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Biomedical subjects

H B Clark

Publications and source records attributed to H B Clark.

At least 55 records · Page 3Linked to original sources
PubMed1989

The adoptive transfer of chronic relapsing experimental allergic encephalomyelitis with lymph node cells sensitized to myelin proteolipid protein.

In this report we describe the transfer of chronic relapsing experimental allergic encephalomyelitis (EAE) with in vitro-stimulated lymph node cells (LNC) from SJL/J mice immunized with human myelin proteolipid protein (PLP). No additional immune enhancing procedures were applied in the transfer recipients. Clinical and histological EAE was transferred with 10-30 X 10(6) LNC to 27/28 mice. The LNC proliferated in vitro to PLP, but not to myelin basic protein (MBP), and induced delayed-type hypersensitivity. Enrichment for lymphoblasts by Ficoll centrifugation was essential for the disease development. The clinical course usually showed an early episode of acute paralytic illness, followed by chronic relapsing disease, and resembled the transfer of EAE using MBP-specific cells, both clinically and histologically.

Animals↗
PubMed1988

Expression of nerve growth factor receptors by Schwann cells of axotomized peripheral nerves: ultrastructural location, suppression by axonal contact, and binding properties.

Axotomy of sciatic nerve fibers in adult rats induces expression of NGF receptor in the entire population of Schwann cells located distal to the injury (Taniuchi et al., 1986b). In the present study we have used immunocytochemistry, with a monoclonal antibody directed against the rat NGF receptor, to examine axotomized peripheral nerves by light and electron microscopy. We have found that (1) the NGF receptor molecules were localized to the cell surface of Schwann cells forming bands of Bungner; (2) axonal regeneration into the distal portion of sciatic nerve coincided temporally and spatially with a decrease in Schwann cell expression of NGF receptor; (3) Schwann cell NGF receptor could be induced by axotomy of NGF-independent neurons, such as motoneurons and parasympathetic neurons; and (4) the presence of axon-Schwann cell contact was inversely related to expression of Schwann cell NGF receptor. Using biochemical assays we have found that, in striking contrast to peripheral nerves, there was no detectable induction of NGF receptor in the spinal cord and brain after axotomy of NGF receptor-bearing fibers. Filtration assays of 125I-NGF binding to the induced NGF receptors of Schwann cells measured a Kd of 1.5 nM and a fast dissociation rate, both characteristics of class II receptor sites. We conclude that Wallerian degeneration induces Schwann cells, but not central neuroglia, to produce and position upon their plasmalemmal surface the class II NGF receptor molecules. The induction is ubiquitous among Schwann cells, irrespective of the type of axon they originally ensheathed. Expression of Schwann cell NGF receptor is negatively regulated by axonal contact, being induced when axons degenerate and suppressed when regenerating axons grow out along the Schwann cell surface. We propose that the induced NGF receptors function to bind NGF molecules upon the Schwann cell surface and thereby provide a substratum laden with trophic support and chemotactic guidance for regenerating sensory and sympathetic neurons.

Animals↗
PubMed1987

Myelin proteolipid protein induces demyelinating disease in mice.

Using two methods of immunization (A and B), 5/10 (A) and 27/45 (B), BALB/c by J mice immunized with human myelin proteolipid protein developed a demyelinating disease with a spectrum of chronic progressive to relapsing-remitting courses. Demyelinative lesions were seen histopathologically in all clinically affected animals that were examined. Many of the clinically unaffected animals also had histopathologic evidence of demyelination. Some of the animals had evidence of multiple ages of foci of activity. Evidence is presented that contamination with myelin basic protein could not account for the disease. This is a new model for multiple sclerosis in mice.

Animals↗
PubMed1987

Demonstration of the retrograde transport of nerve growth factor receptor in the peripheral and central nervous system.

NGF acts on responsive neurons by binding to specific NGF receptors on axonal termini, after which a critical biochemical signal is retrogradely transported to the cell body. The identity of the signal(s) is unknown; candidates include NGF itself or some other "second messenger." A possible second messenger is the NGF receptor. As a first step in assessing the possible role of NGF receptor in the generation of the NGF-dependent signal, and in understanding the economy of NGF receptor synthesis and utilization, we determined whether the NGF receptor is retrogradely transported. Using immunohistochemical staining with a monoclonal antibody (192-IgG) against rat NGF receptor, we looked for accumulation of NGF receptor molecules distal (retrograde transport), as well as proximal (anterograde transport), to sites of axonal ligation or transection. By 10-12 hr in both the ligated sciatic nerve and the lesioned fimbria-fornix, accumulated NGF receptor was detected proximal and distal to the ligation/lesion site. The transported receptor presumably was located in sympathetic and sensory neurons in the sciatic nerve and in forebrain cholinergic neurons projecting from the medial septum to the hippocampus. In both anatomical sites, accumulation of NGF receptor on the proximal (anterograde) side occurred in streams of fine axonal processes, whereas staining on the distal (retrograde) side occurred in varicose or granular configurations. These results raise the possibility that the NGF receptor has a role in the mechanism of NGF beyond the initial binding event at the plasma membrane of the axonal terminus.

Animals↗
PubMed1987

Acrylamide-induced sympathetic autonomic neuropathy resulting in pineal denervation.

The effect of acrylamide on the sympathetic innervation of the rat pineal gland was examined by using ultrastructural, immunohistological, biochemical, and physiological methods. Separation of sympathetic terminals from perivascular pineocyte processes was facilitated by administration of the false neurotransmitter 5-hydroxydopamine, which preferentially labeled sympathetic terminals, shortly before sacrifice. Administration of acrylamide (50 mg/kg/day, intraperitoneally) for 1 to 2 weeks resulted in the near-total loss of pineal parenchymal perivascular axons and axons intercalated between individual pineocytes. More proximal portions of these sympathetic axons located within the capsule of the pineal gland developed markedly enlarged swellings containing neurofilaments and tubulovesicular elements. The ultrastructural appearance of axons swollen by tubulovesicular elements resembled that of regenerating axons and axons whose regenerative progress had been frustrated. The activity of pineal dopamine-beta-hydroxylase, a noradrenergic marker enzyme confined to sympathetic axons and their terminals and absent in pineocytes, was determined in an attempt to develop a quantitative measure of the extent of sympathetic denervation. The loss of 50% of pineal dopamine-beta-hydroxylase activity underestimated the extent of parenchymal denervation due to the marked engorgement of remaining capsular sympathetic axons by immunoreactive dopamine-beta-hydroxylase. The daily rhythm of pineal serotonin N-acetyltransferase (NATase) activity, which is dependent on the circadian variation in the activity of pineal sympathetic axons, was decreased 90% by chronic acrylamide administration. Pineal NATase activity increased 25- to 50-fold in acrylamide intoxicated rats in which isoproterenol was used to stimulate pineocyte beta-adrenergic receptors directly, which is evidence against a nonspecific toxic effect of acrylamide on pineocytes. Administration of N,N'methylene-bis-acrylamide, a non-neurotoxic analog of acrylamide, was without effect on pineal ultrastructure or NATase activity.

Acrylamide↗
PubMed1986

Induction of nerve growth factor receptor in Schwann cells after axotomy.

We have discovered that axotomy of sciatic nerve induces Schwann cells distal to the lesion to express de novo, or at greatly increased levels, receptors for nerve growth factor (NGF). Surgical transection of sciatic nerve was performed on adult Sprague-Dawley rats, and, at various times after the operation, the following tissues were dissected for quantitation of NGF receptor: L4 and L5 dorsal root ganglia, sciatic nerve proximal to the transection, sciatic nerve distal to the transection, tibialis anterior muscle, and skin of the dorsum of the foot. The NGF receptor content of these samples was determined by labeling receptor molecules with radioiodinated NGF (125I-NGF) and then specifically immunoprecipitating the 125I-NGF-receptor complexes with 192-IgG, a monoclonal antibody directed against the rat NGF receptor. We observed a time-dependent increase in the amount of radioligand-labeled NGF receptor proteins found in the distal segment of transected sciatic nerve; by 7 days the density of receptor (crosslinked 125I-NGF molecules per mg of homogenate protein) had increased at least 50-fold. The number of receptor molecules in tibialis anterior muscle and dorsal foot skin, two structures denervated by the transection, also increased, with time courses parallel to that of distal sciatic nerve. There was little increase in the density of NGF receptors in the sciatic nerve proximal to the lesion and in the dorsal root ganglia. Immunohistochemical examination of the distal portion of transected sciatic nerve and of the muscle, with 192-IgG as the primary ligand, revealed prominent and exclusive staining of apparently all Schwann cells of the endoneurium, indicating that these peripheral neuroglial cells were expressing NGF receptors. These results show that axonal damage can induce the expression of NGF receptors in the population of sheath cells thought to promote neuronal regeneration. This dramatic increase in NGF receptors may be a mechanism to facilitate the regeneration of NGF-responsive neurons.

Animals↗
PubMed1985

Electron microscopic immunocytochemical localization of myelin proteolipid protein and myelin basic protein to oligodendrocytes in rat brain during myelination.

Electron microscopic immunocytochemical studies were carried out to localize myelin basic protein and myelin proteolipid protein during the active period of myelination in the developing rat brain using antisera to purified rat brain myelin proteolipid protein and large basic protein. The anti-large basic protein serum was shown by the immunoblot technique to cross-react with all five forms of basic protein present in the myelin of 8-day-old rat brain. Basic protein was localized diffusely in oligodendrocytes and their processes at very early stages in myelination. The immunostaining for basic protein was not specifically associated with any subcellular structures or organelles. The ultrastructural localization of basic protein suggests that it may be involved in fusion of the cytoplasmic faces of the oligodendrocyte processes during compaction of myelin. Immunoreactivity in the oligodendrocyte and myelin due to proteolipid protein appeared at a later stage of myelination than did that due to basic protein. Staining for proteolipid protein in the oligodendrocyte was restricted to the membranes of the rough endoplasmic reticulum, the Golgi apparatus, and apparent Golgi vesicles. The early, uncompacted periaxonal wrappings of oligodendrocyte processes were well stained with antiserum to large basic protein whereas staining for proteolipid protein was visible only after the compaction of myelin sheaths had begun. Our evidence indicates that basic protein and proteolipid protein are processed differently by the oligodendrocytes with regard to their subcellular localization and their time of appearance in the developing myelin sheath.

Animals↗
PubMed1985

Myelin basic protein and P2 protein are not immunohistochemical markers for Schwann cell neoplasms. A comparative study using antisera to S-100, P2, and myelin basic proteins.

Immunohistochemical localization of tissue specific or cell-specific antigenic markers in neoplastic cells has become an increasingly important tool in the pathologic diagnosis of tumors. The myelin-specific proteins of peripheral nervous system myelin, because they are normally synthesized in Schwann cells, are potentially useful markers for neoplasms arising from peripheral nerves. The authors carried out immunohistochemical studies on 18 cases of Schwann cell neoplasms, including schwannomas, neurofibromas, and granular cell tumors, to determine whether two myelin-specific proteins, myelin basic protein and P2 protein, were present in neoplastic Schwann cells. None of these tumors showed immunostaining for either myelin basic protein or P2 protein in neoplastic cells. In contrast, S-100 protein, which is a well established marker for normal and neoplastic Schwann cells, was localized by immunohistochemistry to neoplastic cells in all 18 neoplasms. Therefore, although myelin basic protein and P2 protein are known to be Schwann-cell-specific proteins, they do not appear to be expressed commonly in neoplastic Schwann cells.

Histocytochemistry↗
PubMed1984

Hypertension induced changes in cerebral capillary permeability to water are mediated by afferent neuronal connections from the carotid sinus to the brain.

Mild acute hypertension was induced in rats pharmacologically and non-pharmacologically with metaraminol, angiotensin II or bilateral vagotomy. The extraction fraction of water (Ew) was measured in both hypertensive and normotensive animals. It was demonstrated that cerebral blood flow did not change during hypertensive states. Under constant flow conditions Ew becomes a direct index of the cerebral capillary permeability to water. Ew was significantly decreased (P less than 0.001) in all brain regions examined in the hypertensive animals as compared to normotensive controls. However, the decrease Ew induced by hypertension was not observed if the animal had previously received bilateral carotid sinus denervation. This observation indicated that the capillary permeability response induced by hypertension is mediated by a neurogenic mechanism involving information transfer from the peripheral baroreceptors in the carotid sinus along afferent connections into the central nervous system.

Afferent Pathways↗
PubMed1984

Identification of antibodies in anti-CNS and anti-PNS myelin sera by immunoblot, characterization by immunohistochemistry, and their effect in tissue culture.

Immunoblot analysis of antiserum to rat central nervous system (CNS) myelin revealed antibodies to myelin basic protein (MBP), proteolipid protein (PLP), and numerous high molecular weight proteins. In addition, anti-CNS myelin serum exclusively immunostained 4 basic proteins of rat peripheral nervous system (PNS) myelin. Similarly, anti-PNS myelin sera immunostained many high molecular weight proteins in both CNS and PNS myelin in addition to P0 and 4 basic proteins. Purified MBP and PLP were immunostained by anti-CNS myelin sera and MBP and P0 by anti-PNS myelin sera, indicating that antigenic sites are preserved during protein purification. Immunohistochemical localization with antisera was confined to the myelin sheath except that antisera to CNS myelin also stained oligodendrocytes during the active period of myelination. While anti-CNS myelin sera specifically demyelinated centrally myelinated fibers in culture, none of the anti-PNS myelin sera used here demyelinated organotypic spinal cord-dorsal root ganglion cultures.

Animals↗
PubMed1984

Identification of dystrophic sympathetic axons in experimental diabetic autonomic neuropathy.

Markedly dilated dystrophic post-ganglionic sympathetic axons have been identified by dopamine-beta-hydroxylase immunohistochemistry in the paravascular ileal mesenteric nerves of rats with chronic streptozotocin diabetes. Many axons contained multiple dilatations with interposed axonal segments of near normal dimensions. These axonal abnormalities were absent in control animals. The time course of the development of the axonopathy and its distribution in the alimentary tract correlate with quantitative ultrastructural findings previously reported in this system.

Animals↗
PubMed1984

Immunohistochemistry of nervous system antigens: diagnostic applications in surgical neuropathology.

Immunohistochemical localization of nervous system antigens is becoming an increasingly useful tool in the diagnosis of neuropathologic lesions. Use of antigen markers specific for certain cell types, when properly employed, can confirm suspected diagnoses or solve difficult differential diagnoses. Presently the most useful markers are the glial antigens, glial fibrillary acid protein and S-100 protein, but a number of other markers, such as neurofilament proteins and neuron-specific enolase, have shown some diagnostic utility.

Antigens↗
PubMed1983

Demyelinating and myelination-inhibiting factors induced by chloroform-methanol insoluble proteins of myelin.

A host of proteins was seen when the chloroform-methanol insoluble protein (CMIP) fraction of bovine brain myelin was transferred from polyacrylamide gels to cellulose nitrate sheets. Inoculation of rabbits with the CMIP fraction generated a number of antibodies which were demonstrated by the immunoblot technique. These antisera against CMIP contained antibodies which induced demyelination and inhibited myelin formation in central nervous system cultures. The demyelinating factor was specific for centrally myelinated fibers, and did not demyelinate peripherally myelinated axons.

Animals↗
PubMed1983

Primary melanocytic schwannoma of the stomach. Evidence of dual melanocytic and schwannian differentiation in an extra-axial site in a patient without neurofibromatosis.

Pigmented neoplasms of the peripheral nervous system are uncommon. Such lesions, while well documented in the para-axial region, are infrequently encountered in more peripheral extra-axial sites. The authors present a case of a melanocytic schwannoma arising in the gastric antrum of a 51-year-old woman. Ultrastructural features attesting to the nerve sheath origin of the neoplasm included the presence of a well-developed basal lamina, complex infoldings of the plasma membrane, and intercellular bundles of long spaced collagen ("Luse bodies"). Premelanosomes and melanosomes in all stages of development were identified within the cytoplasm of many of the neoplastic cells, providing morphologic support for a close relationship between Schwann cells and melanocytes. Immunoperoxidase studies with S-100 protein disclosed intense cytoplasmic staining throughout the tumor. The S-100 protein antigen has been documented in both nerve sheath elements and melanocytes; this shared antigenicity lends further support to the close relationship between melanocytic and schwannian elements suggested by ultrastructural observations. Additional immunoperoxidase studies employing antibody to glial fibrillary acidic protein revealed focal cytoplasmic staining, an uncommon phenomenon in peripheral nerve sheath neoplasms. A review of the literature failed to provide any previous well documented cases of melanocytic schwannoma of the stomach. The patient was treated with en bloc resection of the tumor and gastric antrum, and was free of evidence of recurrent or metastatic disease 22 months postoperatively. Theories regarding the histogenesis of pigmented nerve sheath neoplasms and prognostic implications are discussed, based on experience with this and previously reported pigmented nerve sheath tumors.

Cell Differentiation↗
PubMed1983

Gastric stromal tumors. Reappraisal of histogenesis.

Twenty-eight gastric wall tumors classified by light microscopy as leiomyomas or leiomyosarcomas were reevaluated for histogenesis. Each case was analyzed for the presence of S-100 protein, a marker for cells derived from neuroectoderm, by the immunoperoxidase technique. Eight tumors contained cells with positive immunostaining for S-100 protein. Usually this staining was focal, but in one case staining was diffuse. In three additional cases the immunostaining outlined a nerve within the tumor. In contrast, two esophageal and 10 uterine leiomyomas, as well as normal gastric smooth muscle, gave negative reactions for S-100 protein. Twelve cases had tissue processed for electron microscopy. Only two of the tumors, one leiomyoma and one leiomyosarcoma, contained cytoplasmic myofilaments with densities expected in cells derived from smooth muscle; neither of these tumors stained for S-100 protein. In one case, the tumor with diffuse staining for S-100 protein, the cells resembled Schwann cells ultrastructurally. The remaining nine cases had neither smooth muscle nor Schwann cell features. They did contain interposed cell processes, primitive junctions, and large cytoplasmic vacuoles. The results of this study indicate that many gastric wall tumors are not derived from smooth muscle. The presence of S-100 protein suggests a nerve sheath origin in some cases. While the ultrastructure of many gastric tumors does not resemble nerve sheath cells in most peripheral nerves, the myenteric nervous system is a possible source for perineurial or mesenchymal nerve sheath cells with distinctive fine structure. Further study is needed to refine our knowledge of the histogenesis of gastric stromal tumors.

Esophageal Neoplasms↗
PubMed1982

An intravenous technique for the measurement of cerebral vascular extraction fraction in the rat.

An intravenous injection method to measure cerebral vascular extraction fractions of highly diffusible substances in the rat is described. The brain extraction fractions of 3H-labeled water (Ew) and ethanol (Ee) were defined as the ratio of either of those tracers to the freely diffusible reference tracer, 14C-butanol, in the brain, divided by the ratio of the tracers available for the extraction during the time between simultaneous intravenous injection of the tracers and decapitation of the rat. Ew and Ee were measured in five regions of brain, including brainstem and cerebellum, under PaCO2 conditions ranging from 15 to 85 mm Hg. The extraction fractions for both test tracers were shown to vary with PaCO2-induced flow changes according to the equation, ln(1 - E) = -PS/F. When PS/F values calculated from regional measurements of Ew and Ee were plotted versus PaCO2, least squares regression equations of the plots could be used to compare permeabilities of both tracers at any given PaCO2 value. Ratios of the permeabilities of water and ethanol varied regionally but were relatively constant in a given region under different flow states. This intravenous injection method allows for accurate measurement of the extraction fractions of even highly diffusible tracers under varied flow conditions in all brain regions regardless of arterial blood supply.

Animals↗