Inflammatory response in human tick-borne encephalitis: analysis of postmortem brain tissue

In Central European tick-borne encephalitis (TBE) mechanisms of tissue destruction are poorly understood. To evaluate the contribution of immunological mechanisms to tissue injury, the authors immunohistochemically analyzed paraffin-embedded autoptic brain tissue of 26 human TBE cases. In the parenchymal compartment, there was a predominance of macrophages/microglia and cytotoxic T cells. In addition, it was found that granzyme B-expressing lymphocytes were in close contact with TBE-expressing neurons up-regulating caspase-3. These findings indicate that cellular and humoral pathways of the immune system, especially granzyme B-releasing cytotoxic T cells and macrophages/microglia, mainly contribute to tissue destruction in TBE.     

Tick-borne encephalitis in dogs: neuropathological findings and distribution of antigen

Eight dogs originating from different regions of Austria [all of them known as tick-borne encephalitis (TBE) areas] with severe neurological signs were either euthanatized or died spontaneously. Tick-borne encephalitis virus (TBEV) antigen was detected in the brains of five of these dogs by immunohistology, but not in the others. All of the dogs, however, had identical neuropathological changes. There were moderate lymphohistiocytic meningitis, widespread neuronal necroses, karyorrhexis of glial cells, numerous neuronophagic nodules, and extensive microgliosis. In the cerebellum, loss of Purkinje cells and proliferation of microglial cells in the molecular layer were found. All brain regions showed numerous perivascular cuffs consisting of lymphocytes, macrophages, plasma cells and, occasionally, red blood cells. The blood-derived cells were not restricted to the perivascular spaces but diffusely infiltrated the neuropil. The most severe changes were localized in the neuroparenchyma surrounding the fourth ventricle. Lesions were less severe in basal ganglia, thalamus, mesencephalon, nuclei of pons and medulla oblongata. Moderate lesions were found in the gray matter of neocortex and allocortex, hippocampus and molecular and Purkinje cell layers of the cerebellum. White matter was slightly to moderately affected. The choroid plexus was free of inflammation. Due to rapid virus clearance mechanisms in this disease, antigen was not detectable in all cases. Neuropathological changes identical with those of immunohistologically proven cases justified the diagnosis TBE in these cases. In addition, the neuropathological diagnosis was supported by the origin of the affected dogs from endemic areas, the seasonal occurrence of the disease and a clinical history of a highly febrile neurological disease with short duration. Received: 25 July 1997 / Revised: 16 October 1997 / Accepted: 20 October 1997     

Immunohistochemical demonstration of viral antigens in Japanese encephalitis

Summary Japanese encephalitis virus antigens were immunohistochemically demonstrated in formalin-fixed paraffin sections from an autopsied brain. Glial nodules were always associated with antigen-positive cell debris. Glia shrubs in the cerebellar cortex appeared to be formed along the apical dendrite of Purkinje cells. Most, but not all, of the neurons involved in neuronophagia were viral antigen positive. Antigenic masses were occasionally encountered in the center of so-called acellular plaques. Neurons with strong viral antigens were sporadically found in normal-appearing regions in the thalamus and cerebral cortex. Viral antigens were demonstrable only in neurons and not in glial or vascular endothelial cells.Supported in part by grants from the Ministry of Health and Welfare, Japan and the Gonryo Foundation     

Corticobasal degeneration with olivopontocerebellar atrophy and TDP-43 pathology: an unusual clinicopathologic variant of CBD

Corticobasal degeneration (CBD) is a disorder affecting cognition and movement due to a progressive neurodegeneration associated with distinctive neuropathologic features, including abnormal phosphorylated tau protein in neurons and glia in cortex, basal ganglia, diencephalon, and brainstem, as well as ballooned neurons and astrocytic plaques. We identified three cases of CBD with olivopontocerebellar atrophy (CBD-OPCA) that did not have α-synuclein-positive glial cytoplasmic inclusions of multiple system atrophy (MSA). Two patients had clinical features suggestive of progressive supranuclear palsy (PSP), and the third case had cerebellar ataxia thought to be due to idiopathic OPCA. Neuropathologic features of CBD-OPCA are compared to typical CBD, as well as MSA and PSP. CBD-OPCA and MSA had marked neuronal loss in pontine nuclei, inferior olivary nucleus, and Purkinje cell layer. Neuronal loss and grumose degeneration in the cerebellar dentate nucleus were comparable in CBD-OPCA and PSP. Image analysis of tau pathology showed greater infratentorial tau burden, especially in pontine base, in CBD-OPCA compared with typical CBD. In addition, CBD-OPCA had TDP-43 immunoreactive neuronal and glial cytoplasmic inclusions and threads throughout the basal ganglia and in olivopontocerebellar system. CBD-OPCA met neuropathologic research diagnostic criteria for CBD and shared tau biochemical characteristics with typical CBD. These results suggest that CBD-OPCA is a distinct clinicopathologic variant of CBD with olivopontocerebellar TDP-43 pathology.     

Ganglioside GD3 of cerebral neurons and Purkinje cells in aged human brains

Immunocytochemical staining for ganglioside GD3 (II^3α (NeuAcα2–8NeuAc)-LacCer, GD3) in neuronal cells of the cerebral cortices (cerebral neurons), and cerebellar dentate nucleus (dentate neurons) and Purkinje cells, in human autopsy cases of progressive supranuclear palsy, senile dementia of the Alzheimer type, Pick's disease, amyotrophic lateral sclerosis and olivopontocerebellar atrophy (OPCA) was undertaken using mouse IgM anti-GD3 monoclonal antibody. Cerebral neurons and dentate neurons were constantly GD3-immunoreactive and immunoreactivity was observed in the cytoplasm. The peroxidase reaction product for GD3 (RP) in cerebral and dentate neurons was granular in appearance. It appeared that RP was associated with lipofuscin granules. However, immunoreactivity of Purkinje cells varied among cases, and the RP was slightly granular even when they were positive. This study suggests that lipofuscin granules contributed to the neuronal immunoreactivity of GD3 in aged human brains.     

Developmental expression of monocyte chemoattractant protein-1 in the human cerebellum and brainstem

The developmental expression of monocyte chemoattractant protein-1 (MCP-1) in the cerebellum, medulla oblongata and pons was investigated in 26 normal human brains, ranging from 20 weeks of gestation (GW) to adulthood by means of an immunohistochemical method. Immunoreactivity to MCP-1 was observed in neurons of the cerebellum and brainstem, and was mainly distributed in the cytoplasm and dendrites. MCP-1-positive Purkinje cells appeared at 27 GW, reached a peak at 36 GW, and then decreased after 1 month of age, almost completely disappearing by 1-2 years of age. MCP-1-reactive neurons in the dentate nucleus and inferior olivary nucleus showed temporal increases similar to that of Purkinje cells. In the pons, however, MCP-1 reactivity was low in neurons of the pontine nuclei persisting from the fetal to the adult period, and was very low and short in the reticular formation and cranial nerve nuclei during the fetal and/or neonatal period. MCP-1 Western blotting of the cerebellar cortex confirmed the specificity of the immunohistochemistry. Our results suggest that MCP-1 may be related to the maturation of Purkinje cells, the dentate nucleus, the inferior olivary nucleus, and their network, promoting the growth of dendrites and synapses. Furthermore, MCP-1 may also be useful for the study of abnormal neuron development and ischemic damage.     

Herpes simplex virus (HSV) DNA in microglial nodular brainstem encephalitis

Thirty-four brains with microglial nodular brain stem encephalitis were retrospectively investigated for herpes simplex virus (HSV) and cytomegalovirus (CMV) by in situ hybridization (ISH) with biotinylated cDNA probes, and by immunocytochemistry with polyclonal and monoclonal antibodies on formalin fixed paraffin embedded serial tissue sections. In 16 cases (47%), HSV DNA was found by ISH in the nuclei of neurons in microglial nodules or in the adjacent parenchyma of the brainstem, and more rarely at various cerebellar and telencephalic sites. None of the 34 cases was labeled for CMV DNA and none revealed HSV or CMV antigens. Ten control brains without microglial nodules were not labeled. This study suggests an HSV etiology for many cases with microglial nodular brainstem encephalitis.     

Immunohistological study of grumose degeneration of the dentate nucleus in progressive supranuclear palsy

The grumose degeneration observed in the dentate nuclei of 7 cases of progressive supranuclear palsy (PSP) was studied with a panel of antibodies which included 2 neurofilaments, Tau and ubiquitin. Dentate nucleus neurons were negative with all antibodies except ubiquitin which showed a slightly positive homogeneous pattern of staining. The amorphous material surrounding swollen or normal neurons was strongly positive for neurofilament and subunits and numerous torpedoes were observed in the granular layer of the cerebellar cortex. Our results confirm that grumose degeneration consists in degeneration of terminal axons of Purkinje cells in the dentate nucleus. The positivity of dentate nucleus neurons for ubiquitin may support the concept of synaptic dysfunction between Purkinje cells and dentate nucleus neurons.     

Pathologic characterization of a murine model of human enterovirus 71 encephalomyelitis

We describe a model of Enterovirus 71 encephalomyelitis in 2-week-old mice that shares many features with the human central nervous system (CNS) disease. Mice were infected via oral and parenteral routes with a murine-adapted virus strain originally from a fatal human case. The mice succumbed to infection after 2 to 5 days. Vacuolated and normal-appearing CNS neurons showed viral RNA and antigens and virions by in situ hybridization, immunohistochemistry, and electron microscopy; inflammation was minimal. The most numerous infected neurons were in anterior horns, motor trigeminal nuclei, and brainstem reticular formation; fewer neurons in the red nucleus, lateral cerebellar nucleus, other cranial nerve nuclei, motor cortex, hypothalamus, and thalamus were infected. Other CNS regions, dorsal root, and autonomic ganglia were spared. Intramuscular-inoculated mice killed 24 to 36 hours postinfection had viral RNA and antigens in ipsilateral lumbar anterior horn cells and adjacent axons. Upper cord motor neurons, brainstem, and contralateral motor cortex neurons were infected from 48-72 hours. Viral RNA and antigens were abundant in skeletal muscle and adjacent tissues but not in other organs. The distinct, stereotypic viral distribution in this model suggests that the virus enters the CNS via peripheral motor nerves after skeletal muscle infection, and spread within the CNS involves motor and other neural pathways. This model may be useful for further studies on pathogenesis and for testing therapies.     

Behaviour, chemosignals and endocrine functions in male mice infected with tick-borne encephalitis virus

Odour attractiveness, social behaviour and endocrine status of male mice (outbred ICR strain) were examined 6-7 days after inoculation with subclinical dose of tick-borne encephalitis virus (TBE). According to RT-PCR control of efficiency of infection, males injected with TBE were divided on the two subgroups: TBE+ (males with viral RNA) and TBE- (males without viral RNA). Susceptible males (TBE+ subgroup) showed the higher level of plasma testosterone in comparison with both control and nonsusceptible (TBE- subgroup) males. TBE+ males had also more odour attraction for oestrus females and more aggressiveness in social conflict. Higher sexual attractiveness and aggressiveness of the infected host benefit the pathogen's distribution in the host population.     

Neuropathology of the dentate nucleus in developmental disorders

The dentate nucleus was examined histologically and immunohistochemically in 47 cases of nonprogressive developmental disorders. Neuronal loss and/or atrophy was observed in 13 cases, while mild neuronal lesions, characterized by dendritic swelling and/or the appearance of eosinophilic materials around the neurons, were exhibited in 20 cases. The former change was accompanied by diffuse central nervous system involvement, and the etiology was perinatal hypoxic ischemic encephalopathy, acute encephalopathy, and meningoencephalitis in most cases. On the other hand, most of the patients with kernicterus showed the latter change. Immunohistochemically, the mild neuronal lesions mimicking grumose degeneration, described in some neurodegenerative diseases, seemed to reflect the changes of Purkinje cell terminals. It is suggested that secondary structural alteration of the dentate neurons in the absence of severe atrophy can occur in nonprogressive developmental disorders. Received: 11 September 1996 / Revised: 30 December 1996 / Accepted: 7 January 1997     

Difference in pathogenesis between herpes simplex virus type 1 encephalitis and tick-borne encephalitis demonstrated by means of cerebrospinal fluid markers of glial and neuronal destruction

We determined the extent of neuronal and glial cell destruction in 13 patients with herpes simplex type 1 (HSV-1) encephalitis, 15 patients with tick-borne encephalitis (TBE), and 20 noninfectious controls by analyzing the cerebrospinal fluid (CSF) concentrations of neurofilament protein (a marker of neurons, mainly axons), neuron-specific enolase (a marker of neurons, mainly somas), glial fibrillary acidic protein, and S-100 protein (markers of astrocytes). In addition, in patients with HSV-1 encephalitis CSF samples were collected serially before 7, 8-14, and 18-49 days and 3-10 months after the onset of neurological symptoms. In the acute stage of HSV-1 encephalitis we found markedly higher CSF levels of the cell damage markers than in patients with TBE. The concentration of cell damage markers in HSV-1 encephalitis decreased within 45 days after acute infection, except for neurofilament protein. The CSF concentrations of neurofilament protein increased during the second week, remained extremely high throughout the next month, and decrease thereafter. The changes in these markers of neuronal and glial destruction demonstrate the neuronal and astroglial cell damage during the first month after HSV-1 encephalitis. In contrast, most patients with TBE had signs only of slight astrogliosis, except for two patients with paresis.     

Cerebellar ataxia in patients with mitochondrial DNA disease: a molecular clinicopathological study

Cerebellar ataxia is a prominent clinical symptom in patients with mitochondrial DNA (mtDNA) disease. This is often progressive with onset in young adulthood. We performed a detailed neuropathologic investigation of the olivary-cerebellum in 14 genetically and clinically well-defined patients with mtDNA disease. Quantitative neuropathologic investigation showed varying levels of loss of Purkinje cells and neurons of the dentate nucleus and inferior olivary nuclei. Typically, focal Purkinje cell loss was present in patients with the m.3243A>G mutation caused by the presence of microinfarcts, with relative preservation of neuronal cell populations in the olivary and dentate nuclei. In contrast, patients with the m.8344A>G mutation or recessive POLG mutations showed extensive and global neuronal cell loss in all 3 olivary-cerebellum areas examined. Molecular analysis of mutated mtDNA heteroplasmy levels revealed that neuronal cell loss occurred independently of the level of mutated mtDNA present within surviving neurons. High levels of neuronal respiratory chain deficiency, particularly of complex I, were detected in surviving cells; levels of deficiency were greater in regions with extensive cell loss. We found a relationship between respiratory deficiency and neuronal cell density, indicating that neuronal cell death correlates with respiratory deficiency. These findings highlight the vulnerability of the olivary-cerebellum to mtDNA defects.     

Neuropathology of acquired immunodeficiency syndrome (AIDS). Analysis of 22 Brazilian cases

The neuropathologic study of 22 Brazilian cases of acquired immuno-deficiency syndrome (AIDS) was performed. Thirteen cases (59%) showed neuropathologic lesions. These included infection by Toxoplasma (n = 4), Cryptococcus neoformans (n = 3), viral encephalitis (n = 4), primary lymphomas (n = 2), isolated cerebral infarct (n = 1), and reactive gliosis (n = 1). In 2 cases, primary lymphoma and viral encephalitis were associated. Axonal spheroids in the gracilis and cuneatus nuclei were present in a case of toxoplasmosis. Mammillary bodies lesions consistent with Wernicke's encephalopathy were found in a case of viral encephalitis. In addition, circulatory changes (focal cortical infarcts) were associated lesions in 3 cases. These findings were compared with the main series reported in American and European literature.     

Major histocompatibility complex (MHC) antigen expression in HIV encephalitis

In order to study possible immunopathogenic mechanisms in Human Immunodeficiency Virus (HIV) encephalitis, immunocytochemical localization of Class I and Class II major histocompatibility complex (MHC) antigens was studied in formalin-fixed tissue sections from the brains of 10 individuals who had died with this disorder. Using the avidin biotin peroxidase technique and monoclonal antibodies to these antigens, increased expression of Class I antigens was found in five out of 10 and of Class II antigens in six out of 10 cases of HIV encephalitis. This contrasted with results obtained with the HIV-specific anti-P24 antibody which reacted with only a small number of cells in four cases. Class I and II antigens were detected mainly in perivascular monocytes/macrophages and also in multinucleated giant cells. In two cases, slight labelling was also detected in these cells more diffusely in the brain parenchyma. Immune and viral antigens were not detected in glial cells or neurons. Neither normal control cases nor brain sections from patients who had died from other neurological diseases were labelled with any of the antibodies apart from two cases of varicella-zoster virus-associated encephalitis in which increased expression of Class II antigens occurred. These findings support the notion that indirect immune-mediated mechanisms may be important in the pathogenesis of HIV encephalitis.     

Problems of differential diagnosis in tick-borne encephalitis-induced polyradiculitis

In about 10% of cases, tick-borne encephalitis (TBE) presents with additional myeloradiculitic features mimicking acute poliomyelitis, which can rarely appear as the sole symptom. We report on a 59-year-old man infected with TBE in Thuringia,Germany, who developed polyradiculitis with rapidly progressive, predominantly proximal tetraparesis and respiratory failure. We discuss the differential diagnosis and the epidemiological relevance in conjunction with a second typical case of TBE acquired in the same region and time period.     

Disconnection of cerebellar Purkinje cells in Kearns-Sayre syndrome.

Kearns-Sayre syndrome (KSS) is a sporadic multisystem disorder due to rearrangements in mitochondrial DNA (mtDNA). To gain further insight into the pathogenesis of cerebellar dysfunction in KSS, antibodies against synaptophysin (SY) were used to identify presynaptic terminals and antibodies to calbindin D (CB) to identify Purkinje cells in the cerebellar cortex and in the dentate nucleus from two autopsied cases of KSS. By conventional neuropathology we found marked spongiform degeneration and by immunohistochemistry a disruption of presynaptic terminals and of the terminal arborizations of Purkinje cell axons on multipolar neurons of the dentate nucleus in the KSS patients. We suggest that a disconnection of Purkinje cells at the dentate nucleus may play a role in the pathogenesis of cerebellar ataxia in KSS.     

Olivopontocerebellar atrophy: immunocytochemical and Golgi observations

Brain tissue was obtained promptly after death from a patient with autosomal dominant olivopontocerebellar atrophy and studied by immunocytochemistry and a Golgi technique. Antiglutamic acid decarboxylase showed severe loss of Purkinje cells and their terminals in the dentate nucleus. Stains for neuron-specific enolase (NSE) and microtubule-associated proteins (MAP) confirmed the integrity of the dentate nucleus. Basket and stellate cells revealed secondary changes, but Golgi neurons were intact. Methods for NSE and MAP disclosed dendritic alterations and loss of neurons in the basis pontis and inferior olivary nuclei. Golgi impregnation of Purkinje cells showed loss of major dendrites, paucity of spiny branchlets, and axonal expansions.     

Expression of Toll-like receptor 3 in the human cerebellar cortex in rabies, herpes simplex encephalitis, and other neurological diseases

There is recent in vitro evidence that human neurons express the innate immune response receptor, Toll-like receptor-3 (TLR-3), and that expression is enhanced in viral infections. The authors examined the immunohistochemical expression of TLR-3 in the cerebellar cortex of postmortem human brains. Purkinje cells were found to express TLR-3 in all cases of rabies (4 of 4) and herpes simplex encephalitis (2 of 2) as well as in cases of amyotrophic lateral sclerosis (1 of 2), stroke (1 of 2), and Alzheimer's disease (3 of 3). In cases of viral infection, direct viral infection was not necessary for enhanced neuronal TLR-3 expression, suggesting that soluble factors likely play an important role in inducing TLR-3 expression. In addition to neurons, occasional Bergmann glia expressed TLR-3 in some cases. This study has provided evidence that human brain neurons can express TLR-3 in vivo and suggests that neurons may play an important role in initiating an inflammatory reaction in a variety of neurological diseases.