RESUMEN
The aim of the present study was to investigate the overall clinical expression characteristics of the cluster of differentiation (CD)28 family receptors [CD28, inducible T-cell co-stimulator, programmed cell death protein 1 (PD1), cytotoxic T-lymphocyte-associated protein 4 and B and T-lymphocyte attenuator] on T cells in patients with chronic hepatitis B (CHB), analyze the correlations among these receptors and the clinical parameters, and to investigate the effects of PD1 blockade on the receptor expression profiles, Tcell function and other biological effects. The expression characteristics of the CD28 family of receptors, the effects of PD1 blockade on the receptor expression profiles and the levels of interferon (IFN)γ were investigated in the T cells of patients with CHB. In addition, the transcription factor, Tbox 21 (Tbet) and GATA binding protein 3 (GATA3) mRNA expression levels were investigated in the peripheral blood mononuclear cells (PBMCs) of patients with CHB. The expression levels of the CD28 family receptors in the T cells of patients with CHB demonstrated distinct characteristics , for example levels of PD1 and CTLA4 on CD4 T cells and ICOS, PD1, and BTLA on CD8 T cells were increased in cells from patients with CHB compared with those from the healthy individuals. A significant positive correlation was demonstrated among the serum HBV DNA titers and the levels of PD1 on CD8+ T cells with the highest expression of PD1 corresponding to viral levels >106 IU/ml. A significant positive correlation was observed between the serum HBV DNA titers and the expression levels of BTLA on CD8+ T cells with the highest expression of BTLA corresponding to viral levels >106 IU/ml. PD1 blockade altered the expression profiles of CD28 family receptors in the T cells of patients with CHB, partly enhanced T cell function and increased the ratio of Tbet/GATA3 mRNA in PBMCs. Thus, CD28 family receptors are potential clinical indicators for the rapid monitoring of changes in T cell function during CHB treatment. Furthermore, PD1 blockade has a therapeutic potential that may be enhanced by modulating the expression of co-stimulatory and -inhibitory receptors of the CD28 family.
Asunto(s)
Antígenos CD28/metabolismo , Hepatitis B Crónica/inmunología , Hepatitis B Crónica/metabolismo , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismo , Anticuerpos Monoclonales/farmacología , Antígeno B7-H1/metabolismo , Biomarcadores , Antígenos CD28/genética , Estudios de Casos y Controles , Factor de Transcripción GATA3/metabolismo , Expresión Génica , Antígeno HLA-A2/genética , Antígeno HLA-A2/inmunología , Virus de la Hepatitis B/inmunología , Hepatitis B Crónica/genética , Hepatitis B Crónica/virología , Humanos , Inmunofenotipificación , Interferón gamma/biosíntesis , Leucocitos Mononucleares/efectos de los fármacos , Leucocitos Mononucleares/inmunología , Leucocitos Mononucleares/metabolismo , Receptor de Muerte Celular Programada 1/antagonistas & inhibidores , Proteínas de Dominio T Box/metabolismo , Carga ViralRESUMEN
AIM: To construct a prokaryotic plasmid expressing truncated duck hepatitis B virus core protein (DHBc(1-214)), purify the recombinant protein, and to develop polyclonal antibodies against DHBc. METHODS: DHBc(1-214) was cloned into vector pRSET-B, then expressed in E.coli Rosetta(DE3) pLacI induced by IPTG. The recombinant protein was purified using Ni-NTA spin column. Polyclonal antibody was developed by immunizing BALB/c mice with the purified recombinant protein, and their sensitivity and specificity were tested using enzyme-linked immunosorbent assay, immunohistochemical staining and Western blot analysis. RESULTS: Recombinant plasmid expressing truncated DHBc(1-214) was successfully constructed. A protein of 28,000 was expressed and purified. Polyclonal serum antibody with a high specificity was obtained by immunizing BALB/c mice with the purified recombinant protein. CONCLUSION: The truncated recombinant DHBc(1-214) developed in this study is purified and shown strong antigenecity. The polyclonal antibody against DHBc protein is generated by regular immunization method, demonstrating both high sensitivity and specificity. The protein and the antibody can be used for further clinical examination and research of DHBV.