w chorobie Parkinsona.

Current views on the pathogenesis of Parkinson's disease are presented. Studies, particularly those carried out during the last decade, highlight the significance of endogenic processes responsible for a cumulative production of neurotoxic substances, especially free oxygen radicals which exert chronic effect on neurons. In Parkinson's disease, overproduction of free radicals and concomitant failure of protective mechanisms are most likely. An excess of free radicals is cytotoxic because of their very high chemical activity and uncontrolled chain reactions with numerous organic compounds, especially those which are mostly responsible for vital functions of cells. Oxidative stress disturbs metabolism of the cell what finally leads to its death most probably due to damage of cell membrane. That results in increased plasma membrane permeability for calcium ions which activate several subcellular mechanisms and initiate the final phase of cell death. Nonprotein-bound "free" iron ions are the strongest and most dangerous generators of free oxygen radicals. It is thought that ferric (Fe-3+" iron bound to neuromelanin may play a profound role in the overproduction of especially cytotoxic hydroxyl radicals, derivatives of molecular oxygen. Both, oxygen stress inducing factor and the sequence of related biochemical disorders remain still unknown. However, the synergy of the excess of reactive oxygen metabolites (mainly free radicals), nitric oxide, "free" iron ions and neuromelanin may contribute considerably to the generation of oxygen stress.

Qualitative and quantitative analysis of locus coeruleus neurons in Parkinson's disease.

The analysis of qualitative changes in the locus coeruleus (LC) was performed on brains from 21 cases of Parkinson's disease. Eleven cases were selected for quantitative analysis of the loss of LC noradrenergic pigmented neurons. The qualitative studies revealed uneven dissemination of the noradrenergic cells loss of overall structure of the LC. Few preserved neurons showed degenerative changes. Extracellular neuromelanine granules, traces of dying neurons, were also observed. A weak astro- and microglia proliferation corresponded with neuronal loss. Lewy bodies were found in the LC in all cases. The quantitative analysis revealed that the average loss of adrenergic neurons in the LC accounts for about 70% in relation to the control group. The degenerative changes were observed in the whole LC, but they were most intensive in its caudal and next in the middle segment. The results suggest also that the degenerative process began in the middle segment and then it spread towards caudal segment of the LC as the stages of disease advanced.

Development of human fetal substantia nigra grafts in the brain of non-immunosuppressed rats.

The survival of xenogenous tissue after transplantation to the brain of Wistar rats without immunosuppression was studied. Cell suspension was prepared from the ventral mesencephalon of human embryos 7-8 weeks of gestation, and injected either into the striatum or motor cortex of adult rats. After 1, 3, 7, 14 days and 1, 3, 6, 8 months the rats were sacrificed and the brains were processed for tyrosine hydroxylase (TH), glial fibrillary acidic protein (GFAP), ferritin immunocytochemistry and electron microscopy study. The intensity of inflammatory reaction of the host brain strongly affected the viability of grafted cells, the extend of GFAP and ferritin reactions in the tissue around the graft. Grafted human TH-positive neurons were found for 3 month survival only. After transplantation, we observed the grafted cell differentiation similar to that in normally developing mesencephalon. Six and eight months after transplantation glial cells prevailed in the grafts, while most neurons looked abnormal. An extensive bundles of myelinated fibers transpassing the intracortical transplants were seen. We are concluding that human mesencephalic cells can survive and develop in the brain of non-immunosuppressed rats.

Quantitative study of pathological forms of astroglia in Wilson's disease.

The number and distribution of Alzheimer type I and II cells (Alz I and II) as well as Opalski cells (Opl) were estimated in chosen brain regions of seven autopsied cases with Wilson's disease (WD). The authors of this study focused especially on the question whether the kind and intensity of astrocytes is linked to the clinical form of the disease and to the intensity of brain damage. Alz I and II cells were counted by the use of the HE method, whereas the number of Opl cells was calculated using the PAS method. The study revealed that among the three types of cells the number of Alz II cells was the highest and that of Alz I cells was the lowest. The distributional patterns of these three types of cells were different. Alz I cells were found mainly in the putamen. Alz II cells were observed diffusely, although they occurred in different numbers in the whole brain. The highest number of Opl cells was found in the putamen. Alz I cells were found only in the neurological type of the disease. The highest number of Alz II cells was seen in the hepatic type of the disease, whereas the highest number of Opl cells was observed in the neurological "mixed" forms. Moreover, intensity of tissue damage with presence of necroses was greatest in neurological WD. In the hepatic type dispersed areas of status spongiosus were observed, without presence of necroses. Our study revealed that the type and amount of the pathological astroglia may correlate both with the clinical form of WD and intensity of tissue damage. Alz II cells seem to be a characteristic feature of the early stage of astroglial response to the pathogenic factor whereas Alz I and Opl cells occur in WD only in the advanced stage of tissue damage.

Fractal estimation of the senile brain atrophy.

The paper presents an attempt to fractal analysis of senile brain changes. The differences of atrophy velocity in the white and gray matter can be noticed via fractal dimension according to the described one layer approximate model. The young (34 years) and old (82 years) brains are examined and compared.

[The structural organization of suspension grafts of human embryonic midbrain in the rat brain]. / Strukturnaia organizatsiia suspenzial'nykh transplantatov srednego mozga émbrionov cheloveka v mozge krys.

Dynamics of survival, differentiation and migration of human fetal cells after their transplantation into the rat brain without immunosuppression was studied using routine histology, electron microscopy and immunocytochemistry. A cell suspension prepared from the ventral mesencephalon of human embryos 7-8 weeks of gestation was injected into the striatum of rats-recipients. The graft development depends on the intensity of immune reaction. Under a weak reaction, the viability and differentiation of human embryo cells in the rat brain were observed within the three months of the experiment. The grafted cells conserve their mediator specificity, some of them being seen to migrate into the host (rat) tissue beyond the graft. In the transplant neuropil various types of cell contacts were observed, including synapses. The described method makes its possible to study the human nervous tissue histogenesis in an abnormal environment.

[Morphometric assessment of lesions in the dopaminergic nigrostriatal system in Parkinson disease]. / Morfometryczna ocena zmian w nigrostriatlnym ukladzie dopaminergicznym w chorobie Parkinsona.

The qualitative changes in substantia nigra were analysed in the material of 25 cases of Parkinson's disease. A morphometric quantitative study of depletion of pigmented dopaminergic cells of substantia nigra was performed in six long-term cases of the disease. In addition, the number of melanin nodules was assessed as a marker of cell disintegration. The results obtained were compared with the morphometric evaluation of neuronal depletion in the mesocorticolimbic system (ventral tegmental area-VTA). The qualitative study indicated that melanin depletion in dopaminergic cells of substantia nigra in Parkinson's disease is diffuse and it is located mainly in the lateral alfa layer. Neuronal depletion with concomitant numerous extracellular neuromelanin nodules and granules was observed. A slight astrocytic gliosis free of macrophages accompanied cellular changes. The qualitative changes in substantia nigra are similar to those observed in VTA. The morphometric evaluation revealed that depletion of dopaminergic neurons in substantia nigra in Parkinson's disease is 73%, on average, while in VTA it remains under 5%. Hence, depletion in substantia nigra is much more intense. The analysis of the relationship between the number of neurons in substantia nigra and the age of subjects in the control group indicated that the number of neurons decreased proportionally to the age. In the group under study no significant relationship between neurons depletion and duration of disease or patients age was found. In the studied group of patients with Parkinson's disease, the number of melanin nodules in substantia nigra was significantly higher than in controls.

Major histocompatibility complex class I expression on neurons in subacute sclerosing panencephalitis and experimental subacute measles encephalitis.

Lack of major histocompatibility class I antigens on neurons has been implicated as a possible mechanism for viral persistence in the brain since these antigens are required for cytotoxic T-lymphocyte recognition of infected cells. In subacute sclerosing panencephalitis (SSPE), measles virus (MV) persists in neurons, resulting in a fatal chronic infection. MHC class I mRNA expression was examined in formalin-fixed brain tissue from 6 SSPE patients by in situ hybridization. In addition MHC class I protein expression in MV-infected neurons was examined in experimental Subacute Measles Encephalitis (SME) by double immunohistochemistry. MHC class I mRNA expression was found to be upregulated in SSPE tissues studied, and in 5 out of 6 cases the expression was definitively seen on neurons. The percentage of neurons expressing MHC class I mRNA ranged between 20 to 84% in infected areas. There was no correlation between the degree of infection and expression of MHC class I molecules on neurons. Importantly, the number of neurons co-expressing MHC class I and MV antigens was markedly low, varying between 2 to 8%. Similar results were obtained in SME where 20 to 30% of the neurons expressed MHC class I but <8% co-expressed MHC class I and MV antigens. Perivascular infiltrating cells in the infected regions in SME expressed IFNgamma immunoreactivity. The results suggest that MV may not be directly involved in the induction of MHC class I on neurons and that cytokines such as IFNgamma may play an important role. Furthermore, the paucity of neurons co-expressing MHC class I and MV antigens in SSPE and SME suggests that such cells are either rapidly cleared by cytotoxic T lymphocytes (CTL), or, alternatively, lack of co-expression of MHC class I on MV infected neurons favors MV persistence in these cells by escaping CTL recognition.

Changes in dopaminergic neurons of the mesocorticolimbic system in Parkinson's disease.

The qualitative analysis of changes in major nuclei (n. paranigralis left and right, n. interfascicularis) of the mesocorticolimbic system (ventral tegmental area-VTA) was carried out on 25 cases with Parkinson's disease (PD). The cellular depletion with insignificant gliosis without the presence of macrophages was found. In addition, numerous extracellular melanin nodules being the remnants of broken dopaminergic neurons were found. The presence of Lewy bodies observed in all cases confirms the diagnosis of idiopathic Parkinson's disease. The morphometric analysis performed on selected long-lasting 7 PD cases and 6 controls showed that cellular depletion in n. paranigralis and in n. interfascicularis accounts for 42% and 62%, respectively in relation to controls. The number of melanin nodules grows with the age in the control group. While in the group of PD cases the number of nodules in VTA declines with the disease duration. It may indicate that the factor which damages melanin neurons also exerts a destructive effect on extracellular melanin.

Evaluation of survival and maturation of cryopreserved dopaminergic fetal cells transplanted into rat striatum and an analysis of the host brain reaction to graft.

A fetal, cryopreserved ventral mesencephalic rat tissue was transplanted into striatum of healthy adult rats. A stereotactic apparatus was used for transplantation of solid tissue blocks. The survival of transplanted dopaminergic cells in rat striatum was evaluated by means of histological and immunocytochemical methods (TH - thyrosine hydrolase) 1, 3, 7, 14, and 21 days after transplantation. The cellular reaction of the host to graft and to sham-lesion was examined. Glial fibrillary acidic protein (GFAP) was used for the visualization of astroglial reaction and ferritin for microglia. It was found that fetal cells of cryopreserved rat ventral mesencephalon transplanted into adult rat striatum survive though, in a small number. Cellular reactions of the host to both graft of dopaminergic cells and sham-lesion are similar to glial scar and are nonspecific.

Adult schizophrenic-like variant of adrenoleukodystrophy.

A 35-year-old man died after 30 months following the onset of the disease. There was a history of changes in his mental condition, including disturbances of behavior as well as the evidence of progressing dementia. The patient revealed gait disturbances and finally became bed ridden. Bizarre behavior and changes of mood with concurrent growing irritability which predominated during the course of disease, may explain the initial diagnosis of schizophrenia. Then cerebellar and spastic movement disorders leading to paraparesis and sphincters disturbances developed. Clinical symptoms of adrenal failure were not found apart from episodes of arterial pressure fall. After two years a magnetic resonance imaging (MRI) revealed an extensive diffuse demyelinative process in white matter of cerebral and cerebellar hemispheres. Activity of lysosomal enzymes was normal. A general autopsy revealed atrophy of adrenal cortex and the presence of ballooned cells with striated cytoplasm in the reticular and fasciculate zones. Neuropathological examination revealed an extensive demyelination of white matter in cerebral and cerebellar hemispheres and of the long paths of the brain stem, corresponding to changes in MRI examination. Within demyelination areas damage of axons and diffuse cellular and fibrous gliosis were found as well as perivascular lymphocytic infiltrations with the presence of strong PAS (+) and Sudan (+) macrophages. Immunocytochemical reactions with HAM-56 and RCA1 in macrophages were positive. Electron microscopy examination revealed lamellar inclusions in cytoplasm of macrophages. Similar structures were present in the lysosomes of astrocytes. Morphological examination of adrenal glands as well as morphological and ultrastructural study of the brain allowed us to diagnose the cerebral form of adrenoleukodystrophy (ALD). Topography and character of the brain changes seems to be in keeping with a rare schizophrenic-like variant of ALD with progressive dementia. Abnormal plasma profile and increased VLCFA concentration in the patient's 13-year-old daughter confirm the ALD diagnosis.

Primitive neuroectodermal tumor (PNET). A case report.

The authors present a case of relatively rare tumor of the central nervous system (CNS) in a 19-year-old female, who died 18 months after the first manifestation of meningismus, increased intracranial pressure and secondary hydrocephalus. Brain autopsy revealed abundant neoplastic infiltrations, which spread through the subarachnoid space. Neoplastic infiltrations were also present in the third ventricle and in a form of small subependymal nodules along the whole ventricular system. The microscopical examination showed that neoplasm consisted of small cells, which formed neuroblastic Homer Wright rosettes. Immunohistochemical studies (for synaptophysin, chromogranin A, GFAP, vimentin) together with morphology and localization of neoplasm suggested diagnosis of primitive neuroectodermal tumor (PNET) that spread mainly in the leptomeninges and caused obliteration of subarachnoid space.

[Neurotransplantation. III. Intracerebral grafting of fetal substantia nigra in Parkinson's disease]. / Neurotransplantacje. III. Domózgowe przeszczepy plodowej istoty czarnej w chorobie Parkinsona.

The paper reports the results of fetal substantia nigra transplantations into the brain of patients suffering from Parkinson's disease, performed in different neurosurgical centers in the world. Commonly accepted Ethical Guidelines for Fetal Tissue Transplantations are presented. The criteria regarding patients selection for therapeutic transplantation are also discussed. The results of studies concerning the problem of optimal fetal age for transplantations are presented. The optimal age seems to cover the period between the 7th gestational week when dopamine synthesis begins and the 11th week when protoplasmic processes start to grow intensively. The damage of these processes during transplantation procedure is thought to make the fetal cells survival difficult. The discussion concerning controversial problem of immunosuppression treatment after the neurotransplantation is also reported. The paper presents the principles of signs and symptoms assessment before and after operation including CAPIT system which emphasises the role of movement ability tests, PET-scanning with [18F]-6-L-fluorodopa and magnetic resonance imaging. The PET-scanning allows to follow up the changes in patient's brain resulting from the graft survival and its dopamine synthesis ability. The methods of donor woman examination to avoid the transmission of infections into the patient's brain are reviewed. The analysis of yet published results of about 100 operated on parkinsonian patients show marked improvement due to human fetal substantia nigra transplantation. The improvement lasts, at least 46 months postoperatively (the longest period of observation). Many of the grafted patients returned to normal self-dependent living activities and some of them resumed even their professional jobs. Most authors present the opinion that the therapeutic effects of fetal substantia nigra transplantations are more valuable and longer lasting than those after adrenal medulla autografts. However it should be borne in mind that both methods are yet only the experimental approach in Parkinson's disease therapy.

[Mossbauer spectroscopy of iron in substantia nigra in Parkinson disease and controls]. / Spektroskopia Mössbauerowska zaelaza istoty czarnej w chorobie Parkinsona i kontroli.

Mössbauer spectroscopy was used to study iron content, its redox state and binding sites in substantia nigra from parkinsonian and control brains. Measurements performed on fresh frozen samples demonstrated the presence of ferric iron only, both in disease and control. We found no difference in the total amount of iron in substantia nigra between the disease and control. Mössbauer spectra observed at 4.1 K in fresh frozen samples were different from those obtained in formalin fixed samples. In the fresh frozen samples only ferritin like iron was observed, whereas in the formalin fixed samples also non-ferritin iron was detected. It seems that in formalin fixed brains, during years, iron is released from ferritin and bound to an iron chelator or formalin.