Myeloma neuropathy: passive transfer from man to mouse.

Mice were injected daily, for up to 10 weeks, with purified monoclonal immunoglobulin G from patients with myelomatous polyneuropathy or benign gammopathy. The animals developed a demyelinating polyneuropathy with slowed nerve conduction velocities. The putative antinerve factor may be an antibody since injection of Fab fragments from the monoclonal immunoglobulin G produced a similar demyelination. This provides evidence of a circulating factor in the serum of myeloma patients with polyneuropathy that reproduces typical features of the human disease on passive transfer. This disorder is thus distinguished from other neuropathies that occur as remote effects of malignant disease but have no identified pathogenic factors associated with them.

Intracisternal A and C particles in mouse neurons: a thin-section study of normal trigeminal ganglion and C1300 neuroblastoma.

Trigerminal ganglia of 4 adult albino mice of the NMRI outbred stock were examined by electron microscopy. In all animals, about 10% of the neurons contained intracisternal A particles. Isolated structures resembling intracisternal A particles could be detected in atleast 50% of the nerve cells and in a few Schwann cells. Budding at the cell surface and/or extracellular type-C particles were not observed. An intracerebrally transplanted mouse C1300 neuroblastoma was likewise studied. Most tumor cells exhibited large numbers of intracisternal A particles having the same ultrastructure as the particles in trigeminal neurons. In addition, budding and extracellular type-C particles were occasionally observed.

Persistence of nerve sprouting with features of synapse remodelling in soleus muscles of adult mice.

An increase in the number of nerve branches of the unmyelinated axon terminals with increasing age was observed in normal adult mouse motor endplates. In addition, ultrastructural investigation revealed signs of nerve retraction. A combined light and electron microscopic investigation was performed on zinc-iodine-osmium stained endplates in soleus muscles. The number of branch points in a synapse, endplate length and muscle fiber diameter were evaluated in "young adult" (3 months) and adult (6 and 11 months) mice. For all 3 parameters, 3-month-old animals had the lowest values. Eleven-month-old animals had more branch points and larger endplate lengths than 6-month-old animals while there was no significant difference in fiber diameters. Branch point numbers and endplate length were correlated in each muscle while fiber diameters did not correlate with any of the other parameters. The ultrastructure of 15 thin nerve branches--likely candidates for new branches--was investigated in serial section and in 14 of them synaptic contacts were found. Near such contacts, empty gutters, possibly abandoned former synaptic sites, were present in several cases. It is concluded that there is continual nerve sprouting in synapses of adult mice and that sprouts form synaptic contacts. The possible signs of nerve retraction observed indicate that, as in the frog, synaptic contacts in mouse muscles undergo some continual remodeling.

Presence and absence of virus particles in hybridomas secreting monoclonal antibodies against gliomas.

Hybridoma clones were produced by fusion of splenocytes from glioma-immunized hosts and the X63-Ag8.653 mouse myeloma line and Y3-Ag.1.2.3. rat myeloma line. Oncornavirus particles were found in all clones descending from the mouse myeloma line. No virus particles could be found in either the spleens of immunized Balb/c mice and Fischer rats or in the rat myeloma line and the hybridomas derived from it.

Myopathy with tubular aggregates in a patient adrenalectomized for Cushing's syndrome.

We report a patient with attacks of muscle weakness and mild myopathy with tubular aggregates, following bilateral adrenalectomy for adrenal Cushing's syndrome and replacement therapy with cortisone acetate and 9 alpha-fluorohydrocortisone. The replacement of 9 alpha-fluorohydrocortisone therapy by desoxycorticosterone acetate therapy led to the cessation of the attacks.

Light and electron microscopic identification of a nerve sprout in muscle of normal adult frog.

Recent evidence from this laboratory indicates that axonal sprouting (and regression) occurs in neuromuscular junctions of normal adult frogs. In the present investigation, the appearance of a single nerve branch, which from light microscopy was assumed to be a sprout, was studied in ultrathin serial sections. In confirming the light microscopic evidence small synaptic contacts were found, which showed characteristics of new synapse formation. Unexpectedly, the Schwann cell surrounding the axon extended several microns distally from the axon tip. It appears that nerve sprouting (and regression) is a physiological event in adult frog muscles.

Abandoned synaptic sites in muscles of normal adult frog.

In previous investigations light microscopic cholinesterase (ChE) deposits without any nerve were found adjacent to normally occupied parts of a frog neuromuscular synapse [4, 9]. After identifying one such site at the light microscopic level ultrathin sections were cut and viewed with an electron microscope. ChE reaction product-filled secondary, clefts were observed in the region studied but a nerve was invariably missing. From this it is concluded that these loci are former synaptic sites from which nerve and Schwann cell have retracted, When incubating muscles with fluorescence-labelled alpha-bungarotoxin, all 23 abandoned sites found in 6 muscles remained bare of visible amounts of label. This indicate that receptor molecules eventually disappear from the synaptic membrane after retraction of the nerve and Schwann cell. No information as to the underlying time schedule of nerve retraction, turnover of ChE and alpha-bungarotoxin binding sites was obtained. Taken together with the evidence for synapse new formation in untreated frog muscles obtained previously [9, 11] the present observations indicate some ongoing remodeling of frog neuromuscular junction.

A case of adrenoleukodystrophy in a girl. Genetic considerations.

We report the case of a girl who developed leukoencephalopathy and adrenal atrophy and died at 3 years of age. Histologically, demyelination, gliosis, perivascular lymphocytic cuffing and sudanophilia were present in the brain. The adrenal cortex was atrophic. Ultrastructurally, there were numerous cytoplasmic inclusions in brain macrophages, consisting of two leaflets separated by an intervening space of variable low electron density. Brain tissue cholesterol esters contained a high proportion of long chain fatty acids. The findings are discussed and compared with those in the literature. Emphasis is placed on the fact that the disease occurred in a girl in apparent contradiction to the commonly accepted X-linked hereditary transmission of adrenoleukodystrophy. Some possible genetic explanations for our case are put forward.

Morphological, immunocytochemical and biological characteristics of experimental rabbit brain tumors in tissue culture.

Brain tumors were induced in 3-month-old rabbits of either sex by repeated intravenous injections of N-methyl-N-nitrosourea. Twelve brain tumors (6 pleomorphic gliomas, 5 grade 2--3 astrocytomas, 1 grade 2--3 oligodendroglioma) were established in culture and, with the exception of 2 neoplasms, were propagated in vitro as permanent cell lines. The glial nature of all cell lines was ascertained at several passage levels by testing the cells for the production of S-100 and GFA. It could be shown that most cells of all lines fluoresced positively for the S-100 protein, albeit differences in intensity of fluorescence were clearly noted between cells of the same culture and between different cultures. In general, astrocytoma cell lines had the strongest fluorescence. Pleomorphic glioma cells but especially astrocytoma cells reacted positively also for the GFA protein. Surprisingly enough, isolated cells of the oligodendroglioma line also showed evidence of GFA production. Exposure of cultures of rabbit glioma cells to db-cAMP for 8--10 hr resulted in inhibition of cell proliferation and stimulation of process formation. Furthermore, positive fluorescence for the S-100 and GFA proteins was more intense in cells treated with db-cAMP than in untreated cells. The latter observation may indicate that production and/or accumulation of glial proteins also was enhanced during the stationary phase of cell cultures.

Immunofluorescence study of lymphocytic infiltration in gliomas. Identification of T-lymphocytes.

Five human brain tumours (3 glioblastomas and 2 astrocytomas) and 5 rat brain tumours induced in Sprague--Dawley animals by systemic administration of N-methyl-N-nitrosourea (3 pleomorphic gliomas and 2 mixed gliomas) were studied. The human brain tumours were surgical specimens excised from patients with no cranial surgery prior to their disease. The experimental brain tumour had been adapted to tissue culture, propagated in vitro and then transplanted to immunocompetent and immunodeficient rats of the same stock. The above-described material was selected in consideration of the mononuclear cell infiltrates occurring in these tumours. Frozen sections of human and rat gliomas, the latter both primary and transplanted, were prepared and investigated as to the presence of T-lymphocytes within the mononuclear round cell infiltrates. This was done with the indirect immunofluorescence method using rabbit antisera against man and rat T-lymphocytes. With this technique a variable percentage of T-lymphocytes was demonstrated in the cell infiltrates of human and rat gliomas alike. The tumour transplanted in thymectomized rats showed only isolated, scattered, positive-reacting cells, i.e., cells recognizable as T-lymphocytes by the above method. The results can be interpreted as circumstantial evidence for the occurrence of tumour-specific and/or tumour-associated antigens in the parenchymal cells of spontaneous and chemically-induced gliomas.

Definition of a cell clone with astroglial characteristics derived from a chemically induced rabbit brain glioma.

Three-month-old rabbits were started on a fortnightly schedule of intravenous injections of N-methyl-N-nitrosourea. All but two of the central nervous system tumors induced in this manner were propagated in culture as permanent cell lines. On the 76RB-G-414-H line established from a grade 2 astrocytoma of this series of neoplasms, a cloning procedure was carried out using a laser microbeam. The clonal line originated in this way has been maintained in long-term culture and given the 76/RB-G-414-H-C designation. The cells of the clone display invariably a bipolar or multipolar configuration with long processes. Intermediate filaments are common and even abundant in some cells. Positivity for S-100 and GFA proteins is a regular finding in these cells. In addition, dibutyryl cyclic adenosine monophosphate treatment reduces cell division and stimulates cell process formation of these cells. Thus, it appears that we succeeded in establishing in vitro and maintaining in long-term culture a clone of tumor cells with astrocytic characteristics.

Stenosis of central canal of spinal cord in man: incidence and pathological findings in 232 autopsy cases.

The central canal of the spinal cord is generally regarded as a vestigial structure that is obliterated after birth in 70% to 80% of the general population. This report describes the first detailed histological study of the human central canal in 232 subjects ranging in age from 6 weeks' gestation to 92 years. Whole spinal cords were harvested at autopsy and sectioned serially from the conus medullaris to the upper medulla. Histological findings and morphometric analysis of the cross-sectional luminal area were used to grade stenosis at seven levels of the canal. Varying grades of stenosis were present at one or more levels in none (0%) of 60 fetuses, one (3%) of 34 infants, three (18%) of 17 children, 21 (88%) of 24 adolescents and young adults, 67 (96%) of 70 middle-aged adults, and all 27 adults aged 65 years or older (100%). The stenotic process was most pronounced in the thoracic segments of the canal and involved more levels with higher grades of stenosis in older individuals. Histological findings consisted of disorganization of the ependymal epithelium, formation of ependymal rosettes or microcanals, proliferation of subependymal gliovascular buds, and intracanalicular gliosis. These features are consistent with a pathological lesion involving ependymal injury and scarring and are less compatible with an involutional or degenerative process. Stenosis of the central canal probably influences the anatomical features of syringomyelia and may account for variations in cavity formation such as the prevalence of holocord syrinxes in children, the formation of focal and paracentral syrinxes in adults, and the rare incidence of syrinx formation in many older individuals with acquired lesions known to produce syringomyelia.

Pathological basis of spinal cord cavitation in syringomyelia: analysis of 105 autopsy cases.

This report summarizes neuropathological, clinical, and general autopsy findings in 105 individuals with nonneoplastic syringomyelia. On the basis of detailed histological findings, three types of cavities were distinguished: 1) dilations of the central canal that communicated directly with the fourth ventricle (47 cases); 2) noncommunicating (isolated) dilations of the central canal that arose below a syrinx-free segment of spinal cord (23 cases); and 3) extracanalicular syrinxes that originated in the spinal cord parenchyma and did not communicate with the central canal (35 cases). The incidence of communicating syrinxes in this study reflects an autopsy bias of morbid conditions such as severe birth defects. Communicating central canal syrinxes were found in association with hydrocephalus. The cavities were lined wholly or partially by ependyma and their overall length was influenced by age-related stenosis of the central canal. Non-communicating central canal syrinxes arose at a variable distance below the fourth ventricle and were associated with disorders that presumably affect cerebrospinal fluid dynamics in the spinal subarachnoid space, such as the Chiari I malformation, basilar impression, and arachnoiditis. These cavities were usually defined rostrally and caudally by stenosis of the central canal and were much more likely than communicating syrinxes to dissect paracentrally into the parenchymal tissues. The paracentral dissections of the central canal syrinxes occurred preferentially into the posterolateral quadrant of the spinal cord. Extracanalicular (parenchymal) syrinxes were found typically in the watershed area of the spinal cord and were associated with conditions that injure spinal cord tissue (for example, trauma, infarction, and hemorrhage). A distinguishing feature of this type of cavitation was its frequent association with myelomalacia. Extracanalicular syrinxes and the paracentral dissections of central canal syrinxes were lined by glial or fibroglial tissue, ruptured frequently into the spinal subarachnoid space, and were characterized by the presence of central chromatolysis, neuronophagia, and Wallerian degeneration. Some lesions extended rostrally into the medulla or pons (syringobulbia). Although clinical information was incomplete, simple dilations of the central canal tended to produce nonspecific neurological findings such as spastic paraparesis, whereas deficits associated with extracanalicular syrinxes and the paracentral dissections of central canal syrinxes included segmental signs that were referable to affected nuclei and tracts. It is concluded that syringomyelia has several distinct cavitary patterns with different mechanisms of pathogenesis that probably determine the clinical features of the condition.

Reactivity of tumor-infiltrating, blood, spleen and lymph node lymphocytes against syngeneic glioma target cells.

Brain tumors were induced in inbred Fischer rats (F344) by administration of methylnitrosourea in the drinking water. One of the induced tumors, a pleomorphic glioma (78-219), was established in vitro and propagated as 78FR-G-219 permanent cell line. The tumorigenicity of the established line was investigated by intracerebral or subcutaneous inoculation of 1X10 (6) - 10X10(6) cells. Lymphocytes infiltrating secondary tumors (TIL) were enriched by a single step centrifugation on different discontinuous Percoll density gradients, while blood lymphocytes (PBL) and lymphocytes from spleen and lymph node (SL and LNL) were enriched by Ficoll - Isopaque flotation. The reactivity spectrum of the isolated lymphocytes raised against cultured 78FR-G-219 cells was monitored by means of two different assays: Lymphocyte microcytotoxicity test (LMC) and colony inhibition assay (CIA). Reactivity of PBL, SL and LNL of glioma-bearing animals was clearly reduced, while TIL showed no natural killer (NK) activity, no cytotoxicity against syngeneic 78FR-G-219 pleomorphic glioma cells and no colony inhibition in mixed lymphocyte/target cell cocultivation. NK activity of TIL was only slightly reduced against target cells of a non-cross-reacting syngeneic astrocytoma line (78FR-G-299).

Radioimmunodetection of human glioma xenografts by radiolabelled monoclonal antibodies.

Radiolabelled monoclonal antibodies (131I-MUC 8-22, 131I-MUC 2-63) were used for external scintigraphy of human glioma xenografts. To induce transplantation tumors. 5 x 10(6) cells (85HG-66) of an in vitro established human malignant astrocytoma (N66/85) were inoculated s.c. in BALB/c-nu/nu mice. The labelling of the immunoglobulins with 131iodine was carried out according to the iodogen method, the nude mice, bearing xenograft, received 30 m. 131I-labelled intact monoclonal immunoglobulins (200mCi: 7,4MBq) and the imaging was performed on days 4, 8 and 12 after the application. After 4 days, a clear tumor accumulation of iodinated MUC 2-63 antibodies recognizing surface determinants was visible. This enrichment of monoclonal antibodies (MAbs) led to a characteristic tumor presentation on day 8. Obviously, the MUC 2-63 antibodies remain in the tumor tissue for a long time, so that even on day 12 satisfactory tumor imaging is possible. On the other hand, neither with normal mouse IgG nor with MUC 8-22 antibodies - which react with intracellular structures - could a tumor localization be achieved. The result of the studies on the distribution of 131I-MUC 2-63 on day 19 was that the activity in the tumor tissue was about 4.4 times higher than in the blood and even more times higher than in solid organs.

Immunological aspects of experimental brain tumors (review).

Evidence that experimental neural tumors contain glia specific and glioma-associated antigens is reviewed. The fact that glioma cells share antigens with normal glia cells is of crucial importance for the histogenetic immunodiagnosis of intracranial neoplasms. Moreover, the increasing use of in vitro techniques in neuro-oncology has accentuated the necessity for employment of cell-type characteristic antigens. This allows for objective identification of the various types of brain tumor cells, and also for ascertaining the neurological nature of long-term cultured cells. Humoral and cell-mediated immune reactions to gliomas could be demonstrated in autochthonous and syngeneic hosts. Since glioma-associated antigens are rather weak and glioma cells are low immunogens, various approaches for enhancing glioma-cell immunogenicity have been described, such as treatment with membrane-modifying enzymes, or haptenization with various chemicals. Recently, nitrophenylation of glioma cells has become available for artificially increasing the immunogenicity of these cells. Furthermore, methods have recently been worked out by which monoclonal antibodies of predefined specificity can be produced in order to analyze the nature of glioma-associated antigens. Such methods may have a significant impact on clinical immunodiagnostics, and perhaps on the development of new immunotherapeutic approaches.

Virions and virus-associated structures within dendrites in an experimental flavovirus encephalomyelitis.

In experimental yellow fever virus encephalomyelitis of adult albino mice, virions, and virus-associated structures were observed not only inside neuronal perikarya but also within dendrites of varied size. The finding permits the following explanations: (1) either the viral agent is synthesized in the nerve cell bodies and transported intradendritically in a proximodistal direction; or (2) virus morphogenesis takes place in neuronal perikarya and dendrites as well; or (3) both possiblities are equally valid. Some incidental findings were suggestive of virus release at postsynaptic dendrite membranes. They are discussed with reference to a hypothetical long-distance pathway of viral dissemination involving endocytosis of the agent by presynaptic axon terminals, intraaxonal virus decoating, and retrograde axoplasmic transport of the infectious nucleic acid to the cell soma.

The degenerative diseases or syndromes in maturing central nervous system.

The diagnosis of degenerative diseases or syndromes in the nervous system in based on their morphological picture. The changes occur in selected CNS structures or systems being induced in the course of more or less known processes sometimes with known, more often unknown etiology. Degenerative syndromes may be classified according to the topography of changes. They appear often with aging, but also in even greater number in infants. We tried to analyze the problem and find out to what degree the structure and topography of CNS degenerative changes in infants depend on maturity of nervous tissue constituting the background of pathologic process. The cases with two syndromes representative for small infants: progressive poliodystrophy of Alpers type and a degenerative syndrome with cerebral calcifications and disseminated demyelination were examined from this point of view. Our observations revealed that the stage of CNS development stipulates the type and topography of degenerative changes.

The white matter changes in microencephalic HIV infected infants. A preliminary report.

The brains of six infants 14-34 months of age and with microencephaly (brain weight deficit 20-55.5%) were chosen from a group of cases vertically infected with HIV. The center of our investigations was focused on the white matter changes of which two types were observed in the examined brains. Within the periventricular white matter of four cases evident lesions consisting of myelin pallor and concomitant gliosis were recognized as HIV-1 infection related leukoencephalopathy. In all those cases myelination delay was also noted. In one case HIV encephalitis was diagnosed. Our observations suggest that in the majority of HIV infected infants changes resulting in the brain "too small for age" corelate with myelination delay coexisting with early-onset leukoencephalopathy. Because of the small number of cases in this study the results should be considered preliminary, and will require further investigations.

Prominent white matter cavitation in an infant with Alexander's disease.

The clinical history and autopsy findings are reported on a case of infantile Alexander's disease (AD). The patient, a white baby girl, developed seizures at age 4 months accompanied by internal hydrocephalus. She died at age 11 months following a progressive, downhill course of profound psychomotor retardation, recurrent seizures and cachexia. The general autopsy was remarkable for cachexia. The formalin fixed brain and spinal cord were studied by light and electron microscopy (EM). The brain was normal in weight for age but showed diffuse pallor of white matter and marked cavitation involving the cerebral and cerebellar subcortical white matter, most profound in the frontal lobes. Microscopically the CNS showed classic features of AD with diffuse paucity of myelin and massive proliferation of astrocytes bearing Rosenthal fibers (RF). The latter appeared as granular osmiophilic deposits associated with 8-10 nm filaments within astrocytic processes and cell bodies by EM. This case of AD is remarkable for the extreme degree of cavitation. Cavitary changes affect up to one third of typical cases of AD and are invariably present in the frontal white matter. Affected patients are generally much younger and have a shorter clinical course than AD patients without brain cavitation. The dysmyelination of AD inversely parallels the temporal sequence of normal myelination and suggests a relative resistance of early myelinated structures to the presumed astrocytic defect causing AD. Adults with de novo formation of RF's in the CNS have a varied clinical and pathological appearance, rarely show brain cavitation and should probably be distinguished from classic AD in children.