PubMed Health⌕ Search

Biomedical subjects

J C Ramírez

Publications and source records attributed to J C Ramírez.

At least 19 recordsLinked to original sources
PubMed2001

Towards a new generation of vaccines: the cytokine IL-12 as an adjuvant to enhance cellular immune responses to pathogens during prime-booster vaccination regimens.

A main goal of the industrialized world is the development of effective vaccines to control infectious diseases with major health and socio-economic impact. Current understanding of the immune response triggered during infection with pathogens causing malaria, hepatitis C and AIDS emphasizes the importance of cytotoxic T lymphocytes (CTLs) in combating these infections. This has led to the development of new vaccination strategies, some of which are in phase I/II clinical trials. Promising strategies of vaccination are based on highly attenuated viral vectors, such as Vaccinia virus (VV) in combination with heterologous like vectors naked DNA, referred to as priming/booster vaccination. While these immunization schedules increased the production of specific CTLs, there is a need to further expand the CD8+T cell population to control an infection. Among molecules that play a significant role in the modulation of the CTL response is the cytokine IL-12. Immunoregulation by IL-12 is of central importance in cell-mediated immunity (CMI) against those pathogens and tumors that are controlled by cell-mediated mechanisms, supported by Thl cells. The use of this cytokine in combination with highly immunogenic VV-derived vectors is a promising system for development of future vaccination schedules. In this review, we summarize recent data on the use of IL-12 in vaccination procedures, as well as undesired side-effects of the cytokine that can be overcome by accurate use of dose, route and time-window administration of IL-12 encoding vectors. Results described here indicate that VV IL-12-mediated enhancement of the specific CMI response against a model antigen HIV-1 env was time- and dose-dependent and that the antigen and the cytokine could be expresed from two different rVVs modulating the doses of the vectors and allowing for enhancement of a specific CMI response. Moreover, the use of IL-12 during DNA prime/VV boost regimens enhanced the specific anti-HIV-1 env cellular response 20 times compared to that generated after a single rVVenv inoculation. Variables such as: a) dose of the cytokine applied, b) time of its administration and c) routes of inoculation play a critical role in the final outcome of the response. The findings presented here can be extended to other antigens, suggesting that immunomodulatory cytokines can be useful in the development of the future vaccines against numerous infectious diseases and tumors.

Adjuvants, Immunologic↗
PubMed2000

Interleukin-12 (IL-12) enhancement of the cellular immune response against human immunodeficiency virus type 1 env antigen in a DNA prime/vaccinia virus boost vaccine regimen is time and dose dependent: suppressive effects of IL-12 boost are mediated by nitric oxide.

We previously demonstrated that codelivery of interleukin-12 (IL-12) with the human immunodeficiency virus type 1 (HIV-1) Env antigen from a recombinant vaccinia virus (rVV) can enhance the specific anti-Env cell-mediated immune (CMI) response. In the present study, we have investigated the effects of IL-12 in mice when it is expressed in a DNA prime/VV boost vaccine regimen. The delivery of IL-12 and Env product during priming with a DNA vector, followed by a booster with VV expressing the Env gene (rVVenv), was found to trigger the optimal CMI response compared with other immunization schedules studied. Significantly, if IL-12 is also delivered as a booster from the viral vector, an impairment of the effects of IL-12 was observed involving nitric oxide (NO), since it was overcome by specific inhibitors of inducible NO synthase. NO caused transient immunosuppression rather than impairment of viral replication. Moreover, at certain viral doses, coadministration of the NO inhibitor during the booster resulted in IL-12-mediated enhancement of the specific CD8(+) T-cell response. In addition, the dose of the IL-12-encoding plasmid (pIL-12) and the route of administration of both vectors were relevant factors for optimal CMI responses. Maximal numbers of Env-specific CD8(+) gamma interferon-secreting cells were obtained when 50 microg of pIL-12 was administered intramuscularly at priming, followed by an intravenous rVVenv boost. Our results demonstrate, in a murine model, critical parameters affecting the success of vaccination schedules based on a combination of DNA and VV vectors in conjunction with immunomodulators.

Animals↗
PubMed2000

Attenuated modified vaccinia virus Ankara can be used as an immunizing agent under conditions of preexisting immunity to the vector.

A problem associated with the use of vaccinia virus recombinants as vaccines is the existence of a large human population with preexisting immunity to the vector. Here we showed that after a booster with attenuated recombinant modified vaccinia virus Ankara (rMVA), higher humoral and cellular immune responses to foreign antigens (human immunodeficiency virus type 1 Env and beta-galactosidase) were found in mice preimmunized with rMVA than in mice primed with the virulent Western Reserve strain and boosted with rMVA. This enhancement correlated with higher levels of expression of foreign antigens after the booster.

Animals↗
PubMed2000

Biology of attenuated modified vaccinia virus Ankara recombinant vector in mice: virus fate and activation of B- and T-cell immune responses in comparison with the Western Reserve strain and advantages as a vaccine.

The modified vaccinia virus Ankara (MVA) strain is a candidate vector for vaccination against pathogens and tumors, due to safety concerns and the proven ability of recombinants based on this vector to trigger protection against pathogens in animals. In this study we addressed the fate of the MVA vector in BALB/c mice after intraperitoneal inoculation in comparison with that of the replication-competent Western Reserve (WR) strain by measuring levels of expression of the reporter luciferase gene, the capability to infect target tissues from the site of inoculation, and the length of time of virus persistence. We evaluated the extent of humoral and cellular immune responses induced against the virus antigens and a recombinant product (beta-galactosidase). We found that MVA infects the same target tissues as the WR strain; surprisingly, within 6 h postinoculation the levels of expression of antigens were higher in tissues from MVA-infected mice than in tissues from mice infected with wild-type virus but at later times postinoculation were 2 to 4 log units higher in tissues from WR-infected mice. In spite of this, antibodies and cellular immune responses to viral vector antigens were considerably lower in MVA-inoculated mice than in WR virus-inoculated mice. In contrast, the cellular immune response to a foreign antigen expressed from MVA was similar to and even higher than that triggered by the recombinant WR virus. MVA elicited a Th1 type of immune response, and the main proinflammatory cytokines induced were interleukin-6 and tumor necrosis factor alpha. Our findings have defined the biological characteristics of MVA infection in tissues and the immune parameters activated in the course of virus infection. These results are of significance with respect to optimal use of MVA as a vaccine.

3T3 Cells↗
PubMed2000

Phosphorylation status of the parvovirus minute virus of mice particle: mapping and biological relevance of the major phosphorylation sites.

The core of the VP-1 and VP-2 proteins forming the T=1 icosahedral capsid of the prototype strain of the parvovirus minute virus of mice (MVMp) share amino acids sequence and a common three-dimensional structure; however, the roles of these polypeptides in the virus infection cycle differ. To gain insights into this paradox, the nature, distribution, and biological significance of MVMp particle phosphorylation was investigated. The VP-1 and VP-2 proteins isolated from purified empty capsids and from virions containing DNA harbored phosphoserine and phosphothreonine amino acids, which in two-dimensional tryptic analysis resulted in complex patterns reproducibly composed by more than 15 unevenly phosphorylated peptides. Whereas secondary protease digestions and comigration of most weak peptides in the fingerprints revealed common phosphorylation sites in the VP-1 and VP-2 subunits assembled in capsids, the major tryptic phosphopeptides were remarkably characteristic of either polypeptide. The VP-2-specific peptide named B, containing the bulk of the (32)P label of the MVMp particle in the form of phosphoserine, was mapped to the structurally unordered N-terminal domain of this polypeptide. Mutations in any or all four serine residues present in peptide B showed that the VP-2 N-terminal domain is phosphorylated at multiple sites, even though none of them was essential for capsid assembly or virus formation. Chromatographic analysis of purified wild-type (wt) and mutant peptide B digested with a panel of specific proteases allowed us to identify the VP-2 residues Ser-2, Ser-6, and Ser-10 as the main phosphate acceptors for MVMp capsid during the natural viral infection. Phosphorylation at VP-2 N-terminal serines was not necessary for the externalization of this domain outside of the capsid shell in particles containing DNA. However, the plaque-forming capacity and plaque size of VP-2 N-terminal phosphorylation mutants were severely reduced, with the evolutionarily conserved Ser-2 determining most of the phenotypic effect. In addition, the phosphorylated amino acids were not required for infection initiation or for nuclear translocation of the expressed structural proteins, and thus a role at a late stage of MVMp life cycle is proposed. This study illustrates the complexity of posttranslational modification of icosahedral viral capsids and underscores phosphorylation as a versatile mechanism to modulate the biological functions of their protein subunits.

Animals↗
PubMed2000

A beta-stranded motif drives capsid protein oligomers of the parvovirus minute virus of mice into the nucleus for viral assembly.

The determinants of nuclear import in the VP-1 and VP-2 capsid proteins of the parvovirus minute virus of mice strain i (MVMi) synthesized in human fibroblasts were sought by genetic analysis in an infectious plasmid. Immunofluorescence of transfected cells revealed that the two proteins were involved in cooperative cytoplasmic interactions for nuclear cotransport. However, while VP-1 translocated regardless of extension of deletions and did not form capsid epitopes by itself, VP-2 seemed to require cytoplasmic folding and the overall conformation for nuclear transport. The sequence (528)KGKLTMRAKLR(538) was found necessary for nuclear uptake of VP-2, even though it was not sufficient to confer a nuclear localization capacity on a heterologous protein. In the icosahaedral MVMi capsid, this sequence forms the carboxy end of the amphipathic beta-strand I (betaI), and all its basic residues are contiguously positioned at the face that in the unassembled subunit would be exposed to solvent. Mutations in singly expressed VP-2 that either decrease the net basic charge of the exposed face (K530N-R534T), perturb the hydrophobicity of the opposite face (L531E), or distort the betaI conformation (G529P) produced cytoplasmic subviral oligomers. Particle formation by betaI mutants indicated that the basic residues clustered at one face of betaI drive VP oligomers into the nucleus preceding and uncoupled to assembly and that the nuclear environment is required for MVMi capsid formation in the infected cell. The degree of VP-1/VP-2 transport cooperativity suggests that VP trimers are the morphogenetic intermediates translocating through the nuclear pore. The results support a model in which nuclear transport signaling preserves the VP-1/VP-2 stoichiometry necessary for efficient intranuclear assembly and in which the beta-stranded VP-2 nuclear localization motif contributes to the quality control of viral morphogenesis.

Amino Acid Motifs↗
PubMed1996

Parvovirus minute virus of mice strain i multiplication and pathogenesis in the newborn mouse brain are restricted to proliferative areas and to migratory cerebellar young neurons.

Newborn BALB/c mice intranasally inoculated at birth with a lethal dose of the immunosuppressive strain of the parvovirus minute virus of mice (MVMi) developed motor disabilities and intention tremors with a high incidence by the day 6 postinfection (dpi). These neurological syndromes paralleled the synthesis of virus intermediate DNA replicative forms and yield of infectious particles in the brain, with kinetics that peaked by this time. The preferred virus replicative sites in the brain were established early in the infection (2 dpi) and at the onset of clinical symptoms (6 dpi) and were compared with major regions of cellular proliferative activity found after intraperitoneal injection of bromodeoxyuridine 24 h before encephalons were subjected to immunohistochemistry detection. At 2 dpi, viral capsid antigen was located in the laterodorsal thalamic and the pontine nuclei but not in the extensive proliferative regions of the mouse brain at this postnatal day. At 6 dpi, however, the neurotropism of the MVMi was highlighted by its ability to target the subventricular zone of the ventricles, the subependymal zone of the olfactory bulb, and the dentate gyrus of the hippocampus, which are the three main germinal centers of the cerebrum in mouse postbirth neurogenesis. Unexpectedly, in the cerebellum, the MVMi capsid antigen was confined exclusively to cells that have undergone mitosis and have migrated to the internal granular layer (IGL) and not to the proliferative external granular layer (EGL), which was stained with antiproliferative cell nuclear antigen antibody and is the main target in other parvovirus infections. This result implies temporal or differentiation coupling between MVMi cycle and neuroblast morphogenesis, since proliferative granules of the EGL should primarily be infected but must migrate in a virus carrier state into the IGL in order to express the capsid proteins. During migration, many cells undergo destruction, accounting for the marked hypocellularity specifically found in the IGL and the irregular alignment of Purkinje cell bodies, both consistent histopathological hallmarks of animals developing cerebellar symptoms. We conclude that MVMi impairs postmitotic neuronal migration occurring in the first postnatal week, when, through the natural respiratory route of infection, the virus titer peaks in the encephalon. The results illustrate the intimate connection between MVMi neuropathogenesis and mouse brain morphogenetic stage, underscoring the potential of parvoviruses as markers of host developmental programs.

Animals↗
PubMed1995

Transcriptional inhibition of the parvovirus minute virus of mice by constitutive expression of an antisense RNA targeted against the NS-1 transactivator protein.

We have assessed a genetic resistance approach based on antisense RNA to interfere with the prototype Minute Virus of Mice (MVMp), an autonomous parvovirus. MVMp is a cytolytic virus when infecting the permissive A9 mouse fibroblast cell line, and its gene expression is largely regulated at the level of transcription initiation by the nonstructural transactivator NS-1 protein, a multifunctional polypeptide also involved in viral DNA replication and cytotoxicity. An NS-1 specific antisense RNA constitutively expressed in transfected A9 clones increased several fold the proliferative viability of the cells upon high multiplicity virus infection, and cultures infected at low multiplicity reached confluence overcoming virus progression. All clones shared a common phenotype of resistance characterized by a lowered synthesis of viral DNA replicative intermediates and genomic forms, a significant reduction in the accumulation of the three viral messengers in the cytoplasmic and nuclear compartments, and a specific inhibition in viral protein synthesis. These results indicate that the constitutive antisense RNA mediates an overall repression of viral macromolecular synthesis by preventing the onset of NS-1 functions. Therefore, cytocidal parvoviruses may be hampered by engineered antisense RNA targeted against early regulators of virus growth.

Animals↗
PubMed1995

Knowledge, attitudes and practices during a community-level ivermectin distribution campaign in Guatemala.

Community acceptance and participation are essential for the success of mass ivermectin chemotherapy programmes for onchocerciasis (river blindness). To explore the local understanding of the purpose of ivermectin and willingness to continue taking the drug, we performed questionnaire surveys in four communities with hyperendemic onchocerciasis after each of three ivermectin treatment rounds. More than 100 respondents participated in each KAP survey, representing the heads of 30% of the households in each community. The respondents rarely stated that the goal of the ivermectin treatment programme was to prevent visual loss. Instead, they said they were taking the drug for their general well-being, to cure the onchocercal nodule (filaria), or to cure the microfilaria, a term newly introduced by agents of the treatment programme. The principal reason identified for refusal to take ivermectin was anxiety about drug-related adverse reactions, and there were marked differences between communities in acceptance of treatment. In one community over 50% of residents initially refused to take ivermectin, although participation rates improved somewhat after programmatic adjustments. We recommend that ivermectin distribution programmes establish surveillance activities to detect where acceptance is poor, so that timely and community-specific adjustments may be devised to improve participation.

Community Health Planning↗
PubMed1994

[Renal carcinoma with tumor embolism of the supradiaphragmatic inferior vena cava: therapeutic alternatives and review of the literature].

We report a case of a man with a left renal cancer and vena caval thrombus extending to the right atrium, that was not possible to remove because infiltration of the posterior plane was detected intraoperatively. The patient was rehospitalized for intractable haematuria. He underwent selective embolization of the left renal artery with ethanol and coil. There were no complications and the patient is well seven months after the procedure. We review the literature on the diagnostic and therapeutic options in such cases. We can conclude that it is necessary to know the exact stage of the primary cancer in order to choose the best treatment. MRI or dynamic CT should be utilized to determine the presence of infiltration of adjacent structures or distant metastasis. Radical nephrectomy and the removal of the caval thrombus afford the only possibility of survival in these patients.

Diagnosis, Differential↗
PubMed1993

Protein species of the parvovirus minute virus of mice strain MVMp: involvement of phosphorylated VP-2 subtypes in viral morphogenesis.

The pattern of induced protein species of the prototype strain of the parvovirus minute virus of mice was determined in permissive A9 mouse fibroblast cells by high-resolution two-dimensional gel electrophoresis. Identities of the viral proteins in the gels were assigned by probing two-dimensional blots with antisera raised against either purified capsids (recognizing VP-1 and VP-2) or specific coding regions of the nonstructural proteins (NS-1 and NS-2) expressed as beta-galactosidase fusion products in bacteria. All viral proteins showed posttranslational modifications, phosphate being a common substituent. The NS-1 protein migrated as a basic polypeptide in the pI range of 7.4 to 7.8 with multiple stages of modification and as a likely minor but hyperphosphorylated component in the neutral region of the gel. The NS-2 isoforms were resolved at a pI value close to 5.5 as three groups of unevenly phosphorylated polypeptides, each composed of at least two protein species. Both VP-1 and VP-2 structural polypeptides were induced as heterogeneous phosphoproteins. The major VP-2 protein could be resolved in the form of a consistent pattern of three abundant (a to c), two intermediate (d and e), and one meager (f) neutral isoelectric focusing species or subtypes. This posttranslational modification precedes and is uncoupled from viral assembly, and all of the VP-2 subtypes are packaged into empty capsids at the induced stoichiometry. However, intracellular full virions harbored additional phosphorylated subtypes (g to l) and a subtle rearrangement in the whole VP-2 composition, while mature virions purified from lysed cultures lacked these subtypes, coordinately with the emergence of six neutral VP-3 subtypes. Thus, the virion coat undergoes a chemical transition entailed by genome encapsidation, in which phosphates seem to play a major role, triggering the preferential proteolytic cleavage of the more acidic VP-2 subtypes to VP-3. Parvoviruses, with small coding capacity, may regulate some morphogenetic steps, such as assembly, genome encapsidation, and maturation, by posttranslational modifications of their structural proteins.

Amino Acid Sequence↗
PubMed1991

Knowledge, attitudes and perceptions (KAP) of onchocerciasis: a survey among residents in an endemic area in Guatemala targeted for mass chemotherapy with ivermectin.

If ivermectin distribution programs are to have maximum impact on the morbidity and transmission of human onchocerciasis there must be broad and sustained acceptance within the endemic communities. Educational activities, developed with careful consideration of community attitudes, should promote positive treatment seeking behavior while simultaneously addressing local reservations about the control effort. To better understand the ambient knowledge, attitudes, and practices concerning onchocerciasis in the context of ivermectin use in Guatemala, we conducted a survey among 145 heads of households in five endemic communities. Given the country's long-standing nodulectomy program, it was not surprising that 100% of persons interviewed had heard of the disease 'la filaria', which they defined as a skin nodule that could cause blindness. Ninety-five percent of respondents identified surgery as the only cure for the condition. Relatively few (39%) knew that la filaria was caused by a worm, although slightly more (50%) knew that the condition was acquired by the bite of an insect. The term microfilaria was not broadly recognized. We also determined that onchocerciasis was not perceived as a serious health problem: few persons (12%) mentioned la filaria when requested to provide a complete list of illnesses that occurred in the community, and the gravity of infection (based on rank ordering of common illnesses) was similar to that of a bad cold. Recommendations were made which might assist long-term acceptance of a national chemotherapy initiative against onchocerciasis in Guatemala.

Adolescent↗
PubMed1991

[Photon-emission tomography (SPECT) in the detection of hepatic hemangiomas of small size].

By means of ultrasonography it is possible to detect hepatic lesions of few millimeters, however, it doesn't give a precise diagnosis. The role of planar scintigraphy in the diagnosis of hepatic hemangioma has already been established, although it has limitations which are partially solved by means of SPECT. 46 patients with different pathologies and 9 healthy volunteers have been studied. In the 19 confirmed hemangiomas, planar scintigraphy showed a sensitivity of 73.6% versus 84.2% for SPECT, mostly due to lesions less than 2.5 cm. Besides that, SPECT allowed a correct visualization of abdominal and intrahepatic major vessels, although lesions less than 1.5 cm are difficult to evaluate.

Adult↗
PubMed1990

Multigene families in African swine fever virus: family 360.

A group of cross-hybridizing DNA segments contained within the restriction fragments RK', RL, RJ, and RD' of African swine fever virus DNA were mapped and sequenced. Analysis of these sequences revealed the presence of a family of homologous open reading frames in regions close to the DNA ends. The whole family is composed of six open reading frames with an average length of 360 coding triplets (multigene family 360), four of which are located in the left part of the genome and two of which are in the right terminal EcoRI fragment. In close proximity to the right terminal inverted repeat, we found an additional small open reading frame which was homologous to the 5'-terminal portion of the other open reading frames, suggesting that most of that open reading frame has been deleted. These repeated sequences account for the previously described inverted internal repetitions (J.M. Sogo, J.M. Almendral, A. Talavera, and E. Viñuela, Virology 133:271-275, 1984). Most of the genes of multigene family 360 are transcribed in African swine fever virus-infected cells. A comparison of the predicted protein sequences of family 360 indicated that several residues are conserved, suggesting that an overall structure is maintained for every member of the family. The transcription direction of each open reading frame, as well as the evolutionary relationships among the genes, suggests that the family originated by gene duplication and translocation of sequences between the DNA ends.

Amino Acid Sequence↗
PubMed1987

[Prune belly syndrome. Etiopathogenic considerations in 4 cases].

Four cases of prune belly syndrome of 19, 24, and 32 weeks of gestational age, and 14 days of postnatal life are described. All of them show urethral atresia, muscle defects of abdominal wall and undescended testes. Analysis of these four cases at different developmental stages brings authors to approach etiopathogenic basis of this syndrome.

Female↗
PubMedDate not supplied

[Uncoordinated urination syndrome].

Fourteen male and female patients aged 8 to 21 (mean 12.57 years) underwent clinical and urodynamic evaluation. These patients had different lower urinary tract symptoms, and flow changes and increased perineal muscle activity during voiding were demonstrated urodynamically. The urodynamic work up consisted of flowmetry, cystomanometry, detrusor pressure-flow test, videocystourethrography and electromyography of the periurethral sphincter. The urinary symptoms were nocturnal enuresis in 6 cases (42.85%), urinary infection in 6 (42.85%), difficulty in voiding in 4 (28.57%), urgency incontinence in 4 (28.57%), voiding urgency in 2 (14.28%) and diurnal urgency and frequency in 1 case (7.18%). The following 3 voiding bladder behaviour patterns were observed; 1) involuntary micturition from bladder instability (7 cases, 50%), 2) micturition from voluntary bladder contraction (2 cases, 14.28%) and 3) absence of bladder contractile activity with voiding achieved using the abdominal muscles (5 cases, 35.71%). Decompensated voiding was observed in 21% of the overall group. Of these, 75% belonged to the group that used the abdominal muscles to void. Dilatation of the proximal or prostatic urethra was frequently accompanied by bladder instability (57%). The urodynamic data may be useful in determining the outcome of treatment.

Abdominal Muscles↗
PubMedDate not supplied

[Effect of dopamine on lung vascular mechanics. Its study in the isolated canine lobe model embolized with gel foam].

After inducing an increase in pulmonary vascular resistance (Rp) in an isolated in situ lobe (LAIS) embolized with gelfoam (GE), the effect of dopamine infused at 12 mcg/kg/min in eight mongrel dogs was evaluated. We studied the rectilinear pressure-flow (QL/PL) relationship, the hemodynamic and blood gas before (condition A), 15 minutes later GE (condition B) and after dopamine infusion (condition C). After GE a decrease in QL and an increase in inflow (PL) and mean critical closing pressures (PLi) were noticed (p less than 0.01). Also decrease in pulmonary vascular conductance (1/Rp) was noted. At condition C a rise in QL and PL were noticed (p less than 0.01), but 1/Rp and PLi did not change. We concluded that dopamine in the LAIS with increased Rp at the used doses does not have vasoactive action for those vessels responsible for the phenomenons where flow resistance or mean critical closing pressure occurs.

Animals↗