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. 2000 Apr;74(7):3205-16.
doi: 10.1128/jvi.74.7.3205-3216.2000.

Pathogenesis of primary R5 human immunodeficiency virus type 1 clones in SCID-hu mice

R M Scoggins  1 , J R Taylor Jr, J Patrie, A B van't Wout, H Schuitemaker, D Camerini
Affiliations

Pathogenesis of primary R5 human immunodeficiency virus type 1 clones in SCID-hu mice

R M Scoggins et al. J Virol. 2000 Apr.

Abstract

We studied the replication and cytopathicity in SCID-hu mice of R5 human immunodeficiency virus type 1 (HIV-1) biological clones from early and late stages of infection of three patients who never developed MT-2 cell syncytium-inducing (SI; R5X4 or X4) viruses. Several of the late-stage non-MT-2 cell syncytium-inducing (NSI; R5) viruses from these patients depleted human CD4(+) thymocytes from SCID-hu mice. Earlier clones from the same patients did not deplete CD4(+) thymocytes from SCID-hu mice as well as later clones. We studied three R5 HIV-1 clones from patient ACH142 in greater detail. Two of these clones were obtained prior to the onset of AIDS; the third was obtained following the AIDS diagnosis. In GHOST cell infection assays, all three ACH142 R5 HIV-1 clones could infect GHOST cells expressing CCR5 but not GHOST cells expressing any of nine other HIV coreceptors tested. Furthermore, these patient clones efficiently infected stimulated peripheral blood mononuclear cells from a normal donor but not those from a homozygous CCR5Delta32 individual. Statistical analyses of data obtained from infection of SCID-hu mice with patient ACH142 R5 clones revealed that only the AIDS-associated clone significantly depleted CD4(+) thymocytes from SCID-hu mice. This clone also replicated to higher levels in SCID-hu mice than the two earlier clones, and a significant correlation between viral replication and CD4(+) thymocyte depletion was observed. Our results indicate that an intrinsic property of AIDS-associated R5 patient clones causes their increased replication and cytopathic effects in SCID-hu mice and likely contributes to the development of AIDS in patients who harbor only R5 quasispecies of HIV-1.

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Figures

FIG. 1
FIG. 1
CD4 and CD8 expression of thymocytes isolated from two SCID-hu thymus/liver grafts infected with the R5-AIDS HIV-1 biological clone AMS198-4A2 and from one mock-infected thymus/liver graft. Cells were isolated 13 weeks postinfection and stained with fluorochrome-conjugated CD4 and CD8 MAbs. Expression of the corresponding cell surface antigens was measured by flow cytometry. Percentages of CD4 SP CD8 SP and CD4 CD8 DP cells are shown.
FIG. 2
FIG. 2
HIV-1 capsid (p24) production by PBMC infected with HIV-1 ACH142-8G9, ACH142-32D2, ACH142-*E11, or NL4-3, at an MOI of 0.01. PBMC were isolated from a CCR5+ homozygous donor (A) and a CCR5Δ32 homozygous donor (B). p24 concentration was measured on supernatants collected on days 3, 6, 9, and 12 postinfection using a commercial ELISA kit (NEN Life Science Products).
FIG. 3
FIG. 3
GFP expression of CCR5-expressing GHOST cells infected with HIV-1 clone ACH142-8G9, ACH142-32D2, ACH142-*E11, NL4-3, or JR-CSF. GHOST cells were infected with 0.5 ml of viral stocks in the presence of Polybrene (4 μg/ml). Forty-eight hours after infection, cells were harvested and fixed, and GFP expression was measured by flow cytometry with a FACScan.
FIG. 4
FIG. 4
Representative CD4 and CD8 expression on thymocytes from thymus/liver grafts 3, 6, and 12 weeks after infection with HIV-1 ACH142-8G9, ACH142-32D2, or ACH142-*E11. Thymus/liver grafts were biopsied, a single-cell suspension was made, and the cells were stained with fluorochrome-conjugated CD4 and CD8 MAbs. Expression of the corresponding cell surface antigens was measured by flow cytometry. The data are representative of those summarized in Table 3.
FIG. 5
FIG. 5
Plot comparing percent CD4 CD8 DP thymocytes (A) or CD4 SP/CD8 SP thymocyte ratio (B) at 6 weeks after infection of thymus/liver grafts infected with HIV-1 ACH142-8G9, ACH142-32D2, ACH142-*E11, or NL4-3 or mock infected.
FIG. 6
FIG. 6
HIV-1 DNA copies per 105 thymus/liver cells in infected thymus liver grafts derived from SCID-hu mice, determined by quantitative PCR. Genomic DNA was isolated and subjected to 22 cycles of quantitative PCR using a primer pair specific for HIV-1 DNA and a primer pair specific for the human β-globin gene. In each case, one primer was end labeled with 32P. PCR products were resolved on a 6% polyacrylamide gel. Quantitation was performed by comparison to standard curves of the PCR products of known amounts of both HIV-1 plasmid DNA and human genomic DNA, using a PhosphorImager.
FIG. 7
FIG. 7
Scatter plot of percent CD4 CD8 DP thymocytes (A) or CD4 SP/CD8 SP thymocyte ratio (B) versus HIV-1 DNA copies per 105 thymus/liver cells recovered 6 weeks after infection with the indicated HIV-1 clones. A nonlinear regression was determined and plotted using a four-parameter logistic equation and Prism software (GraphPad Software).
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