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New Research Finds HIV causes rapid aging in key infection-fighting cells: "Successful ART treatment does not fully reconstitute the CD31-CD4+ naïve T-cell subset"- pdf attached
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"in individuals infected with HIV-1, these cells underwent unexpectedly rapid aging - the equivalent of 20 to 30 years of aging within three years of infection. They also found that the number of CD31- T-cells, which are more quickly pulled into the fight against new pathogens, had fallen drastically......The researchers also investigated whether appropriate treatment could reverse this aging effect. They examined cells from HIV-positive individuals who had been on antiretroviral therapy for two years and whose therapy had successfully kept HIV-1 under control. They found that while the therapy kept their viral loads at undetectable levels, it did not entirely restore their immune systems, suggesting a reason why younger HIV-positive people still become ill with conditions more common to older people....our results help to explain some of the clinical observations that have been documented in HIV-infected people and emphasize the need for developing therapeutic approaches directed at improving the naive immune cell compartment"
The Dual Impact of HIV-1 Infection and Aging on Naïve CD4+ T-Cells: Additive and Distinct Patterns of Impairment Tammy M Rickabaugh, Ryan D Kilpatrick, Lance E Hultin, Patricia M Hultin, Mary Ann Hausner, Catherine A Sugar, Keri N Althoff, Joseph B Margolick, Charles R Rinaldo, Roger Detels, John Phair, Rita B Effros, Beth D Jamieson Impact of HIV-1 Infection and Aging on Naïve CD4 + T ... The Dual Impact of HIV-1 Infection and Aging on Naïve CD4+ T-Cells: Additive and Distinct Patterns of Impairment ... Effects of HIV-1 and Aging on Naïve CD4 + T-Cells PLoS ONE: Research Article, published 26 Jan 2011 10.1371/journal.pone.0016459
"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."
"HIV-1 infection also exerts a distinct negative effect on CD31-CD4+ T-cell number that is not seen with aging.......Loss of the CD31-CD4+ naïve T-cells subset, and failure of this subset to reconstitute, is likely to have long-term deleterious effects on the immune response to neoantigens in individuals treated with ART."
"Our results lead us to hypothesize that the additive effects of HIV-1 and aging on the CD31+CD4+ naïve T-cell subset, and the HIV-1 associated loss of the CD31- naïve CD4+ T-cells, contribute to accelerated HIV-1 disease progression in HIV-1-infected adults over the age of 50 [66], [67], and to the decreased response to anti-retroviral therapy in older HIV-1-infected adults [68], [69]. Moreover, there is accumulating evidence that ART-treated HIV-1-infected individuals clinically present with malignancies and infectious diseases more consistent with older SN adults [3]-[5], [70], [71]. Further elucidation of the mechanisms contributing to naïve CD4+ T-cell loss, telomere shortening, and delayed or arrested reconstitution of the CD31- naïve CD4+ T-cell subset may lead to the identification of novel therapeutic targets to enhance the immune response against HIV-1 and other pathogens, as well as strategies to retard the formation of neoplasms in ART-treated individuals."
"Our results demonstrate that the negative effects of HIV-1 infection and aging on naïve CD4+ T-cells are predominantly additive. However, HIV-1 infection also exerts a distinct negative effect on CD31-CD4+ T-cell number that is not seen with aging. Together, these results suggest a role for impaired helper T-cell responses, particularly to neoantigens, in the more rapid progression to AIDS observed in individuals 50 years of age or older......Since telomere length is associated with proliferative capacity, those naïve CD4+ T-cells still present after the initial HIV-1 infection would be unlikely to generate, or support, robust CD8+ T- and B-cell responses to HIV-1 or opportunistic infections. Not only would HIV-1 be allowed to replicate more rapidly in older adults, but there would also be less CD4+ T-cell reserves, due to the reduced numbers of CD4+ T-cells and their progeny. These effects would undoubtedly contribute to the more rapid progression to AIDS."
"Successful ART treatment does not fully reconstitute the CD31-CD4+ naïve T-cell subset"
"the effect of HIV-1 infection on CD31+CD4+ T-cell numbers appears to be additive to the effects of aging alone.....HIV-1 infection, but not age, was associated with a significant reduction of CD31-CD4+ naïve T-cells......The number of CD31-CD4+ naïve T-cells was 2.9 times lower in young SP participants than in young SN participants (p = 0.0070). This is in contrast to a 1.8- and a 1.9-fold difference in the numbers of CD31+CD4+ naïve and CD45RA- effector/memory T-cell subsets, respectively, between the same two groups of participants (Figure 1B). There was also a larger difference in CD31-CD4+ naïve T-cell number between older SN and older SP participants than in the CD31+CD4+ subset for these two groups, 2.5-fold versus 2.0-fold difference respectively, but it did not reach significance. Whereas the loss of either naïve CD4+ T-cell subsets is likely to be detrimental to the host, our previous study has shown that the CD45RA+CD31- subset is particularly important for maintaining the naïve CD4+ T-cell pool during aging.....HIV-1 infection was associated with significant telomere shortening within both subsets of naïve CD4+ T-cells across both age groups (younger: p = 0.0004 for CD31+, p = 0.0096 for CD31-; older: p<0.0002 for both naive subsets)......the level of CD31 expression was, in fact, strongly associated with TREC number in CD31+CD4+ naive T-cells. The CD31+brightCD4+ naïve T-cells showed an average of 80% more TREC than were found within total CD31+CD4+ naïve T-cells, while the CD31+dimCD4+ naïve T-cell subset showed an average of 60% fewer TREC (p = 0.0088)."
"in individuals infected with HIV-1, these cells underwent unexpectedly rapid aging - the equivalent of 20 to 30 years of aging within three years of infection. They also found that the number of CD31- T-cells, which are more quickly pulled into the fight against new pathogens, had fallen drastically......The researchers also investigated whether appropriate treatment could reverse this aging effect. They examined cells from HIV-positive individuals who had been on antiretroviral therapy for two years and whose therapy had successfully kept HIV-1 under control. They found that while the therapy kept their viral loads at undetectable levels, it did not entirely restore their immune systems, suggesting a reason why younger HIV-positive people still become ill with conditions more common to older people......Taken together, our results help to explain some of the clinical observations that have been documented in HIV-infected people and emphasize the need for developing therapeutic approaches directed at improving the naive immune cell compartment," said senior investigator Beth D. Jamieson, an associate professor of medicine in the division of hematology and oncology at the David Geffen School of Medicine at UCLA"
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The Dual Impact of HIV-1 Infection and Aging on Naïve CD4+ T-Cells: Additive and Distinct Patterns of Impairment
Tammy M. Rickabaugh1, Ryan D. Kilpatrick1,7, Lance E. Hultin1, Patricia M. Hultin7, Mary Ann Hausner1, Catherine A. Sugar3, Keri N. Althoff4, Joseph B. Margolick5, Charles R. Rinaldo6, Roger Detels7, John Phair8, Rita B. Effros1,2, Beth D. Jamieson1*
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.
Introduction
The lifespan of an HIV-1-infected North American or European individual is shortened by an average of 10 years, despite antiretroviral therapy (ART) [1]. Many of the causes of morbidity and mortality in these individuals are similar to those more commonly observed in uninfected older adults (50-65 years of age) and the elderly (>65 years of age), and include frailty [2], non-Hodgkin's lymphoma [3], anal and cervical carcinomas [4], [5], osteoporosis [6], [7], liver [8]-[10] and renal impairment [11], cardiovascular disease [12], [13], diabetes [14] and hypertension [14], [15]. The diminished lifespan and higher prevalence of these diseases in HIV-1-infected individuals, in comparison to age-matched uninfected controls, has led to the theory that HIV-1 infection causes accelerated aging in multiple organ systems. As it is not clear whether HIV-1 contributes to age-inappropriate clinical manifestations through mechanisms distinct from aging, a better understanding of the effects of HIV-1 infection and aging on various organ systems is essential to future treatment of HIV-1-infected individuals.
Survival time for HIV-1-infected adults both pre-and post-ART is closely correlated with CD4+ T-cell counts. The life-expectancy of an untreated HIV-1-infected individual with 200 CD4+ T-cells/mm3 is approximately one to two years [16]. An ART-treated, 20-year-old adult with a CD4+ T-cell count under 200 cells/mm3 at ART initiation is predicted to survive 32 years, compared to 50 years for an age-matched individual who initiates ART at a higher CD4+ T-cell count [16]. An increased risk for frailty is also associated with decreased CD4+ T-cell counts pre- and post-ART initiation, as is the risk for non-Hodgkin lymphoma [2], [3]. Poor CD4+ T-cell recovery upon initiation of ART is also correlated with an increased risk for both AIDS and non-AIDS diseases [17], emphasizing the important role of the CD4+ T-cell compartment in maintaining good health.
Although HIV-1 infection of naïve CD4+ T-cells occurs at low frequency in comparison to that of activated effector/memory CD4+ T-cells, HIV-1 infection is associated with quantitative and qualitative changes within the naïve CD4+ T-cell compartment in both children and adults [18]-[21]. In HIV-1-infected adults, a loss of naïve CD4+ T-cells precedes the loss of T-cell homeostasis and progression to AIDS [20], and inverted naïve to effector/memory ratios are not always restored upon administration of ART [17], [22]. Since reconstitution of the naïve T-cell compartment contributes to reconstitution of overall CD4+ T-cell counts, a continued deficit in naïve CD4+ T-cell numbers would have downstream implications for the effector/memory compartment. In addition, functional defects, such as diminished antigen-specific proliferative responses [23], persist in the naïve CD4+ T-cell compartment, despite treatment. As naïve CD4+ T-cell proliferative responses post-ART predict immune responses to immunization with neoantigens [24], it is possible that an impaired naïve CD4+ T-cell compartment may contribute to the clinical observations regarding poor health and age-associated pathologies post-ART.
Aging, in the absence of HIV infection, is also associated with quantitative and qualitative changes within the naïve CD4+ T-cell compartment [25]-[27]. Decreased numbers of recent thymic emigrants (RTE), shortened telomeres, hyporesponsiveness to stimulation, decreased proliferative capacity, reduced IL-2 production, alterations in signal transduction and changes in cell surface phenotype [26]-[28] have all been reported. These changes likely contribute to the poor response to vaccines and increased susceptibility to infectious diseases and neoplasms reported for older adults [29]-[31] and possibly also contribute to the more rapid disease progression in HIV-1-infected individuals over 50 years of age.
Many factors, including co-infections and toxicity, or negative side effects, of antiretroviral drugs, are likely to contribute to the decreased lifespan and increased morbidities seen in HIV-1-infected individuals. In the current study, we address the question of whether HIV-1 infection accelerates the aging process by characterizing the effects of HIV-1 infection and aging on the naïve CD4+ T-cell compartment. We investigate whether the negative effects of HIV-1 infection and aging on the naïve CD4+ T-cell compartment are additive or interactive. To this end, we subdivided the naïve CD4+ T-cell compartment into two biologically disparate subsets based on the surface expression of PECAM-1 (CD31), which distinguishes TREC high (CD31+) naïve CD4+ T-cells from their proliferative progeny, the TREC low (CD31-) naïve CD4+ T-cell subset [27], [32]. We had previously shown that aging is associated with changes in the relative proportions of these two subsets and with telomere shortening within cells from both subsets [27]. Therefore, in the current study we compared the two naïve CD4+ T-cell subsets within younger (20-32 years old) and older (39-58 years old) HIV-1 seropositive (SP) ART-naïve adults relatively early in infection, as well as with HIV-1 seronegative (SN) age-matched adults. Our results suggest that the effects of HIV-1 infection and aging on the the naïve CD4+ T-cell compartment are both additive and distinct, and that HIV-1 induced impairments are not fully restored to an age-appropriate status by ART.
Participants
Cross-sectional study. Twenty-eight HIV-1 SN participants aged 19-30 years, nineteen SN participants aged 47-60 years, nine HIV-1 SP participants aged 20-32 years, and ten SP participants aged 39-58 years. All SP participants were within 1-3 years of infection by self-report and were treatment-naïve.
Longitudinal study. From the Multicenter AIDS Cohort Study (MACS), a study of the natural and treated history of HIV-1 infection in men who have sex with men [33], [34], we selected ten SP men who initiated ART while enrolled in the MACS. ART is self-reported during the semi-annual MACS study visits. Selection criteria included the following characteristics at the visit prior to initiating ART: an age of 40-50 years, an absolute T-cell count >250 cells/mm3 of blood, a viral load >50 copies/ml (90% of those selected had >5000 copies/ml), and a successful response to treatment, defined by a viral load of <400 RNA copies/ml approximately one year post-ART (90% of those selected had a viral load of <50 RNA copies/ml). The average increase in absolute CD4+ T-cell count one year post-ART was 147 cells/mm3. Ten SN MACS participants, age-matched (within 6 months to a year of age) to the pre-ART donor visit, were selected as controls. We analyzed cryopreserved PBMC provided by these men at 6 months to one year prior to ART initiation and at 1 and 2 years after that time.
Results
Preferential loss of CD31- CD4+ naïve T-cells during HIV-1 infection
We first quantified the percentage and absolute numbers of CD31+CD45RA+CD27+CD3+CD4+ (CD31+CD4+ naïve) and CD31-CD45RA+CD27+ CD3+CD4+ (CD31-CD4+ naïve) T-cells in the peripheral blood of young (20-32 years of age) and older (39-58 years of age) HIV-1-infected adults and age matched SN controls. As shown in Figure 1A, both older age and HIV-1 infection were associated with significantly decreased numbers of CD31+CD4+ T-cells. Young HIV-1-infected individuals demonstrated an average of 139 fewer CD31+CD4+ T-cells/mm3 than SN age-matched controls (317 cells/mm3 versus 178 cells/mm3, respectively; p<0.0002), and in older SP individuals, HIV-1 was associated with an additional difference of 113 cells/mm3 in this subset beyond what was accounted for by aging alone (223 cells/mm3 versus 110 cells/mm3, respectively; p = 0.0004). Since there was no evidence of an interaction between age and HIV-1 infection (p = 0.6423), the effect of HIV-1 infection on CD31+CD4+ T-cell numbers appears to be additive to the effects of aging alone. Notably, the absolute number of CD31+ CD4+ naïve T-cells in young SP individuals closely matched those of SN participants who were 17 to 28 years their senior (Figure 1A).
HIV-1 infection, but not age, was associated with a significant reduction of CD31-CD4+ naïve T-cells. While younger SN individuals had an average of 178 cells/mm3 CD31-CD4+ naïve T-cells, a decrease of 117 cells/mm3 was observed in the younger SP individuals (p<0.0002). However, this effect was independent of age; younger and older SP men had similar absolute cell numbers (61 cells/mm3 and 75 cells/mm3, respectively; interaction p = 0.9455). Aging in the absence of HIV-1 infection was not significantly associated with a difference in the number of CD31-CD4+ naïve T-cells (an average of 178 cells/mm3 and 188 cells/mm3 in the younger and older groups respectively; interaction p = 0.7476), consistent with previous reports by our group [27] and others [45], demonstrating that HIV-1 infection is associated with negative effects on this subset that are distinct from aging.
Of note, the decrease in the absolute number of CD31-CD4+ naïve T-cells associated with HIV-1 infection was greater than the decrease observed for any other CD4+ T-cell phenotype tested (Figure 1B). The number of CD31-CD4+ naïve T-cells was 2.9 times lower in young SP participants than in young SN participants (p = 0.0070). This is in contrast to a 1.8- and a 1.9-fold difference in the numbers of CD31+CD4+ naïve and CD45RA- effector/memory T-cell subsets, respectively, between the same two groups of participants (Figure 1B). There was also a larger difference in CD31-CD4+ naïve T-cell number between older SN and older SP participants than in the CD31+CD4+ subset for these two groups, 2.5-fold versus 2.0-fold difference respectively, but it did not reach significance. Whereas the loss of either naïve CD4+ T-cell subsets is likely to be detrimental to the host, our previous study has shown that the CD45RA+CD31- subset is particularly important for maintaining the naïve CD4+ T-cell pool during aging [27].
HIV-1 infection is associated with shortened telomeres within naïve CD4+ T-cells
As shown in Figure 2, HIV-1 infection was associated with significant telomere shortening within both subsets of naïve CD4+ T-cells across both age groups (younger: p = 0.0004 for CD31+, p = 0.0096 for CD31-; older: p<0.0002 for both naive subsets). In agreement with our previous findings [27], aging was also associated with telomere shortening in both naïve CD4+ T-cell subsets, (younger vs. older SN: p = 0.0132 for CD31+ and p = 0.0018 for CD31- subsets; younger vs. older SP: p = 0.0036 for CD31+ and p<0.0002 for CD31-). There was no evidence of an interaction between age and HIV-1 infection in either naïve subset (interaction p = 0.9564 for CD31+ and p = 0.5492 for CD31- CD4+ naïve T-cell subsets) indicating that, although age and HIV-1 serostatus each contribute to telomere shortening, these effects appear to be additive.
CD31 expression correlates with TREC number in CD31+CD4+ naive T-cells
Due to the challenge of obtaining the quantity of CD31+ CD4+ naïve T-cells from both older adults and HIV-1-infected individuals required for TREC and telomere analysis, we investigated alternative methods to determine the proliferative history of CD31+CD4+ naïve T-cells for use in the experiments described below. Based on the observation that CD31 is eventually lost from the surface of CD31+ naïve CD4+ T-cells [32], [46], we investigated whether the relative fluorescence intensity of CD31 on CD31+CD4+ naïve T-cells was associated with differing levels of TREC content. PBMC from five seronegative participants were sorted into three subsets based on the relative number of CD31 molecules on the surface of the cells: CD31+brightCD4+, CD31+dimCD4+, and total CD31+CD4+ (Figure 3A). As shown in Figure 3B, the level of CD31 expression was, in fact, strongly associated with TREC number in CD31+CD4+ naive T-cells. The CD31+brightCD4+ naïve T-cells showed an average of 80% more TREC than were found within total CD31+CD4+ naïve T-cells, while the CD31+dimCD4+ naïve T-cell subset showed an average of 60% fewer TREC (p = 0.0088). These data suggest that CD31+brightCD4+ naive T-cells, as compared to CD31+dimCD4+ naïve T-cells, are more highly enriched in RTE that have not undergone proliferation. Thus, relative fluorescence intensity can be used as an indirect measure of proliferative history within RTE in situations where cell numbers are limiting.
Successful ART treatment does not fully reconstitute the CD31-CD4+ naïve T-cell subset
Since we determined that HIV-1 infection has detrimental effects on the absolute numbers of both naïve CD4+ T-cell subsets, we evaluated whether ART was able to restore these subsets to age-appropriate levels. Using the CD31 marker as an indicator of emigration of naïve cells from the thymus, we performed a longitudinal study on cryopreserved PBMC obtained from MACS participants before, and after, initiation of ART. One year post-ART, the absolute number of CD31+brightCD4+ naïve T-cells significantly increased from an average of 60 cells/mm3 to an average of 180 cells/mm3 (Figure 4, p = 0.0013). In fact, 2 years post-ART there was no significant difference in cell number between SP men and their SN age-matched controls (p = 0.4904).
Since CD31+brightCD4+ naïve T-cells appear to be enriched for non-proliferated RTE (Figure 3B), the data in Figure 4 are consistent with previous reports of measurable thymic emigration after only one year of ART [43], [47] and continuing after that time. Interestingly, although there was no significant difference between the CD31+CD4+ naïve T-cell subset and SN controls after 2 years of ART (p = 0.2670), the absolute number of CD31-CD4+ naïve T-cells at the same time point showed no significant increase (Figure 5). The ultimate outcome, therefore, is a significant difference in CD31-CD4+ T-cell numbers between SP and SN age-matched controls two years post-ART (p = 0.0022).
Discussion
Our results demonstrate that the negative effects of HIV-1 infection and aging on naïve CD4+ T-cells are predominantly additive. However, HIV-1 infection also exerts a distinct negative effect on CD31-CD4+ T-cell number that is not seen with aging. Together, these results suggest a role for impaired helper T-cell responses, particularly to neoantigens, in the more rapid progression to AIDS observed in individuals 50 years of age or older.
Older SN individuals demonstrate diminished absolute numbers of CD31+CD4+ naïve T-cells and shortened telomeres within those subsets [27]. Both of these deficits undoubtedly contribute to the well-documented reduced responses to vaccines and other neoantigens well documented in older adults [29]-[31]. Indeed, initial HIV-1 infection of older individuals is likely to be met with a diminished CD4+ T-cell response, which, in turn, would affect both B- and T-cell responses to HIV-1, allowing the virus to spread quickly. Moreover, our results suggest that once infection is established, the naïve CD4+ T-cell compartment is further impaired by an accelerated loss of CD31+CD4+ naïve T-cells, and a loss of CD31- naïve T-cells that is not normally associated with SN adults in their fifties [27]. Compounding this cell loss is the additive effect of aging and HIV-1 infection on telomere shortening within both subsets of naïve CD4+ T-cells (Figure 2). Since telomere length is associated with proliferative capacity, those naïve CD4+ T-cells still present after the initial HIV-1 infection would be unlikely to generate, or support, robust CD8+ T- and B-cell responses to HIV-1 or opportunistic infections. Not only would HIV-1 be allowed to replicate more rapidly in older adults, but there would also be less CD4+ T-cell reserves, due to the reduced numbers of CD4+ T-cells and their progeny. These effects would undoubtedly contribute to the more rapid progression to AIDS.
The underlying mechanisms for the accelerated loss of the naïve CD4+ T-cells during HIV-1 disease most likely include increased recruitment into the effector/memory pool as well as direct infection with HIV-1 and eventual cell death. The latter is supported by previous reports showing that naïve CD4+ T-cells can be infected by HIV-1 [48], [49]. Our own unpublished observation that the CD31- naïve subset harbors HIV-1 in vivo in ART-naïve individuals (data not shown) may also partially explain why this subset declines in HIV-1-infected, but not age matched SN, individuals.
Loss of the CD31-CD4+ naïve T-cells subset, and failure of this subset to reconstitute, is likely to have long-term deleterious effects on the immune response to neoantigens in individuals treated with ART. We previously demonstrated that homeostasis of the naïve CD4+ T-cell subset in adults is largely maintained by proliferation of CD31-CD4+ naïve T-cells and not by recent thymic emigrants (CD31+CD4+ naïve T-cells) [27]. Furthermore, the presence of clonal expansions of naïve CD31-CD4+ T-cells in HIV-1-infected individuals, similar to those in the effector/memory CD4+ T-cell pool, suggest that it is the CD31-CD4+ T-cells which are recruited into the effector/memory pool in response to antigen (Kilpatrick, et al. unpublished results). Failure of this subset to fully reconstitute in the older SP group within 2 years after ART initiation (Figure 5) suggests that, despite reconstitution of the CD31+ naïve CD4+ T-cell compartment shown by us and others [50], fewer naïve CD4+ T-cells are available for recruitment into the effector/memory pool, as compared to uninfected peers. The lack of reconstitution could be due to a true "block" in differentiation from CD31+CD4+ to CD31-CD4+, or by CD31-CD4+ T-cells being rapidly recruited into the effector/memory pool to fill the "immunologic space". These two mechanisms would have very different implications for the health of ART-treated individuals and could suggest very different therapeutic strategies for enhancing overall CD4+ T-cell reconstitution.
To our knowledge, this is the first study demonstrating shortened telomere lengths in naïve CD4+ T-cells sorted from HIV-1-infected participants and subdivided by the surface markers CD31 and CD27. While these data accord with the majority of studies examining telomere length in total, or effector/memory, CD4+ T-cell populations during HIV-1 infection [51]-[54], they do conflict with the findings of Miedema and colleagues [55]. Using CD45RA and CD45RO to sort naïve and effector/memory cells, Wolthers et al. [55] failed to find evidence of telomere shortening in either CD4+ T-cell subset during HIV-1 infection. The difference between the Wolthers et al. results and our own study may be due, at least in part, to a difference in flow cytometry gating strategies. For example, expression of CD45RA is not a stringent phenotype for naïve CD4+ T-cells; other cell types, such as terminally differentiated cells would be included in this population. In addition, in contrast to the Wolthers, et al. study, our own HIV-1 infected cohort was age-matched to the SN controls to avoid the confounding effects of aging on the telomere lengths within our HIV-1 infected individuals.
Our observation of telomere shortening in even the least differentiated naïve CD4+ T-cells (i.e.,the CD31+CD4+ T-cell subset) in response to both aging and HIV-1 infection (Figure 2) is intriguing and implies possible telomere shortening in an earlier progenitor cell. In support of this hypothesis, both HIV-1 infection and aging have been linked to decreased telomerase activity within hematopoietic progenitors [56]-[58]. Alternatively, it is possible that extensive cellular proliferation, oxidative stress, or a deficit in telomerase directly contributes to telomere shortening within the peripheral CD4+ naïve T-cells. Our data do not support a major role for extensive proliferation of the naïve T-cells alone in the telomere shortening. Based on the correlation between TREC levels and CD31 expression (Figure 3B), we performed a cross-sectional analysis of CD31 expression levels on naïve CD31+ CD4+ T-cells from older vs. younger SN and HIV-1 infected age matched adults, but did not find a diminution of the CD31 relative fluorescence intensity that would be consistent with extensive cellular turnover during aging or HIV-1 infection (data not shown). In addition, in a previous cross-sectional study of the naïve CD4+ T-cell compartment during aging, we observed a significant decline in TREC in naïve CD31+ T cells over a 1-2 decade period [27]. However, the T-cell expansion was subtle and unlikely to completely account for the significant telomere shortening associated with aging, suggesting that additional factors, possibly oxidative stress, may be involved. Indeed, oxidative stress is associated with chronic inflammation, which is well documented in both HIV-1 infection [59] and aging [60]-[63] and is known to accelerate telomere shortening in both the presence and absence of proliferation [64], [65].
Our results lead us to hypothesize that the additive effects of HIV-1 and aging on the CD31+CD4+ naïve T-cell subset, and the HIV-1 associated loss of the CD31- naïve CD4+ T-cells, contribute to accelerated HIV-1 disease progression in HIV-1-infected adults over the age of 50 [66], [67], and to the decreased response to anti-retroviral therapy in older HIV-1-infected adults [68], [69]. Moreover, there is accumulating evidence that ART-treated HIV-1-infected individuals clinically present with malignancies and infectious diseases more consistent with older SN adults [3]-[5], [70], [71]. Further elucidation of the mechanisms contributing to naïve CD4+ T-cell loss, telomere shortening, and delayed or arrested reconstitution of the CD31- naïve CD4+ T-cell subset may lead to the identification of novel therapeutic targets to enhance the immune response against HIV-1 and other pathogens, as well as strategies to retard the formation of neoplasms in ART-treated individuals.
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