亲代L7811及体外培养小鼠白血病细胞系L7811—85...

L7811 was an ascitic form of lymphocytic leukemia induced by myleran in 615 mice. Under electron microscope, cytoplasmic and intracisternal type A virus-like particles were observed. L7811-85 was a cell line established in vitro from the parental in vivo L7811 line. Instead of type A particles, mature and immature type C murine virus-like particles could be observed budding from the plasma membrane to extracellular space. With the in vitro subpassage of L7811-85 cells, more intracellular and extracellular type C particles appeared with an increase in its tumorigenicity. When L7811-85 cells, were inoculated IP to normal 615 mice, ascites tumor developed. The type A virus particles appeared again budding into cisterna of endoplasmic reticulum but not to extracellular space. The mechanism of virus particle type transformation remains to be studied.     

Oncornavirus-like particles in squirrel monkey (Saimiri sciureus) placenta and placenta culture.

Oncornavirus-like particles similar in morphology to type D particles were observed in 1 of 2 squirrel monkey (Saimiri sciureus) placentas. Intracytoplasmic type A particles, immature virus particles, and mature viruses with eccentric or occasionally centric nucleoids were associated with placental syncytiotrophoblasts. A spike layer typical of type B viruses was not detected in viral envelopes. Onvornaviruses, identical to those previously isolated from squirrel monkey tissues and similar to Mason-Pfizer monkey virus, were seen in cultures derived from the virus-positive squirrel monkey placenta cocultivated with a mink lung culture. The major morphologic difference between the in vivo and the in vitro squirrel monkey virus was in the nucleoid position of mature virus particles.     

Type-C particles in human tissues. II. Electron microscopic study of embryonic cultures infected with a murine leukemia virus

Type-C virus particles are synthesized by cells from a number of human embryonic cultures infected with the Rauscher murine leukemia virus (RV). These particles have the characteristic morphology of the murine leukemia viruses. In some cultures, type-C particles were also observed budding into the cisternae of rough endoplasmic reticulum of a few cells. Virus particles in the vicinity of such cells were of “immature” type. In contrast, particles in the vicinity of those cells which did not synthesize intracisternal type-C particle were of the “mature” type. It is not known if the “mature” intracisternal particles have the morphology of the “mature” extracellular type-C particles.     

Electron microscopic studies of intracisternal virus particles in Soehner-Dmochowski murine sarcoma virus-induced bone tumors of New Zealand Black rats.

Soehner-Dmochowski murine sarcoma virus (Moloney)-induced bone tumors of New Zealand Black rats carry two morphologically different types of virus particles, namely, extracellular type C and intracisternal virus particles, which have thus far not been reported. These two types of virus particles have also been observed in the tissue culture cells derived from normal prostate tissues of A/Dm and BALB/c/Dm mice after inoculation of cell-free extracts of these bone tumors. The intracisternal virus particles, 90 to 120 nm in diameter, have always been found in the rough endoplasmic reticulum; they have two inner concentric layers with a relatively electron-lucent center, frequently showing cylindrical, chain-like, or multipolar budding forms. Type C virus particles produced by Soehner-Dmochowski murine sarcoma virus (Moloney)-infected prostate tissue culture cells from A/Dm and BALB/c/Dm mice belong to the murine sarcoma-murine leukemia virus group, as revealed by the fixed immunofluorescence test and by immunoelectron microscopy. The morphological and immunological relationship of intracisternal virus particles and other types of virus particles (such as type C, type H, and intracisternal type A virus particles) and intracisternal virus particles in guinea pig leukemia are defined by routine electron microscopy observations and by immunoelectron microscopy studies.     

The early appearance and subsequent distribution of murine leukaemia virus in NZB embryos.

Information concerning both spontaneous autoimmune haemolytic disease and a uniquely high expression of solely xenotropic murine leukaemia virus (MuLV) is transmitted genetically in New Zealand Black (NZB) mice. The actual production, location, and dissemination of MuLV has now been monitored in NZB embryos by electronmicroscopy, which is the most suitable method for detecting such virus very early in prenatal life i.e. in eggs before they become implanted approximately 4 1/2 days after mating. A systematic search was made for intracisternal type-A and type-C virus particles in NZB embryos from the one-cell stage to the 14th day of gestation. AKR, Balb/c, and F¹ (BALB/cx NZB) eggs were included for comparison of preimplantation phases, and five combinations of blastocysts fused in vitro as 6- to 8-celI eggs were also examined. Four variants of type-A particles were distinguished, characteristically associated with the cisternae of the endoplasmic reticulum. All replicated at some stage of preimplantation development in NZB and AKR eggs, as well as in fused AKRAKR and BALB/c?BALB/c blastocysts, and three of them were still to be found in 8- or 14-day NZB embryos. Moreover, the smallest type-A variant also appeared in one-cell unfertilized NZB eggs, indicating that its production did not depend on the stimulus of fertilization. No type-A virus was identified in normal BALB/c and F¹ (BALB/cx NZB) blastocysts, or in NZBB?LB c, CBA/H-T6CBA/H-T6, and AKRCBA/H-T6 combinants. By comparison, immature type-C virus was first seen in 5-day NZB implants. But it was not possible to decide whether implantation or a particular stage of differentiation determined its emergence, or whether it had only then reached detectable levels. Thereafter, immature C and type-A particles coexisted and replicated until at least the 14th day of gestation, but mature C virus was not evident at this age. Both ecotropic and xenotropic MuLV can be represented morphologically as type-C particles. The known lack of infectivity of the type-A particles, coupled with our evidence of their genetic transmission and of their ubiquity in NZB embryos which do not harbour ecotropic Mulv, could indicate that they, too, are xenotropic. Coincidentally, huge numbers of immature and mature type-C MuLV were found distending the uterine glands of pregnant NZB females carrying 5- or 7-day implants. But a similarly extravagant picture was seen in agematched NZB virgins. These findings are to be investigated further.     

Particles resembling C-type oncornaviruses in normal rat prostate.

A large number of particles resembling oncornaviruses were observed in the ventral lobe of a normal rat prostate. The particles were seen budding into the intercellular spaces of the epithelial lumen cells. They exhibited features morphologically similar to immature C-type particles, with an electron-lucent center and a diameter of approximately 100 nm. Neither intracellular A-type particles nor mature C-type particles were observed. This is the first report of particles resembling oncornaviruses in normal rat prostatic tissue.     

Cytoplasmic leucine naphthylamidase activity expressed in signet-ring cell carcinoma of the stomach.

Histochemical studies were performed on signet-ring cells in surgically resected gastric carcinomas from 64 patients. On the basis of periodic acid-Schiff reactivity of the intracellular mucin, the signet-ring cells were classified as type A (immature), type B (intermediate), or type C (mature). Different staining reactions to Alcian blue at pH 2.5 enabled type C cells to be subclassified as C-1 or C-2. In 63 cases (98.4%), intense leucine naphthylamidase (LNAase) activity was found histochemically in the cytoplasm of nearly all cancer cells except type C-2. Localization of LNAase activity in the cytoplasm was confirmed by conventional biochemical assay. From the present results, histochemical characteristics of signet-ring cell carcinoma are discussed.     

Induction of type C virus-related functions in normal rat embryo fibroblasts by treatment with 5-iododeoxyuridine

Secondary cultures of rat embryo fibroblasts, derived from six different strains of normal laboratory and wild rats, were treated with 5-iododeoxyuridine in an attempt to establish the presence of endogenous type C viruses in rat cells. All the cell lines tested responded with a transient appearance of RNA-dependent DNA polymerase (RDP) activity which reached a peak 3 days after the beginning of treatment. However, no viral particles or rat gs-antigen could be detected at this stage. In the cells derived from two highly inbred strains, a spontaneous second burst of RDP activity was observed after 10–12 days, and on subculturing these cells a 20- to 40-fold increase of enzyme activity could be obtained. During this peak of activity both gs-antigen and viral particles capable of incorporating labelled uridine were detected. Electron microscopic examination revealed the presence of complete or immature virions in one cell strain. The virus induced was non-infective for monolayer cultures of 10 different animal species. Attempts to rescue infective virus by complementation with murine leukemia or sarcoma viruses failed. Prolonged cultivation of treated cultures did yield cells with trasformed morphology, but no oncogenicity could be demonstrated for these cells.     

Morphologic, cytogenetic and virologic studies in vitro of a malignant lymphoma from an African child

A cell line of human lymphoblasts (AL1), derived from a malignant lymphoma involving the jaw of an African child, has been established in a medium containing 20 per cent fetal bovine serum. The cells grow in aggregates in suspension and are morphologically similar to previously described human lymphoma cell lines grown in media containing human serum. The chromosome number distribution was wide, ranging from 40 to 77. The modal number was 46 with a pseudodiploid karyotype. Electron microscopically, herpes-like virus particles were found in less than 1 per cent of the cells during the first 4 months of culture, but subsequently no cells containing virus particles have been seen. Although the medium was free of human serum which might contain antibodies to many human viruses, numerous and varied attempts to isolate viruses from the cells were unsuccessful. Following the report of interferon production by other cells lines derived from lymphomas of African children, we were able to demonstrate interferon in the supernatant fluids of AL1 cultures. The interferon was demonstrable even when virus particles could no longer be found in the cells electron microscopically. Attempts to serially pass the antiviral activity in the absence of lymphoma cells were unsuccessful. The significance of the interferon production by the lymphoma cells is not known. Cultures of AL1 cells infected with SV40 became SV40 carrier cultures but there were no morphological changes in the lymphoma cells and there was no increase in herpes-like particles electron microscopically. AL1 cells infected with herpes simplex virus developed degenerative changes, and typical immature and mature herpes particles were seen by electron microscopy in most of the cells. AL1 cultures infected with adenoviruses 7 and 12, vaccinia, reovirus 3 and cytomegalovirus showed no evidence of growth of these viruses in the lymphoma cells and no increase in numbers of herpes-like virus particles by electron microscopy.     

Maturation of dendritic cells is a prerequisite for inducing immune responses in advanced melanoma patients.

PURPOSE: We have investigated the capacity of immature and mature monocyte-derived DCs pulsed with melanoma-associated peptides (gp100 and tyrosinase) to induce a primary cytotoxic T-lymphocyte response in vivo. EXPERIMENTAL DESIGN: Advanced HLA-A2.1(+) melanoma patients were vaccinated with peptide- and keyhole limpet hemocyanin (KLH)-pulsed DCs, either immature (9 patients) or matured by monocyte-conditioned medium/tumor necrosis factor alpha/prostaglandin E(2) (10 patients). RESULTS: All patients vaccinated with mature DCs showed a pronounced proliferative T-cell and humoral response against KLH. By contrast, KLH responses were absent in most of the patients vaccinated with immature DCs. Delayed-type hypersensitivity (DTH) reactions against antigen-pulsed DCs were only observed in patients vaccinated with mature DCs and not in patients vaccinated with immature DCs. MHC-peptide tetramer staining of DTH-derived T cells revealed the presence of specific T cells recognizing the melanoma-associated peptides in 1 patient. In a second patient, DTH-derived T cells showed specific lysis of tumor cells expressing the antigens used for DC pulsing. Only patients vaccinated with mature DCs showed objective clinical responses. Interestingly, both patients with long-term progression-free survival (22 and >40 months) were both vaccinated with mature DCs and demonstrated antigen-specific T-cell reactivity of DTH-derived T cells. CONCLUSIONS: We conclude that mature DC are superior to immature DC in the induction of immunological responses in melanoma patients, which may translate into improved clinical results.     

A new glucocorticoid receptor detected in host rat liver but not in various hepatomas

Previously a new glucocorticoid receptor (Peak C), which eluted with 0.12 to 0.14 M NaCl from DEAE-cellulose column, was identified in addition to another receptor (Peak B), a classic type of glucocorticoid receptor, which eluted with 0.05 to 0.08 M NaCl. Peak C appeared after stress or injection of a high dose (20 micrograms/100 g body weight) of dexamethasone into rats. Peak C was also detected in the liver of rats bearing various tumors, but it was not found in malignant tumors (Yoshida sarcoma and Yoshida ascites hepatoma AH 130), a less malignant Yoshida ascites hepatoma (LY-5), or in minimal deviation-type hepatomas (Morris hepatomas 7316A and 7794A). The absence of Peak C in these tumors coincided with the inability of the glucocorticoid to induce tryptophan oxygenase in these tumors and in the liver of rats during early postnatal development. Peak B was consistently observed in various hepatomas and immature rat liver with capability to induce tyrosine aminotransferase. Thus Peak C appeared to be a highly differentiated type of glucocorticoid receptor mediating specific hormone actions and to be present in mature liver cells, but not in immature liver or tumor cells, even of the minimal deviation type.     

Plasma cell maturity as a predictor of prognosis in multiple myeloma

In this study, the impact of plasma cell maturity on the prognoses of multiple myeloma (MM) patients in the era of novel agents was investigated. Myeloma cell maturity was classified via immunophenotyping: myeloma cells showing mature plasma cell 1 (MPC-1)-positive and CD49e-positive cells were considered mature type; MPC-1-positive and CD49e-negative cells were considered intermediate type; and MPC-1-negative cells were considered immature type. This study included 87 newly diagnosed MM patients who were initially treated with bortezomib and/or chemotherapy. Myeloma cell maturity was a critical factor affecting overall survival (OS) in the cohort, with median OS not reached in mature-type, 50 months in intermediate-type, and 20 months in immature-type cells. Multivariate analysis showed that immature type and stage III according to the International Staging System were both independent prognostic factors affecting OS. The findings of this study demonstrate the clinical importance of myeloma cell classification according to immunophenotyping using MPC-1 and CD49e antibodies to determine patient prognosis in this era of novel therapeutic agents.     

Virus-specific markers and virus-like particles in cell lines of tumors produced by CELO virus in Syrian golden hamsters.

Cell lines were established from 2 primary hepatocellular carcinomas (HC's) and 3 sarcomas produced in Syrian golden hamsters inoculated as newborns with chicken embryo lethal orphan (CELO) virus. Cell lines from 2 sarcomas (COT, CMT) and 1 HC (CEHEP) produced CELO virus-specific T-antigen. The antigen was not detected in cells of the third sarcoma line (RCT) until they had undergone more than 34 passages in vitro. Although 5-10% of cells in the second HC line (CILT/2) contained T-antigen during early passages, it was not demonstrable after the fifth subculture. Nevertheless, cells of both HC lines possessed CELO virus tumor-specific transplantation antigen. All 5 cell lines also contained hamster type R particles, and both HC lines had type C and intracytoplasmic type A particles. The percentage of carcinoma cells producing type R particles increased during cultivation in vitro, whereas the number of cells with type A particles decreased. Treatment with dibutyryl cyclic AMP and theophylline enhanced the number of cells producing type C particles in 1 HC line and type R particles in 2 sarcoma lines.     

Detection of provirus in an HTLV-II producer CD8+ T cell line by polymerase chain reaction combined with digoxigenin-ELISA method

A human T-cell leukemia virus type II (HTLV-II) producer cell line, designated HTLV-IIA, was established by cocultivation with leukocytes from an anti-human T-cell leukemia type I (HTLV-I) antibody-positive white male intravenous drug abuser and a healthy Japanese female. The cell line was examined for viral antigens by the indirect immunofluorescence method. The cytoplasm of over 80% of the cells was brilliantly stained. Cytogenetically, the cell line has a normal female karyotype. Electron microscopy of the HTLV-IIA disclosed many C-type retrovirus particles of mature, immature and non-cored types in the extracellular spaces. The surface markers of the transformed cells are CD2+, CD3+, CD4- and CD8+. To distinguish between HTLV-I and HTLV-II infection in the cell line, a method for detection of the HTLV-II provirus was developed by combining the polymerase chain reaction method with digoxigenin-enzyme-linked immunosorbent assay method.     

Detection of Provirus in an HTLV-II Producer CD8+ T Cell Line by Polymerase Chain Reaction Combined with Digoxigenin-ELISA Method

A human T-cell leukemia virus type II (HTLV-II) producer cell line, designated HTLV-IIA, was established by cocultivation with leukocytes from an anti-human T-cell leukemia type I (HTLV-I) antibody-positive white male intravenous drug abuser and a healthy Japanese female. The cell line was examined for viral antigens by the indirect immunofluorescence method. The cytoplasm of over 80% of the cells was brilliantly stained. Cytogenetically, the cell line has a normal female karyotype. Electron microscopy of the HTLV-IIA disclosed many C-type retrovirus particles of mature, immature and non-cored types in the extracellular spaces. The surface markers of the transformed cells are CD2+, CD3+, CD4– and CD8+. To distinguish between HTLV-I and HTLV-II infection in the cell line, a method for detection of the HTLV-II provirus was developed by combining the polymerase chain reaction method with digoxigenin-enzyme-linked immunosorbent assay method.     

Activation in vitro by BUdR of a productive EB virus infection in the epithelial cells of nasopharyngeal carcinoma.

Material from a nasopharyngeal carcinoma (NCP) has been passaged in athymic (nude) mice to eliminate non-malignant infiltrating cells. The human origin and derivation from NPC malignant epithelial cells of the nude mouse tumours have been confirmed by chromosome examination, electron microscopy showing desmosomes and keratin fibrils, and postive EB virus nuclear antigen (EBNA) testing. Samples of the mouse-grown tumours were cultured and pure monolayers of epithelial cells were obtained which still expressed EBNA and contained desmosomes and keratin; these cultures grew well for about 3 weeks. Extensive electron microscope searches failed to reveal herpes virus particles. In contrast, cultures treated with BUdR showed typical immature and mature herpes virus particles in epithelial, keratin-containing cells, and immunofluorescence tests for virus capsid antigen with a battery of human sera identified this agent as EB virus. EB virus has thus, for the first time, been activated in NPC epithelial cells and shown to be capable of replication in a cell type other than a primate B-lymphocyte.     

Viral inclusions and other cytoplasmic components in a Leydig cell murine tumor: an electron microscopic study

Cytoplasmic inclusion bodies which were clearly visible under light microscopy have recently been described in estrogen-induced testicular interstitial cell tumors of mice. An ultrastructural study has revealed that the inclusions were formed of considerable numbers of type A virus particles which measured approximately 76 mμ in external diameter. Despite the abundance of intracytoplasmic viral particles, there was no sign of extracellular release or budding of mature particles from cellular membranes. Among the unusual cytoplasmic components of the tumor cells were bundles of filaments with a wavy appearance and a distinctive cross-sectional pattern. These fibrillar structures may be a cellular manifestation of the presence of virus. In discussion, the inclusions of the testicular intestitial cell tumors are compared to inclusions found in other murine tumors.     

Demonstration of type-R and type-C virus particles in hamster pancreatic adenocarcinomas

Two transplanaable solid tumors (CBP and PDP) and two continuous cultured cell lines (CBP-TC and PDP-TC) were established from nitrosamine-induced pancreatic ductal adenocarcinomas in inbred Syrian hamsters. Electron microscopic examination of both CBP and PDP solid tumors as well as CBP-TC cultured cells revealed the presence of type-R virus particles, which consisted of a 100 nm envelope containing a central 50 nm nucleoid with characteristic radiating ‘spokes’. Type-R particles were abundant in cultured CBP-TC cells, moderately frequent in CBP solid tumors, and rare in PDP solid tumors; they were not present in PDP-TC cultured cells. The CBP and PDP solid tumors and cultured PDP-TC cells exhibited type-C viral particles, having a 150 nm envelope containing a 75 nm nucleoid. Mature, immature and budding forms of Type-C viruses were present and quite prominent in PDP tumors. Type-C particles were present but rare in CBP solid tumors and in PDP-TC cells. C-particles were not observed in cultured CBP-TC cells. Examination of normal hamster pancreas, liver, duodenum, muscle and connective tissue failed to reveal evidence of type-R or type-C virus particles. It is important to recognize the presence of virus particles in hamster pancreatic carcinoma lines, since viruses could potentially affect biochemical or immunological studies on the carcinomas through the production of viral proteins that could be mistaken for tumor-associated antigens.