The dual impact of HIV-1 infection and aging on naïve CD4 T-cells: additive and distinct patterns of impairment

Abstract

HIV-1-infected adults over the age of 50 years progress to AIDS more rapidly than adults in their twenties or thirties. In addition, HIV-1-infected individuals receiving antiretroviral therapy (ART) present with clinical diseases, such as various cancers and liver disease, more commonly seen in older uninfected adults. These observations suggest that HIV-1 infection in older persons can have detrimental immunological effects that are not completely reversed by ART. As naïve T-cells are critically important in responses to neoantigens, we first analyzed two subsets (CD45RA(+)CD31(+) and CD45RA(+)CD31(-)) within the naïve CD4(+) T-cell compartment in young (20-32 years old) and older (39-58 years old), ART-naïve, HIV-1 seropositive individuals within 1-3 years of infection and in age-matched seronegative controls. HIV-1 infection in the young cohort was associated with lower absolute numbers of, and shorter telomere lengths within, both CD45RA(+)CD31(+)CD4(+) and CD45RA(+)CD31(-)CD4(+) T-cell subsets in comparison to age-matched seronegative controls, changes that resembled seronegative individuals who were decades older. Longitudinal analysis provided evidence of thymic emigration and reconstitution of CD45RA(+)CD31(+)CD4(+) T-cells two years post-ART, but minimal reconstitution of the CD45RA(+)CD31(-)CD4(+) subset, which could impair de novo immune responses. For both ART-naïve and ART-treated HIV-1-infected adults, a renewable pool of thymic emigrants is necessary to maintain CD4(+) T-cell homeostasis. Overall, these results offer a partial explanation both for the faster disease progression of older adults and the observation that viral responders to ART present with clinical diseases associated with older adults.

Figure 1. ART-naïve seropositive individuals 1-3 years post-infection have significantly fewer naïve CD4⁺ T-cells than seronegative controls.

PBMC from each individual in our cross-sectional study were analyzed for the CD4⁺ naïve T-cell subsets, defined as CD45RA⁺CD27⁺ and either CD31⁺ or CD31^-, and for the CD4⁺ T-cell effector/memory subset, defined as CD45RA^-CD27^-, using flow cytometry. A.) The distribution of the absolute number of naïve and effector/memory CD4⁺ T-cells was determined for each serotype and age group. B.) The average absolute number of cells of each subset was determined for each serotype and age group. For each subset, the fold difference between the seronegative and seropositive age-matched groups is shown above the bars. The asterisks signify the following values, * p<0.05, ** p<0.01, ***p<0.005.

Figure 2. Telomeres are significantly shortened in naïve CD4⁺ T-cells within 1-3 years of HIV-1 infection.

PBMC from each individual in the cross-sectional study were sorted into CD31⁺CD4⁺and CD31^-CD4⁺ T-cell subsets. The sorted cells were then subjected to telomere length Real-Time PCR. Telomere length values are expressed as a percentage of telomere lengths within a human T-cell leukemia cell line, 1301 cells. Each symbol represents a sample from a single individual within the indicated serostatus and age group. The asterisks signify the following values, * p≤0.05, ** p≤0.01, *** p<0.005.

Figure 3. TREC content correlates with levels of CD31 expression on CD31⁺CD4⁺ naive T-cells.

Fresh PBMC from five HIV-1 seronegative individuals, ranging from 25-56 years of age, were sorted into total CD31⁺CD4⁺, CD31^+brightCD4⁺ and CD31^+dimCD4⁺ naïve T-cell subsets. A.) Pre-sort gates for determining the CD31^+brightCD4⁺, CD31^+dimCD4⁺, and CD31^-CD4⁺ naïve T-cell subsets are shown. The bottom three plots depict representative post-sort analysis of the subsets. B.) Genomic DNA was extracted from each subset and subjected to quantitative Real-Time PCR to quantify TREC content. The data is expressed as a percentage of TREC number in the indicated subset shown relative to TREC number in total CD31⁺CD4⁺ naïve T-cells. The asterisks signify the following value, ** p = 0.009.

Figure 4. Seropositive individuals two years post-ART have significantly greater numbers of CD31^+bright cells.

Cryopreserved PBMC from ten HIV-1 seropositive MACS participants collected at the indicated times pre- and post-ART were analyzed by flow cytometry for expression of CD4, CD45RA, and CD31. Absolute numbers of CD31^+brightCD4⁺ naïve T-cells were determined for each participant at the indicated time points. The data is shown in a box plot format where the top and bottom of the box represent the 25^th and 75^th percentile and middle band represents the 50^th percentile. The asterisks signify the following value, *** p<0.005.

Figure 5. Seropositive individuals demonstrate evidence of post-ART reconstitution in CD31⁺CD4⁺, but not CD31^-CD4⁺ T-cells.

Cryopreserved PBMC from ten seropositive MACS participants collected at the indicated time points pre- and post-ART, were analyzed for the CD31⁺ and CD31^- naïve CD4⁺ T-cell subsets by flow cytometry. Cryopreserved PBMC from ten age-matched seronegative participants in the MACS were used as controls. The data is shown in a box plot format where the top and bottom of the box represent the 25^th and 75^th percentile and middle band represents the 50^th percentile. The asterisks signify the following value, *** p<0.005.