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Persistent resistance to HIV-1 infection in CD4 T cells from exposed uninfected Vietnamese individuals is mediated by entry and post-entry blocks.

Sáez-Cirión A, Versmisse P, Truong LX, Chakrabarti LA, Carpentier W, Barré-Sinoussi F, Scott-Algara D, Pancino G - Retrovirology (2006)

Bottom Line: In two cases this resistance was associated with low CCR5 surface expression, which was itself associated with heterozygous CCR5 mutations.The restriction was not overcome by a high viral inoculum, suggesting that it was not mediated by a saturable inhibitory factor.Various constitutive mechanisms of CD4 T cell resistance to HIV-1 infection, affecting entry or post-entry steps of viral replication, are associated with resistance to HIV-1 in subjects who remain uninfected despite long-term high-risk behavior.

View Article: PubMed Central - HTML - PubMed

Affiliation: Unité de Régulation des Infections Rétrovirales, Institut Pasteur, Paris, France. asiersc@pasteur.fr

ABSTRACT

Background: We have previously reported that CD4 T cells from some exposed uninfected (EU) Vietnamese intravenous drug users are relatively resistant to HIV infection in vitro. Here, we further characterized the restriction of viral replication in CD4 T cells from five EUs and assessed its persistence in serial samples.

Results: CD4 T cells and/or PBMC sampled during a period of between 2 and 6 years were challenged with replication-competent HIV-1 and other retroviral particles pseudotyped with envelope proteins of various tropisms. CCR5 expression and function in resistant CD4 T cells was evaluated. The step at which HIV-1 replication is restricted was investigated by real-time PCR quantification of HIV-1 reverse transcripts. We identified three patterns of durable HIV-1 restriction in EU CD4 T cells. CD4 T cells from four of the five EU subjects were resistant to HIV-1 R5 infection. In two cases this resistance was associated with low CCR5 surface expression, which was itself associated with heterozygous CCR5 mutations. In the other two cases, CD4 T cells were resistant to HIV-1 R5 infection despite normal CCR5 expression and signaling function, and normal beta-chemokine secretion upon CD4 T cell activation. Instead, restriction appeared to be due to enhanced CD4 T cell sensitivity to beta-chemokines in these two subjects. In the fifth EU subject the restriction involved post-entry steps of viral replication and affected not only HIV-1 but also other lentiviruses. The restriction was not overcome by a high viral inoculum, suggesting that it was not mediated by a saturable inhibitory factor.

Conclusion: Various constitutive mechanisms of CD4 T cell resistance to HIV-1 infection, affecting entry or post-entry steps of viral replication, are associated with resistance to HIV-1 in subjects who remain uninfected despite long-term high-risk behavior.

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Role of β-chemokines in HIV-1 restriction in CD4 T cells from subjects W278 and B195. A. CD4 T cells from subjects W278 and B195 were challenged with the HIV-BaL pseudotype in the absence (white bars) or presence of a combination of neutralizing anti-RANTES (5 μg/ml), anti-MIP1α (15 μg/ml) and anti-MIP1β (25 μg/ml) (R&D systems, France) mAbs (black bars) or with an isotype control antibody (45 μg/ml) (patterned bars). The antibodies were added 30 minutes before challenge and maintained throughout the time course of infection. Results are expressed as relative luciferase activity, compared to the maximal activity found in the presence of the neutralizing anti-β-chemokines in each case, and are the mean of three independent infections ± standard deviation. ** Significant difference (P < 0.001 and P = 0.008 for W278 and B195, respectively, independent sample t-test). B. Challenge with the HIV-BaL pseudotype (n = 3, mean ± SD) of CD4 T cells from EUs W278 and B195, stimulated with PHA three days before (white bars) or two hours after (black bars) challenge. Luciferase activity in cell lysates from a representative control challenged in the same conditions was attributed a value of 100%. C. Sensitivity of CD4 T cells to recombinant chemokines. Non mitogen-stimulated CD4 T cells from EU B195 (filled circles) and one control (open squares) were exposed to various concentrations of a mixture of the recombinant β-chemokines RANTES, MIP-1α and MIP-1β (R&D systems, France) for 30 minutes prior to and during infection. The mixtures contained the three chemokines at concentrations ranging from 500 ng to 2 ng each. Results (n = 3, mean ± SD) are expressed as luciferase activity per second in cell lysates. Points were fitted to a four-parameter logistic curve (r2 were 0.845 and 0.826 for B195 and control, respectively). Statistical analysis and curve-fitting were performed with Sigmaplot software (Systat Software, Inc, CA, USA).
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Figure 3: Role of β-chemokines in HIV-1 restriction in CD4 T cells from subjects W278 and B195. A. CD4 T cells from subjects W278 and B195 were challenged with the HIV-BaL pseudotype in the absence (white bars) or presence of a combination of neutralizing anti-RANTES (5 μg/ml), anti-MIP1α (15 μg/ml) and anti-MIP1β (25 μg/ml) (R&D systems, France) mAbs (black bars) or with an isotype control antibody (45 μg/ml) (patterned bars). The antibodies were added 30 minutes before challenge and maintained throughout the time course of infection. Results are expressed as relative luciferase activity, compared to the maximal activity found in the presence of the neutralizing anti-β-chemokines in each case, and are the mean of three independent infections ± standard deviation. ** Significant difference (P < 0.001 and P = 0.008 for W278 and B195, respectively, independent sample t-test). B. Challenge with the HIV-BaL pseudotype (n = 3, mean ± SD) of CD4 T cells from EUs W278 and B195, stimulated with PHA three days before (white bars) or two hours after (black bars) challenge. Luciferase activity in cell lysates from a representative control challenged in the same conditions was attributed a value of 100%. C. Sensitivity of CD4 T cells to recombinant chemokines. Non mitogen-stimulated CD4 T cells from EU B195 (filled circles) and one control (open squares) were exposed to various concentrations of a mixture of the recombinant β-chemokines RANTES, MIP-1α and MIP-1β (R&D systems, France) for 30 minutes prior to and during infection. The mixtures contained the three chemokines at concentrations ranging from 500 ng to 2 ng each. Results (n = 3, mean ± SD) are expressed as luciferase activity per second in cell lysates. Points were fitted to a four-parameter logistic curve (r2 were 0.845 and 0.826 for B195 and control, respectively). Statistical analysis and curve-fitting were performed with Sigmaplot software (Systat Software, Inc, CA, USA).

Mentions: R5 virus entry into CD4 T cells can be blocked by endogenously produced β-chemokines [20]. We therefore investigated whether the restriction of HIV-1 R5 replication in CD4 T cells from subjects W278 and B195 could be overcome by neutralizing monoclonal antibodies (MAbs) to RANTES, MIP1α and MIP1β. The addition of anti-β-chemokine mAbs, but not of irrelevant IgGs, strongly enhanced the infection of both W278 and B195 cells by the HIV-1 pseudotype, while no significant enhancement was observed in CCR5-wt control cells with similar CCR5 surface expression (fig. 3A). These results suggest that endogenously produced β-chemokines may be responsible for the inhibition of HIV-1 infection in CD4 T cells from subjects W278 and B195.


Persistent resistance to HIV-1 infection in CD4 T cells from exposed uninfected Vietnamese individuals is mediated by entry and post-entry blocks.

Sáez-Cirión A, Versmisse P, Truong LX, Chakrabarti LA, Carpentier W, Barré-Sinoussi F, Scott-Algara D, Pancino G - Retrovirology (2006)

Role of β-chemokines in HIV-1 restriction in CD4 T cells from subjects W278 and B195. A. CD4 T cells from subjects W278 and B195 were challenged with the HIV-BaL pseudotype in the absence (white bars) or presence of a combination of neutralizing anti-RANTES (5 μg/ml), anti-MIP1α (15 μg/ml) and anti-MIP1β (25 μg/ml) (R&D systems, France) mAbs (black bars) or with an isotype control antibody (45 μg/ml) (patterned bars). The antibodies were added 30 minutes before challenge and maintained throughout the time course of infection. Results are expressed as relative luciferase activity, compared to the maximal activity found in the presence of the neutralizing anti-β-chemokines in each case, and are the mean of three independent infections ± standard deviation. ** Significant difference (P < 0.001 and P = 0.008 for W278 and B195, respectively, independent sample t-test). B. Challenge with the HIV-BaL pseudotype (n = 3, mean ± SD) of CD4 T cells from EUs W278 and B195, stimulated with PHA three days before (white bars) or two hours after (black bars) challenge. Luciferase activity in cell lysates from a representative control challenged in the same conditions was attributed a value of 100%. C. Sensitivity of CD4 T cells to recombinant chemokines. Non mitogen-stimulated CD4 T cells from EU B195 (filled circles) and one control (open squares) were exposed to various concentrations of a mixture of the recombinant β-chemokines RANTES, MIP-1α and MIP-1β (R&D systems, France) for 30 minutes prior to and during infection. The mixtures contained the three chemokines at concentrations ranging from 500 ng to 2 ng each. Results (n = 3, mean ± SD) are expressed as luciferase activity per second in cell lysates. Points were fitted to a four-parameter logistic curve (r2 were 0.845 and 0.826 for B195 and control, respectively). Statistical analysis and curve-fitting were performed with Sigmaplot software (Systat Software, Inc, CA, USA).
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
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Figure 3: Role of β-chemokines in HIV-1 restriction in CD4 T cells from subjects W278 and B195. A. CD4 T cells from subjects W278 and B195 were challenged with the HIV-BaL pseudotype in the absence (white bars) or presence of a combination of neutralizing anti-RANTES (5 μg/ml), anti-MIP1α (15 μg/ml) and anti-MIP1β (25 μg/ml) (R&D systems, France) mAbs (black bars) or with an isotype control antibody (45 μg/ml) (patterned bars). The antibodies were added 30 minutes before challenge and maintained throughout the time course of infection. Results are expressed as relative luciferase activity, compared to the maximal activity found in the presence of the neutralizing anti-β-chemokines in each case, and are the mean of three independent infections ± standard deviation. ** Significant difference (P < 0.001 and P = 0.008 for W278 and B195, respectively, independent sample t-test). B. Challenge with the HIV-BaL pseudotype (n = 3, mean ± SD) of CD4 T cells from EUs W278 and B195, stimulated with PHA three days before (white bars) or two hours after (black bars) challenge. Luciferase activity in cell lysates from a representative control challenged in the same conditions was attributed a value of 100%. C. Sensitivity of CD4 T cells to recombinant chemokines. Non mitogen-stimulated CD4 T cells from EU B195 (filled circles) and one control (open squares) were exposed to various concentrations of a mixture of the recombinant β-chemokines RANTES, MIP-1α and MIP-1β (R&D systems, France) for 30 minutes prior to and during infection. The mixtures contained the three chemokines at concentrations ranging from 500 ng to 2 ng each. Results (n = 3, mean ± SD) are expressed as luciferase activity per second in cell lysates. Points were fitted to a four-parameter logistic curve (r2 were 0.845 and 0.826 for B195 and control, respectively). Statistical analysis and curve-fitting were performed with Sigmaplot software (Systat Software, Inc, CA, USA).
Mentions: R5 virus entry into CD4 T cells can be blocked by endogenously produced β-chemokines [20]. We therefore investigated whether the restriction of HIV-1 R5 replication in CD4 T cells from subjects W278 and B195 could be overcome by neutralizing monoclonal antibodies (MAbs) to RANTES, MIP1α and MIP1β. The addition of anti-β-chemokine mAbs, but not of irrelevant IgGs, strongly enhanced the infection of both W278 and B195 cells by the HIV-1 pseudotype, while no significant enhancement was observed in CCR5-wt control cells with similar CCR5 surface expression (fig. 3A). These results suggest that endogenously produced β-chemokines may be responsible for the inhibition of HIV-1 infection in CD4 T cells from subjects W278 and B195.

Bottom Line: In two cases this resistance was associated with low CCR5 surface expression, which was itself associated with heterozygous CCR5 mutations.The restriction was not overcome by a high viral inoculum, suggesting that it was not mediated by a saturable inhibitory factor.Various constitutive mechanisms of CD4 T cell resistance to HIV-1 infection, affecting entry or post-entry steps of viral replication, are associated with resistance to HIV-1 in subjects who remain uninfected despite long-term high-risk behavior.

View Article: PubMed Central - HTML - PubMed

Affiliation: Unité de Régulation des Infections Rétrovirales, Institut Pasteur, Paris, France. asiersc@pasteur.fr

ABSTRACT

Background: We have previously reported that CD4 T cells from some exposed uninfected (EU) Vietnamese intravenous drug users are relatively resistant to HIV infection in vitro. Here, we further characterized the restriction of viral replication in CD4 T cells from five EUs and assessed its persistence in serial samples.

Results: CD4 T cells and/or PBMC sampled during a period of between 2 and 6 years were challenged with replication-competent HIV-1 and other retroviral particles pseudotyped with envelope proteins of various tropisms. CCR5 expression and function in resistant CD4 T cells was evaluated. The step at which HIV-1 replication is restricted was investigated by real-time PCR quantification of HIV-1 reverse transcripts. We identified three patterns of durable HIV-1 restriction in EU CD4 T cells. CD4 T cells from four of the five EU subjects were resistant to HIV-1 R5 infection. In two cases this resistance was associated with low CCR5 surface expression, which was itself associated with heterozygous CCR5 mutations. In the other two cases, CD4 T cells were resistant to HIV-1 R5 infection despite normal CCR5 expression and signaling function, and normal beta-chemokine secretion upon CD4 T cell activation. Instead, restriction appeared to be due to enhanced CD4 T cell sensitivity to beta-chemokines in these two subjects. In the fifth EU subject the restriction involved post-entry steps of viral replication and affected not only HIV-1 but also other lentiviruses. The restriction was not overcome by a high viral inoculum, suggesting that it was not mediated by a saturable inhibitory factor.

Conclusion: Various constitutive mechanisms of CD4 T cell resistance to HIV-1 infection, affecting entry or post-entry steps of viral replication, are associated with resistance to HIV-1 in subjects who remain uninfected despite long-term high-risk behavior.

Show MeSH
Related in: MedlinePlus