Russell H. Neubauer

Transforming activity and antigenicity of an Epstein-Barr-like virus from lymphoblastoid cell lines of baboons with lymphoid disease.

Two lymphoblastoid cell lines were established from baboons with lymphoid disease. Cells of these lines were positive for complement and Fc receptors but lacked sheep cell receptors, theraby indicating B-cell origin. The cells contained antigens which cross-reacted with Epstein-Barr virus (EBV), viral capsid antigen (VCA), early antigen (EA) and membrane antigen (MA). Both lines released virus with in vitro transforming activity for lymphocytes of several primate species including humans. Cells of the original lines and transformed cells showed no staining for EB nuclear antigen (EBNA). The virus was neutralized by anti-MA positive baboon and human sera. Baboon virus and EBV had different but overlapping in vitro host-cell ranges.

Comparisons of nuclear antigens of Epstein-Barr virus (EBV) and EBV-like simian viruses.

Nuclear antigens (NA) of EBV (EBNA), Herpesvirus gorilla, H. papio, H. pongo and H. pan were tested with sera of human, gorilla, chimpanzee, orangutan, gibbon and baboon origins. Both conventional anticomplement immunofluorescence (ACIF) and acid-fixed nuclear binding of antigen followed by ACIF (AFNB) procedures were used. Comparisons of antibody titres by ACIF and AFNB suggested that human sera detected the same antigenic determinants on EBNA by the two procedures but gorilla sera measured different determinants on H. gorilla NA. Asymmetric cross-reactions were found with gorilla, chimpanzee and baboon sera but individual human and orangutan sera were found which had extensive cross-reactivities. Absorption experiments with these broadly reactive sera with H. gorilla NA and comparisons of antibody titres of human sera with EBNA, H. gorilla and H. papio NA suggested the presence of an EBV-specific determinant as well as a broadly reactive determinant on EBNA.

Comparisons of Surface Markers on Herpesvirus-associated Lymphoid Cells of Nonhuman Primates and Established Human Lymphoid Cell Lines

Herpesvirus saimiri (HVS)-owl monkey lymphoid cells were found to have high levels of surface receptor for sheep erythrocytes and erythrocytes of 3 other species. These HVS-lymphoid cells lacked a receptor for modified complement. Lymphoid cells of one HVS-owl monkey line showed evidence for the presence of surface immunofluorescent staining with anti-kappa chain serum. Cells of an established HVS-marmoset lymphoid line had similar surface markers. Of 4 established human lymphoid cell lines, all lacked a receptor for sheep erythrocytes, 3 showed evidence for the presence of receptor for modified complement, and 3 showed immunofluorescent evidence for the presence on the surface of both heavy and light chain immunoglobulin. Preliminary data on an established Epstein-Barr virus (EBV)-owl monkey lymphoid cell line indicated a lack of receptor for sheep erythrocytes, presence of receptor for modified complement, and surface immunofluorescent staining with both anti-heavy and anti-light chain sera.

Cytotoxic activity of normal and Herpesvirus saimiri-transformed nonhuman primate cells

Owl monkey mononuclear cells were separated from peripheral blood by centrifugation on Ficoll gradients, removal of adherent cells, and subsequent separation on discontinuous Percoll gradients. Lymphocytes recovered from the various fractions were tested for cytotoxic reactivity immediately after isolation. Low-density cells, enriched in large granular lymphocytes (LGL), demonstrated cytotoxic activity against the human natural killer-susceptible cell lines MOLT 4 and K562. In addition, IL-2-independent T-cell lines which had been obtained by immortalization with the primate herpesvirus Herpesvirus saimiri showed cytotoxicity, even after prolonged culture in vitro, similar to that demonstrated by fresh LGL. Cytotoxic activity of these lines was regulated by IL-2 in a fashion which appeared to be independent of the growth-promoting effects of this lymphokine. These results indicate a function for IL-2 beyond its role in supporting cellular proliferation. Cytotoxic activity could also be demonstrated in culture fluids from one of these cell lines (70N2). In addition, these results indicate the usefulness of immortalized cell lines (like 70N2) as a potential source for studies of the biochemical characterization and purification of supernatants containing cytotoxic factors.

Biochemical characterization of Epstein-Barr Virus membrane antigen associated glycoproteins

Abstract The Epstein-Barr Virus membrane antigen has been associated with at least four proteins on the basis of radioimmunoprecipitation experiments. The largest of these proteins has now been purified to electrophoretic homogeneity by a combination of lentil lectin - Sepharose chromatography and electrophoresis on acrylamide gel, and the amino acid composition of this protein has been determined.

Selective stimulation and differentiation of early antigens in lymphoblastoid cell lines producing Epstein-Barr-like viruses.

Comparisons of early antigens (EA) of EBV-related lymphotropic herpes virus of Old World primates have proved difficult to accomplish because antigen levels are low in infected cells and the ratio of virus capsid antigen to EA is generally high. To overcome these difficulties, we have developed a procedure which combines antigen stimulation with inhibition of late virus functions and results in the selective stimulation of EA. Using this procedure, we have examined the EA of EBV, herpesvirus papio, h. pongo, and h. pan. The results show that the EA of these viruses are composed of both R (restricted) and D (diffuse) components and that the EA, while related, are distinguishable.

Characterization of a spontaneous undifferentiated carcinoma from an African green monkey (Cercopithecus aethiops)

SummaryAn adult male African green monkey (Cercopithecus aethiops) with an undifferentiated carcinoma, probably originating from the nasal mucosa, was received from the Akron, Ohio, zoo. Cultivation of this tumor in vitro resulted in a mixture of fibroblastic and epithelial cells which was subsequently separated using differential trypsinization. The neoplastic nature of the cultured epithelial cells was verified by their ability to transplant into athymic nude, or antithymocyte serum-treated mice, where poorly differentiated carcinomas were produced, and cultures of the tumors that arose in nude mice were morphologically similar to pretransplantation cultures. Early cultures showed a normal male karyotype characteristic of the species; however, in long-term cultures, a clearly defined, small submetacentric Y chromosome was not observed. Electron microscopic examination of tumor tissue and cultured tumor cells revealed desmosomes and the presence of cytoplasmic (keratin-type) fibrils, which tended to be organized around the nucleus. In addition to the keratin-type fibrils, the cultured tumor cells also contained a large amount of cytoplasmic inclusion material that may represent keratohyalin granules. There was no evidence of a viral association with tumor material or cultured cells. The cultures were susceptible to infection by vesicular stomatitis virus,Herpesvirus hominis type 1, andH. saimiri, but were resistant to the Epstein-Barr virus.