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Biomedical subjects

B N Fields

Publications and source records attributed to B N Fields.

At least 109 records · Page 6Linked to original sources
PubMed1983

Determinants of reovirus interaction with the intestinal M cells and absorptive cells of murine intestine.

Reovirus type 1 penetrates the gastrointestinal tract in suckling mice via specialized epithelial cells, designated membranous cells, or M cells, located in the epithelium overlying Peyer's patches. We have examined whether the interaction of reovirus with murine mucosa of in situ closed ileal loops is influenced by mouse age or strain or reovirus serotype. Neither mouse age (suckling or adult), strain (C3H/HeJ or Balb/cJ), nor reovirus serotype (types 1 and 3) affected reovirus adherence to and transport through M cells. In all conditions, reovirions adhered to the M-cell surface and were transported across M cells in endocytic vesicles. The adherence to and endocytosis by M cells of type 1 reovirus and reassortants with the viral hemagglutinin of type 1 were selective in suckling mice; type 1 virus was not adherent to nor endocytosed by absorptive cells. In adult mice, type 1 reovirions adhered to the surface of a minority of absorptive cells but were never seen within absorptive cell cytoplasm. In contrast, type 3 reovirus and reassortants with the viral hemagglutinin of type 3 adhered to and were endocytosed not only by M cells but also by absorptive cells of suckling mice. Virions accumulated within lysosomelike bodies in absorptive cells but transport of virions across absorptive cells was not observed. These studies indicate that (a) adherence of reovirus to the apical surface of and transcellular transport by M cells is independent of viral serotype or viral surface proteins, (b) adherence of reovirus to and transcellular transport by M cells is independent of mouse age after 9 days and comparable in two mouse strains, and (c) adherence of reovirus to and their endocytosis by absorptive cells of suckling mice is determined by the viral hemagglutinin (sigma 1 protein).

Age Factors↗
PubMed1983

Cell receptors for the mammalian reovirus. I. Syngeneic monoclonal anti-idiotypic antibody identifies a cell surface receptor for reovirus.

We have prepared and characterized a syngeneic monoclonal anti-idiotype directed against the idiotype (Id3) of the anti-reovirus type 3 hemagglutinin B cell hybridoma 9BG5. We have shown this monoclonal antibody to be anti-idiotypic in a series of inhibition studies with purified sigma 1 (hemagglutinin) protein and with absorption studies on the idiotypic R1.1 cell line. That the idiotype Id 3 defined by the monoclonal anti-idiotype is distinct from H-2 was demonstrated by its presence on an idiotype-expressing H-2 negative cell line (R1.E). Binding of the anti-idiotype is relatively tunicamycin resistant and pronase/trypsin sensitive, which suggests that the recognition site of the anti-idiotypic receptor is poorly glycosylated. The presence of idiotype-like receptors on reovirus type 3 susceptible neurons may have important implications for the presence of related gene families between neural and lymphoid tissue. The implications of recognition by neuronal cells of an anti-idiotypic monoclonal antibody directed against the reovirus 3 attachment receptor for the pathogenesis of viral-induced auto-immunity are discussed.

Animals↗
PubMed1983

Cell receptors for the mammalian reovirus. II. Monoclonal anti-idiotypic antibody blocks viral binding to cells.

A monoclonal anti-idiotypic antibody directed against an idiotypic determinant on a monoclonal anti-reovirus type 3 hemagglutinin antibody with viral neutralization activity is capable of binding to the surface of lymphoid cells (BW5147, R.1.1, and R1.E) and of inhibiting viral binding to cell surface receptors. The inhibition of viral binding was specific for the anti-idiotype antibody; viral binding was not inhibited by antibodies bound to the H-2k and Thy-1.2 antigens on the surface. Viral binding to idiotype-negative cells (P-815, EL-4) was not inhibited by the anti-idiotypic antibody, although these cells are susceptible to type 3 reovirus infection. These data suggest that there are at least two structural classes of type 3 reovirus receptors on murine cells. It is probable that anti-idiotypic antibodies of this type will be useful in studying the structure and regulation of viral receptors on cell surfaces and for the purification of these receptors, and may provide a way to block viral infection.

Animals↗
PubMed1982

Genetic and molecular mechanisms of viral pathogenesis: implications for prevention and treatment.

The pathogenesis of infection of mice by the mammalian reoviruses involves several discrete steps. Each of the three viral outer capsid proteins has a highly distinct and specialized role: one protein (sigma 1) binds to cell surface receptors; a second protein (mu 1C) determines the capacity for viral growth at mucosal surfaces; and the third protein (sigma 3) is responsible for inhibiting cell macromolecular synthesis. A detailed picture of the molecular basis of reovirus virulence and attention is now emerging.

Animals↗
PubMed1982

Identification of a hemagglutinin-specific idiotype associated with reovirus recognition shared by lymphoid and neural cells.

A xenogeneic antiserum raised to antireovirus immunoglobulin was used to define an idiotypic determinant present on antibodies to reovirus type 3 hemagglutinin. The same idiotype was identified on nonimmune lymphoid cells and on neuronal cells that specifically bind the hemagglutinin of type 3 reovirus. This idiotypic determinant, called Id3, is shared by (a) a monoclonal antibody to the neutralization site of hemagglutinin from type 3 reovirus; (b) BALB/c serum antibodies to the hemagglutinin of reovirus type 3; (c) R1.1, a murine thymoma cell line that binds reovirus type 3; (d) primary cultures of murine neuronal cells. The presence of an idiotype shared by antihemagglutinin antibodies and by structures on nonlymphoid cells suggests a general relationship between disparate receptors that recognize a common determinant. Furthermore, this suggests a novel approach for the study of viral receptor interactions and for analysis of mechanisms of autoimmune responses.

Animals↗
PubMed1982

Role of the S4 gene in the establishment of persistent reovirus infection in L cells.

We have studied the viral genes playing a role in persistence of reovirus in L cells. We established persistent infections by coinfecting L cells using wild-type reovirus type 2 and defective reovirus type 3. An analysis of the genomic double-stranded RNA pattern of the virus population selected during persistent infection revealed that the S4 gene was derived from the defective parent in three independently established lines, suggesting that the S4 gene plays an essential role in the establishment of persistent infection. The selection of recombinant viruses containing the S4 gene derived from the defective virus was specific for persistent infection, since it was not selected in lytic infections. Following the establishment of persistent infection, mutations in the S1 gene appeared in two of three cell lines. Thus mutations in the S4 gene play a critical role in the establishment of persistent infection, while mutations in the S1 gene play a role in the maintenance of the persistent infection.

Animals↗
PubMed1982

Identification of idiotypic receptors on reovirus-specific cytolytic T cells.

Cytolytic T lymphocytes (Tc) specific for cells infected with reovirus type 3 were shown to lyse an uninfected B-cell hybridoma line (designated 87.92.6). This hybridoma expresses and secretes an anti-idiotypic antibody that reacts with a monoclonal antibody (termed G-5). G-5 recognizes a domain on the hemagglutinin of the reovirus that is relevant to virus tropism. The Tc cell response was H-2 restricted and could be inhibited by G-5. As shown by limiting dilution analysis, identical clones lysed reovirus-infected and anti-idiotype-bearing target cells. Tc cells induced by a variant of wild-type reovirus type 3 (immunologically selected by resistance to neutralization by G-5) were unable to recognize the anti-idiotype-bearing cells although they lysed reovirus type 3-infected tumor cells. We conclude that Tc cells must bear determinants that bind anti-idiotype molecules and, furthermore, that B cells and cytolytic T cells can share these idiotypic determinants.

Animals↗
PubMed1982

Molecular basis of reovirus neurovirulence: role of the M2 gene in avirulence.

A number of field isolates of reovirus 3 were examined to determine their relative neurovirulence after intracerebral inoculation. One isolate was found that had decreased neurovirulence. This "avirulent" strain showed the typical type 3 neural tropism but grew significantly less well in brain tissue than T3 (Dearing) and the other type 3 reoviruses. The avirulent virus was not temperature-sensitive, and its yield in mouse L cells in vitro was similar to that of the laboratory strains. To determine the reason that this clone was avirulent, we isolated a series of reassortant progeny clones from crosses between the avirulent strain and T1 (Lang) and T3 (Dearing). Using these reassortants, we showed that avirulence was a property of the M 2 gene segment. The M2 segment was also responsible for conferring greater sensitivity to chymotryptic digestion on the avirulent strain, compared to more virulent strains. Prior studies have determined that the localization of virus in different cell types in the brain (tropism) is a property of the viral hemagglutinin, the product of the S1 RNA genome segment. Our studies thus indicate that the basis for relative neurovirulence does not reside in the viral hemagglutinin and clearly illustrate the multigenic nature of neurovirulence.

Animals↗
PubMed1982

Genetic studies on the mechanism of chemical and physical inactivation of reovirus.

The three serotypes of reovirus differ markedly in their response to a variety of chemical inactivating agents. We used intertypic recombinants containing various combinations of genes derived from the parental serotypes to study the basis of these differences. In addition to recombinants derived from types 1 and 3, and 2 and 3, we were able to isolate recombinants derived from types 1 and 2, suggesting that these two serotypes also undergo unrestricted reassortment. The intertypic recombinants behaved like one parent or the other in the presence of the inactivating agents and allowed us to determine the genes responsible for each difference. Recombinants derived from crosses between wild-type parental serotypes produced straightforward results, while recombinants derived from mutagenized, temperature-sensitive parents often did not. Sensitivity to 2.5 M-guanidine-HCl and pH 11 was determined by the S1 gene, sensitivity to 55 degrees C and 1% SDS was determined by the S4 gene, and sensitivity to 33% ethanol and to 1% phenol was determined by the M2 gene. Thus, relatively nonspecific chemical agents appear to have their predominant effect on specific proteins of the reovirus virion.

Animals↗
PubMed1982

Biochemical studies on the mechanism of chemical and physical inactivation of reovirus.

We have examined the effects of heat and several chemical inactivating agents on the buoyant density, particle-associated polypeptides and ultrastructure of reovirus particles. Treatment at pH 11 removed polypeptide sigma 1 from the outer capsid of reovirus type 2 but not from type 1; resultant particles were unchanged in their buoyant density and morphology. Treatment of reovirus types 2 and 3 with 2.5 M-guanidine-HCl produced particles with unchanged polypeptide content but an increased buoyant density, and caused aggregation of type 3 but not type 2. Treatment with 1% SDS removed polypeptide sigma 3 from both types 1 and 2 and increased the buoyant density of the virus particles. The outer capsid of SDS-treated virions was greatly altered and often indistinct. Treatment of type 3 with either 1% phenol or 33% ethanol produced particles that had a full complement of polypeptides, were unaltered in buoyant density, but were greatly aggregated. Thus, these inactivating agents affect reovirus particles in specific and distinct ways. The differential effects of such treatments can thus be used to study the structure and function of the reovirus capsid components.

Capsid↗
PubMed1982

Activation and characterization of the reovirus transcriptase: genetic analysis.

We studied the ability of chymotrypsin to activate the transcriptases of the three serotypes of reovirus. When we used conditions that reproducibly caused the activation of type 3 transcriptase by chymotrypsin alone, type 2 transcriptase was sometimes activated, and type 1 transcriptase was never activated. Using intertypic recombinants containing various combinations of genome segments from reovirus types 3 and 1, we showed that the M2 segment determined this difference. Biochemical experiments indicated that the digestion of reovirus type 1 by chromotrypsin was blocked at an intermediate stage in uncoating. We found conditions which reproducibly activated the transcriptases of all three serotypes. This allowed us to compare the biochemical properties of the three transcriptases. Although the monovalent cation preferences, divalent cation preferences and optima, and temperature optima of type 1, 2, and 3 transcriptases were indistinguishable, the pH activity curves were reproducibly different. The largest difference was between type 2 and 3 transcriptases; the pH optimum of type 2 transcriptase was lower than the pH optimum of type 3 transcriptase. Using intertypic recombinants containing various combinations of genome segments from reovirus types 2 and 3, we demonstrated that the L1 segment specified this difference.

Cations↗
PubMed1982

Host immune response to reovirus: CTL recognize the major neutralization domain of the viral hemagglutinin.

Previous studies have demonstrated the importance of the hemagglutinin (HA) of reovirus in determining virus-specific immune responses. In this study, we have used anti-HA monoclonal antibodies and reoviruses with antigenically altered HA proteins to further analyze the CTL response against the HA. We showed that reovirus-specific CTL primarily recognize one distinct antigenic domain on the HA. These results emphasize the fine specificity of the T cell receptor that recognizes viral determinants. The CTL response is directed against the region of the HA that is also responsible for neurotropism and binding to neutralizing antibody.

Animals↗