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Dendritic cell-mediated infection of primary B cells with KSHV

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Circulating B lymphocytes are the major reservoir of KSHV infection in infected subjects. However, B cell lines and primary B cells are resistant to direct KSHV infection in vitro. In addition, primary B cells are difficult to propagate for more than a few days. In this study, we combined a novel primary B cell propagation method with efficient infection mediated by dendritic cells to study KSHV de novo infection of primary B cells in vitro.

Primary monocyte-derived dendritic cells (MDDCs) or plasmacytoid dendritic cells (pDCs) were pulsed with KSHV for 4 hours at which point KSHV DNA was readily detectable. Uptake of KSHV was significantly reduced by pre-incubating cells with antibodies to integrins α3, β1 and DC-SIGN. Autologous B cells were grown on a feeder layer of irradiated NIH3T3 cells transduced with a human CD40L retroviral vector. KSHV+ DCs were co-cultivated with primary B cells for 4–8 hours and then separated by CD19+ immunomagnetic isolation. B cell cultures were maintained on feeder cells for >30 days and monitored for KSHV infection.

Efficient KSHV infection of primary B cells was mediated by both MDDCs and pDCs. KSHV LANA protein (ORF73) was detected by IFA in 2–15 percent of B cells through day 14. Viral gene expression analysis using a KSHV whole genome virus array showed establishment of latent KSHV infection followed by spontaneous reactivation of lytic viral replication in the primary B cell cultures.

These studies suggest that dendritic cells play an important role in the transmission and pathogenesis of KSHV in infected subjects as well as demonstrating a powerful in vitro model for studying KSHV infection of B cells.

Acknowledgements

Funded by NCI Contract N01-CO-12400.

Author information

Correspondence to R Bagni.

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Open Access This article is published under license to BioMed Central Ltd. This is an Open Access article is distributed under the terms of the Creative Commons Attribution 2.0 International License (https://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Keywords

  • Dendritic Cell
  • NIH3T3 Cell
  • Viral Gene Expression
  • Plasmacytoid Dendritic Cell
  • Spontaneous Reactivation