AIM: To study persistence and replication of hepatitis C virus (HCV) in patients' peripheral blood mononuclear cells (PBMC) cultured in vitro. METHODS: Epstein Barr virus (EBV) was used to transform the hepatitis ...AIM: To study persistence and replication of hepatitis C virus (HCV) in patients' peripheral blood mononuclear cells (PBMC) cultured in vitro. METHODS: Epstein Barr virus (EBV) was used to transform the hepatitis C virus from a HCV positive patient to permanent lymphoblastoid cell lines (LCL). Positive and negative HCV RNA strands of the cultured cells and growth media were detected by reverse transcriptase-polymerase chain reaction (RT-PCR) each month. Core and NS5 proteins of HCV were further tested using immunohistochemical SP method and in situ RT-PCR. RESULTS: HCV RNA positive strands were consistently detected the cultured cells for one year. The negative-strand RNA in LCL cells and the positive-strand RNA in supernatants were observed intermittently. Immunohistochemical results medicated expression of HCV NS3 and C proteins in LCL cytoplasm mostly. The positive signal of PCR product was dark blue and mainly localized to the LCL cytoplasm. The RT-PCR signal was eliminated by overnight RNase digestion but not DNase digestion. CONCLUSION: HCV may exist and remain functional in a cultured cell line for a long period.展开更多
AIM:To clone and identify genes differentially expressed in the EB virus transformed human B cell.METHODS:Suppression subtractive hybridization was used to construct the library which contains the differentiately expr...AIM:To clone and identify genes differentially expressed in the EB virus transformed human B cell.METHODS:Suppression subtractive hybridization was used to construct the library which contains the differentiately expressed cDNAs in EB virus transformed human B cell.Then the isolated genes were cloned and sequenced and identifed by RT PCR.Nucleic acid homology searches were performed using the BLAST program.RESULTS:By this technique,4 differentiately expressed gene cDNA fragments of EB virus transformed human B cell were obtained.CONCLUSION:SSH is an effective method to isolate differentiately expressed genes.展开更多
Background:Previous studies have examined the bulk transcriptome of peripheral blood immune cells in acquired immunodeficiency syndrome patients experiencing immunological non-responsiveness.This study aimed to invest...Background:Previous studies have examined the bulk transcriptome of peripheral blood immune cells in acquired immunodeficiency syndrome patients experiencing immunological non-responsiveness.This study aimed to investigate the characteristics of specific immune cell subtypes in acquired immunodeficiency syndrome patients who exhibit immunological non-responsiveness.Methods:A single-cell transcriptome sequencing of peripheral blood mononuclear cells obtained from both immunological responders(IRs)(CD4^(+)T-cell count>500)and immunological non-responders(INRs)(CD4^(+)T-cell count<300)was conducted.The transcriptomic profiles were used to identify distinct cell subpopulations,marker genes,and differentially expressed genes aiming to uncover potential genetic factors associated with immunological non-responsiveness.Results:Among the cellular subpopulations analyzed,the ratios of monocytes,CD16^(+)monocytes,and exhausted B cells demonstrated the most substantial differences between INRs and IRs,with fold changes of 39.79,11.08,and 2.71,respectively.In contrast,the CD4^(+)T cell ratio was significantly decreased(0.39-fold change)in INRs compared with that in IRs.Similarly,the ratios of natural killer cells and terminal effector CD8^(+)T cells were also lower(0.37-fold and 0.27-fold,respectively)in the INRs group.In addition to several well-characterized immune cell-specific markers,we identified a set of 181 marker genes that were enriched in biological pathways associated with human immunodeficiency virus(HIV)replication.Notably,ISG15,IFITM3,PLSCR1,HLA-DQB1,CCL3L1,and DDX5,which have been demonstrated to influence HIV replication through their interaction with viral proteins,emerged as significant monocyte marker genes.Furthermore,the differentially expressed genes in natural killer cells were also enriched in biological pathways associated with HIV replication.Conclusions:We generated an atlas of immune cell transcriptomes in HIV-infected IRs and INRs.Host genes associated with HIV replication were identified as markers of,and were found to be differentially expressed in,different types of immune cells.展开更多
基金The paper was support by a grant from the Ministry Youth Research of China,No.98-1-269
文摘AIM: To study persistence and replication of hepatitis C virus (HCV) in patients' peripheral blood mononuclear cells (PBMC) cultured in vitro. METHODS: Epstein Barr virus (EBV) was used to transform the hepatitis C virus from a HCV positive patient to permanent lymphoblastoid cell lines (LCL). Positive and negative HCV RNA strands of the cultured cells and growth media were detected by reverse transcriptase-polymerase chain reaction (RT-PCR) each month. Core and NS5 proteins of HCV were further tested using immunohistochemical SP method and in situ RT-PCR. RESULTS: HCV RNA positive strands were consistently detected the cultured cells for one year. The negative-strand RNA in LCL cells and the positive-strand RNA in supernatants were observed intermittently. Immunohistochemical results medicated expression of HCV NS3 and C proteins in LCL cytoplasm mostly. The positive signal of PCR product was dark blue and mainly localized to the LCL cytoplasm. The RT-PCR signal was eliminated by overnight RNase digestion but not DNase digestion. CONCLUSION: HCV may exist and remain functional in a cultured cell line for a long period.
文摘AIM:To clone and identify genes differentially expressed in the EB virus transformed human B cell.METHODS:Suppression subtractive hybridization was used to construct the library which contains the differentiately expressed cDNAs in EB virus transformed human B cell.Then the isolated genes were cloned and sequenced and identifed by RT PCR.Nucleic acid homology searches were performed using the BLAST program.RESULTS:By this technique,4 differentiately expressed gene cDNA fragments of EB virus transformed human B cell were obtained.CONCLUSION:SSH is an effective method to isolate differentiately expressed genes.
基金supported by the Joint Research Project of Health and Education in Fujian Province(No.2019-WJ-15)Natural Science Foundation of Fujian Province(Nos.2021J011295 and 2020J011165)Fujian Chinese Traditional Medicine University research funding(No.XB2020147)
文摘Background:Previous studies have examined the bulk transcriptome of peripheral blood immune cells in acquired immunodeficiency syndrome patients experiencing immunological non-responsiveness.This study aimed to investigate the characteristics of specific immune cell subtypes in acquired immunodeficiency syndrome patients who exhibit immunological non-responsiveness.Methods:A single-cell transcriptome sequencing of peripheral blood mononuclear cells obtained from both immunological responders(IRs)(CD4^(+)T-cell count>500)and immunological non-responders(INRs)(CD4^(+)T-cell count<300)was conducted.The transcriptomic profiles were used to identify distinct cell subpopulations,marker genes,and differentially expressed genes aiming to uncover potential genetic factors associated with immunological non-responsiveness.Results:Among the cellular subpopulations analyzed,the ratios of monocytes,CD16^(+)monocytes,and exhausted B cells demonstrated the most substantial differences between INRs and IRs,with fold changes of 39.79,11.08,and 2.71,respectively.In contrast,the CD4^(+)T cell ratio was significantly decreased(0.39-fold change)in INRs compared with that in IRs.Similarly,the ratios of natural killer cells and terminal effector CD8^(+)T cells were also lower(0.37-fold and 0.27-fold,respectively)in the INRs group.In addition to several well-characterized immune cell-specific markers,we identified a set of 181 marker genes that were enriched in biological pathways associated with human immunodeficiency virus(HIV)replication.Notably,ISG15,IFITM3,PLSCR1,HLA-DQB1,CCL3L1,and DDX5,which have been demonstrated to influence HIV replication through their interaction with viral proteins,emerged as significant monocyte marker genes.Furthermore,the differentially expressed genes in natural killer cells were also enriched in biological pathways associated with HIV replication.Conclusions:We generated an atlas of immune cell transcriptomes in HIV-infected IRs and INRs.Host genes associated with HIV replication were identified as markers of,and were found to be differentially expressed in,different types of immune cells.
