Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2014 Feb 25;111(8):3146-51.
doi: 10.1073/pnas.1318175111. Epub 2014 Feb 10.

Humanized-BLT mouse model of Kaposi's sarcoma-associated herpesvirus infection

Lin-Xu Wang  1 , Guobin Kang, Pankaj Kumar, Wuxun Lu, Yue Li, You Zhou, Qingsheng Li, Charles Wood
Affiliations

Humanized-BLT mouse model of Kaposi's sarcoma-associated herpesvirus infection

Lin-Xu Wang et al. Proc Natl Acad Sci U S A. .

Abstract

Lack of an effective small-animal model to study the Kaposi's sarcoma-associated herpesvirus (KSHV) infection in vivo has hampered studies on the pathogenesis and transmission of KSHV. The objective of our study was to determine whether the humanized BLT (bone marrow, liver, and thymus) mouse (hu-BLT) model generated from NOD/SCID/IL2rγ mice can be a useful model for studying KSHV infection. We have tested KSHV infection of hu-BLT mice via various routes of infection, including oral and intravaginal routes, to mimic natural routes of transmission, with recombinant KSHV over a 1- or 3-mo period. Infection was determined by measuring viral DNA, latent and lytic viral transcripts and antigens in various tissues by PCR, in situ hybridization, and immunohistochemical staining. KSHV DNA, as well as both latent and lytic viral transcripts and proteins, were detected in various tissues, via various routes of infection. Using double-labeled immune-fluorescence confocal microscopy, we found that KSHV can establish infection in human B cells and macrophages. Our results demonstrate that KSHV can establish a robust infection in the hu-BLT mice, via different routes of infection, including the oral mucosa which is the most common natural route of infection. This hu-BLT mouse not only will be a useful model for studying the pathogenesis of KSHV in vivo but can potentially be used to study the routes and spread of viral infection in the infected host.

Keywords: HHV-8; humanized mice; mucosa transmission.

PubMed Disclaimer

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.
Fig. 1.
Levels of human immune reconstitution in hu-BLT mice at 12 wk after surgery. (A) PBLs were incubated with antibodies and then analyzed by flow cytometry. Approximately 2 × 104 total live cells were evaluated for the coexpression of either hCD45, mCD45, hCD3, or hCD19. (B) Splenic single-cell suspensions were analyzed by multicolor flow cytometry. Approximately 5 × 104 total live cells were evaluated for the coexpression of either hCD45, mCD45, hCD3, or hCD19. (C) Skin tissues were analyzed by immunofluorescence using rabbit antibodies against hMHC-I (red) or hCD45 (red). DAPI (blue) was used as counterstain. (Scale bars, 20 μm.)
Fig. 2.
Fig. 2.
Determination of KSHV infection of the spleens of hu-BLT mice infected by KSHV orally at 4 wk after infection by flow cytometry. (A) Determination of KSHV infection efficiencies of the spleen tissues at 4 wk after infection (n = 6). (B) Splenic single-cell suspensions were measured by flow cytometry. Approximately 3 × 104 total live cells were evaluated for the coexpression of GFP and hMHC-I.
Fig. 3.
Fig. 3.
Expression of KSHV latent and lytic genes and proteins in the spleens of hu-BLT mice infected orally. (A) KSHV latent and lytic gene expression in the spleen of a hu-BLT mouse by ISH at 12 wk after infection. Probe for latent gene LANA was exposed for 8 d (a, LANA anti-sense probe; b, LANA sense probe), and 16 d for the lytic gene probes (c, K8.1 and gB antisense probes; d, K8.1 and gB sense probes). (B) KSHV latent and lytic proteins expression in the spleens at 4 wk after infection. The spleens were from infected (a, b, d, and e) or from PBS control mice (c and f). The splenic sections were incubated with antibodies to either LANA (a and c) or K8.1 (d and f) or mouse IgG (b and e). (Scale bars, 100 μm in larger panels, 25 μm in Insets.)
Fig. 4.
Fig. 4.
KSHV latent and lytic genes and proteins expression in the skins of hu-BLT mice inoculated orally by KSHV at 12 wk after infection. (A) KSHV latent and lytic gene expression in the skin of a hu-BLT mouse by ISH. Thin sections of the spleen were hybridized with 35S-labeled riboprobe specific for KSHV LANA gene after 8 d exposure (a, LANA antisense probe; b, LANA sense probe), and 16 d exposure for lytic genes (c, K8.1 and gB antisense probes; d, K8.1 and gB sense probes). (B) Detection of KSHV latent and lytic proteins expression by IHCS in the skins. The skins were collected from the infected (a, b, d, and e) or control PBS mice (c and f). Skin sections were incubated with antibodies to either LANA (a and c) or K8.1 (d and f) or mouse IgG (b and e). Arrows indicate LANA+ cells. (Scale bars, 100 μm in larger panels, 25 μm in Insets.)
Fig. 5.
Fig. 5.
Identification of KSHV-infected cell types in the spleens of hu-BLT mice at 4 wk after oral infection by flow cytometry. (A) KSHV infection efficiencies and infected cell populations in the spleen tissue at 4 wk after infection (n = 6). (B) Splenic single-cell suspensions were measured. The splenic single-cell suspensions were incubated with hCD45, hCD19, and hCD31 mouse monoclonal antibodies and rabbit anti-GFP antibody. Approximately 3 × 104 total live cells were evaluated using a combination of two antibodies.
Fig. 6.
Fig. 6.
Identification of KSHV-infected cell types in the spleens of hu-BLT mice inoculated orally using double-label IFA. (A) Identification of the cell types infected by KSHV latently (LANA+) in the spleen tissues. (B) Identification of the cells types infected by KSHV lytically (K8.1+) in the spleen tissues. Double-labeled IFA was performed using mouse antibodies to LANA (green) at 1:100 dilution or to K8.1 at 1:1,000 dilution (green), and rabbit monoclonal or polyclonal antibodies to human MHC-I, CD45, CD20, CD68, or CD31 (red). DAPI (blue) was used as counterstain. (Scale bars, 10 μm.)
Fig. 7.
Fig. 7.
Identification of the cell types infected by KSHV latently in the skins of hu-BLT mice inoculated orally using double-label IFA. Double-labeled IFA was performed using mouse antibody to LANA (green) at 1:500 dilution, and rabbit monoclonal or polyclonal antibodies to human MHC-I, CD45, CD68, or CD31 (red). DAPI (blue) was used as counterstain. (Scale bars, 10 μm.)
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Antman K, Chang Y. Kaposi’s sarcoma. N Engl J Med. 2000;342(14):1027–1038. - PubMed
    1. Fukumoto H, Kanno T, Hasegawa H, Katano H. Pathology of Kaposi’s sarcoma-associated herpesvirus infection. Front Microbiol. 2011;2:175. - PMC - PubMed
    1. Tamburro KM, et al. Vironome of Kaposi sarcoma associated herpesvirus-inflammatory cytokine syndrome in an AIDS patient reveals co-infection of human herpesvirus 8 and human herpesvirus 6A. Virology. 2012;433(1):220–225. - PMC - PubMed
    1. Picchio GR, et al. The KSHV/HHV8-infected BCBL-1 lymphoma line causes tumors in SCID mice but fails to transmit virus to a human peripheral blood mononuclear cell graft. Virology. 1997;238(1):22–29. - PubMed
    1. Foreman KE, et al. Injection of human herpesvirus-8 in human skin engrafted on SCID mice induces Kaposi’s sarcoma-like lesions. J Dermatol Sci. 2001;26(3):182–193. - PubMed

Publication types