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B Chesebro

Publications and source records attributed to B Chesebro.

At least 109 records · Page 6Linked to original sources
PubMed1988

Genetic control of T cell responsiveness to the Friend murine leukemia virus envelope antigen. Identification of class II loci of the H-2 as immune response genes.

T cells primed specifically for the envelope glycoprotein of Friend murine leukemia helper virus (F-MuLV) were prepared by immunizing mice with a recombinant vaccinia virus that expressed the entire env gene of F-MuLV. Significant proliferative responses of F-MuLV envelope-specific, H-2a/b T cells were observed when the T cells were stimulated with antigen-pulsed peritoneal exudate cells (PEC) having the b allele at the K, A beta, A alpha, and E beta loci of the H-2. On the other hand, PEC having only the kappa allele at these loci did not induce the envelope-specific T cell proliferation, even when the PEC had the b allele at the E alpha, S, or D loci. F-MuLV envelope-specific proliferation of H-2a/b T cells under the stimulation of antigen-pulsed, H-2a/b PEC was specifically blocked with anti-I-Ab and anti-I-Ek mAbs but not with anti-Kb, anti-Kk, or anti-I-Ak mAbs. Moreover, (B10.MBR x A/WySn)F1 mice that have the b allele only at the K locus but not in I-A subregion were nonresponders to the envelope glycoprotein, and the bm12 mutation at the A beta locus completely abolished the T cell responsiveness to this antigen. These results indicate that proliferative T cells recognize a limited number of epitopes on F-MuLV envelope protein in the context of I-Ab, hybrid I-Ak/b, and/or hybrid I-Ek/b class II MHC molecules but fail to recognize the same envelope protein in the context of I-Ak or I-Ek molecules. This influence of the H-2I region on T cell recognition of the envelope glycoprotein appeared to control in vivo induction of protective immunity against Friend virus complex after immunization with the vaccinia-F-MuLV env vaccine. Thus, these results provide, for the first time, direct evidence for Ir gene-controlled responder/nonresponder phenotypes influencing the immune response to a pathogenic virus of mice.

Alleles↗
PubMed1988

In vitro expression of cloned PrP cDNA derived from scrapie-infected mouse brain: lack of transmission of scrapie infectivity.

A cDNA for the prion protein (PrP) derived from scrapie-infected mouse brain was expressed in C127 mouse cells in vitro under the control of the mouse metallothionein promoter. PrP synthesis was detected by immunoprecipitation using a rabbit antibody specific for a 15 amino acid PrP peptide. Homogenates of cells expressing the cloned PrP cDNA inoculated into weanling mice failed to induce clinical scrapie during 190 days of observation. We conclude that either PrP is not the transmissible agent of scrapie or the PrP is not processed appropriately in this cell system to create the infectious agent.

Animals↗
PubMed1988

In vitro expression in eukaryotic cells of a prion protein gene cloned from scrapie-infected mouse brain.

It has been proposed that the causative agent of scrapie represents a class of infectious particle that is devoid of nucleic acid and that an altered form of the endogenous prion protein (PrP) is the agent. However, it has been difficult to exclude the possibility that PrP purified from scrapie tissues might be contaminated with a more conventional viral agent. To obtain PrP uncontaminated by scrapie-infected tissues, PrP cDNA cloned from a scrapie-infected mouse brain was expressed in mouse C127 cells in vitro. mRNA and protein encoded by the cloned PrP gene were identified. The expressed PrP polypeptides appeared to be glycosylated and were released from the cell surface into the medium. Homogenates of the cells expressing the cloned PrP gene were inoculated into susceptible mice but failed to induce clinical signs of scrapie. Thus, either PrP is not the transmissible agent of scrapie or the expressed PrP requires additional modification to be infectious.

Animals↗
PubMed1988

Detection of prion protein mRNA in normal and scrapie-infected tissues and cell lines.

Prion protein (PrP) forms the fibrils or prion rods isolated from scrapie-infected brain and has been proposed as the major component of the infectious agent of this slowly progressive spongiform encephalopathy. In previous Northern blot analyses PrP-specific mRNAs have been found in both normal and scrapie-infected brains but not in spleen, an organ which harbours large titres of infectivity. In the present study, mouse PrP DNA was used to probe for PrP mRNA in assorted tissues and cells. A reexamination of mouse and hamster spleens revealed that they contained low levels of PrP mRNA (approx. 0.8% of that in brain mRNA). No consistent differences were observed between normal and scrapie-infected tissues. Also positive for PrP mRNA under stringent hybridization conditions were mouse epithelial, neuroblastoma, erythroid, B-lymphocytic and embryo fibroblast tissue culture cell lines, a hamster ovary cell line, a rat glioma cell line, and human T lymphocytic and neuroblastoma cell lines. In contrast, no PrP mRNA was detected in two mouse myeloid cell lines and one T cell lymphoma. These results provide evidence that PrP is a protein common to numerous, but not all, cell types besides those of the brain.

Animals↗
PubMed1988

Development of a sensitive quantitative focal assay for human immunodeficiency virus infectivity.

Accurate and sensitive quantitation of infectious human immunodeficiency virus (HIV) has been difficult to achieve. In this report, a quantitative focal immunoassay (FIA) for HIV was developed using human HeLa cells rendered susceptible to HIV infection by introduction of the CD4 gene via a retrovirus vector. Infected cells were identified by using human anti-HIV antibodies or mouse monoclonal antibodies specific for HIV together with secondary fluorescein- or peroxidase-conjugated antibody specific for mouse or human immunoglobulins. The assay identified cells infected with either wild-type or culture-adapted HIV isolates and was capable of detecting 1 positive cell in 10(6) cells. The FIA was also effective at detecting cell-free HIV, and in contrast to assays using A3.01, CEM, and other human leukemia cells, the FIA detected most wild-type HIV isolates. HIV neutralization could be determined by using the FIA, and two monoclonal antibodies reactive with HIV gp120 were found to neutralize only the LAV-IIIB strain of HIV. These monoclonal antibodies, as well as antibodies in serum samples from patients with acquired immune deficiency syndrome, were able to inhibit the spread of HIV infection in human lymphocyte suspension cultures but not in CD4-positive HeLa cells growing attached to plastic dishes.

Antibodies, Monoclonal↗
PubMed1988

Analyses of frequency of infection, specific infectivity, and prion protein biosynthesis in scrapie-infected neuroblastoma cell clones.

Scrapie, a spongiform encephalopathy of sheep and goats, is caused by a poorly understood transmissible agent in which no nucleic acid has been conclusively identified. Biochemical characterization of agent derived from animal tissues has not been precise because of the tenacious association of the agent with tissue components. As an approach toward obtaining homogeneous preparations of agent generated in vitro, we cloned scrapie-infected neuroblastoma cells. By frequency analysis, nearly every cell in expanded cultures contained scrapie agent. We also analyzed cell-dose infectivity relationships and developed a standard curve which allowed various cultures to be compared. Since a proteinase K (PK)-resistant form of a protein designated prion protein (PrP) has been found in partially purified preparations of scrapie agent from infected animal spleens and brains, we sought to identify this protein in cell cultures. No PK-resistant PrP was found in infected or uninfected cultures, although the PK-sensitive PrP was readily detected. These results suggested that PK-resistant PrP may not be an essential component of the infectious scrapie agent.

Animals↗
PubMed1987

Characterization of scrapie infection in mouse neuroblastoma cells.

A mouse neuroblastoma cell line was successfully infected with scrapie agent. Agent derived from infected mouse brain or spleen infected cultures. However, agent from infected hamsters did not infect mouse cell cultures, suggesting that species specificity influenced the infection process in vitro. Positive cultures supported scrapie replication for as many as 47 passages in vitro. Agent was shown to be cell-associated and between 631 and 7943 unselected culture cells constituted 1 mouse LD50. However, fluctuation analysis indicated that only one of 144 cells in unselected cultures was actually infected. Thus, agent was confined to a small percentage of cells and only 4.4 to 55.1 positive cells were needed to confer a mouse LD50.

Animals↗
PubMed1987

Role of the host immune response in selection of equine infectious anemia virus variants.

Equine infectious anemia virus was isolated from peripheral blood leukocytes collected during two early febrile cycles of an experimentally infected horse. RNase T1-resistant oligonucleotide fingerprint analyses indicated that the nucleotide sequences of the isolates differed by approximately 0.25% and that the differences appeared randomly distributed throughout the genome. Serum collected in the interval between virus isolations was able to distinguish the isolates by membrane immunofluorescence on live cells. However, no neutralizing antibody was detected in the interval between virus isolations. In fact, multiple clinical cycles occurred before the development of a neutralizing antibody response, indicating that viral neutralization might not be the mechanism for selection of antigenic variants. The ability of early immune sera to recognize variant specific antigens on the surface of infected cells suggested that immune selection occurs through recognition and elimination of certain virus-infected cells. Alternately, the random distribution of the genomic differences observed between the two isolates may indicate that equine infectious anemia virus variants emerge as a result of nonimmunological selection processes.

Animals↗
PubMed1986

Hemolytic anemia and erythroleukemia, two distinct pathogenic effects of Friend MuLV: mapping of the effects to different regions of the viral genome.

Two different pathogenic effects of the Friend ecotropic murine leukemia virus (F-MuLV) were distinguished by serial examinations of hematocrits and reticulocyte counts of IRW mice inoculated as newborns. F-MuLV induced hemolytic anemia with increased levels of erythropoiesis, which was detectable as early as 13 days of age, whereas blocked erythroid differentiation, associated with erythroleukemia, was apparent only after 30 days of age. Using strains of Friend-MuLV with different virulences, we constructed recombinant viruses that allowed us to map the hemolytic effect and the ability to induce rapid erythroleukemia to different regions of the viral genome. Moreover, the ability of the virus to induce rapid erythroleukemia appeared to be independent of the presence of severe early hemolytic anemia.

Anemia, Hemolytic↗
PubMed1986

T-lymphocyte priming and protection against Friend leukemia by vaccinia-retrovirus env gene recombinant.

The current prevalence of the acquired immune deficiency syndrome in humans has provoked renewed interest in methods of protective immunization against retrovirus-induced diseases. In this study, a vaccinia-retrovirus recombinant vector was constructed to study mechanisms of immune protection against Friend virus leukemia in mice. The envelope (env) gene from Friend murine leukemia virus (F-MuLV) was inserted into the genome of a vaccinia virus expression vector. Infected cells synthesized gp85, the glycosylated primary product of the env gene. Processing to gp70 and p15E, and cell surface localization, were similar to that occurring in cells infected with F-MuLV. Mice inoculated with live recombinant vaccinia virus had an envelope-specific T-cell proliferative response and, after challenge with Friend virus complex, developed neutralizing antibody and cytotoxic T cells (CTL) and were protected against leukemia. In contrast, unimmunized and control groups developed a delayed neutralizing antibody response, but no detectable CTL, and succumbed to leukemia. Genes of the major histocompatibility complex influenced protection induced by the vaccinia recombinant but not that induced by attenuated N-tropic Friend virus.

Animals↗
PubMed1986

Influence of the murine MHC (H-2) on Friend leukemia virus-induced immunosuppression.

Friend murine leukemia virus complex (FV)-induced immunosuppression was studied by assaying splenic anti-SRBC PFC responses and plasma antibody titers in mice at various times after FV inoculation. Genes located within the H-2 complex were found to influence resistance to FV-induced immunosuppression. Near normal responses were observed in mice having the H-2a/b or H-2b/b genotype, whereas mice having the H-2a/a genotype were suppressed. This H-2 effect was observed not only in mice having heterozygous C57BL/10 X A background genes, including Rfv-3r/s, but also was apparent in mice having homozygous A-strain background genes, including Rfv-3s/s. Therefore, the Rfv-3 gene did not appear to convey resistance to FV-induced immunosuppression. The suppression in susceptible H-2a/a mice was characterized by a partial suppression of the IgM response and a profound suppression of both the primary and secondary IgG responses. Neither splenomegaly nor viremia alone appeared to be sufficient for the induction or maintenance of the immunosuppression. The mechanism of suppression was unclear, but both B lymphocyte and T lymphocyte functions appeared to be altered.

Animals↗
PubMed1986

Molecular cloning and complete sequence of prion protein cDNA from mouse brain infected with the scrapie agent.

The prion protein (PrP) is a scrapie-associated fibril protein that accumulates in the brains of hamsters and mice infected with the scrapie agent, and also in the brains of persons affected with kuru or Creutzfeldt-Jakob disease. It has been previously proposed that PrP could be either the primary transmissible agent of scrapie or a secondary component involved in the pathogenesis of scrapie. At present, the second possibility seems more likely, for the PrP-specific mRNA is present in both infected and uninfected brains. We have isolated and sequenced the complete PrP-specific cDNA from mRNA isolated from infected mouse brains. Comparison of the mouse PrP with the hamster PrP reveals a high homology in the amino acid sequence and the presence of a conserved octapeptide repeated four times, whose function is unknown at present. Structural features are discussed and compared with other proteins. Except for its homology with the hamster PrP, mouse PrP has no significant homology to any known protein sequence, including neurofilaments, neuropeptides, and amyloid proteins of Alzheimer disease. Some features of the PrP, however, are similar to structures found in aggregating proteins, such as the wheat glutenin, keratin, and collagen.

Amino Acid Sequence↗
PubMed1986

Monoclonal antibodies against Aleutian disease virus distinguish virus strains and differentiate sites of virus replication from sites of viral antigen sequestration.

Monoclonal antibodies (mAbs) were used to study antigenic differences among strains of Aleutian disease virus (ADV) and to characterize viral proteins in vitro and in vivo. A number of ADV field strains could be discriminated, and highly virulent Utah I ADV was clearly delineated from the tissue culture-adapted avirulent ADV-G strain. This specificity could be demonstrated by indirect immunofluorescence against infected cultures of Crandell feline kidney cells or against tissues of Utah I ADV-infected mink. Viral antigens were demonstrated in both the nuclei and the cytoplasm of infected tissue culture cells. However, in mink mesenteric lymph node, spleen, and liver, viral antigen was observed only in the cytoplasm. Absence of nuclear fluorescence suggested that the detected antigen represented phagocytized viral antigens rather than replicating virus. This conclusion was supported by the finding that mAbs reactive only against low-molecular-weight polypeptides derived from intact viral proteins gave the same pattern of in vivo fluorescence as mAbs with broad reactivity for large or small (or both) viral polypeptides. The distribution of infected cells was the same as that described for macrophages in these tissues and suggested that cells of the reticuloendothelial system had sequestered viral antigens.

Aleutian Mink Disease↗
PubMed1986

Analysis of two strains of Friend murine leukemia viruses differing in ability to induce early splenomegaly: lack of relationship with generation of recombinant mink cell focus-forming viruses.

Friend murine leukemia helper viruses (F-MuLV) 57 and B3 were indistinguishable by genomic structural analyses with RNase T1-resistant oligonucleotide fingerprinting and by antigenic reactivity with a panel of 31 monoclonal antibodies directed against murine leukemia viruses. Nevertheless, F-MuLV 57 and B3 had strikingly different virulences. Approximately 2 months after inoculation, IRW and NFS/N mice inoculated as newborns with F-MuLV 57 had gross splenomegaly caused by erythroid proliferation. In contrast, an equivalent dose of F-MuLV B3 induced spleen or lymph node enlargement 4 to 13 months after inoculation. Although most cases of spleen enlargement in F-MuLV B3-inoculated mice were due to erythroid proliferation, lymphoid or myeloid proliferation was also frequently observed. The replication of both F-MuLV 57 and B3 was equally efficient, and both viruses generated recombinant dual-tropic mink cell focus-forming (MCF) viruses with the same kinetics and efficiency. Moreover, MCF viruses induced by F-MuLV 57 and B3 had the same antigenic patterns. Therefore, the ability of F-MuLV to induce early splenomegaly did not correlate with the generation of recombinant MCF viruses.

Animals↗
PubMed1985

Cloning of a gene whose expression is increased in scrapie and in senile plaques in human brain.

A complementary DNA library was constructed from messenger RNA's extracted from the brains of mice infected with the scrapie agent. The library was differentially screened with the objectives of finding clones that might be used as markers of infection and finding clones of genes whose increased expression might be correlated with the pathological changes common to scrapie and Alzheimer's disease. A gene was identified whose expression is increased in scrapie. The complementary DNA corresponding to this gene hybridized preferentially and focally to cells in the brains of scrapie-infected animals. The cloned DNA also hybridized to the neuritic plaques found with increased frequency in brains of patients with Alzheimer's disease.

Alzheimer Disease↗
PubMed1985

Pseudotyping of dual-tropic recombinant viruses generated by infection of mice with different ecotropic murine leukemia viruses.

Using a new focal immunofluorescence assay (FIA) with monoclonal antibodies, dual-tropic recombinant mink cell focus-inducing (MCF) retroviruses were specifically detected directly in mouse cells. With the FIA, MCF and ecotropic murine leukemia virus (MuLV) infectious centers were quantitated in noninoculated AKR/J mice and in mice inoculated with either ecotropic Friend- or Moloney-MuLV. Comparison of the titers obtained in mouse and mink cells suggested that pseudotyping of MCF viruses with ecotropic MuLV envelopes occurred at different levels in these three mouse models. Adult and newborn IRW mice inoculated with Friend-MuLV produced MCF viruses which were mostly pseudotyped with ecotropic MuLV envelopes. Newborn IRW mice inoculated with Moloney-MuLV produced MCF viruses in both spleen and thymus. Most Moloney-MCF viruses from spleens were pseudotyped with ecotropic MuLV envelopes, whereas thymic Moloney-MCF viruses were not. Noninoculated AKR/J mice spontaneously produced MCF virus at least 2 months before the onset of organ enlargement, and the majority of MCF viruses in both spleens and thymuses of these mice did not appear to be pseudotyped.

Animals↗
PubMed1985

Use of a focal immunofluorescence assay on live cells for quantitation of retroviruses: distinction of host range classes in virus mixtures and biological cloning of dual-tropic murine leukemia viruses.

A rapid and sensitive focal immunofluorescence assay (FIA) using monoclonal antibodies or heterologous antisera was employed for detection and biological cloning of viruses capable of inducing viral antigens on cell surfaces. The FIA was performed directly on a variety of live cells in tissue culture dishes and was used successfully with C-type murine leukemia viruses (MuLV) of different tropism including ecotropic, xenotropic, amphotropic, and dual-tropic recombinant mink cell focus-inducing (MCF) viruses. With the FIA, we were able to titrate and distinguish ecotropic Friend-MuLV and Friend-MCF viruses present in mixtures. Dual-tropic MCF viruses could be specifically detected directly in mouse cells by using MCF-specific monoclonal antibodies. These antibodies replaced the requirement for production of typical MCF cytopathic effect in mink cells for MCF virus detection, and also allowed efficient titration in mouse cells of MCF virions pseudotyped with ecotropic envelope proteins. Furthermore, by picking foci of fluorescent cells and using their cell-free viral progeny, MCF viruses were cloned from complex pseudotypic mixtures. This allowed the cloning of viruses present at low frequency in heterogeneous mixtures obtained from leukemic tissues.

Animals↗