RESUMO
OBJECTIVE: To estimate the prevalence of endometrial polyps and to investigate associated abnormal uterine bleeding in a Danish population aged 20-74 years. METHODS: This was a study of a random selection of women from the Danish Civil Registration System: 1660 women were invited of whom 686 were included (429 pre- and 257 postmenopausal). AUB was assessed by a validated questionnaire. The women underwent transvaginal sonography (TVS) and saline contrast sonohysterography (SCSH). Hysteroscopic resection was performed in cases with suspected focal intrauterine pathology. Full evaluation was performed in 619 women (two failures of TVS and 60 failures of SCSH, in two women SCSH was contraindicated (endometrial cancer), in two women hysteroscopy was contraindicated, and one polyp was lost before histology). World Health Organization histopathological criteria were used for diagnosing true endometrial polyps. RESULTS: On final diagnosis there were 48 women with polyps, eight with submucosal myomas, four with other benign findings and one with polypoidal growing endometrial cancer. Complex hyperplasia without atypia was diagnosed in two women with polyps. The prevalence of endometrial polyps was 7.8% (48/619; 95% CI, 5.6-9.9%). The prevalence was influenced significantly by age (P<0.005); in women below the age of 30 years, the prevalence was 0.9%. Polyps were diagnosed in 5.8% of pre- and 11.8% of postmenopausal women (P<0.01). Thirty-nine (82%) of the women who had histopathologically verified polyps were asymptomatic. In asymptomatic premenopausal women the prevalence of polyps was 7.6%, while it was 13% in asymptomatic postmenopausal women. AUB, in particular intermenstrual bleeding, was more frequent among women without polyps (38%). By ultrasound examination, submucosal myomas were diagnosed in 4.2% (26/622; 95% CI, 2.6-5.8%) and intramural myomas in 11.1% (76/684; 95% CI, 8.8-13.5%) of women. Polyps were diagnosed in 2% of oral-contraceptive and 25% of hormone-therapy users. CONCLUSIONS: The overall prevalence of endometrial polyps was 7.8% and the prevalence increased with age. Polyps were rare (0.9%) in women below the age of 30 years. Surprisingly, AUB was less frequent among women with polyps than among those without polyps.
Assuntos
Neoplasias do Endométrio/epidemiologia , Pólipos/epidemiologia , Hemorragia Uterina/epidemiologia , Adulto , Idoso , Dinamarca/epidemiologia , Neoplasias do Endométrio/diagnóstico por imagem , Endossonografia , Feminino , Humanos , Histeroscopia , Pessoa de Meia-Idade , Pólipos/diagnóstico por imagem , Prevalência , Inquéritos e Questionários , Hemorragia Uterina/diagnóstico por imagem , Adulto JovemRESUMO
OBJECTIVE: To assess the diagnostic value of transvaginal sonographic (TVS) measurement of endometrial thickness for diagnosing focal intrauterine pathology in women without abnormal uterine bleeding (AUB). METHODS: A random selection from the Danish Civil Registration System was made: 1660 women aged 20-74 years were invited to participate and 686 women were eligible and accepted inclusion (429 pre- and 257 postmenopausal). The women underwent TVS measurement of endometrial thickness and saline contrast sonohysterography (SCSH). Hysteroscopic resection with histopathology (gold standard) was performed when focal intrauterine pathology was suspected at SCSH. We excluded women with AUB (n = 237), failure of SCSH (n = 50), a scan that was not in the follicular phase (n = 11), hysteroscopy contraindicated (n = 2), and users of sequential hormone therapy (n = 9) or selective estrogen receptor modulators (n = 2). Thus, 375 women without AUB were included (217 pre- and 158 postmenopausal). Receiver-operating characteristics (ROC) curves for endometrial thickness and focal lesion were analyzed. RESULTS: Focal intrauterine pathology was confirmed in 41 women (35 with polyps, five with submucosal myomas and one with polypoidal growing cancer). For premenopausal women, the area under the ROC curve (AUC) was 0.79 (95% CI, 0.68-0.89) and for postmenopausal women it was 0.84 (95% CI, 0.76-0.92). For premenopausal women, the best negative likelihood ratio (LR- = 0.11) was obtained at an endometrial thickness of 5.2 mm, with a negative predictive value (NPV) of 99% and a positive predictive value (PPV) of 10%. For postmenopausal women the best LR- (0.08) was obtained at an endometrial thickness of 2.8 mm, with a NPV of 99% and a PPV of 26%. CONCLUSIONS: In women without AUB, TVS measurement of endometrial thickness is a poor diagnostic test, but is apparently efficacious in excluding focal intrauterine pathology, especially in postmenopausal women. The 4-5-mm threshold conventionally used to exclude endometrial malignancy in women with postmenopausal bleeding is not transferable to women without AUB for excluding focal intrauterine pathology.
Assuntos
Endométrio/diagnóstico por imagem , Pólipos/diagnóstico por imagem , Hemorragia Uterina/diagnóstico por imagem , Neoplasias Uterinas/diagnóstico por imagem , Adulto , Idoso , Detecção Precoce de Câncer/métodos , Endométrio/patologia , Endossonografia/métodos , Feminino , Humanos , Pessoa de Meia-Idade , Pólipos/patologia , Pós-Menopausa , Valor Preditivo dos Testes , Pré-Menopausa , Sensibilidade e Especificidade , Hemorragia Uterina/patologia , Neoplasias Uterinas/patologia , Adulto JovemRESUMO
Two peptides, designated L and K, covering a sequence near the NH-terminal end of the S1 subunit of pertussis toxin (PT) were conjugated to the PPD (purified protein derivative) of M. tuberculosis by either glutaraldehyde (GLUT) or succinimidyl 4-(N-maleimidomethyl) cyclohexane-1-carboxylate (SMCC) and N-succinimidyl 3-(2-pyridyldithio) propionate (SPDP) and injected into groups of mice and guinea pigs. Initially, the effect of priming the animals with BCG vaccine and the use of aluminium hydroxide as adjuvant for the anti-peptide antibody response was studied. The group of BCG-primed mice immunized with adsorbed peptide conjugates showed the highest anti-peptide conjugate antibody response. Based on this finding, groups of BCG-primed mice were immunized four times with either adsorbed peptide L-GLUT, peptide L-SMCC/SPDP or peptide K-SMCC/SPDP conjugates and the fine peptide specificity as well as the PT and S1 cross-reactivity was investigated in ELISA. Mice immunized with peptide L-GLUT showed a significant antibody response to the homologous conjugate, only, whereas the group injected with the peptide L-SMCC/SPDP conjugate gave a significant response to both peptide K and L conjugated by the SMCC-SPDP method. Likewise, mice immunized with the peptide K-SMCC/SPDP conjugate reacted with the homologous and peptide L-SMCC/SPDP conjugate, although only the response to the former conjugate was significantly greater than the response to PPD. All groups showed a strong anti-PPD response. The anti-PT/S1 cross-reactivity of the antisera varied considerably within each group but was found to be highest in the peptide L-GLUT-immunized animals. The results of the present study not only stress the importance of BCG priming and use of aluminium hydroxide adjuvants for the immunogenicity of the peptides in question but also point to the specificity of the conjugation methods employed as low cross-reactivity between the anti-peptide L-GLUT and L-SMCC/SPDP antisera was noted. Moreover, it appeared that the choice of conjugation method may have an effect on the ability of the peptide conjugates to induce an antibody response cross-reacting with the native protein.
Assuntos
Adjuvantes Imunológicos/química , Anticorpos Antibacterianos/biossíntese , Toxina Pertussis , Tuberculina/imunologia , Fatores de Virulência de Bordetella/química , Fatores de Virulência de Bordetella/imunologia , Sequência de Aminoácidos , Animais , Especificidade de Anticorpos , Vacina BCG/imunologia , Ensaio de Imunoadsorção Enzimática , Feminino , Cobaias , Masculino , Camundongos , Dados de Sequência Molecular , Fragmentos de Peptídeos/síntese química , Fragmentos de Peptídeos/imunologia , Especificidade da Espécie , Tuberculina/químicaRESUMO
The effect of detoxification of pertussis toxin (PT) for vaccine usage by either genetic manipulation, hydrogen peroxide or formaldehyde treatment on epitope recognition by a large collection of murine monoclonal pertussis toxin antibodies (PT MAbs) was assessed in a solid-phase and a soluble phase enzyme-linked immunosorbent assay (ELISA). The MAb binding patterns were found to be different in the two assays as the immobilization step appeared to cause conformational alterations in the native as well as the toxoided forms of PT. According to the solid-phase ELISA, genetic, hydrogen peroxide and 0.35% formaldehyde detoxification of PT resulted in reduced epitope binding in 2.9, 31.4 and 78.1% of the MAbs, respectively. In the soluble-phase ELISA, in which the MAbs were allowed to react with the toxoids or native toxin in solution, the percentages of MAbs showing decreased binding activity were 9.1, 50.0 and 71.4%, respectively. Stabilization of native PT and the genetically inactivated PT by 0.035% formaldehyde reduced the epitope binding activity in 50.0 and 8.7% of the MAbs, respectively. Increased antibody binding in the soluble-phase ELISA was observed in some of the toxoids: this ranged from 0% in the 0.35% formaldehyde-treated PT to 13.6% in the hydrogen peroxide-treated and 27.3% in the genetically detoxified PT. Regarding the effects of detoxification on epitopes recognized by PT-neutralizing MAbs in the soluble-phase ELISA, we found that treatment of PT with either 0.035%, 0.35% formaldehyde or hydrogen peroxide induced impairment of epitope binding in 72.7, 81.8 and 45.5% of the MAbs, respectively. In the genetically inactivated PT, the epitopes recognized by the neutralizing MAbs either appeared to remain intact or to show increased MAb binding activity. The epitope-binding patterns of several PT MAbs with mouse-protective properties varied considerably and were shown to be dependent on the detoxification procedure employed. The relevance of epitope alterations on PT as a vaccine component is discussed. The results of the present study may have important implications for future quality assessment of PT for use in acellular pertussis vaccines.
Assuntos
Anticorpos Monoclonais/imunologia , Epitopos , Formaldeído/farmacologia , Peróxido de Hidrogênio/farmacologia , Toxina Pertussis , Fatores de Virulência de Bordetella/imunologia , Animais , Células CHO , Cricetinae , Ensaio de Imunoadsorção Enzimática , Camundongos , Camundongos Endogâmicos BALB C , Fatores de Virulência de Bordetella/toxicidadeRESUMO
The amounts of pertussis toxin (PT), filamentous haemagglutinin (FHA), 69 kDa outer membrane protein (69 kDa OMP) and agglutinogens (AGG) 2 and 3 in extracts from the Danish whole-cell pertussis vaccine were studied in quantitative capture ELISA. With the exception of PT, the most effective extraction of these antigens was by heating the bacteria at 60 degrees C for 30 min in 2 M urea followed by sonication for 45 s. Extraction by 1 M sodium chloride prior to sonication resulted in higher levels of antigenic and biologically active PT. On average, a single human dose of pertussis vaccine (approximately 16 opacity units) was found to contain 5520 ng FHA, 63 ng PT, 1061 ng 69 kDa OMP, 397 ng AGG 2, 534 ng AGG 3 and 4840 ng lipopolysaccharide (LPS). The antigen content of one dose of the Danish pertussis vaccine appears to be low compared with the amounts found in the acellular vaccines currently in use. These findings may have important implications for the evaluation of the protective substances and the immunogenicity of whole-cell as opposed to acellular pertussis vaccines.
Assuntos
Adesinas Bacterianas , Vacina contra Coqueluche/química , Aglutininas/análise , Antígenos de Bactérias/análise , Proteínas da Membrana Bacteriana Externa/análise , Bordetella pertussis/imunologia , Ensaio de Imunoadsorção Enzimática , Hemaglutininas/análise , Lipopolissacarídeos/análise , Toxina Pertussis , Vacina contra Coqueluche/imunologia , Fatores de Virulência de Bordetella/análiseRESUMO
The cell mediated immune response (CMI) against pertussis antigens following vaccination with the traditional Danish whole cell pertussis vaccine (WC-P) and the Japanese acellular pertussis vaccine (A-PV) JNIH-3 was studied in four adult human volunteers. Vaccination with the A-PV induced an in vitro proliferative response of peripheral blood lymphocytes to pertussis toxin (PT) subunits S2-S4, S3-S4 and S5 and the filamentous hemagglutinin (FHA), and a better serological response to native PT, detoxified PT (dPT) and FHA than the WC-PV. The induced CMI and serological response were followed over a period of 17 weeks, and were not seen to decline during this period. Further, an in vitro proliferative response to Bordetella pertussis agglutinogen 2 and 3 were demonstrated using lymphocytes from recently and not-so-recently pertussis-vaccinated adults.
Assuntos
Adesinas Bacterianas , Formação de Anticorpos , Imunidade Celular , Vacina contra Coqueluche/imunologia , Fatores de Virulência de Bordetella , Adulto , Divisão Celular/efeitos dos fármacos , Ensaio de Imunoadsorção Enzimática , Hemaglutininas/imunologia , Humanos , Técnicas In Vitro , Ativação Linfocitária/efeitos dos fármacos , VacinaçãoRESUMO
The main purpose of the present study was to identify B-cell epitopes on the S4 subunit of pertussis toxin (PT) by the synthetic peptide approach. Two strategies were followed: (i) screening of two series of overlapping peptides (12- and 25-residue peptides) covering the entire S4 sequence by a panel of murine monoclonal anti-PT antibodies and various polyclonal anti-PT antisera in an enzyme-linked immunosorbent assay (ELISA), and (ii) analysis of the S4 amino acid sequence by a predictive algorithm followed by synthesis and immunization of mice with the predicted peptides coupled to diphtheria toxoid. The anti-peptide conjugate antisera were tested in an ELISA for cross-reactivity with native PT, B oligomer, and S4. Screening of the free peptides in an ELISA by the PT antisera indicated the presence of six B-cell epitope-containing domains covered by residues 18 to 32, 33 to 46, 39 to 52, 51 to 65, 71 to 84, and 91 to 106. None of the peptides, however, were recognized by the monoclonal anti-PT antibodies in an ELISA. Immunization with six computer-predicted peptides (B1 to B6) and three potential T-cell epitopes (T1 to T3) gave rise to very high antibody responses towards the homologous conjugates. With the exception of the anti-T1/diphtheria toxoid antisera, all anti-peptide conjugate antisera cross-reacted with PT in an ELISA at different levels. None of these anti-peptide conjugate antisera, however, showed any PT-neutralizing effect as measured by the Chinese hamster ovary cell assay and the leukocytosis-promoting activity test. The results of the present study suggest that discontinuous epitopes are predominant in the S4 subunit of native PT.
Assuntos
Linfócitos B/imunologia , Epitopos/análise , Toxina Pertussis , Fatores de Virulência de Bordetella/imunologia , Sequência de Aminoácidos , Animais , Células CHO , Cricetinae , Ensaio de Imunoadsorção Enzimática , Feminino , Imunização , Leucocitose/etiologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Dados de Sequência Molecular , Fragmentos de Peptídeos/síntese química , Fragmentos de Peptídeos/imunologia , CoelhosRESUMO
The aim of the present study was to identify murine T-cell epitopes on pertussis toxin subunit S4. Six mouse strains with five different haplotypes at the H-2 locus were immunized with the pertussis toxin B oligomer. Lymph node lymphocytes were isolated and stimulated in an in vitro proliferation assay with pertussis toxin components and 11 overlapping synthetic peptides synthesized on the basis of the primary sequence of S4. In vitro proliferative responses to the synthetic peptides revealed the presence of four distinct murine T-cell epitopes on subunit S4. The recognition of the peptides was major histocompatibility complex restricted. Immunizing four of the six mouse strains with the synthetic peptides showed that the peptides which were demonstrated to contain T-cell epitopes following immunization with the B oligomer were able to induce proliferative responses to detoxified pertussis toxin and pertussis toxin components containing subunit S4. One of the identified murine T-cell epitopes corresponded to one of the major human T-cell epitopes previously identified on subunit S4. It is hoped that this murine model system will facilitate the development of a synthetic immunogen mimicking the protective properties of pertussis toxin.
Assuntos
Epitopos/isolamento & purificação , Toxina Pertussis , Linfócitos T/imunologia , Fatores de Virulência de Bordetella/imunologia , Animais , Técnicas In Vitro , Ativação Linfocitária/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos , Fragmentos de Peptídeos/imunologia , VacinaçãoRESUMO
A group of mice was aerosol infected with live, virulent Bordetella pertussis bacteria. During a period of 7 weeks following the infection, with intervals of 1 week, lymphocytes were isolated from the tracheobroncheal lymph nodes (TBL) and the spleens (SPL) of the infected mice. The in vitro proliferative responses as well as the gamma interferon and tumor necrosis factor production levels of the isolated lymphocytes in response to stimulation with whole killed B. pertussis bacteria were measured as parameters for cell-mediated immunity (CMI). The course of the infection was monitored by counting of CFU in the lungs of the mice. Moreover, antibody responses in serum against a range of B. pertussis antigens were assessed. The results showed that a vigorous proliferative response of the TBL and SPL to stimulation with whole killed B. pertussis bacteria was induced by the infection. The proliferative response of the TBL was significantly higher than the response of the SPL. The proliferative responses were maximal 3 to 4 weeks after the infection and were paralleled by in vitro gamma interferon and tumor necrosis factor production upon specific stimulation. The development of the CMI was observed simultaneously with the clearance of the infection from the lungs. Antibody responses became measurable in the sera only after the infection was cleared. A specific CMI against pertussis toxin, the filamentous hemagglutinin, the 69-kDa outer membrane protein, and the agglutinogens 2 and 3, antigens which are under consideration for inclusion in future acellular pertussis vaccines, was successfully demonstrated in mice 3 weeks after the infection.
Assuntos
Bordetella pertussis/imunologia , Interferon gama/biossíntese , Subpopulações de Linfócitos/imunologia , Fator de Necrose Tumoral alfa/biossíntese , Coqueluche/imunologia , Aerossóis , Animais , Anticorpos Antibacterianos/biossíntese , Antígenos de Bactérias/imunologia , Brônquios/imunologia , Linfonodos/citologia , Linfonodos/imunologia , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL , Baço/citologia , Baço/imunologia , Fatores de Tempo , Traqueia/imunologiaRESUMO
Ten adult humans were vaccinated with the Japanese acellular pertussis vaccine JNIH-3, containing detoxified pertussis toxin (PT), formaldehyde, and filamentous hemagglutinin. The vaccination induced a specific antibody response to PT and filamentous hemagglutinin, and a Western blot (immunoblot) analysis of the antibody response to PT revealed antibodies to PT subunits S1, S2, S3, S4 and S5. The response of peripheral lymphocytes to PT was assessed in an in vitro proliferation assay. A proliferative response to detoxified PT and PT dimers S2-S4 and S3-S4 was found, and it was further demonstrated that the proliferative response to detoxified PT and dimer S2-S4 was mediated by T cells of the CD4+ phenotype. The specificity of the proliferative response to subunit S4 was analyzed with a range of synthetic peptides synthesized on the basis of the primary sequence of subunit S4. The proliferative response to the peptides revealed two major and one minor T-cell epitope located in the NH2-terminal end of subunit S4.
Assuntos
Epitopos/análise , Fragmentos de Peptídeos/imunologia , Toxina Pertussis , Linfócitos T/imunologia , Fatores de Virulência de Bordetella/imunologia , Adulto , Sequência de Aminoácidos , Anticorpos Antibacterianos/análise , Feminino , Humanos , Ativação Linfocitária , Depleção Linfocítica , Masculino , Dados de Sequência Molecular , Vacinação , Vacinas Sintéticas/imunologiaRESUMO
The specificity of the cell-mediated immune response to Bordetella pertussis following immunization of C57B1 mice with a whole-cell pertussis vaccine was assessed in a proliferation assay. A proliferative response of lymph node lymphocytes to the filamentous haemagglutinin, the 69 kDa outer membrane protein and the agglutinogens 2 and 3 was demonstrated. The proliferative cells were T cells of the CD4+ phenotype. In addition, several as yet uncharacterized antigens expressed by B. pertussis were shown to induce a proliferative response, also mediated by T cells of the CD4+ phenotype. Although a range of different immunization schedules and preparations of pertussis toxin were used, no specific proliferative responses to pertussis toxin, which is regarded as a protective antigen of major importance from B. pertussis, were found.