RESUMO
Streptomyces species belong to the phylum Actinobacteria and can only grow with oxygen as a terminal electron acceptor. Like other members of this phylum, such as corynebacteria and mycobacteria, the aerobic respiratory chain lacks a soluble cytochrome c. It is therefore implicit that direct electron transfer between the cytochrome bc1 and the cytochrome aa3 oxidase complexes occurs. The complex developmental cycle of streptomycetes manifests itself in the production of spores, which germinate in the presence of oxygen into a substrate mycelium that greatly facilitates acquisition of nutrients necessary to support their saprophytic lifestyle in soils. Due to the highly variable oxygen levels in soils, streptomycetes have developed means of surviving long periods of hypoxia or even anaerobiosis but they fail to grow under these conditions. Little to nothing is understood about how they maintain viability under conditions of oxygen limitation. It is assumed that they can utilise a number of different electron acceptors to help them maintain a membrane potential, one of which is nitrate. The model streptomycete remains Streptomyces coelicolor A3(2), and it synthesises three nonredundant respiratory nitrate reductases (Nar). These Nar enzymes are synthesised during different phases of the developmental cycle and they are functional only under oxygen-limiting (<5% oxygen in air) conditions. Nevertheless, the regulation of their synthesis does not appear to be responsive to nitrate and in the case of Nar1, it appears to be developmentally regulated. This review highlights some of the novel aspects of our current, but somewhat limited, knowledge of respiration in these fascinating bacteria.
Assuntos
Metabolismo Energético/fisiologia , Nitrato Redutase/metabolismo , Nitratos/metabolismo , Oxigênio/metabolismo , Streptomyces coelicolor/metabolismo , Aerobiose/fisiologia , Anaerobiose/fisiologia , Transporte de Elétrons , Complexo I de Transporte de Elétrons/metabolismo , NADH Desidrogenase/metabolismo , Oxirredução , Microbiologia do SoloRESUMO
Active p53 stimulates the transcription of a number of key genes, including the pro-apoptotic gene bax, as well as p21, a cell cycle regulator. In this study we constructed novel chimeric zinc finger-p53 DNA binding domain (DBD) transcription factors designed to bind to the promoters of specific p53 regulated genes. In order to selectively increase the expression of Bax, we coupled a pre-selected three-zinc finger (Zif) peptide targeted to a sequence in the bax promoter to a minimal p53 DBD. This chimeric protein could increase reporter gene transcription from a minimal bax promoter (up to 10-fold) but not from a minimal p21 promoter in p53-deficient Saos-2 cells. However, fusion proteins carrying longer p53 DBDs displayed entirely different selectivity and potency. Thus, Zif-p53 DBD chimeras containing N- and C-terminal extensions of the minimal DBD could increase transcription driven by a minimal p21 promoter up to 800-fold. These chimeras preferred the minimal p21 promoter up to 500-fold over the minimal bax promoter. Additionally, endogenous p21 message and protein levels were increased in cells expressing the p21 selective Zif-p53 DBD chimera and expression of the chimeric proteins resulted in partial cell cycle arrest. Cell fractionation experiments indicated that the Zifs enhanced nuclear localization of the Zif-p53 DBD chimera. These studies suggest that it is possible to create chimeric transcription factors able to strongly and selectively activate genes downstream of p53.
Assuntos
Proteínas de Ligação a DNA/genética , Proteínas Recombinantes de Fusão/genética , Fatores de Transcrição/genética , Transcrição Gênica , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Dedos de Zinco/genética , Animais , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/patologia , Células COS , Ciclo Celular , Proteínas de Ciclo Celular/metabolismo , Núcleo Celular , Células Cultivadas , Chlorocebus aethiops , Inibidor de Quinase Dependente de Ciclina p21 , Humanos , Rim/metabolismo , Luciferases/metabolismo , Osteossarcoma/metabolismo , Osteossarcoma/patologia , Regiões Promotoras Genéticas , Transporte Proteico , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Fatores de Transcrição/metabolismo , Ativação Transcricional , Transfecção , Proteína X Associada a bcl-2RESUMO
The tumor suppressor p53 is the most commonly mutated gene in human cancers. Active p53 is able to stimulate the transcription of a variety of genes including the pro-apoptotic gene bax, as well as p21, a cell cycle regulator. In this study we produced novel zinc finger transcription factors that would selectively increase the expression of bax, but not of other p53 targets. Reporter gene assays in p53-negative Saos-2 cells showed that the novel zinc finger proteins stimulated transcription driven by a minimal bax promoter, but not that driven by a minimal p21 promoter. Moreover, electromobility shift assays demonstrated that the novel transcription factors could bind the bax promoter sequence with high affinity and selectivity. Expression of a five zinc finger protein (5ZFAV) in COS-7 cells resulted in an increase in Bax protein levels, indicating that this novel transcription factor could act on endogenous gene expression. Expression of 5ZFAV also drastically reduced Saos-2 cell survival; this effect could be reversed by the general caspase inhibitor B-D-FMK. These data suggest that 5ZFAV is able to induce apoptosis through increased Bax expression. Further, while expression of 5ZFAV in p53-deficient Saos-2 cells reduced cell survival, there was little effect on U-2 OS cells which have wild-type p53. Thus the selective induction of the pro-apoptotic bax gene may be a valuable adjunct to cancer chemotherapy by diminishing survival of p53-deficient tumor cells.
Assuntos
Apoptose , Proteínas Proto-Oncogênicas c-bcl-2 , Proteínas Proto-Oncogênicas/genética , Fatores de Transcrição/genética , Ativação Transcricional , Animais , Sequência de Bases , Sítios de Ligação , Células COS , Caspases/fisiologia , Humanos , Regiões Promotoras Genéticas , Estrutura Terciária de Proteína , Proteínas Proto-Oncogênicas/biossíntese , Fatores de Transcrição/química , Fatores de Transcrição/metabolismo , Transfecção , Células Tumorais Cultivadas , Proteína Supressora de Tumor p53/fisiologia , Dedos de Zinco , Proteína X Associada a bcl-2RESUMO
The postgenome era offers a plethora of potential therapeutic targets. Many of these targets will be addressable using small organic molecules as drug candidates. However, certain aspects of cell function, particularly those that rely on protein-protein or protein-nucleic acid interactions, will be difficult to influence using small molecules. Thus, the possibility of using highly specific macromolecules as potential therapeutic agents is an intriguing concept. Recent developments in several areas of research have brought this possibility closer to fruition. Peptide and nucleic acid combinatorial libraries allow the generation of novel molecules having exquisite selectivity. Structural information and molecular modeling also contribute to the design of new macromolecules with therapeutic potential. Perhaps most importantly, approaches for delivering macromolecules into the cell interior have been developed and applied with considerable success. Thus, the therapeutic use of macromolecules, including oligonucleotides, peptides, and proteins, may be an idea whose time has come.
Assuntos
Genômica , Substâncias Macromoleculares , Terapêutica , Animais , Técnicas de Química Combinatória , Desenho Assistido por Computador , Desenho de Fármacos , Previsões , Humanos , Modelos Moleculares , Ácidos Nucleicos/uso terapêutico , Oligonucleotídeos/uso terapêutico , Peptídeos/uso terapêuticoRESUMO
An internal adenylyltransferase gene (glnE) fragment from Streptomyces coelicolor was amplified using heterologous PCR primers derived from consensus motifs. The sequence had significant similarity to bacterial glnE genes, and included a motif typical of the C-terminal adenylyltransferase domain of glnE. glnE from S. coelicolor lies on the Asel-C fragment of the chromosome and is localized near glnA (encoding glutamine synthetase I, GSI) and glnII (encoding GSII). To analyse the function of glnE in S. coelicolor, glnE (S. coelicolor E4) and glnA (S. coelicolor HT107) gene replacement mutants were constructed. The GSI activity of the glnE mutant was not down-regulated after an ammonium shock. However, the GSI activity of the wild-type cells decreased to 60% of the original activity. The glnA mutant is not glutamine auxotrophic, but in the gamma-glutamyltransferase assay no GSI activity was detected in unshifted and shifted HT107 cells. By snake venom phosphodiesterase treatment the GSI activity in the wild-type can be reconstituted, whereas no alteration is observed in the E4 mutant. Additionally, the loss of short-term GSI regulation in the E4 mutant was accompanied by an increased glutamine:glutamate ratio.
Assuntos
Glutamato-Amônia Ligase/metabolismo , Nitrogênio/metabolismo , Nucleotidiltransferases/genética , Nucleotidiltransferases/metabolismo , Streptomyces/enzimologia , Streptomyces/genética , Sequência de Aminoácidos , Southern Blotting , Mapeamento Cromossômico , Clonagem Molecular , DNA Bacteriano/genética , Ácido Glutâmico/metabolismo , Glutamina/metabolismo , Dados de Sequência Molecular , Nucleotidiltransferases/química , Plasmídeos/genética , Reação em Cadeia da Polimerase/métodos , Análise de Sequência de DNA , Streptomyces/crescimento & desenvolvimentoRESUMO
Permanently transfected mouse cell lines which expressed different levels of the human autoantigen La/SS-B were infected with different strains of herpes simplex virus type 1, including the strains ANG, HSZP, 17syn+ and HFEM. During infection the localization of the human La protein was followed using an anti-La MoAb, which recognized only the human La protein but did not cross-react with either the endogenous mouse La protein or any viral encoded protein. After infection La protein was transported from the nucleus to the cytoplasm. The time course of translocation was dependent on the amount of human La protein expressed in the respective cell line. Moreover, acceleration of viral replication was dependent on the level of expression of human La protein, suggesting that La protein is a cellular factor that facilitates virus replication.
Assuntos
Autoantígenos/imunologia , Herpesvirus Humano 1/fisiologia , Ribonucleoproteínas/imunologia , Replicação Viral/imunologia , Células 3T3 , Animais , Regulação da Expressão Gênica/imunologia , Humanos , Camundongos , Transfecção , Antígeno SS-BRESUMO
The vagina and medulla of the adrenal gland of mice vaginally infected with herpes simplex virus (HSV) types 1 and 2 were examined in the latent stage of infection (5 to 51 weeks post-infection). RNA in situ hybridization with HSV-1 and -2 latency-associated transcript (LAT) RNA probes resulted in positively stained neuronal cell nuclei in the uterovaginal plexus, but not in the medulla of the adrenal gland. These organs were chosen because HSV antigens can be detected not only in the vaginal epithelium, but also in neurons of the uterovaginal plexus and in the medulla of the adrenal gland at the acute stage of genital infection. To our knowledge, this is the first report describing LATs in neurons of the uterovaginal plexus in the genital tract of latently HSV-infected mice.
Assuntos
Herpes Genital/virologia , Herpes Simples/virologia , Herpesvirus Humano 1/isolamento & purificação , Herpesvirus Humano 2/isolamento & purificação , Neurônios/virologia , Animais , DNA Viral/análise , Modelos Animais de Doenças , Feminino , Herpes Genital/patologia , Herpes Simples/patologia , Herpesvirus Humano 1/genética , Herpesvirus Humano 1/fisiologia , Herpesvirus Humano 2/genética , Herpesvirus Humano 2/fisiologia , Humanos , Camundongos , Camundongos Endogâmicos BALB C , RNA Mensageiro/análise , Útero/virologia , Vagina/patologia , Vagina/virologia , Vaginite/patologia , Vaginite/virologia , Latência Viral , Vulvite/patologia , Vulvite/virologiaRESUMO
Vaginal infections of BALB/c Ann mice with herpes simplex virus type 1 (HSV-1) were studied. Mice were inoculated with virulent strains ANG path and 17 syn+ or low-virulent recombinant strains 27/III and 17-syn3 that differ from parental strains in their glycoprotein B (gB) gene sequences. When low-virulent strains were inoculated separately, no vaginitis/vulvitis was produced despite replication in the vagina. In contrast, after coinfection of mice with the two low-virulent strains, vaginitis/vulvitis was produced and virus could be recovered from the central nervous system (CNS). Two of the CNS isolates produced vaginitis/vulvitis, neuroinvasiveness and death of mice after vaginal infection. Restriction fragment analysis and sequencing were used to assess recombination events in the gB gene sequence of the CNS isolates. After mixed vaginal infection recombination between non-virulent HSV strains occurs, resulting in vaginitis/vulvitis and neuroinvasiveness. No correlation was detected between the syncytial phenotype and local vaginal virulence. Virulence of HSV is not solely dependent on gB function; it seems to be more probable that several genes act in concert to induce virulence and neuroinvasiveness.
Assuntos
Sistema Nervoso Central/virologia , Herpesvirus Humano 1/patogenicidade , Vulvovaginite/virologia , Sequência de Aminoácidos , Animais , Encefalomielite/virologia , Feminino , Herpesvirus Humano 1/genética , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Recombinação Genética , Análise de Sequência de DNARESUMO
Glycoprotein C-negative (gC-) mutants of herpes simplex virus type 1 (HSV-1) derived from strains KOS and ANGpath were used to analyse the influence of soluble heparin on the phase of adsorption/attachment of HSV-1 to cells. A dose of 200 micrograms/ml heparin given 20 mins after infection of cells with the gC- positive (gC+) strains KOS and ANGpath at 4 degrees C reduced the adsorption of infective particles to 20-30% of the controls. A weaker heparin effect was observed with gC- mutants. However, also the gC- mutants exhibited a short heparin-sensitive phase. Mutations in amino acids of gB or gD at positions 854 or 25 and 27, respectively, did not alter the attachment capacities of these HSV mutants in the presence of heparin despite their peculiar fusion properties and resistance to soluble gD. We conclude that HSV-1 strains exhibit a heparin-resistant phase of attachment, which is determined by gC. Lack of gC delays the heparin-resistant attachment phase of HSV-1 to cells.
Assuntos
Heparina/farmacologia , Herpesvirus Humano 1/efeitos dos fármacos , Proteínas do Envelope Viral/metabolismo , Animais , Linhagem Celular , Chlorocebus aethiops , Cricetinae , Resistência Microbiana a Medicamentos , Herpesvirus Humano 1/genética , Herpesvirus Humano 1/isolamento & purificação , Herpesvirus Humano 1/metabolismo , Humanos , Cinética , Mutagênese , Cloreto de Potássio/farmacologia , Temperatura , Células Vero , Proteínas do Envelope Viral/genéticaRESUMO
Apoptosis has been suggested to account for loss of CD4+ T-cells in HIV infected individuals. Aside from MHC II dependent superantigens no mediator for preferential apoptosis of CD4+ T-cell has been described. However, the expression of TNF-alpha is increased in HIV+ patients. Additionally, TNF-alpha is known as a potent inducer of apoptosis in a variety of cell types. We therefore investigated the capacity of TNF-alpha to mediate apoptosis in vitro preferentially in CD4+ T-cells from HIV+ individuals. In the presence of TNF-alpha, CD4+ T-cells from HIV+ individuals with more than 200 CD4+ T-cells/microl (classified CDC A1, A2, B1, B2) could be significantly depleted by apoptosis without a reduction of CD8+ T-cells. In cells from patients with less than 100 CD4+ T-cells/microl (classified CDC C3), TNF-alpha mediated apoptosis was not apparent due to an already immensely elevated rate of apoptosis observed in the absence of TNF-alpha. Here we demonstrate cord blood mononuclear cells as a model for apoptosis since these cells develop apoptosis at a similar rate as that of PBMC in HIV+ patients. More than 50% of TNF-alpha stimulated CD4+ cord blood T-cells were depleted within 3 days by apoptosis, whereas CD8+ T-cells, B-cells and NK-cells were not affected. In PBMC from healthy adults, a preferential loss of CD4+ T-cells mediated by TNF-alpha was not observed. A reduced production of IFN-gamma was observed in mononuclear cells from newborns and from HIV+ patients. Moreover, IFN-gamma and IL-2 could prevent TNF-alpha mediated apoptosis. Therefore, a reduced Th1-cell-function may contribute to TNF-alpha mediated apoptosis of CD4+ T-cells from these donor groups. Taken together, the data suggest that TNF-alpha probably is a mediator of the loss of CD4+ T-cells in HIV infected patients in vivo.
Assuntos
Apoptose , Linfócitos T CD4-Positivos/imunologia , Infecções por HIV/imunologia , Soronegatividade para HIV/imunologia , Soropositividade para HIV/imunologia , Fator de Necrose Tumoral alfa/farmacologia , Fator de Necrose Tumoral alfa/fisiologia , Adulto , Anticorpos Monoclonais/farmacologia , Apoptose/efeitos dos fármacos , Linfócitos B/imunologia , Relação CD4-CD8 , Linfócitos T CD4-Positivos/citologia , Células Cultivadas , Sangue Fetal/imunologia , Citometria de Fluxo , HIV-1 , Humanos , Recém-Nascido , Interferon gama/biossíntese , Interleucina-2/biossíntese , Células Matadoras Naturais/imunologia , Depleção Linfocítica , Modelos Imunológicos , Valores de Referência , Fator de Necrose Tumoral alfa/biossínteseRESUMO
Previous studies have shown that certain strains of herpes simplex viruses type 1 (HSV-1) are able to induce "fusion from without" (FFWO) which means no transcription or translation of the viral genome happens. The main determinants for FFWO in BHK cells are mutations in the C-terminal part of gB-1. But single mutations in this part of the genome are not sufficient to transfer the FFWO phenotype also to Vero cells. Here, we report that FFWO of HSV strains indeed need additional mutations in the N-terminal part of gD in order to produce the FFWO phenotype in BHK and Vero cells. By marker transfer we are able to show that loss of mutations in the N-terminal part of gD influences the ability to induce FFWO in Vero cells but not in BHK cells. We assume that a mutated gD allows the entrance of a multiple number of virus particles into the cell and enhances therefore the fusion activity of the mutated gB. Mutations in gD alone are not sufficient for fusion activity of HSV.
Assuntos
Herpesvirus Humano 1/genética , Proteínas do Envelope Viral/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Linhagem Celular , Chlorocebus aethiops , Cricetinae , DNA Viral , Genoma Viral , Células Gigantes/virologia , Herpesvirus Humano 1/fisiologia , Humanos , Dados de Sequência Molecular , Mutação , Fenótipo , Transfecção , Células VeroRESUMO
After vaginal infections of mice with neuroinvasive strains of herpes simplex virus type 1 and 2 (HSV-1, HSV-2) virus replicates in the epithelium of the vagina, in the paravaginal ganglia, in the spinal cord and finally in the brain and in the adrenal glands. However, viral antigens could be demonstrated only in the medulla of the adrenal glands but not in the cortex, as assessed by immunohistochemistry (IHC). HSV could not be isolated from liver, spleen, uterus, and ovaries. This contrasts to the intraperitoneal (i.p) route of infection with replication in different visceral organs including the adrenal gland's cortex.
Assuntos
Medula Suprarrenal/virologia , Antígenos Virais/análise , Herpesvirus Humano 1/isolamento & purificação , Herpesvirus Humano 2/isolamento & purificação , Medula Suprarrenal/patologia , Animais , Córtex Cerebral/patologia , Córtex Cerebral/virologia , Chlorocebus aethiops , Feminino , Herpesvirus Humano 1/imunologia , Herpesvirus Humano 1/fisiologia , Herpesvirus Humano 2/imunologia , Herpesvirus Humano 2/fisiologia , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Medula Espinal/patologia , Medula Espinal/virologia , Vagina/patologia , Vagina/virologia , Células Vero , Replicação ViralRESUMO
Syncytial mutations of herpes simplex virus type 1 (HSV-1) strains ANG, ANG path, HFEM, tsB5 and HSZP cause extensive cell fusion and were mapped to the cytoplasmic domain of glycoprotein B (gB), within the syn 3 locus. These strains are so far the only ones which show the phenotype 'fusion from without' (FFWO): 60 min after infection with high m.o.i., cells in a tissue culture are fused without transcription and translation of the viral genome. In this report we detected, using the recombinants 27/III and K-7, that an amino acid exchange from Ala to Val at aa position 854 of gB is the main determinant for FFWO activity of strains ANG, ANG path and recombinant K-7. The transfer of this mutation to wild-type strains KOS and 17 syn+ by co-transfection results in recombinants KOS-854Q, 17-syn3, 17-syn3a and 17-syn3b. As a selection marker we used the cyclosporin A resistance of fusion which was shown to be a unique characteristic of syn 3 locus mutants. The recombinants show the FFWO phenotype in BHK cells but not in Vero cells. FFWO was shown to be cell-type dependent by comparing the number of p.f.u. needed to induce FFWO in various cell types.
Assuntos
Aminoácidos/genética , Herpesvirus Humano 1/química , Herpesvirus Humano 1/genética , Proteínas do Envelope Viral/genética , Alanina/genética , Sequência de Aminoácidos , Animais , Fusão Celular/genética , Chlorocebus aethiops , Cricetinae , Efeito Citopatogênico Viral/genética , Ensaio de Imunoadsorção Enzimática , Herpesvirus Humano 1/fisiologia , Rim/virologia , Fenótipo , Recombinação Genética , Mapeamento por Restrição , Valina/genética , Células Vero , Proteínas do Envelope Viral/químicaRESUMO
The number of TNF-alpha and IL-1 beta producing cells was investigated during the acute replication phase of herpes simplex virus (HSV) in trigeminal ganglia after intranasal infection with strains of different virulence. The highly virulent strain WAL replicated strongly and induced many cytokine producing cells early in the ganglia. The low virulent strain HFEM replicated less, only few cytokine producing cells were detected late. The thymidine-kinase negative (TK-) virus 1301 did not replicate but produced some lymphocytic inflammation. The higher the virulence of strains of HSV-1 or -2 was, the stronger was the extent of histopathological lesions; moreover, a dissociation in time between replication and cellular reaction (granulocytic and lymphocytic) could be observed after infection with strains HFEM and TK- virus 1301. CD4 and CD8 positive cells could be detected mainly at the rim of necrotic areas, TNF-alpha and IL-1 beta producing cells, however, were scattered throughout the ganglia.
Assuntos
Herpes Simples/imunologia , Interleucina-1/biossíntese , Simplexvirus/patogenicidade , Subpopulações de Linfócitos T/imunologia , Fator de Necrose Tumoral alfa/biossíntese , Replicação Viral , Administração Intranasal , Animais , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Necrose , Neurônios/microbiologia , Neurônios/patologia , Fatores de Tempo , Gânglio Trigeminal/microbiologiaRESUMO
In former studies, we described herpes simplex virus type 1 (HSV-1) strains ANG, ANG path and HSZP as strains with special fusion activities, caused by mutations in the syn 3 locus. In this report, we describe the special penetration properties of these strains: their ability to penetrate at 4 degrees and to overcome the infection block caused by soluble glycoprotein D-1 (gD-1) in the medium. Using intertypic recombinants of HSV-1 strains ANG path and KOS, we showed that these penetration properties are the result of two mutations in amino acids 25 (Leu-Pro) and 27 (Gln-Arg) in the N-terminal part of the mature gD-1 glycoprotein.
Assuntos
Simplexvirus/fisiologia , Proteínas Virais/fisiologia , Aminoácidos/fisiologia , Animais , Sítios de Ligação , Chlorocebus aethiops , Cricetinae , Cricetulus , Feminino , Fusão de Membrana , Mutação , Proteínas Recombinantes de Fusão , Mapeamento por Restrição , Simplexvirus/genética , Temperatura , Células Vero , Ensaio de Placa Viral , Proteínas Virais/genéticaRESUMO
The influence of tumor-necrosis-factor-alpha (TNF-alpha), granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukine-1 (IL-1) and IL-3 on the in vitro reactivation frequency and replication rate of trigeminal ganglia of mice latently infected with herpes simplex virus (HSV) strain KOS was studied. It could be demonstrated that TNF-alpha and possibility GM-CSF, but not IL-1 and IL-3, enhanced the reactivation frequency and replication of HSV. Interferon alpha/beta (IFN alpha/beta) prevented reactivation and replication.
Assuntos
Herpesvirus Humano 1/efeitos dos fármacos , Gânglio Trigeminal/virologia , Fator de Necrose Tumoral alfa/farmacologia , Ativação Viral/efeitos dos fármacos , Animais , Chlorocebus aethiops , Fator Estimulador de Colônias de Granulócitos e Macrófagos/farmacologia , Herpesvirus Humano 1/fisiologia , Humanos , Interferon-alfa/farmacologia , Interferon beta/farmacologia , Interleucina-1/farmacologia , Interleucina-3/farmacologia , Camundongos , Camundongos Endogâmicos BALB C , Células Vero , Latência Viral/efeitos dos fármacos , Replicação ViralRESUMO
Primary infections with herpes simplex virus type 2 (HSV-2) suppress the antibody response to secondary HSV-1 and -2 infections in the BALB/c mouse. In contrast, a challenge by the i.p. route using a vaccinia virus-HSV-1 glycoprotein B (VV gB1) recombinant induces a significant enhancement of the antibody response. This booster reaction is also observed if a challenge with a formalin-inactivated HSV-1 vaccine is performed. Although no or low humoral and vaginal antibodies are detectable after a single i.p. infection with the VV gB1 recombinant or the HSV-1 vaccine, protection against vaginal challenge with HSV-2 is induced. This points to the important role of cellular immunity for vaginal infections.
Assuntos
Herpes Genital/imunologia , Herpesvirus Humano 1/imunologia , Herpesvirus Humano 2/imunologia , Tolerância Imunológica , Proteínas do Envelope Viral/imunologia , Vacinas Virais/imunologia , Animais , Anticorpos Antivirais/imunologia , Feminino , Herpes Genital/prevenção & controle , Herpesvirus Humano 1/genética , Camundongos , Camundongos Endogâmicos BALB C , Proteínas Recombinantes de Fusão/imunologia , Vacinas de Produtos Inativados , Vaccinia virus/genética , Vagina/virologia , Vaginite/imunologia , Vaginite/prevenção & controle , Proteínas do Envelope Viral/genética , Replicação ViralRESUMO
Addition of heparin-Na+ as well as related substances of high and intermediate MW (Arteparon and polyanion SP54) 3 h after infection inhibit fusion from within (FFWI) induced by HSV strains with mutations in the syn 3 locus only. The concentration of heparin-Na+ required to inhibit FFWI is 10-fold higher (1 mg/ml) than that needed to inhibit adsorption. Instead of fusion, cell rounding is observed. The effect is readily reversible. A low MW heparin disaccharide is ineffective. Neomycin, at a concentration of 8 mM, inhibits FFWI induced by all HSV-1 but not HSV-2 strains, whereas adsorption is inhibited at 3 mM. We conclude from our observations that cell-cell fusion (FFWI) induced by syn 3 locus mutants of HSV-1 depends on a multistep mechanism. One may be constituted by pre-existing cell-cell connections or microfusions leading to cell rounding, whereas another may be active using newly appearing cell bridges during FFWI; also the three-dimensional structure of the cell membrane may be of importance. Moreover, the molecular mechanisms of FFWI induced by mutations in the syn 3 locus compared to the other 5 syn loci should be different.