KSHV infects a subset of human tonsillar B cells, driving proliferation and plasmablast differentiation
Lynn M. Hassman, … , Thomas J. Ellison, Dean H. Kedes
Lynn M. Hassman, … , Thomas J. Ellison, Dean H. Kedes
Published February 1, 2011; First published January 18, 2011
Citation Information: J Clin Invest. 2011;121(2):752-768. https://doi.org/10.1172/JCI44185.
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Research Article

KSHV infects a subset of human tonsillar B cells, driving proliferation and plasmablast differentiation

Abstract

Kaposi sarcoma–associated herpesvirus (KSHV; also known as HHV8) is the causative agent of two B cell tumors, multicentric Castleman disease (MCD) and primary effusion lymphoma (PEL). However, little is known about the nature of the specific B cell subtype(s) most susceptible to infection. Identifying these cells would provide direct insight into KSHV transmission and virus-induced transformation. To identify this subset and to determine whether infection alters its cellular phenotype, we exposed human tonsillar cells to KSHV and characterized infected cells using high-throughput multispectral imaging flow cytometry (MIFC). Stable expression of the virally encoded latency-associated nuclear antigen (LANA), a marker of latent KSHV infection, was observed predominantly in cells expressing the l light chain of the B cell receptor. These LANA+ B cells proliferated and exhibited similarities to the cells characteristic of MCD (IgMl-expressing plasmablasts), including blasting morphology with elevated expression of Ki67, variable expression of CD27, and high levels of IgM and IL-6 receptor. Furthermore, the proportion of infected cells showing a blasting phenotype increased upon addition of exogenous IL-6. Our data lead us to propose that oral transmission of KSHV involves the latent infection of a subset of tonsillar IgMl-expressing B cells, which then proliferate as they acquire the plasmablast phenotype characteristic of MCD.

Authors

Lynn M. Hassman, Thomas J. Ellison, Dean H. Kedes

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Figure 1

Levels of viral genome and PAN transcript associated with human tonsillar B cells following ex vivo inoculation with KSHV.

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Levels of viral genome and PAN transcript associated with human tonsilla...
Tonsillar B cells from 6 individual donors, represented by each symbol, were exposed to KSHV for 2 hours and washed extensively, and genomic DNA was purified at the indicated times. (A) Left panel: Relative levels of genomic KSHV DNA normalized to GAPDH for each time point were determined by qPCR and represent the approximate copies of ORF73/10e3 cells. Right panel: Relative levels of internalized viral genomes for all of the donors at 2.5 hpi determined as in the left panel but after first treating KSHV-exposed samples with trypsin to discriminate KSHV entry from surface binding. Values for each donor are mean of triplicate PCR reactions. Horizontal bars represent mean values for all donors at each time point. Dashed line represents mean level of background signal detected in 2.5-hour cultures unexposed to KSHV. (B) Levels of viral PAN RNA were similarly quantified and normalized to GAPDH using qRT-PCR from total RNA. Values represent the approximate number of PAN transcripts per 37.5 ng of input RNA. Open squares and filled triangles correspond to those in A; light and dark gray circles in B represent results of separate experiments on samples from the same tonsil donor. Horizontal bars represent mean values for all donors at each time point. Signal was below detection in no-RT reactions or uninfected samples (not shown).