RESUMEN
Glycosylation defects occur in several human diseases. In IgA nephropathy, IgA1 contains O-glycans that are galactose-deficient and consist mostly of core 1 alpha2,6 sialylated N-acetylgalactosamine, a configuration suspected to prevent beta1,3 galactosylation. We confirmed the same aberrancy in IgA1 secreted by the human DAKIKI B cell line. Biochemical assays indicated CMP-NeuAc:GalNAc-IgA1 alpha2,6-sialyltransferase activity in this cell line. However, a candidate enzyme, ST6-GalNAcI, was not transcribed in DAKIKI cells, B cells isolated from blood, or Epstein-Barr virus (EBV)-immortalized IgA1-producing cells from the blood of IgAN patients and healthy controls. Instead, ST6-GalNAcII transcription was detected at a high level. Expression of the ST6-GalNAcII gene and activity of the CMP-NeuAc:GalNAc-IgA1 alpha2,6-sialyltransferase were higher in IgA1-producing cell lines from IgAN patients than in such cells from healthy controls. These data are the first evidence that human cells that lack ST6-GalNAcI can sialylate core 1 GalNAc-Ser/Thr.
Asunto(s)
Inmunoglobulina A/biosíntesis , Sialiltransferasas/metabolismo , Línea Celular , Línea Celular Transformada , Ensayo de Inmunoadsorción Enzimática , Regulación Enzimológica de la Expresión Génica , Glicosilación , Células HT29 , Herpesvirus Humano 4/metabolismo , Humanos , Lectinas/metabolismo , Leucocitos Mononucleares/enzimología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Transcripción Reversa/genética , Sialiltransferasas/genética , beta-D-Galactósido alfa 2-6-SialiltransferasaRESUMEN
Heterosubtypic immunity (HSI) is defined as cross-protection to infection with an influenza A virus serotype other than the one used for primary infection. Although HSI has been thought to be mediated by serotype cross-reactive cytotoxic T lymphocytes (CTL) that recognize conserved epitopes of structural proteins, recent studies suggest that antibodies (Abs) may make a significant contribution. In this study, we provide further evidence for the role of Abs in HSI using transgenic mice lacking terminal deoxyribonucleotidyltransferase (TdT), which adds N nucleotides to V-D and D-J junctions of the complementary determining region 3 (CDR3) (TdT(-/-)) and mice with altered Ab repertoires due to replacement of the complete locus of heavy chain diversity segments (D(H)) with an altered D(H) segment (namely, Delta D-iD). Both types of mice failed to generate complete HSI, although they were able to mount protective immunity to a homologous challenge. Lower levels of virus-specific antibodies along with more severely impaired HSI were observed in TdT(-/-) mice compared to those in Delta D-iD mice, while CTL activity remained unchanged in both types of mice. These findings indicate that a properly diversified antibody repertoire is required for HSI and that N addition by TdT is a more effective mechanism in the induction of a properly diversified antibody repertoire and, therefore, complete HSI. The results suggest that the diversity of the antibody repertoire as determined by the composition of the D region of HCDR3 and by N addition are among the mechanisms selected for in evolution to create a favorable environment to resolve infections with mutated viruses.
Asunto(s)
Anticuerpos Antivirales/inmunología , Reacciones Cruzadas , Virus de la Influenza A/inmunología , Gripe Humana/inmunología , Receptores de Antígenos de Linfocitos T/inmunología , Animales , Peso Corporal , ADN Nucleotidilexotransferasa/deficiencia , Modelos Animales de Enfermedad , Humanos , Cadenas Pesadas de Inmunoglobulina/genética , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Eliminación de Secuencia , Análisis de Supervivencia , Subgrupos de Linfocitos T/inmunología , Linfocitos T Citotóxicos/inmunología , Linfocitos T Colaboradores-Inductores/inmunologíaRESUMEN
OBJECTIVES: To address a question whether immune responses to HPV infection play a role in control of cervical cancer, we analyzed systemic and mucosal immune responses to HPV in women who underwent radical hysterectomy for cervical cancer (HCC) or loop conization due to cervical dysplasia (LOOP), or had hysterectomy for other reasons (HNN). METHODS: HPV-specific antibodies in sera and vaginal washes were determined by ELISA using recombinant HPV 16 E7 oncoprotein. Cytokines in vaginal washes were assayed by Linco cytokine multiplex method using Luminex technology. Differential gene expression profiling in cervical tumor was determined by microarray analysis and Real-time RT-PCR. RESULTS: While levels of HPV-16 E7-specific IgG in vaginal wash were significantly higher in women undergoing HCC and HNN, the levels of the HPV-16 E7-specific IgA in vaginal wash of women with cervical cancer and cervical dysplasia were lower as compared to patients in HNN. Proinflammatory cytokines, such as IL-6 and IL-8, were dominant in vaginal washes of all subjects studied. However, no pattern of Th1-type and Th2-type cytokine induction was observed as demonstrated by protein analysis as well as differential gene expression profiling in cervical tumor. CONCLUSIONS: These results suggest a selective down-regulation of local HPV-specific IgA responses in women with cervical cancer.