RESUMO
<p><b>BACKGROUND</b>To investigate the biological functions of a novel hepatitis B virus e antigen (HbeAg) interacting protein AK026018, and to use cDNA microarray technique to screen genes regulated by the protein.</p><p><b>METHODS</b>The AK026018 coding DNA fragment was amplified by reverse transcription polymerase chain reaction (RT-PCR) technique from HepG2 cell. The expressive vector of pcDNA3.1-AK was constructed by routine molecular biological methods. The HepG2 cells were transfected with pcDNA3.1 and pcDNA3.1-AK, respectively by using lipofectamine. The total RNA was isolated and reverse transcribed. The cDNA of each sample was subjected to microarray screening with 8,464 cDNA probes and analyzed by bioinformatics.</p><p><b>RESULTS</b>The expressive vector was constructed and confirmed by DNA sequencing analysis and restriction enzyme digestion. High quality mRNA and cDNA of transfected HepG2 cells had been prepared and successful microarray screening conducted. From the scanning results, there were 122 differential expression genes, of which 36 genes were down-regulated, and 16 genes were up-regulated.</p><p><b>CONCLUSION</b>Microarray technique was successfully used to screen the genes trans-regulated by AK026018. The expression of AK026018 protein affects the expression spectrum of HepG2 cells.</p>
Assuntos
Humanos , Proteínas de Transporte , Genética , Metabolismo , Fisiologia , Linhagem Celular Tumoral , Biologia Computacional , Regulação Neoplásica da Expressão Gênica , Antígenos E da Hepatite B , Metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Métodos , Ligação Proteica , Reação em Cadeia da Polimerase Via Transcriptase Reversa , TransfecçãoRESUMO
<p><b>OBJECTIVE</b>Test the cytokines secretion by peripheral blood mononuclear cells (PBMC) from hepatitis C patients after stimulation with highly variable region (HVR1) synthetic peptides.</p><p><b>METHODS</b>ELISA was used to test the cytokines secretion, flow cytometry to group the proliferated cells, and the antigenicity of synthetic peptide was predicted by the computer modeling.</p><p><b>RESULTS</b>(1)There were PBMC proliferation when stimulated by HVR1 antigen synthetic peptides. (2) There was a rise of IFN-gamma, IL-4 and IL-10. But no rise of IL-2 and TNF-gamma was found. (3) The proliferated cells were mainly CD4+ lymphocytes.</p><p><b>CONCLUSION</b>PMBC from hepatitis C patients tend to secret Th2 cytokines after stimulation with HVR1 designed by the authors.</p>