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We investigated interactions between HIV and aging on brain function demands using functional magnetic resonance imaging (fMRI). A multiple regression model studied the association and interaction between fMRI measures, HIV serostatus, and age for 26 HIV infected (HIV+) and 25 seronegative (HIV−) subjects. While HIV serostatus and age independently affected fMRI measures, no interaction occurred. Functional brain demands in HIV+ subjects were equivalent to ~15–20 year older HIV− subjects. Frailty parallels between HIV and aging could result from continued immunological challenges depleting resources and triggering increased metabolic demands. fMRI could be a non-invasive biomarker to assess HIV in the brain.
Biological similarities exist between aging and human immunodeficiency virus (HIV) infection. Both cause a frailty phenotype characterized by generalized decreased physical function due to enhanced vulnerability to immunological stressors and inflammatory dysregulation . The introduction of highly active antiretroviral treatment (HAART) has increased the prevalence of older HIV infected (HIV+) subjects (> 50 years old) . The physiological impact of aging and HIV have been characterized within multiple organs, but not the brain.
Functional magnetic resonance imaging (fMRI) provides a noninvasive method to study brain metabolic demands. In HIV uninfected (HIV−) controls aging decreases functional activity within certain brain regions while causing recruitment of additional structures . Significant differences in functional brain activity occur between younger (< 40 years old) HIV infected (HIV+) compared to HIV− subjects . To date, no fMRI studies have investigated the possible interaction between HIV and aging. If fMRI is potentially to be used as a preclinical biomarker to assess HIV in the brain, a broader spectrum of ages is required for both HIV+ and HIV− subjects.
HIV− (n=25) and HIV+ (n=26) subjects (20–62 years old) provided written consent approved by the University of California San Diego Institutional Review Board. A detailed medical history was obtained with individuals excluded if they had a previous history of other neurological illness or infections, cerebrovascular disease or strokes, major psychiatric disorders, or substance abuse 3 months prior to imaging. Subjects underwent imaging if reported substance abstinence was confirmed to be negative by urine toxicology. HIV+ subjects were either not taking HAART or were on a stable regimen for > 3 months. All HIV+ patients had hematocrit, CD4 cell counts, and plasma HIV viral load measured. The plasma HIV viral was determined by a reverse transcriptase-polymerase chain reaction (Amplicor®, Roche Diagnostic Systems, Indianapolis, IN) using an ultrasensitive assay (lower quantification limit < 50 copies/mL). CD4 nadir values were obtained by self report or by direct measurement if the measured CD4 count was lower than the self-report measure.
Imaging was performed on a 3 Tesla whole body system (3-T GE Excite, Milwaukee, WI) using an eight-channel receive head coil. High-resolution structural images for anatomical confirmation were acquired using an inversion recovery prepared 3D fast spoiled pulse sequence (TI=450 ms, TR=7.9 ms, TE=3.1 ms, flip angle=12°, FOV=25 × 25 × 16 cm, matrix 256 × 256 × 124). Images were reviewed to ensure absence of structural lesions. For the activation task a block design was used with functional changes in cerebral blood flow and blood oxygen level dependent determined within the visual cortex for a flickering black and white radial checkerboard (8Hz). Activation periods were 20 seconds in length while rest portions were 60 seconds in duration and consisted of an isoluminant gray screen with a center fixation square. A single-shot proximal inversion sequence that controlled for off-resonance effects by using quantitative imaging of perfusion with a single subtraction (TR=2.5 s, TI1=700 ms, TI2=1500 ms, 20-cm tag width, and a 1-cm tag-slice gap), dual-echo gradient echo (GRE) readout, and spiral acquisition of k-space (TE1=9.4 ms, TE2=30 ms, flip angle=90°, field of view (FOV)=24 cm, 64 × 64 matrix) allowed for the alternate acquisition of ‘tag’ and ‘control’ images . The difference between images provided cerebral blood flow values, while the average of images yielded the blood oxygen level dependent signal. An additional resting cerebral blood flow scan was acquired to quantify baseline values using previously described methods .
Images were co-registered to correct for subject movement. The visual cortex was defined as the region between parietal-occipital sulci and was manually delineated on anatomical images. This region was re-sampled to match functional image scans and corrected for possible white matter involvement and partial volume loss. A general linear model used the stimulus function, acquired cardiac and respiratory data, and constant and linear terms as additional nuisance regressors to identify activated visual cortex voxels with a significance threshold (p=0.05) that corrected for multiple comparisons . From these activated voxels, functional changes in cerebral blood flow and the blood oxygen level dependent signal were determined for the task using previously described methods . The mean baseline cerebral blood flow was calculated by averaging all time points within activated visual cortex voxels.
Wilcoxon rank tests assessed for differences in fMRI measures between HIV+ and HIV− groups. The association and interaction between fMRI measures and HIV status and age (in years) were investigated using a multiple regression model. Each fMRI variable baseline cerebral blood flow, functional changes in cerebral blood flow and blood oxygen level dependent signal) was log10-transformed to improve normality and homoscedasticity and back-transformed to the natural scale for plotting. An additive regression model was used with p-values determined from the Wald test. The effects of age and HIV serostatus were determined for each fMRI outcome. The mean change in the fMRI outcome on the log-scale was transformed back to the response scale and expressed as proportional increase (+) or decrease (−) in the outcome with the effect size computed as the regression effect on the logarithmic scale divided by the residual standard deviation. The age-equivalent effect of HIV infection was determined by dividing the HIV effect size by the yearly effect size of age. The age effect was expressed per 15 years of aging for comparison.
Both groups were comparable in regards to age (mean: 41 year old for HIV− and 39 years old for HIV+), sex (% men: 56% for HIV− and 77% for HIV+) and education (mean: 15 years for HIV− and 16 years for HIV+). HIV+ subjects had a median CD4 cell count of 486 cells/ mm3 and a median CD4 nadir of 278 cells/ mm3. Approximately 60% (15/26) of the HIV+ group were on stable HAART.
HIV+ subjects had a lower baseline cerebral blood flow compared to HIV− subjects (Figure 1 top). Increasing age and HIV infection caused significant decreases in baseline cerebral blood flow but no interaction occurred (Table 1). For a given age baseline cerebral blood flow values in HIV+ subjects were equivalent to 15 year older HIV− subjects. Functional cerebral blood flow changes were also greater for HIV+ compared to HIV− subjects (Figure 1 middle). Both aging and HIV infection caused significantly increases in functional changes in cerebral blood flow but no interaction was present. HIV infection was equivalent to a 21 year increase in brain age compared to HIV− controls (Table 1). Functional changes in the blood oxygen level dependent signal were reduced for HIV+ compared to HIV− subjects (Figure 1 bottom). Aging and HIV infection independently caused significant decreases in functional blood oxygen level dependent signal. While the age-related regression lines intersected for the two groups no significant interaction was observed (p=0.16). Functional changes in the blood oxygen level dependent signal for HIV+ subjects were equivalent to 15 years older HIV− controls for a given age (Table 1).
As the number of older HIV+ individuals continues to rise, a growing need exists to understand the potential interactions between HIV and aging within the brain. Our results suggest that HIV infection and aging independently affect brain functional demands measured by fMRI.
While surprising, the lack of a synergistic interaction between age and HIV infection is comparable to larger a neuropsychological study that showed that age and HIV serostatus were independent risk factors for the development of HIV associated neurocognitive disorders . While our sample size is significantly smaller than the previous descriptive study, our cohort included women which may better reflect the gender diversity of the disease.
Conflicting results concerning the relationship between HIV and aging have been noted with structural neuroimaging [9, 10]. An interaction occurred in HAART naïve HIV+ subjects using magnetic resonance spectroscopy with a fivefold increase present in frontal white matter inflammatory and glial metabolites . In contrast, diffusion tensor imaging of tissue water molecules did not observe age dependent changes within HIV+ subjects on stable HAART . Discrepancies in results could reflect differences in neuroimaging techniques or the effect of HAART.
While most neuroimaging studies of HIV have investigated the role of subcortical or frontal areas in HIV associated neurocognitive disorders, primary cortical areas have been shown to be affected . Our analysis was not focused on studying the effects of HIV associated neurocognitive disorders on fMRI measures; but instead we investigated the impact of HIV itself on a cortical structure- the visual cortex. The effects of “normal” aging have been previously characterized in this region . Observed decreases in baseline cerebral blood flow in HIV− controls with increasing aging are similar to previous neuroimaging studies [12, 13]. In general, a decrease in baseline cerebral blood flow occurs with HIV infection . Derangements within primary cortical areas may cause disconnections among higher processing networks despite the absence of abnormalities on traditional structural imaging. The etiology of observed decreases in baseline cerebral blood flow remains unknown but could result from deleterious effects of HIV on platelet function, endothelial function, or dendritic arborization .
In this cross sectional cohort observed parallel trends for HIV and aging in fMRI measures suggest pathophysiological similarities not only throughout the body but in the brain. Body frailty due to aging and HIV results from a complex interplay among viral factors, host responses, and immune dysfunction causing: 1) a shift in T-cell phenotypes from naïve to memory, 2) an increase in replicative senescence by reducing T-cell proliferation, and 3) an increase in the production and release of cytokines . Within the brain continued immunological challenges from “normal” aging could deplete available resources triggering increased functional metabolic demands. Although we did not directly correlate neuroimaging measures with markers of brain immune dysfunction, it is reasonable to hypothesize that persistent low levels of HIV and inflammation may deplete brain reserves augmenting functional metabolic requirements. Observed increases in metabolic demands could result from excessive neurotransmission from oxidative stress signaling pathways . Multiple neurodegenerative and neuroimmune mechanisms may converge in older HIV+ individuals. Future longitudinal analyses are needed as HIV and aging could proceed in parallel making it difficult to distinguish their individual effects.
A number of limitations exist with this study. Co-morbid substance abuse, such as alcoholism or stimulant dependence could influence fMRI outcomes. Many of our subjects had a previous history of drug abuse or used illegal drugs recreationally. However, approximately equal numbers within the HIV− controls (11/25) and HIV+ subjects (10/26) met these criteria. Future studies investigating the interaction between substance abuse, HIV infection, and age are needed. Second, although our cross sectional cohort was modest in size; a larger sample may have demonstrated even greater differences. Despite somewhat large variability within HIV+ patients, observed effect sizes for fMRI measures were quite robust (Table 1). Longitudinal studies of older HIV+ patients are required to confirm these initial findings. Third, the impact of HAART on aging could not be analyzed as most of the older HIV+ subjects were on medications. Prospective studies within older HIV+ subjects are needed to address the appropriateness of current HAART regimens, susceptibility to HAART side effects, and the potential for increased vulnerability to adverse drug reactions due to aging.
Despite these limitations our results highlight the potential role for fMRI in the evaluation of possible impact of HIV and aging on brain function. We found that both HIV-infection and advancing age challenge brain perfusion and functional metabolic pathways within the visual cortex. This may represent diminished capacity and functional frailty within this brain region of HIV+ and older adults. We are not advocating the routine use of fMRI for routine diagnostic evaluation of HIV+ subjects due to its relatively limited availability and expense. However, non-invasive fMRI in research settings could provide valuable information concerning basic pathophysiology of HIV in certain brain areas; the potential neurotoxic actions of HAART; and assist in the evaluation of neuroprotective therapeutic strategies for older HIV+ subjects.
The authors are also deeply indebted to Thomas Liu and Kal Restom for their invaluable suggestions and assistance with code used for analyses. The authors are appreciative of constructive input from David Clifford and Kevin Yarasheski.
Financial Support: This work was supported by a Dana Foundation Brain Immuno Imaging Award (DF3857-41880) (BA) and NIH grants (1K23MH081786) (BA), (NS-36722 and NS-42069) (RB and CL), and P30 MH62512 (HNRC Group). In addition, support was provided by AI27670, AI43638, and the UCSD Center for AIDS Research AI 36214, AI29164, AI47745, AI57167, AI55276, MH62512 from the National Institutes of Health (A.M., S.L., and R.E.) and MH22005, AI47033 (F.V.).
Potential Conflicts of Interest: The authors do not have a commercial or other association that might pose a conflict of interest (e.g., pharmaceutical stock ownership, consultancy, advisory board membership, relevant patents, or research funding).
Previous Presentation: This data was presented at the Conference on Retrovirus and Opportunistic Infections (CROI), February 2009, Montreal, Canada, Abstract # 157.
Conflict of interest
Beau M. Ances- no conflict of interest
Florin Vaida – no conflict of interest
Melinda J. Yeh- no conflict of interest
Christine L. Liang- no conflict of interest
Richard B. Buxton- no conflict of interest
Scott Letendre- no conflict of interest
J. Allen McCutchan - no conflict of interest
Ronald J. Ellis- no conflict of interest