There is a pressing need for a more accurate system of classifying human astrocytomas, one that is based on morphologic characteristics and that could also make use of distinctive biochemical markers. However, little is known about the phenotypic characteristics of astrocytomas. Recent studies have shown that the expression of proteins comprising the intermediate filament (IF) cytoskeleton of astrocytic cells is developmentally regulated. It is our hypothesis that this changing protein profile can be used as the basis of a system for clearly and objectively classifying astrocytomas. A spectrum of human astrocytomas has been examined by immunofluorescence microscopy employing antibodies to several IF structural subunit proteins (GFAP, vimentin, and keratins) and an IF-associated protein, IFAP-300kDa. These proteins occupy unique temporal niches in the cytogenesis of the astrocytic cells: keratins in cells of the neuroectoderm; vimentin and IFAP-300kDa in radial glia and immature glia; GFAP in mature astrocytes; and vimentin in some mature astrocytes. In agreement with previous reports, our immunofluorescence studies have revealed both GFAP and vimentin in all astrocytoma specimens. Two new observations, however, are of particular interest: IFAP-300kDa is detectable in all astrocytic tumors, and the proportion of keratin-containing cells present in the astrocytomas is in direct relationship to the degree of the malignancy. Because IFAP-300kDa is not present in either normal mature or reactive astrocytes, this protein appears to represent a specific marker of transformed (malignant) astrocytes. If it is presumed that higher malignancy grades represent the most dedifferentiated cellular state of the astrocytes, the presence of keratin-containing cells is not totally unexpected, given the ectodermal (epithelial) origin of the CNS. Specific developmentally regulated proteins of the IF cytoskeleton thus appear to hold great potential as diagnostic markers of astrocytomas and as tools for investigating the biology of these tumors.

Astrocytes↗

PubMedDate not supplied

Origin of afferent projections into bovine chromaffin cell implants in the rat periaqueductal gray determined by retrograde and anterograde tracing.

We have previously described long-term survival of isolated bovine chromaffin cell suspension grafts in the periaqueductal gray of adult rats. Electron microscopic analysis of the graft sites revealed synapses on the transplanted chromaffin cells. The origin of these synapses is not known, but they are probably derived from the host since the initial grafts were suspensions of chromaffin cells that were essentially free of other cell types. In order to determine the origin of the observed synapses, retrograde and anterograde tracer analyses were performed on grafted rats at 4 and 8 weeks after transplantation. Following injection of the retrograde tracer (Fluoro-Gold) into graft sites, four major host sites were labeled: hindbrain reticular formation, substantia nigra, lateral hypothalamus, and cingulate cortex. Injection of anterograde tracer (rhodamine-conjugated dextranamine) into the substantia nigra, lateral hypothalamus, and cingulate cortex produced labeled fibers and terminals in and around 4 and 8 weeks old chromaffin cell graft sites. An increase in both the number of retrogradely labeled cells, as well as in the density of anterogradely labeled fibers and terminals within the graft site, was observed from 4 to 8 weeks. This study shows that graft innervation from the host is primarily from areas that normally project afferent fibers to the periaqueductal gray. The increase in labeled fibers and terminals over 8 weeks suggests that de novo synapse formation on grafted bovine chromaffin cells is a continuous process that is dependent on the regenerative capacity and plasticity of the host neuronal network and the grafted bovine chromaffin cells.

Afferent Pathways↗

PubMedDate not supplied

Expression of type VI collagen in psammoma bodies: immunofluorescence studies on two fresh human meningiomas.

OBJECTIVE: To demonstrate that psammoma bodies in human meningiomas contain type VI collagen and laminin. STUDY DESIGN: Two fresh human meningiomas were obtained from surgery and the frozen sections examined for type VI collagen and laminin with specific antibodies. RESULTS: Type VI collagen was found on the surface of the psammoma bodies. In addition, it was detected in the interstitial tissues surrounding the tumor cells and on the tumor cells themselves. Laminin was detected mainly in the walls of capillaries and large blood vessels. In contrast, in some psammoma bodies, laminin was detectable internally. Laminin was not found in the interstitial tissues or tumor cells. CONCLUSION: This is the first report to describe the involvement of type VI collagen in psammoma bodies and whorl formations in meningiomas. This specific type of collagen may serve an anchoring function in psammoma body development.

Brain Neoplasms↗

PubMedDate not supplied

Long-term viability of isolated bovine adrenal medullary chromaffin cells following intrastriatal transplantation.

Adrenal medullary grafts generally exhibit poor viability when grafted into the striatum. Previous work in our laboratory demonstrated that chromaffin cells can survive well for up to 2 mo following grafting into the intact rat striatum after cells are isolated from the nonchromaffin supporting cells (fibroblasts and endothelial cells) of the adrenal medulla. The aim of the present study was to assess the long-term viability of isolated bovine chromaffin cells following grafting into the intact rat striatum. The viability of grafted bovine adrenal medullary chromaffin cells was compared in rats receiving either (a) perfused adrenal medulla; (b) isolated chromaffin cells; or (c) isolated chromaffin cells that were subsequently recombined with their nonchromaffin supporting cells. One year postimplantation, all graft types which included fibroblasts and endothelial cells were infiltrated with macrophages and demonstrated an abundance of cellular debris. No viable chromaffin cells were observed. In contrast, healthy tyrosine hydroxylase (TH) and dopamine beta hydroxylase (D beta H) immunoreactive chromaffin cells survived for 1 yr posttransplantation when grafted in isolation from the nonchromaffin constituents of the adrenal medulla. Good xenograft survival was achieved in this group despite the fact that these rats were only immunosuppressed for 1 mo postimplantation. Grafted cells demonstrated morphological characteristics of chromaffin cells in situ and these implants were not accompanied by macrophage infiltration. These data demonstrate that long-term survival of chromaffin cells can be achieved following intrastriatal implantation and the viability of grafted chromaffin cells is dependent upon the removal of the nonchromaffin supporting cells.

Adrenal Medulla↗

PubMedDate not supplied

Bovine chromaffin cells for CNS transplantation do not elicit xenogeneic T cell proliferative responses in vitro.

Adrenal chromaffin cells have been utilized for several neural grafting applications, but limitations in allogeneic donor availability and dangers inherent in autografting limit the widespread use of this approach clinically. While xenogeneic donors offer promise as a source for cell transplantation in the central nervous system (CNS), immunologic responses to cellular components of the adrenal medulla have not been well characterized. To further study the host T cell xenogeneic response to chromaffin and passenger cells of the adrenal medulla, an in vitro lymphocyte proliferation assay was used. Lymphocyte proliferation was determined by mixing rat lymphocytes with potential stimulator cell subpopulations of the bovine adrenal medulla: isolated chromaffin cells, isolated endothelial cells, or passenger nonchromaffin cells, which include a mixture of fibroblasts, smooth muscle cells, and endothelial cells. As a positive control, bovine aortic endothelial cells were also used. 3[H]-thymidine incorporation, corresponding to lymphocyte proliferation, was measured. Results indicated that isolated bovine chromaffin cells produce only a mild, statistically insignificant stimulation of rat lymphocytes. In contrast, there was a significant response to passenger nonchromaffin cells of the adrenal medulla, especially endothelial cells. The inclusion of low levels of cyclosporin A in the cultures completely eliminated the mild proliferative response to isolated bovine chromaffin cells, while near toxic levels were necessary to abrogate the response to endothelial cells. Immunocytochemical analysis revealed that routine chromaffin cell isolation procedures result in the inclusion of a small percentage of endothelial cells, which may be responsible for the slight lymphocyte stimulation. The results of this study indicate that isolated chromaffin cells possess low immunogenicity, and suggest that passenger cells in the adrenal medulla, particularly endothelial cells, may be primarily responsible for progressive rejection in CNS grafts. Thus, removal of passenger nonchromaffin cells from xenogeneic donor tissues prior to transplantation may produce a more tolerated graft in rodent models of neural transplantation.

Adrenal Medulla↗

PubMedDate not supplied

One-year chromaffin cell allograft survival in cancer patients with chronic pain: morphological and functional evidence.

The control of chronic pain through transplantation of chromaffin cells has been reported over the past few years. Analgesic effects are principally due to the production of opioid peptides and catecholamines by chromaffin cells. Clinical trials have been reported with allografts consisting of whole-tissue fragments implanted into the subarachnoid space of the lumbar spinal cord (14,19,36). In the present study, allogeneic grafts were successfully used to control chronic pain in two patients over a period of 1 yr based on patient reported pain scores, morphine intake, and CSF levels of Met-enkephalin. Macroscopic examination at autopsy located the transplanted tissue fragments in the form of multilobulated nodules at the level of the spinal axis and cauda equina. Immunocytochemical microscopy showed neuroendocrine cells are positive for chromagranin A (CGA), and enzymes tyrosine hydroxylase (TH) and dopamine-beta-hydroxylase (DbetaH). The results suggest that there is a relationship between analgesic effect, Met-enkephalin levels in CSF, and the presence of chromaffin cells surviving in spinal subarachnoid space.

Adult↗

PubMedDate not supplied

Purification of adrenal chromaffin cells increases antinociceptive efficacy of xenotransplants without immunosuppression.

We have found that immunosuppression is necessary for the survival of xenogeneic adrenal medullary transplants. Because chromaffin cells are essentially nonimmunogenic, it is likely that the highly immunogenic "passenger" cells in the transplant preparation bring about rejection. This article describes a procedure that produces an essentially pure preparation of chromaffin cells for transplantation. Bovine adrenal medullary cells were isolated and differentially plated, resulting in a semipurified preparation of chromaffin cells. Ferromagnetic beads were added to the cell suspension, some of which were phagocytized by endothelial cells, which allowed their removal by exposure to a magnet. The remaining cells were then exposed to ferromagnetic beads coated with isolectin B4 from Griffonia simplicifolia and once again to a magnetic field. The "semipurified" preparation contained approximately 90% chromaffin cells, whereas the "highly purified" preparation was > 99.5% chromaffin cells as determined immunohistochemically. The immunogenicity of the two cell preparations was assessed in vitro by determining their capacity to evoke lymphocyte proliferation. Rat spleen lymphocytes were mixed with either a highly purified or semipurified population of bovine chromaffin cells. The results of this assay demonstrated that the highly purified preparation was a much weaker stimulant of lymphocyte proliferation than was the semipurified preparation and may demonstrate better graft survival in vivo. Transplantation via intrathecal catheter of either 80,000 or 250,000 cells from the highly or partially purified preparations onto the lumbar spinal cord of nonimmunosuppressed and non-nicotine-stimulated rats produced a cell number-dependent antinociception for both A(delta) and C fiber-mediated thermonociception at 6 days after transplantation. After 6 days and up to 28 days, only the "highly purified" preparation showed antinociception. These results suggest that nearly complete purification of bovine chromaffin cells minimizes immunorejection of xenogeneic transplants of these cells.

Adrenal Medulla↗