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E Paoletti

Publications and source records attributed to E Paoletti.

At least 73 records · Page 4Linked to original sources
PubMed1994

Avipox virus-vectored Japanese encephalitis virus vaccines: use as vaccine candidates in combination with purified subunit immunogens.

An avipox virus, canarypox (ALVAC), which is naturally host-range restricted, was used to construct recombinants encoding the Japanese encephalitis virus (JEV) prM, E and NS1 genes (vCP107) and prM and E genes (vCP140). Mice immunized with these recombinant viruses produced JEV neutralizing antibodies and were protected from lethal JEV challenge. Protection was also observed in mice immunized with a subunit vaccine candidate, consisting of extracellular particles (EPs; RNA-free subviral membrane vesicles containing prM/M and E proteins) derived from HeLa cell cultures infected with a JEV-vaccinia recombinant. Mice primed with vCP107 and boosted with EPs had higher antibody levels than mice immunized twice with EPs alone, although the levels were comparable to that obtained in mice immunized twice with the recombinant virus. Mice immunized with a mixture of recombinant virus (vCP107) plus EPs had neutralizing antibody titres higher than mice immunized with the recombinant virus or EPs alone.

Animals↗
PubMed1994

Nucleotide sequence of the genes encoding the canine herpesvirus gB, gC and gD homologues.

The nucleotide sequence of the genes encoding the canine herpesvirus (CHV) gB, gC and gD homologues was determined. These genes are predicted to encode polypeptides of 879, 459 and 345 amino acids, respectively. Comparison of the predicted amino acid sequences of CHV gB, gC and gD with the homologous sequences from other herpesviruses indicates that CHV is an alphaherpesvirus, a conclusion that is consistent with the previous classification of this virus according to biological properties. Alignment of the homologous gB, gC and gD amino acid sequences indicates that most of the cysteine residues are conserved, suggesting that these glycoproteins possess similar tertiary structures. The nucleotide sequence of the open reading frame downstream from the CHV gC gene was also determined. The predicted amino acid sequence of this putative polypeptide appears to be homologous to a family of proteins encoded downstream from the gC gene in most, although not all, alphaherpesviruses.

Amino Acid Sequence↗
PubMed1994

Characterization of a cleavage mutant of the measles virus fusion protein defective in syncytium formation.

Membrane fusion caused by measles virus (MV) is a function of the fusion (F) protein. This process is essential for penetration into the host cell and subsequent initiation of the virus replicative cycle. The biological activity of the MV F protein is generated by endoproteolytic cleavage of a precursor protein (F0) into a large F1 subunit and a smaller F2 subunit held together by disulfide bonds. The cleavage site consists of a cluster of five basic amino acids (amino acids 108 to 112) within the predicted primary structure of the F protein. To investigate the role of the arginine residue at the carboxy terminus of the F2 subunit (arginine 112), site-directed mutagenesis was used to construct a cleavage mutant of the MV F protein in which this arginine residue was changed to a leucine residue. The mutated F gene, encoding four out of the five basic amino acids at the cleavage site, was inserted into the genome of vaccinia virus. The resulting recombinant virus was used to study expression of the mutant F protein in infected cells. Analysis of the Leu-112 mutant protein made in infected cells demonstrated that this single-amino-acid substitution resulted in a reduced rate of transport of the mutant protein to the cell surface, despite its efficient cleavage to yield F1 and F2 subunits. However, the electrophoretic mobilities of the Leu-112 polypeptides suggested that the protein was cleaved incorrectly. This aberrant cleavage appears to have abolished the ability of the F protein to cause syncytium formation. The data indicate that the arginine 112 residue is critical for the correct proteolytic cleavage that is required for the membrane fusion activity of the MV F protein.

Amino Acid Sequence↗
PubMed1994

Functional analysis of N-linked glycosylation mutants of the measles virus fusion protein synthesized by recombinant vaccinia virus vectors.

The role of N-linked glycosylation in the biological activity of the measles virus (MV) fusion (F) protein was analyzed by expressing glycosylation mutants with recombinant vaccinia virus vectors. There are three potential N-linked glycosylation sites located on the F2 subunit polypeptide of MV F, at asparagine residues 29, 61, and 67. Each of the three potential glycosylation sites was mutated separately as well as in combination with the other sites. Expression of mutant proteins in mammalian cells showed that all three sites are used for the addition of N-linked oligosaccharides. Cell surface expression of mutant proteins was reduced by 50% relative to the wild-type level when glycosylation at either Asn-29 or Asn-61 was abolished. Despite the similar levels of cell surface expression, the Asn-29 and Asn-61 mutant proteins had different biological activities. While the Asn-61 mutant was capable of inducing syncytium formation, the Asn-29 mutant protein did not exhibit any significant cell fusion activity. Inactivation of the Asn-67 glycosylation site also reduced cell surface transport of mutant protein but had little effect on its ability to cause cell fusion. However, when the Asn-67 mutation was combined with mutations at either of the other two sites, cleavage-dependent activation, cell surface expression, and cell fusion activity were completely abolished. Our data show that the loss of N-linked oligosaccharides markedly impaired the proteolytic cleavage, stability, and biological activity of the MV F protein. The oligosaccharide side chains in MV F are thus essential for optimum conformation of the extracellular F2 subunit that is presumed to bind cellular membranes.

Animals↗
PubMed1994

Safety and immunogenicity of recombinants based on the genetically-engineered vaccinia strain, NYVAC.

NYVAC-based recombinants expressing pertinent immunogens from equine influenza virus (EIV), pseudorabies virus (PRV), Japanese encephalitis virus (JEV) and human immunodeficiency virus (HIV) were used to evaluate the safety and immunogenicity of this vector. Administration of either NYVAC recombinants or parenteral virus to mice, horses and swine was well tolerated with no notable local or systemic reactivities. Further, despite a highly attenuated phenotype, NYVAC was found to function effectively as an immunization vehicle capable of eliciting both humoral and cell-mediated immune responses.

AIDS Vaccines↗
PubMed1994

Applications of canarypox (ALVAC) vectors in human and veterinary vaccination.

This paper presents data derived from safety and efficacy studies of ALVAC-based rabies and feline leukemia virus (FeLV) vaccine candidates in target species. Inoculation of the ALVAC-RG recombinant was well tolerated in all species including humans and very young dogs. Protection induced in dogs against rabies challenge was long-lasting and could be elicited in the face of high levels of maternally-derived neutralizing antibody. Parenteral inoculation of cats with an ALVAC-FeLV recombinant was safe and induced protection against persistent infection following oro-nasal FeLV challenge.

Adult↗
PubMed1994

Safe and effective poxvirus vectors--NYVAC and ALVAC.

This paper describes the development and in vitro/in vivo characterization of two highly attenuated recombinant poxvirus vectors. The vectors, NYVAC and ALVAC, exhibit a restricted host range and a greatly enhanced degree of attenuation following inoculation into a range of animal species by a variety of routes. The safety profile of these vectors, together with their demonstrated ability to induce both humoral and cell-mediated immune responses to extrinsic antigens, makes them ideal candidates for use as vaccines in both human and veterinary disease situations.

Animals↗
PubMed1993

Mutations in ORF D13L and other genetic loci alter the rifampicin phenotype of vaccinia virus.

Phenotypic analysis of vaccinia virus temperature-sensitive mutants identified several virus isolates that were either resistant or hypersensitive to rifampicin. Temperature-sensitive mutants 1085, 7743, 1085R, and 7743R were found to be rifampicin resistant (RifR). Each virus contained at least one mutation in ORF D13L which resulted in a change of amino acid sequence. Analysis of each point mutation by marker rescue demonstrated that mutations gly to asp at residue 80, pro to phe at 458, lys to asn at 484, and ile to val at 485 conferred the resistance phenotype, while the mutation changing ser to leu at 176 did not affect the rifampicin phenotype. Temperature-sensitive mutants C6, C17, and C43 were found to be rifampicin hypersensitive. Only C43, however, was found to contain a lesion in ORF D13L. This mutation, ser to asn at 169 was confirmed by marker rescue to confer the hypersensitive phenotype. The results presented here along with those from previous reports show that known mutations conferring rifampicin resistance cluster at the C or N-terminus of D13L while a mutation mapping at residue 169 confers rifampicin hypersensitivity. Further, the finding that the hypersensitive mutants C6 and C17 contain a wild-type D13L ORF suggests a possible role of other viral functions in the interaction of rifampicin with vaccinia virus.

Codon↗
PubMed1993

Induction of cytotoxic T lymphocytes by recombinant canarypox (ALVAC) and attenuated vaccinia (NYVAC) viruses expressing the HIV-1 envelope glycoprotein.

Successful immunization against many viruses, including retroviruses such as HIV-1, is thought to depend upon the roles of both antibody and cytotoxic T-lymphocyte responses. With safety a major concern, we developed two poxvirus recombinants expressing the envelope glycoprotein of HIV-1 IIIB. Canarypox (ALVaC), which is not known to replicate in mammalian cells, and a highly attenuated vaccinia (NYVAC) virus deleted of 18 open reading frames associated with virulence and host range were used as vectors. Upon inoculation into BALB/c mice, both the ALVAC and NYVAC recombinants were capable of inducing antibody responses to HIV gp120 and provoking remarkable levels of primary and memory Thy1.2+, CD4-, CD8+ cytotoxic T-lymphocyte responses to the hypervariable V3 loop of the HIV-1 envelope glycoprotein.

Amino Acid Sequence↗
PubMed1993

An update on the vaccinia virus genome.

This communication is intended as a single source update of the initial report (Goebel et al., 1990a,b) which described the complete DNA sequence of the vaccinia virus genome. We have integrated published information as well as unpublished data. Our understanding of the complexities of the genetic functional organization of poxviruses is increasing at a remarkable rate. While some previously unknown identities have since been elucidated, the fact that the majority of vaccinia-encoded gene products still lack assigned functions lends excitement to the immediate future of poxvirus research.

Genes, Viral↗
PubMed1993

Vaccination of pigs against pseudorabies with highly attenuated vaccinia (NYVAC) recombinant viruses.

Poxvirus recombinants, based on the highly attenuated NYVAC strain of vaccinia virus (Tartaglia et al., 1992), containing single gene inserts encoding the pseudorabies virus (PRV) gII, gIII, or gp50 glycoproteins were tested for their immunogenicity in pigs. Twenty-four pigs were randomly divided into six groups of four. Groups 1-3 were inoculated with 10(7) CCID50 of NYVAC/PRV gII, NYVAC/PRV gIII, or NYVAC/PRV gp50, respectively, while groups 4 and 5 received the NYVAC parent virus or an inactivated PRV vaccine control, respectively. Group 6 represented the sham vaccinated control group. All inoculations were given by the intramuscular route on weeks 0 and 4. The candidate vaccines were shown to be safe with no local or systemic reactions. At 4 weeks following the second inoculation, all pigs were challenged by an oronasal administration of a virulent PRV strain. Pigs were monitored before and after challenge for clinical manifestations resulting from vaccination and challenge exposure, respectively. Sera were analyzed for PRV neutralizing activity. Virological analyses after challenge included assessment of virus shedding and the development of latent PRV infections. All but one animal developed latent PRV infection following challenge exposure; however, significant protection against PRV-induced signs was afforded by vaccination with either the NYVAC/PRV gp50 or NYVAC/PRV gII recombinant viruses, as well as with the inactivated PRV vaccine. The NYVAC/PRV gp50 also reduced overall virus shedding after challenge. The extent of protection against PRV-induced clinical signs, in general, was associated with the level of pre-challenge virus neutralizing activity.

Animals↗
PubMed1993

Regression of left ventricular hypertrophy in hypertensive dialyzed uremic patients on long-term antihypertensive therapy.

There have been no studies of the possibility of reversing the left ventricular hypertrophy (LVH) of chronically hemodialyzed hypertensive uremics (HDH) with long-term antihypertensive therapy. We have measured left ventricular sizes of eight (6 male, 2 female, aged 29 to 61 years) HDH with M-mode echocardiography, before and 12, 18 and 24 months after the start of a combined antihypertensive therapy which included ACE-inhibitors, beta-blockers and calcium-antagonists. Pre-treatment values for mean blood pressure (MBP), 116.6 +/- 2.9 mm Hg, end diastolic diameter (EDD), 62.6 +/- 6.6 mm, interventricular septum (IVS), 14.2 +/- 3.0 mm, and left ventricular mass index (LVMi), 239 +/- 61 g/m2, were all significantly higher than those for nine sex- and age-matched hemodialyzed normotensive subjects (HDN) with comparable hemoglobin (Hb) levels. During the antihypertensive treatment, both the systolic and diastolic BP decreased steadily (P = 0.0001; P = 0.0003; ANOVA) and significantly by the third month (P < 0.05; P < 0.01), reaching levels comparable to those of the HDN group after 12 months. At this time the LVMi (204 +/- 67) and the IVS (13.1 +/- 2.7), although both significantly lower than baseline, were still higher than in the HDN group, while the EDD was similar. After 24 months, however, both the IVS (12.3 +/- 3.1) and the LVMi (161 +/- 65) were no longer different from those of the HDN group.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1993

Murine cytotoxic T cell response specific for human cytomegalovirus glycoprotein B (gB) induced by adenovirus and vaccinia virus recombinants expressing gB.

A murine model of the cytotoxic T lymphocyte (CTL) response to glycoprotein B (gB) of human cytomegalovirus (HCMV) was developed based on the use of adenovirus (Ad) and vaccinia virus (Vac) recombinants expressing gB. Mice of different major histocompatibility haplotypes [CBA (H-2k), BALB/k (H-2k) and BALB/c (H-2d)] infected with the Ad-gB recombinant developed an Ad-specific CTL response. However, only the H-2k mice developed a significant HCMV gB-specific CTL response, as indicated by the major histocompatibility complex class I-restricted lysis of Vac strain Copenhagen (VacC)-gB recombinant-infected target cells by H-2k mouse immune spleen cells. The VacC-gB recombinant elicited only a weak gB-specific CTL response in these mice, indicating that the observed gB-specific CTL response in mice is dependent on the expression vector used for immunization. The gB-specific cytotoxicity observed in H-2k mice was mediated by the CD8 lymphocyte subset.

Adenoviridae↗
PubMed1993

Immunogenicity of the Plasmodium falciparum serine repeat antigen (p126) expressed by vaccinia virus.

cDNA encoding the serine repeat antigen (SERA) (also called p126) of Plasmodium falciparum has been isolated from the FCR3 strain and inserted into a recombinant vaccinia virus designated vP870. Expression analysis of vP870-infected Vero cells by immunoprecipitation has demonstrated several intracellular forms of SERA and a single secreted SERA peptide. Endoglycosidase digestion of these immunoprecipitated SERA peptides indicated that the intracellular SERA peptides contain simple, high-mannose N-linked oligosaccharides and that the secreted SERA peptide contains complex N-linked oligosaccharides. Pulse-chase experiments indicate that the multiple intracellular SERA peptides in infected Vero cells represent a trafficking pathway whereby the smallest SERA peptide is converted into larger peptides by co- and posttranslational modifications, including glycosylation, and eventually secreted from the cell with complex N-linked oligosaccharides. To study the immunogenicity of vaccinia virus-expressed SERA, rabbits were immunized with vP870 and their sera were analyzed for reactivity with authentic, parasite-derived SERA protein. The anti-vP870 rabbit sera reacted with P. falciparum-infected erythrocytes by immunofluorescence analysis, recognized authentic SERA from schizonts by both immunoprecipitation and Western blot (immunoblot) analyses, and recognized proteolytically processed fragments of SERA secreted into the culture medium by Western blot analysis. These results indicate that when expressed by vaccinia virus, SERA is glycosylated and secreted from infected cells and that in immunized rabbits, vaccinia virus-expressed SERA can stimulate a humoral immune response against SERA derived from blood-stage parasites.

Animals↗
PubMed1993

Protection of cats against feline leukemia virus by vaccination with a canarypox virus recombinant, ALVAC-FL.

Two ALVAC (canarypox virus)-based recombinant viruses expressing the feline leukemia virus (FeLV) subgroup A env and gag genes were assessed for their protective efficacy in cats. Both recombinant viruses contained the entire gag gene. ALVAC-FL also expressed the entire envelope glycoprotein, while ALVAC-FL(dl IS) expressed an env-specific gene product deleted of the putative immunosuppressive region. Although only 50% of the cats vaccinated with ALVAC-FL(dl IS) were protected against persistent viremia after oronasal exposure to a homologous FeLV isolate, all cats administered ALVAC-FL resisted the challenge exposure. Significantly, protection was afforded in the absence of detectable FeLV-neutralizing antibodies. These results represent the first effective vaccination of cats against FeLV with a poxvirus-based recombinant vector and have implications that are relevant not only to FeLV vaccine development but also to developing vaccines against other retroviruses, including human immunodeficiency virus.

Animals↗
PubMed1993

Effects of a protein meal on blood amino acid profile in patients with chronic renal failure.

Arterial whole blood levels of amino acids were determined in patients with chronic renal failure and in healthy subjects before and after 270 min after the ingestion of a grilled beefsteak (4 g/kg). In patients, total nonessential amino acids increased significantly more (+46%) than in controls owing to an exaggerated rise of serine, glutamine, proline, glycine, cyst(e)ine and alanine. Total essential amino acids increased as much as in controls; however, threonine, histidine and phenylalanine showed greater increases, while tryptophan had a smaller increment. Abnormalities in amino acid levels were even more evident in the postprandial period than in the postabsorptive state owing to reduced levels of valine, leucine, tryptophan, tyrosine, aspartate and glutamate and higher levels of glutamine, proline, glycine, cyst(e)ine, threonine, histidine and phenylalanine. Moreover, after the meal, the ratios total essential amino acids/total nonessential amino acids, valine/glycine, and branched-chain amino acids/total amino acids rose but persisted to be reduced whereas tryptophan/total amino acids and tyrosine/phenylalanine ratios increased in controls, but did not change in patients. In conclusion, in chronic renal failure, protein ingestion enhances the imbalance in amino acid levels already present in the postabsorptive state. The all data indicate that in patients with chronic renal failure, the metabolism of exogenous protein is impaired and the flow of amino acids to the organs is altered during the phase of body nitrogen replenishment.

Adult↗
PubMed1993

Peripheral metabolism of branched-chain keto acids in patients with chronic renal failure.

Peripheral tissue metabolism of branched-chain amino acids (BCAA) and branched-chain keto acids (BCKA) in the postabsorptive state was evaluated in 8 patients with chronic renal failure (CRF) and 7 controls by measuring the arterial-deep forearm venous differences for BCAA and BCKA. Arterial whole blood levels of BCAA and BCKA were also measured in an additional 7 patients and 11 controls. In CRF, total BCKA levels are reduced owing to a decrease in ketoisocaproic acid (KICA) and ketoisovaleric acid (KIVA) levels, parallel to changes in BCAA levels, whereas levels of ketomethylvaleric acid (KMVA) are not different from controls. Both in normal conditions and in patients, arterial levels of individual BCAA are directly correlated with arterial levels of the corresponding BCKA. However, in CRF, the ratios of leucine to KICA and of isoleucine to KMVA are increased. A direct correlation between KICA and HCO3- levels is observed. In CRF, the release of leucine and valine as well as of KICA and KMVA from peripheral tissues is reduced, whereas KIVA is neither released nor taken up by the forearm. The lack of KICA release from peripheral tissues likely accounts for its low circulating levels. The depressed peripheral release of leucine associated with the lack of KICA release suggests an increased degradation of leucine which proceeds beyond the transamination step.

Adult↗