Browsing by keyword "B cell"
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Dynamics of dengue virus-specific B cells in the response to dengue virus-1 infections using flow cytometry with labeled virionsBACKGROUND: The development of reagents to identify and characterize antigen-specific B cells has been challenging. METHODS: We recently developed Alexa Fluor-labeled dengue viruses (AF DENV) to characterize antigen-specific B cells in the peripheral blood of DENV-immune individuals. RESULTS: In this study, we used AF DENV-1 together with AF DENV-2 on PBMC from children in Thailand undergoing acute primary or secondary DENV-1 infections to analyze the phenotypes of antigen-specific B cells that reflected their exposure or clinical diagnosis. DENV serotype-specific and cross-reactive B cells were identified in PBMC from all subjects. Frequencies of AF-DENV+ class switched memory B cells (IgD-CD27+ CD19+ cells) reached up to 8% during acute infection and early convalescence. AF DENV-labeled B cells expressed high levels of CD27 and CD38 during acute infection, characteristic of plasmablasts, and transitioned into memory B cells (CD38-CD27+) at the early convalescent time point. There was higher activation of memory B cells early during acute secondary infection suggesting reactivation from a previous DENV infection. CONCLUSIONS: AF DENV reveal changes in the phenotype of DENV serotype-specific and cross-reactive B cells during and after natural DENV infection and could be useful in analysis of the response to DENV vaccination.
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Kaposi Sarcoma-associated Herpesvirus Glycoprotein H is Indispensable for Infection of Epithelial, Endothelial, and Fibroblast Cell TypesKaposi sarcoma-associated herpesvirus (KSHV) is an emerging pathogen and is the causative infectious agent of Kaposi sarcoma and two malignancies of B cell origin. To date, there is no licensed KSHV vaccine. Development of an effective vaccine against KSHV continues to be limited by a poor understanding of how the virus initiates acute primary infection in vivo in diverse human cell types. The role of glycoprotein H (gH) in herpesvirus entry mechanisms remains largely unresolved. To characterize the requirement for KSHV gH in the viral life cycle and in determination of cell tropism, we generated and characterized a mutant KSHV in which expression of gH was abrogated. Using a bacterial artificial chromosome containing a complete recombinant KSHV genome and recombinant DNA technology, we inserted stop codons into the gH coding region. We used electron microscopy to reveal that the gH-null mutant virus assembled and exited from cells normally, compared to wild-type virus. Using purified virions, we assessed infectivity of the gH-null mutant in diverse mammalian cell types in vitro Unlike wild-type virus or a gH-containing revertant, the gH-null mutant was unable to infect any of the epithelial, endothelial, or fibroblast cell types tested. However, its ability to infect B cells was equivocal, and remains to be investigated in vivo due to generally poor infectivity in vitro Together, these results suggest that gH is critical for KSHV infection of highly permissive cell types including epithelial, endothelial, and fibroblasts. MPORTANCE: All homologues of herpesvirus gH studied to date have been implicated in playing an essential role in viral infection of diverse permissive cell types. However, the role of gH in the mechanism of KSHV infection remains largely unresolved. In this study, we generated a gH-null mutant KSHV and provided evidence that deficiency of gH expression did not affect viral particle assembly or egress. Using the gH-null mutant, we showed that gH was indispensable for KSHV infection of epithelial, endothelial, and fibroblast cells in vitro. This suggests that gH is an important target for the development of a KSHV prophylactic vaccine to prevent initial viral infection.
