RESUMO
Infection of mouse and rat cells by the murine sarcoma virus (Moloney isolate) showed two-hit kinetics for focus production in mouse cells but one-hit kinetics in rat cells. Antiserum added to cultures after infection suppressed focus formation in mouse cells but not in rat cells. These studies suggest that, in rat cells infected with murine sarcoma virus, leukemina virus is not needed for focus formation and that these foci result from proliferation of the transformed rat cell; in mouse cells, on the other hand, leukemia virus is needed as "helper," and focus formation requires spread of virus. The term "defectiveness" then, if used, should not be applied to RNA tumor viruses without qualification for the viral function studied and the cell system employed.
Assuntos
Vírus da Leucemia Murina de Moloney/patogenicidade , Animais , Linhagem Celular , Transformação Celular Neoplásica , Testes de Fixação de Complemento , Técnicas de Cultura , Efeito Citopatogênico Viral , Vírus Defeituosos/patogenicidade , Embrião de Mamíferos/citologia , Cinética , Camundongos , RatosRESUMO
Virus-transformed rat and hamster embryonic cells formed larger cell aggregates than those formed by normal counterpart cells within 1-3 days. This aggregate property correlated with growth in soft agar and tumorigenicity.
Assuntos
Agregação Celular , Transformação Celular Neoplásica , Animais , Células Cultivadas , Cricetinae , Ratos , Vírus do Sarcoma Murino , Sarcoma Experimental/patologia , Vírus 40 dos SímiosRESUMO
Various purine and pyrimidine analogs and methotrexate were tested to determine whether they induce morphologic and functional myeloid differentiation in HL-60, a human promyelocytic leukemia cell line. Functional maturity was assessed by nitro blue tetrazolium reduction assays. 3-Deazauridine caused nearly all of the cells to differentiate during 6 days of treatment. Pyrazofurin, virazole, puromycin aminonucleoside, and the tricyclic nucleoside 3-amino-1,5-dihydro-5-methyl-1-beta-D-ribofuranosyl-1,4,5,6,8-pentaazaacenaphthylene induced maturation in 44-64% of the cells, whereas 5-azacytidine, 5-bromo-2'-deoxyuridine, 5-iodo-2' deoxyuridine, thymidine, and the antimetabolite methotrexate induced maturation in 28-36% of the cells. In terms of effective concentration, the most potent inducer was methotrexate (10-8 M). The predominant cell types after treatment with all of these compounds were the metamyelocyte and banded neutrophilic granulocyte.
Assuntos
Diferenciação Celular/efeitos dos fármacos , Leucemia Mieloide Aguda/patologia , Metotrexato/farmacologia , Nucleosídeos de Purina/farmacologia , Nucleosídeos de Pirimidina/farmacologia , Linhagem Celular , Humanos , Cinética , Nitroazul de TetrazólioRESUMO
Specific tumor rejection was obtained with the use of simian virus 40 (SV40)-transformed cells from several species including man, rat, ape, sheep, and hamster. Growth of the syngeneic sarcoma mKSA in BALB/c mice was strikingly inhibited following a single immunization with as few as 10(3) intact, viable cells. Non-SV40-transformed cells did not induce tumor rejection activity nor did SV40-transformed lines induce immunity against the 3-methylcholanthrene-induced sarcoma Meth A, syngeneic with BALB/c mice. A close relationship existed between the tumor rejection antigen, the tumor-specific transplantation antigen (TSTA) located on the plasma membrane, and the intranuclear tumor antigen (T-ag). Both were associated with the DNA sequence of the early region of the SV40 genome, and TSTA activity was found in the nucleus. However, we did not observe a close parallelism between T-ag activity and TSTA. Neverthesless, the results strongly suggested that TSTA, like T-ag, was encoded by the virus.
Assuntos
Antígenos de Neoplasias/administração & dosagem , Transformação Celular Neoplásica , Rejeição de Enxerto , Antígenos de Histocompatibilidade/administração & dosagem , Sarcoma Experimental/imunologia , Vírus 40 dos Símios/imunologia , Infecções Tumorais por Vírus/imunologia , Animais , Linhagem Celular , Cricetinae , Reações Cruzadas , Feminino , Camundongos , Camundongos Endogâmicos BALB C , Transplante de Neoplasias , Ratos , Sarcoma Experimental/etiologia , Especificidade da Espécie , Transplante IsogênicoRESUMO
The ability of cell populations to survive in the aggregate form was compared to colony formation in soft agar and tumorigenicity in nude mice. Nontumorigenic human osteogenic sarcoma (HOS) cells, which formed colonies in soft agar, could not survive in the aggregate form. Tumorigenic HOS cell lines, which also formed colonies in soft agar, survived and proliferated in the aggregate form. Other cell types were tested with the same results. This approach, based on cell survival in the aggregate form, may provide an additional, reliable method for predicting the tumorigenic status of a cell population.
Assuntos
Agregação Celular , Transformação Celular Neoplásica , Animais , Divisão Celular , Linhagem Celular , Sobrevivência Celular , Meios de Cultura , Humanos , Camundongos , Camundongos Nus , Necrose , Transplante de Neoplasias , Osteossarcoma/patologia , Transplante HeterólogoRESUMO
Cultured human hematopoietic cells from several normal and leukemic sources, including those cells initiated after exposure to primate type C retroviruses were tested for their capacity to induce tumors in young athymic BALB/c (nu/nu) mice after sc inoculation. An attempt was made to correlate these results with virus expression and chromosome patterns. Progressively growing tumor formation was observed in 5 of 18 normal diploid B-lymphoblast lines from normal peripheral blood and in one of three diploid B-lymphoblast lines from leukemic donors established after infection with primate type C viruses (gibbon ape leukemia virus or simian sarcoma virus). In contrast, none of eight spontaneously transformed B-lymphoblast lines with normal diploid karyotypes formed progressively growing tumors, although one formed a tumor that remained the same size (0.5 cm) for several months. Progressive tumor formation occurred in four of seven previously established cell lines of different cell types that had abnormal karyotypes. Of the normal diploid B-lymphoblast cultures exposed to type C viruses, 12 were tested for the presence of viral RNA and structural proteins (p12, p30, gp70), and this information was correlated with tumorigenicity. Four of the six cultures expressing viral RNA or proteins were tumorigenic, whereas only one of six cultures that did not express virus information was positive. The results of this study suggest that expression of type C viral RNA and proteins by human B-lymphoblasts increases their tumorigenicity in nude mice. It is also apparent that caution must be used in attempts to correlate cell tumorigenicity and chromosome abnormalities in nude mice.
Assuntos
Transformação Celular Viral , Neoplasias Experimentais/etiologia , Retroviridae , Animais , Linfócitos B , Linhagem Celular , Células Cultivadas , Aberrações Cromossômicas , Células-Tronco Hematopoéticas/ultraestrutura , Humanos , Cariotipagem , Camundongos , Camundongos Nus , Transplante de Neoplasias , Neoplasias Experimentais/ultraestruturaRESUMO
Human cervical cancers are often associated with human papillomavirus (HPV). In HPV-positive cervical cancers, the oncoproteins E6 and E7 are consistently expressed. In this study, the effects of antisense inhibition of both proteins were examined. Phosphorothioate oligonucleotides (ODNs) AE6 and AE7 complementary to regions flanking the start codons of HPV16 E6 and E7 genes, respectively, were synthesized. These anti-HPV ODNs inhibited the growth of cervical cell lines CaSki and SiHa, which harbor HPV16 but had little effect on cells that do not. Both ODNs also affected the ability of CaSki cells to form colonies in soft agar. In nude mice, treatment with either AE6, AE7, or a mixture of both led to substantially smaller tumors. AE7 was observed to inhibit E7 synthesis. The AE6 ODN probably exerts its effect by suppressing the expression of E6 as well as E7. Cell cultures and tumors treated with AE6 showed a decrease in E7 expression. In addition, an antisense ODN targeted at the retinoblastoma gene was able to reverse some of the inhibitory effect of AE6 on CaSki cells, indicating that AE6 inhibited E7 synthesis. This study further demonstrates that anti-HPV ODNs may be useful therapeutically.
Assuntos
Regulação Viral da Expressão Gênica , Genes Virais , Papillomaviridae/genética , Proteínas Repressoras , Neoplasias do Colo do Útero/virologia , Proteínas Estruturais Virais/genética , Animais , Antineoplásicos/farmacologia , Sequência de Bases , Adesão Celular , Feminino , Inibidores do Crescimento/farmacologia , Humanos , Técnicas In Vitro , Camundongos , Camundongos Nus , Dados de Sequência Molecular , Transplante de Neoplasias , Oligonucleotídeos Antissenso/química , Oligonucleotídeos Antissenso/farmacologia , Proteínas Oncogênicas Virais/metabolismo , Proteínas E7 de Papillomavirus , RNA Mensageiro/genética , Proteína do Retinoblastoma/metabolismo , Células Tumorais Cultivadas , Neoplasias do Colo do Útero/patologiaRESUMO
Normal and transformed rat liver epithelial cell lines exhibited differences in the ability to survive in the aggregate form. Normal rat liver epithelial cells in the aggregate form underwent a rapid decline in the number of viable cells, while counterpart transformed epithelial cells exhibited an ability to survive and proliferate in the aggregate form. This survival ability was found to correlate with colony formation in soft agar and tumorigenicity in nude mice. Cell survival in the aggregate form could possibly serve as a criterion for in vitro transformation of epithelial cells derived from rat liver.
Assuntos
Transformação Celular Neoplásica , Fígado/patologia , Agregação Celular , Linhagem Celular , Sobrevivência Celular , Células Epiteliais , Epitélio/patologia , Fígado/citologiaRESUMO
Syrian hamster embryo fibroblasts transformed in vitro with benzo(a)pyrene were analyzed for the presence of type C viral components, including extra- and intracellular reverse transcriptase activity, intracellular type C hamster virus-related RNA, and cellular hamster virus group-specific antigen. No evidence could be obtained for the presence of any of these components, although they were easily detectable in hamster fibroblasts producing either B-34 virus (a hamster virus pseudotype of Harvey murine sarcoma virus which contains an excess of helper type C hamster virus) or Harvey virus itself. In addition, intracellular viral RNA could not be detected in normal hamster embryo fibroblasts, in hamster fibroblasts transformed with simian virus 40, or in newborn hamster kidney and liver. Thus the detectable expression of the indigenous hamster type C virus is not required to maintain the transformed phenotype of these cells.
Assuntos
Benzopirenos , Transformação Celular Neoplásica , Retroviridae/isolamento & purificação , Antígenos Virais , Células Cultivadas , Hibridização de Ácido Nucleico , RNA Viral/análise , DNA Polimerase Dirigida por RNA/análise , Retroviridae/imunologia , Retroviridae/metabolismoRESUMO
Sera from 50 Japanese hemophiliacs were screened for antibodies to human T-lymphotropic retrovirus types I and III (HTLV-I and -III). As a whole, antibody to HTLV-I, antibody to HTLV-III, and antibodies to HTLV-I and -III were detected in sera from 2, 17, and 6 hemophiliacs, respectively. Among them, two hemophiliacs developed acquired immunodeficiency syndrome who were positive for both antibodies to HTLV-I and -III in sera. All of the others were asymptomatic. Most of the blood products transfused into these hemophiliacs were imported from abroad, whence the source of HTLV-III infection presumably originated. However, since quite a high percentage of these antibody-positive hemophiliacs was positive for antibody to HTLV-I, even though they are native residents in HTLV-I nonendemic areas of Japan, some special factors may have participated in HTLV-I infection. These special factors should be investigated in the future.
Assuntos
Anticorpos Antivirais/análise , Hemofilia A/imunologia , Síndrome da Imunodeficiência Adquirida/imunologia , Adolescente , Adulto , Transfusão de Sangue , Criança , Pré-Escolar , Fator IX/uso terapêutico , Fator VIII/uso terapêutico , Feminino , Anticorpos Anti-HIV , Hemofilia A/sangue , Humanos , Hipersensibilidade Tardia , Imunoglobulinas/análise , Japão , Células Matadoras Naturais/imunologia , Contagem de Leucócitos , Ativação Linfocitária , Masculino , Pessoa de Meia-Idade , Infecções por Retroviridae/imunologiaRESUMO
The murine malaria parasite Plasmodium berghei contains a plastid-like extrachromosomal genome. This genome is 30.7 kb in size and is transcriptionally active as shown by RT-PCR. DNA sequence analysis of the genome reveals 69.9-95.5% homology to sequences of the 35-kb extrachromosomal circle found in the human malaria species Plasmodium falciparum. Homologous sequences include regions of genes for the ssu-rRNA, lsu-rRNA, rpo B and clusters of t-RNAs. Sequence variation between the two Plasmodium species exists in the non-coding interspacing regions. A physical map has been constructed for the P. berghei circle, indicating the EcoRI and HindIII restriction sites as well as the arrangement of the rRNA, rpo B and tRNA genes. Arrangement of these genes is similar to that found on the P. falciparum 35-kb circle. The P. berghei circular element is distinct from the mitochondrial 6-kb DNA of both the murine and the human Plasmodium species. Preliminary results indicate that the circle may be a useful target for drug therapy.
Assuntos
DNA de Protozoário/química , Plasmodium berghei/genética , Animais , Sequência de Bases , DNA Mitocondrial/química , DNA Mitocondrial/genética , DNA de Protozoário/genética , DNA de Protozoário/ultraestrutura , Variação Genética , Humanos , Camundongos , Plasmodium falciparum/genética , Plastídeos/ultraestrutura , Reação em Cadeia da Polimerase , RNA de Protozoário/biossíntese , Mapeamento por Restrição , Homologia de Sequência do Ácido NucleicoRESUMO
Action mechanisms of a newly synthesized polysaccharide, curdlan sulfate (CRDS), on human immunodeficiency virus type 1 (HIV-1) infection were investigated in vitro using syncytium formation microassay and p24 antigen capture enzyme-linked immunosorbent assay. These assays measured the titer of infectious virions and the amounts of HIV-1 core antigen p24 in soluble, intraviral, and intracellular forms. CRDS treatments were performed for 1 h at 37 degrees C. H9 cells pretreated with 0.1 to 100.0 micrograms/ml of CRDS appreciably inhibited HIV-1 infection. CRDS-treated HIV-1 virions were less able to infect H9 cells than untreated virions. The simultaneous treatment of H9 cells and HIV-1 virions with CRDS induced a significant inhibition of HIV-1 infection, resulting in the temporary disappearance of virions at the highest dose of CRDS. In contrast, CRDS treatment of newly HIV-1-infected H9 cells caused a significant decrease in the titer of infectious HIV-1 and the p24 amounts of all three forms, but no absolute elimination. Taken together, these results indicate that CRDS may block the binding of the HIV-1 envelope to the H9 cell surface, with emphasis on the high affinity of CRDS to the HIV-1 envelope.
Assuntos
Antivirais/farmacologia , Linfócitos T CD4-Positivos/microbiologia , Glucanos/farmacologia , Infecções por HIV/microbiologia , HIV-1/crescimento & desenvolvimento , Replicação Viral/efeitos dos fármacos , beta-Glucanas , Linhagem Celular , Ensaio de Imunoadsorção Enzimática , Produtos do Gene gag/análise , Proteína do Núcleo p24 do HIV , HIV-1/efeitos dos fármacos , Humanos , Proteínas do Core Viral/análise , Vírion/efeitos dos fármacosRESUMO
Short-term (1 h) treatment with a newly synthesized sulfated polysaccharide, curdlan sulfate (CRDS), showed relatively weak blocking effects on the binding of human immunodeficiency virus type 1 (HIV-1) to the surface of H9 cells. To investigate whether long-term treatment with CRDS could strengthen this effect, CRDS in various doses (0.1, 1, 10, and 100 micrograms/ml) was used in 2-week treatment periods in four separate protocols or "Procedures." SF titers and p24 antigen levels were partially suppressed during long-term CRDS treatment but returned to control levels after the treatment was terminated. In addition, no direct cytotoxicity of CRDS to H9 cells or H9/HIV-1 cells was observed in vitro in the course of continuous exposure to 100 micrograms/ml CRDS for 2 weeks. These results demonstrate the effectiveness of long-term treatment of cells infected with HIV-1 in inhibiting virus expression. The most dramatic inhibition results were obtained when the compound was present both at the time of exposure of cells to virus and during a long-term follow-up treatment. These results show that CRDS inhibits both the cell-free and cell-associated transmission of HIV-1 to host cells and interferes with early events in virus infection. In contrast, CRDS exhibits no significant virucidal activity and has little effect on already infected cells.
Assuntos
Antivirais/farmacologia , Glucanos/farmacologia , HIV-1/efeitos dos fármacos , beta-Glucanas , Antivirais/toxicidade , Sequência de Bases , Southern Blotting , Divisão Celular/efeitos dos fármacos , Linhagem Celular , DNA Viral , Glucanos/toxicidade , Humanos , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Fatores de TempoRESUMO
The Plasmodium parasite possesses two extrachromosomal genomes; the mitochondrial genetic element and the extrachromosomal plastid-like DNA. The latter has only been fully described for one culture strain of P. falciparum. In this study, a rapid procedure for amplifying plastid DNA from dried blood spots of blood infected with different malaria species was developed. PCR amplification of a 595 bp fragment within the plastid-like large subunit ribosomal-RNA (LSU-rRNA) gene was achieved using primers derived from the P. falciparum sequence. The PCR product was observed in all Plasmodium species examined. Sequence analysis of amplified products homologous to an LSU-rRNA fragment of the plastid-like extrachromosomal circle revealed extensive conservation between Plasmodium species including P. falciparum, P. vivax, P. malariae and P. berghei.