Paraneoplastic pemphigus associated with Castleman tumor, myasthenia gravis and bronchiolitis obliterans.

BACKGROUND: Cases of paraneoplastic pemphigus (PNP) have been reported associated with various lymphoproliferative malignancies and benign Castleman tumors, with the most severe course and fatal outcome seen in patients with bronchiolitis obliterans. OBJECTIVE: The aim was to establish immunologic associations by coexistence of Castleman tumor, myasthenia gravis, and bronchiolitis obliterans and to evaluate the treatment modalities. METHODS: Clinical studies included computed tomography of the mediastinum, computed tomography and magnetic resonance imaging of the abdominal cavity, and quantitative electromyography. Direct and indirect immunofluorescence on various substrates, immunoblot analysis, immunoprecipitation, and specific enzyme-linked immunosorbent assay using recombinant desmogleins (Dsg) were performed as immunologic assays. RESULTS: Direct and indirect immunofluorescence including rat bladder showed intercellular antibodies. Immunoblotting disclosed antibodies to envoplakin (210 kd protein) and periplakin (190 kd protein); in addition, immunoprecipitation detected antibodies to desmoplakin I (250 kd protein). Antibodies to Dsg3 (pemphigus vulgaris antigen) were detected by specific enzyme-linked immunosorbent assay. Myasthenia gravis was controlled by drugs; however, mucocutaneous changes were not fully responsive to corticosteroids and cyclophosphamide pulses, cyclosporine, and intravenous immunoglobulins. The surgical removal of Castleman tumor did not change the course of the disease. The fatal outcome was the result of bronchiolitis obliterans that occurred after the surgery and was only transitionally controlled by plasmapheresis. CONCLUSION: This is the first case of paraneoplastic pemphigus associated with Castleman tumor, myasthenia gravis, and bronchiolitis obliterans. Despite a benign character of the tumor the patient died, as do all patients with bronchiolitis obliterans. Massive plasmapheresis has only a transient effect. We confirmed the presence of antibodies to Dsg 3, in addition to the set of specific paraneoplastic pemphigus antibodies against various proteins of plakin family.

The developmental changes of the "paraclaustral reservoir" of migrating cells in the rat brain: a study using morphometric and in situ DNA end labeling techniques.

A distinct group of small cells lying in the ventral part of the external capsule in the rat brain is clearly visible at birth. On the basis of its location (medially to the prepiriform claustrum) and probably its function (as a source of neurons for adjacent structures), we define this nucleus as the "paraclaustral reservoir". The present study reveals the cellular changes of the paraclaustral reservoir during postnatal development of the rat brain using unbiased morphometry and in situ DNA end labeling. During the first 4 days after birth the density and total number of cells in the paraclaustral reservoir were stable; after this period a decrease of these parameters was observed until the complete disappearance of this structure at the end of first postnatal week. The rather low number of TUNEL (TdT mediated dUTP nick end labeling of fragmented DNA) positive nuclei in the paraclaustral reservoir suggests that apoptosis is not a crucial mechanism leading to decay of this structure.

Postnatal development of the rat striatum--a study using in situ DNA end labeling technique.

We have examined the development of rat striatum for evidence of cells dying in the process of physiological cell death. In present study we have indicated apoptotic cells in sections stained with cresyl violet (cell death characterized by pyknosis) or with DNA end labeling assay (TUNEL method). Our results demonstrated that cell loss during maturation of the rat striatum had the characteristics of apoptosis rather than necrosis. The greatest number of TUNEL-positive and pyknotic cells in the striatum were found during the first postnatal days; after 7th day of postnatal life a rapid decrease of its number was observed after the second postnatal week no TUNEL-positive cells were observed in the striatum. Our analysis suggests that apoptotic cell death occurring during the development of striatal neuronal population takes place during the first week of postnatal life.

Neuronal changes in the basolateral complex during development of the amygdala of the rat.

Neuronal changes in the amygdala basolateral complex were studied during development and maturation in fetal and postnatal rat brains using morphometrical methods. Forty brains of animals of various ages were fixed in formalin, frozen and cut into 25 microm thick sections and stained with cresyl violet or haematoxylin and eosin (H&E). In cresyl violet preparations, the complex appeared for the first time on embryonic day (E)17 and was composed of two homogeneous nuclei lateral and basolateral. On about the seventh postnatal day, each of these nuclei was divided into two parts the first one into the dorsolateral and ventromedial and the second one into the anterior and posterior. Morphometric investigations showed a different increase of the neuronal and nuclear size in various parts of the basolateral complex up to postnatal day (P)14; after that time these parameters did not change significantly. The neuronal density and the total number of neurons stabilized at P7 in all parts of this complex, except for the dorsolateral part of the lateral nucleus in which a 30% decrease of the total number of cells was observed. From P14, in all nuclei under study, the total number of neurons did not change significantly.

Volume and topographical changes of the basolateral complex during the development of the rat's amygdaloid body.

Volume and topography of basolateral complex during development and maturation in fetal and postnatal rat brains were studied using morphometrical methods. 39 rat brains of various ages were fixed in formalin, frozen and cut into 25-micron-thick sections and stained with cresyl violet. In cresyl violet preparation the basolateral complex appeared first on 17th prenatal day and it was composed of two homogenous parts--lateral and basolateral nuclei. On about 7th postnatal day each of these nuclei divided into two parts--the first one into the dorsolateral and ventromedial part, while the second one--into the anterior and posterior parts. Morphometric investigations showed that volume of the basolateral complex and its parts underwent the biggest changes up to 14th postnatal day, and we suspected that during this time it was more sensitive to pathological changes. After that day the volume of the basolateral complex had changed only a little.

Acetylcholinesterase activity as a marker of maturation of the basolateral complex of the amygdaloid body in the rat.

Development and maturation of the basolateral complex of the amygdaloid body were studied in fetal and postnatal rat brains. In cresyl violet-stained sections the basolateral complex was distinguishable at the 17th prenatal day. On about the 14th postnatal day, it showed all the features of the adult structure. The acetylcholinesterase activity appeared in neuropil of the basolateral complex on the seventh postnatal day; it increased slowly up to the 60th day. After that age, the acetylcholinesterase activity was stable in all parts of the basolateral complex and corresponded to its cytoarchitectonic differentiation. Our results suggest that the process of maturation of the amygdaloid basolateral complex, being related to the ingrowing of the cholinergic fibers from the basal forebrain, lasts at least up to the end of the second month of postnatal life.

Loss of neurons in the claustrum of aging brain.

The study was performed on 19 brains of nondemented patients with age ranging from 36 to 89 years. After embedding in paraffin, coronal 8-microns-thick serial sections were cut and stained either with cresyl violet or with immunocytochemical methods for amyloid and tangles. Morphometrical studies were performed in all parts of the claustrum along its whole extension. Changes related to aging (neuronal loss and decrease in volume) were found in all parts of the claustrum, but the time of origin of these changes is different in various parts of this structure. We did not observe neurofibrillary pathology in any parts of the claustrum. In the oldest subjects a small number of amyloid plaques was found in the paraamygdalar part of the claustrum. We suggest that the neurons of the claustroneocortical loop are affected severely and earlier than those in the claustroentorhinal loop.

Division of the human claustrum according to its architectonics and morphometric parameters.

The topography and cytoarchitectonics of the claustrum as well as morphometric parameters of its neurons were studied in 10 human brains obtained from patients without any detectable neuropathological changes. We distinguished four parts of the claustrum: dorsal, orbital, temporal and paraamygdalar. The dorsal and orbital parts contain larger cells, than those of the temporal and paraamygdalar parts, although these differences were statistically non significant. The highest neuronal density was observed in the paraamygdalar part. The nucleus and nucleus@cell body area ratio was significantly smaller in the dorsal part than in other parts of the claustrum. We described three types of neurons in the claustrum: (1) medium-sized either fusiform or triangular cells with darkly stained cytoplasm; they predominate in the dorsal and temporal parts, (2) medium-sized as well as large cells, either multipolar or pyramidal-like with lightly stained cytoplasm; they are most numerous in the orbital and paraamygdalar parts, (3) small, multipolar or oval neurons with darkly stained ring of cytoplasm; these types of neurons are uniformly distributed throughout all parts of the claustrum. The subdivision of the human claustrum is in accordance with our observations that each of these parts possesses connections with different cortical regions.

Influence of fixation and histological procedure on the morphometric parameters of neuronal cells.

The influence of fixation and histological procedure on morphometric parameters of human and rat brains were analyzed. We found negative correlation of brain weight with the fixation time both in human and rat material. Human brains fixed for shorter period of time than one year showed large differences in weight. The group of brains fixed for longer than one year was characterized by much smaller dispersion. Moreover, we found that during histological preparation the shrinkage of brain tissue of older human subjects is statistically significantly smaller than younger ones (below 65 years). In order to correct these artifacts easily we propose three simple formulas.

Amyloid deposits and loss of neurones in the claustrum of the aged dog.

In dogs under 17 years of age no amyloid deposits were observed in the claustrum. After that age amyloid deposits were found in all the brains studied. The number of amyloid deposits increases rapidly between 17 and 19 years of age. In none of the animals were neurofibrillary tangles found. However, in almost all claustra with amyloid deposits the number of neurones was smaller than in those without amyloid pathology. The loss of neurones resulted in a decrease in the volume of claustrum, which is about 22% smaller in dogs over 17 years of age than in younger animals. No considerable age-related differences were found in the average size of neurones and their nuclei.

Insular claustrum of the dog--a morphometric investigation of cellular structure.

Investigation of the dog's insular claustrum was performed using morphometric methods. The brains of 14 dogs (7-14 year-old) were studied. The size, shape and density of neurons are different in various parts of the insular claustrum. The posterior part, related to the visual system, is heterogenous and possesses medium-sized and small densely packed neurons, the average cross-sectional area of cell body being 175.8 +/- 45.5 micron2. The similar average cross-sectional area of cell body (174.1 +/- 55.3 micron2) was found in the central part of the insular claustrum. However, neuronal architecture is here less heterogenous and neurons are less densely arranged. The anterior part, connected mainly with the frontal and motor cortex, is more homogenous and contains loosely arranged neurons of a rather large size (average size--198.7 +/- 53.46 micron2).

Variability of the parahippocampal surface of the transverse fissure in the human brain.

The parahippocampal (ventral) surface of the lateral part of the transverse fissure (LTF), formed by the parahippocampal gyrus, was examined in 53 human brain hemispheres without pathological changes. Cytoarchitectonics of this region was studied on the frontal histological sections stained either with cresyl violet or with Weil method. Four types of the shape of the parahippocampal surface of LTF were distinguished. In type 1 the surface is either flat or elevated in its medial part (medial eminence; 34.0%). In type 2 the surface is elevated in its lateral part (lateral eminence, 24.5%). In type 3 there are two eminences--the lateral and the medial; both of similar height (20.8%). In type 4 the lateral eminence is distinctly higher than the medial one (20.7%). The eminences contain cytoarchitectonically different structures. The lateral eminence is formed mainly by the subiculum and CA1 area, the medial by the presubicular region. The shape of the parahippocampal surface of LTF may be of importance in assessment of the Alzheimer's disease pathology.

The second layer neurones of the entorhinal cortex and the perforant path in physiological ageing and Alzheimer's disease.

The hippocampal formation was studied in 5 brains of younger (29 to 52 years of age) and 6 brains of elderly (61 to 89 years of age) subjects without signs of dementia, as well as in 11 brains of patients with Alzheimer's disease (65 to 91 years of age). The 8-microns-thick sections were stained either with cresyl violet, Weil method or with immunocytochemical methods for amyloid (4G8) and neurofibrillary tangles (Tau-1). Cell bodies, senile plaques and tangles were counted in all brains. In brains of patients with Alzheimer's disease a significant neuronal loss (about 56%) was observed in the second layer of the entorhinal cortex. The tangles/neurones ratio was very high (62.79.1%) in this layer. A great number of senile plaques were present in the whole hippocampal formation, especially in the molecular layer of the dentate gyrus (22.91.5 plaques/mm2) which is the termination zone of the perforant path. It seems therefore, that pathological alterations in Alzheimer's disease disrupt the main input to the hippocampal formation. In "physiological" ageing we did not observe changes in the density of neurones, although single tangles and plaques were found in all hippocampal areas. In elderly individuals 3.81.3% of neurones of the second layer revealed neurofibrillary pathology; a few plaques were found in various areas of the hippocampal formation. These observations may suggest only a slight decrease in number of neurones in the hippocampal formation. However, these changes cause a slight impairment of memory and learning often found in elderly individuals without dementia.

The insular claustrum in the methylazoxymethanol acetate (MAM) treated rats shows two different populations of neurons.

Methylazoxymethanol acetate (MAM) is a substance that inhibits migration of neurons in the embryonic brain. After intraperitoneal injection of two different doses of MAM to pregnant rats, microcephaly with or without complete development of the cerebral cortex was observed in every litter. High MAM doses (30 mg/kg) resulted in the lack of superficial layers (II-IV) of the cerebral cortex when the deep layers (V, VI) were seen. The claustrum was present but composed of loosely packed, medium-size, triangular or fusiform neurons with anarchic oriented long axes. After administration of low MAM doses (14 mg/kg) two different parts (medial and lateral) of the insular claustrum were observed. Our results suggest that neurons of the insular claustrum create two different subpopulations of cells, which were similar to that observed in primitive insectivore (e.g., hedgehog), but fuse in development.

The role of lipases in the removal of dormancy in apple seeds.

It was found that the temperature optimum for apple (Malus domestica Borb.) seed acid lipase is the same as that for seed after-ripening process. The activity of the enzyme occurs between the 40th and 70th days of stratification, whereas the activity of alkaline lipase very low at that time appears about 20 days later. The changes of both enzyme activities were also studied during dark and light culture of embryos isolated from seeds after different times of stratification. Only the alkaline enzyme activity is under the control of light. It was concluded that essentially the same process, i.e. the hydrolysis of reserve fats is catalysed by two different enzymes: acid lipase acting during the cold-mediated breaking of embryo dormancy and alkaline lipase acting during the germination of dormant embryos, thus being under light control.