To evaluate the possibility that the observed increases in CD4 cell counts were the result of selective drop-out of patients experiencing minimal CD4 T-cell gains, we compared the slopes of CD4 T-cell changes from month 3 to month 18 across groups of patients who were censored at different time points (Fig. 3). There was no evidence that patients who remained in the analysis at year 4 were selectively enriched for those experiencing the highest CD4 T-cell gains. Instead, there was a trend toward decreased CD4 T-cell slopes in those who remained at year 4.
We next evaluated factors associated with early and late CD4 T-cell changes. The median increase in CD4 T-cell counts from pre-therapy to month 3 was 65 × 106/l (interquartile range, 22–128 × 106/l) and the median increase in CD4 T-cells from month 3 to year 4 was 273 × 106/l (interquartile range, 144–404 × 106/l). The CD4 T-cell changes from pre-therapy to month 3 were associated with the pre-therapy plasma HIV RNA level (+ 20 × 106/l per log10 copies/ml increase; P < 0.001), but not age, pre-therapy CD4 T-cell count, sex, or hepatitis C serostatus in a multivariable model (P > 0.10 for all associations).
Several factors were independently associated with CD4 T-cell count changes from month 3 to year 4. Of the 153 patients who maintained a plasma HIV RNA level ≤ 1000 copies/ml for 4 years, 135 had available CD4 cell count determinations at month 3 and year 4. Pre-therapy CD4 cell counts were < 50 × 106/l in 35, 50 × 106–199 × 106/l in 37, 200 × 106–349 × 106/l in 34, and ≥ 350 × 106/l in 29. Among these patients and after adjustment for other significant factors, every 100 × 106/l increase in the pre-therapy CD4 cell count was associated with 63 × 106 fewer CD4 T-cells/l (P = 0.005) gained from month 3 to year 4 (Table 3). Likewise, for every 10-year increase in age, there were 60 × 106 fewer CD4 T-cells/l (P = 0.02) gained from month 3 to year 4. Women gained a mean of 188 × 106 CD4 T-cells/l (P = 0.03) more than men. Moderate (3.5–4.5 log10 copies/ml) but not high (> 4.5 log10 copies/ml) pre-therapy plasma HIV RNA levels were associated with increased CD4 T-cell gains compared to patients with low pre-therapy plasma HIV RNA levels (< 3.5 log10 copies/ml). However, hepatitis C virus serostatus was not predictive. Interestingly, patients with infrequent low-level viremia between week 48 and year 4 (defined as those having between 1 and 20% of all plasma HIV RNA levels during this time between 50 and 1000 copies/ml) gained a mean of 134 × 106 more cells/l (P = 0.006) than patients who had no episodes of detectable viremia during this time. Patients with infrequent low-level viremia also gained a mean of 173 × 106 more cells/l (P = 0.005) than patients with frequent low-level viremia (defined as those having > 20% of all plasma HIV RNA levels during this time between 50 and 1000 copies/ml). Adjustment for total number of plasma HIV RNA determinations did not alter this association (data not shown). Finally, there was a trend toward an association between CD4 cell count at month 3 of HAART and subsequent CD4 T-cell gains (P = 0.07). Substituting the absolute change in CD4 cell count from baseline to month 3 for the month 3 CD4 cell count in the multivariable model did not change any of the adjusted associations and was also positively associated with subsequent CD4 T-cell gains (P = 0.004, data not shown).
The capacity for CD4 T-cell regeneration during long-term HAART has not been well defined. Specifically, there remains a concern that CD4 cell counts often reach a plateau after the first few years of viral suppression [4,9,10]. This concern has been put forth as one reason to consider administering HAART earlier rather than later in the course of HIV infection. To assess the immune system's capacity for CD4 T-cell restoration, we studied HIV-infected patients experiencing durable viral suppression on HAART and observed that most patients continue to experience significant CD4 cell count increases for at least 4 years. There was no evidence of a plateau in CD4 T-cell gains in the majority of patients with pre-therapy CD4 counts < 350 × 106 cells/l who maintained viral suppression ≤ 1000 copies/ml. Consequently, irreversible depletion of circulating CD4 T-cells appears to be uncommon in patients with advanced immunodeficiency as long as durable treatment-mediated viral suppression can be maintained.
While we present robust evidence that CD4 cell counts continue to increase through year 4 of suppressive HAART, it is notable that the rate of recovery is slow and that steady state has not yet been reached by 4 years. This slow rate of recovery may reflect the inherent slow repopulation rate of naive CD4 T-cells by the adult thymus as has been suggested by others [22–24]. Indeed, the rate of CD4 T-cell gains we observed after year 1 of HAART is comparable to the slow rate of naive CD4 T-cell increases experienced by HIV-uninfected adults in the first 2 years following intensive chemotherapy . Alternatively, the slow rate of CD4 T-cell recovery may be the result of a countervailing HIV-induced process that either limits T-cell production or enhances destruction, such as ongoing T-cell activation despite viral suppression [26,27]. Since T-cell activation is independently associated with clinical progression in untreated HIV infection , a slow decay in levels of T-cell activation with long-term viral suppression might explain the slow rate of CD4 T-cell recovery.
We also determined several factors associated with HAART-mediated CD4 T-cell changes. Lower pre-therapy CD4 cell counts were associated with greater CD4 T-cell gains, an effect most prominent in the first 2 years of suppressive HAART. This suggests homeostatic mechanisms regulating CD4 T-cell synthesis or apoptosis that are sensitive to the degree of CD4 T-cell depletion. Alternatively, the redistributive phase of T-cells may be prolonged in patients with advanced pre-HAART immunodeficiency. Consistent with others’ findings [9,29,30], we also found that younger age predicted greater late CD4 T-cell gains on suppressive HAART, supporting the importance of thymic function (or its surrogate) in immune reconstitution. While few women were in our sample, the increased CD4 T-cell gains we observed in women are consistent with other recent reports [31–33], and may indicate an important sex difference in immune reconstitution that needs further exploration. That patients with infrequent low-level viremia have higher CD4 T-cell gains than patients with sustained undetectable plasma HIV RNA levels has recently been shown by another cohort . The mechanism for this is not known, but conceivably, higher CD4 T-cell gains may provide a larger pool of infectable cells, thereby driving rather than being a consequence of detectable bursts of viral replication.
A potential limitation of this study is the possibility that the continued CD4 cell count gains we observed were due to selective drop-out of patients with the worst immunologic responses, thereby enriching the sample over time with patients with the best immunologic responses. However, there was a trend toward decreased CD4 T-cell changes from month 3 to month 18 in the patients who remained at year 4 compared to patients who were censored earlier. Furthermore, assuming that patients who leave the analysis would exhibit similar CD4 T-cell changes as those who stay in the analysis after controlling for prior changes, our use of random slopes and intercepts in the repeated measures model should prevent inferences being driven solely by those who remain in the analysis [16,17].
Despite the sustained CD4 T-cell increases over 4 years of HAART observed in this study, our results should not be interpreted as evidence supporting the delay of antiretroviral therapy in patients with CD4 cell counts < 200 × 106/l. Indeed, several cohort studies have reported an increased risk of progression to AIDS and death in patients initiating HAART at CD4 cell counts below < 200 × 106/l [35–37]. Patients with low pre-HAART nadirs may also have persistent defects in immune function despite achieving normal CD4 cell counts on suppressive HAART . Moreover, we evaluated a selected group of patients who achieved and maintained long-term viral suppression. Since prior reports from clinic-based cohorts have found that half of HIV-infected patients experience virologic failure within the first year of HAART [11,39,40], our results likely overestimate the CD4 T-cell gains experienced by all patients initiating HAART. However, our results provide definitive evidence that the immune system is capable of slowly repopulating circulating CD4 T lymphocytes, even after advanced HIV-associated immune depletion, at least through 4 years of suppressive HAART.
Sponsorship: Supported in part by the University of California AIDS Research Center (CC99-SF-001), the UCSF-Gladstone Institute of Virology & Immunology Center for AIDS Research (P30 MH59037), the Case Western Reserve University Center for AIDS Research (P30 AI 36219), the NIH (RO1 AI052745), the Center for AIDS Prevention Studies (P30 MH62246) and the General Clinical Research Center at San Francisco General Hospital (5-MO1-RR00083-37).
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