RESUMO
Human papillomavirus (HPV) consists in a great clinical importance worldwide, being involved in the most frequent sexually transmitted viral infection, genital warts, with high incidence in both men and women. Furthermore, HPV can also develop a broad spectrum of cancers, the most common is cervical cancer, being one of the main causes of morbidity and mortality in women worldwide, especially in low-and middle-income countries (LMICs). Here we discuss the importance of cervical cancer prevention and others related cancers caused by HPV, information about HPV vaccines, evaluation of single-dose use, low vaccination coverage, as well as other topics regarding viral transmission, biology, and epidemiology, aiming for the reduction of cervical carcinoma cases and others HPV associated diseases, reducing infections and death in the world as regarding to HPV, through the wide people immunization.
RESUMO
Human papillomavirus (HPV) infection is a leading cause of morbidity and mortality in women worldwide. The virus is associated with benign warts and a broad spectrum of malignancies, including cervical cancer, considered a disease of high clinical relevance, especially in developing countries. In this study we developed the production of recombinant proteins HPV16 L1 and HPV16 L2 in human cells in suspension (293-F), which were transiently co-transfected with the pUF3L1h and pUF3L2h vectors. Expressions of recombinant HPV16 L1 and L2 capsid proteins was detected by laser scanning confocal microscopy and flow cytometry. Both proteins were identified intracellularly in the nucleus and cytoplasm of cells. The presence of these heterologous proteins and VLPs formation were detected by transmission electron microscopy (TEM) through colloidal gold immunolabeling and negative staining. Cell extracts containing recombinant proteins were purified by affinity chromatography and immunization of Balb/c mice with the formulation HPV16 L1/L2 VLPs containing adjuvant was able to induce higher titer of anti-HPV16 L1, when compared to HPV16 L2 antibodies by indirect ELISA assay. These data indicate that transient expression in 293-F cells was efficiently established. The results are promising for obtain recombinant proteins of the HPV capsid for future studies involving human papillomavirus, as well as to contribute for the development of other vaccine strategies for prevention against HPV.
RESUMO
Aprotinin, the most studied serine proteinase inhibitor, was isolated from porcine lung for the first time. The purified porcine aprotinin had an Mr value of approximately 7 kDa. It cross-reacted with polyclonal serum anti-commercial aprotinin. About 1 microg porcine aprotinin inhibited 6 microg trypsin whereas 1 microg commercial soybean inhibitor inhibited only 1 microg trypsin. The aprotinin gene was also isolated from porcine lung: the deduced amino acid sequence showed 74% identity to bovine aprotinin.
Assuntos
Aprotinina/genética , Aprotinina/isolamento & purificação , Pulmão/metabolismo , Sequência de Aminoácidos , Animais , Aprotinina/metabolismo , Sequência de Bases , Bovinos , Cromatografia de Afinidade , DNA/genética , Eletroforese em Gel de Poliacrilamida , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Homologia de Sequência de Aminoácidos , Suínos , Tripsina/metabolismoRESUMO
Aprotinin, the most studied serine proteinase inhibitor, was isolated from porcine lung for the first time. The purified porcine aprotinin had an Mr value of ¡7 kDa. It cross-reacted with polyclonal serum anti-commercial aprotinin. About 1 ¥ìg porcine aprotinin inhibited 6 ¥ìg trypsin whereas 1 ¥ìg commercial soybean inhibitor inhibited only 1 ¥ìg trypsin. The aprotinin gene was also isolated from porcine lung: the deduced amino acid sequence showed 74% identity to bovine aprotinin.
Assuntos
Masculino , Feminino , Animais , Aprotinina/isolamento & purificação , SuínosRESUMO
In order to develop a combined recombinant Mycobacterium bovis BCG (rBCG) vaccine against diphtheria, pertussis and tetanus (DPT), we have constructed different strains of rBCG expressing tetanus toxin fragment C (FC), driven by the up-regulated M. fortuitum beta-lactamase promoter, pBlaF*. Tetanus toxin FC was expressed in comparable levels in native form or in fusion with the beta-lactamase exportation signal sequence; however, in both constructs it was localized to the cytosol. Immunization of mice with rBCG-FC or its combination with rBCG expressing CRM197, induced anti-tetanus toxin antibodies with a Th2 immunoglobulin profile. Administration of a subimmunizing dose of the diphtheria-tetanus toxoid vaccine showed that rBCG-FC primed mice for production of an intense humoral response. Interestingly, the combination of rBCG-FC and rBCG-CRM197 reduced the time required for maturation of the immune response and increased anti-tetanus toxin antibody levels, suggesting adjuvant properties for rBCG-CRM197; this combination induced 75% protection in mice challenged with 100 minimum lethal doses (MLD) of tetanus toxin. Antisera from guinea pigs immunized with this combination were shown to neutralize tetanus toxin and diphtheria toxin. Our results suggest reciprocal adjuvant effects of rBCG-FC and rBCG-CRM197, which may contribute to induction of a more effective immune response against both diseases.
Assuntos
Adjuvantes Imunológicos , Anticorpos Antibacterianos/imunologia , Vacina BCG/imunologia , Proteínas de Bactérias/biossíntese , Mycobacterium bovis/genética , Mycobacterium bovis/imunologia , Toxina Tetânica/imunologia , Animais , Formação de Anticorpos/imunologia , Proteínas de Bactérias/genética , Western Blotting , Chlorocebus aethiops , Toxina Diftérica/antagonistas & inibidores , Vacina contra Difteria, Tétano e Coqueluche/imunologia , Ensaio de Imunoadsorção Enzimática , Escherichia coli/genética , Vetores Genéticos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Mycobacterium bovis/crescimento & desenvolvimento , Testes de Neutralização , Sorotipagem , Toxina Tetânica/antagonistas & inibidores , Toxina Tetânica/biossíntese , Vacinas Combinadas , Vacinas Sintéticas/imunologia , Células VeroRESUMO
In order to develop a combined recombinant Mycobacterium bovis BCG (rBCG) vaccine against diphtheria, pertussis and tetanus (DPT), we have constructed different strains of rBCG expressing tetanus toxin fragment C (FC), driven by the up-regulated M. fortuitum â-lactamase promoter, pBlaF∗. Tetanus toxin FC was expressed in comparable levels in native form or in fusion with the â-lactamase exportation signal sequence; however, in both constructs it was localized to the cytosol. Immunization of mice with rBCG-FC or its combination with rBCG expressing CRM197, induced anti-tetanus toxin antibodies with a Th2 immunoglobulin profile. Administration of a subimmunizing dose of the diphtheriatetanus toxoid vaccine showed that rBCG-FC primed mice for production of an intense humoral response. Interestingly, the combination of rBCG-FC and rBCG-CRM197 reduced the time required for maturation of the immune response and increased anti-tetanus toxin antibody levels, suggesting adjuvant properties for rBCG-CRM197; this combination induced 75% protection in mice challenged with 100 minimum lethal doses (MLD) of tetanus toxin. Antisera from guinea pigs immunized with this combination were shown to neutralize tetanus toxin and diphtheria toxin. Our results suggest reciprocal adjuvant effects of rBCG-FC and rBCG-CRM197, which may contribute to induction of a more effective immune response against both diseases.
Assuntos
Animais , Ratos , Mycobacterium bovis , Vacina BCGRESUMO
Samples from 20 lots of diphtheria-tetanus (adult use dT) vaccine and from 20 lots of diphtheria-tetanus-pertussis (DTP) vaccine were used to standardize and validate the in vitro toxin binding inhibition (ToBI) test for the immunogenicity test of the tetanus component. The levels of tetanus antitoxin obtained by ToBI test were compared to those obtained using the toxin neutralization (TN) test in mice routinely employed to perform the quality control of the tetanus component in adsorbed vaccines. The results ranged from 1.8 to 3.5 IU/ml for dT and 2 to 4 IU/ml for DTP by ToBI test and 1.4 to 3 IU/ml for dT and 1.8 to 3.5 IU/ml for DTP by TN in mice. These results were significantly correlated. From this study, it is concluded that the ToBI test is an alternative to the in vivo neutralization procedure in the immunogenicity test of the tetanus component in adsorbed vaccines. A substantial refinement and a reduction in use of animals can be achieved.
Assuntos
Vacina contra Difteria e Tétano/imunologia , Vacina contra Difteria, Tétano e Coqueluche/imunologia , Antitoxina Tetânica/imunologia , Animais , Toxoide Diftérico , Cobaias , Camundongos , Camundongos Endogâmicos BALB C , Testes de Neutralização , Controle de Qualidade , Antitoxina Tetânica/sangue , Toxina Tetânica , Toxoide TetânicoRESUMO
Samples from 20 lots of diphtheria-tetanus (adult use dT) vaccine and from 20 lots of diphtheria-tetanus-pertussis (DTP) vaccine were used to standardize and validate the in vitro toxin binding inhibition (ToBI) test for the immunogenicity test of the tetanus component. The levels of tetanus antitoxin obtained by ToBI test were compared to those obtained using the toxin neutralization (TN) test in mice routinely employed to perform the quality control of the tetanus component in adsorbed vaccines. The results ranged from 1.8 to 3.5 IU/ml for dT and 2 to 4 IU/ml for DTP by ToBI test and 1.4 to 3 IU/ml for dT and 1.8 to 3.5 IU/ml for DTP by TN in mice. These results were significantly correlated. From this study, it is concluded that the ToBI test is an alternative to the in vivo neutralization procedure in the immunogenicity test of the tetanus component in adsorbed vaccines. A substantial refinement and a reduction in use of animals can be achieved
Assuntos
Animais , Cobaias , Camundongos , Antitoxina Tetânica , Vacina contra Difteria, Tétano e Coqueluche , Vacina contra Difteria e Tétano , Controle de Qualidade , Toxina Tetânica , Testes de Neutralização , Toxoide Diftérico , Antitoxina Tetânica , Toxoide Tetânico , Camundongos Endogâmicos BALB CRESUMO
Samples from 20 lots of dT vaccine and from 20 lots of DTP vaccine were used to standardize and validate the Vero cell and the toxin binding inhibition (ToBI) tests for the potency control of diphtheria component. For the Vero cell method, violet crystal solution was used to stain the cells and estimate the endpoint of diluted diphtheria antitoxin. Diphtheria anatoxin was used for performing the ToBI test instead of toxin. The results obtained by both in vitro tests were similar to those obtained by in vivo toxin neutralization test in guinea pigs. The various analysis and the chi(2) test applied to evaluate the reproducibility and homogeneity, respectively, among in vitro tests and in vivo toxin neutralization test did not detect statistical significant difference for both analysed vaccines. An excellent correlation among in vitro tests and in vivo neutralization test was observed by Spearman's correlation coefficient.
Assuntos
Vacina contra Difteria e Tétano/imunologia , Vacina contra Difteria, Tétano e Coqueluche/imunologia , Adsorção , Animais , Chlorocebus aethiops , Toxoide Diftérico , Relação Dose-Resposta Imunológica , Cobaias , Imunoglobulina G/metabolismo , Testes de Neutralização , Peroxidases/metabolismo , Controle de Qualidade , Células VeroRESUMO
Samples from 20 lots of dT vaccine and from 20 lots of DTP vaccine were used to standardize and validate the Vero cell and the toxin binding inhibition (ToBI) tests for the potency control of diphtheria component. For the Vero cell method, violet crystal solution was used to stain the cells and estimate the endpoint of diluted diphtheria antitoxin. Diphtheria anatoxin was used for performing the ToBI test instead of toxin. The results obtained by both in vitro tests were similar to those obtained by in vivo toxin neutralization test in guinea pigs. The various analysis and the ÷2 test applied to evaluate the reproducibility and homogeneity, respectively, among in vitro tests and in vivo toxin neutralization test did not detect statistical significant difference for both analysed vaccines. An excellent correlation among in vitro tests and in vivo neutralization test was observed by Spearman's correlation coefficient.