Publications

2021
Lorenz DR, Mukerji SS, Misra V, Uno H, Gelman BB, Moore DJ, Singer EJ, Morgello S, Gabuzda D. Predictors of Transition to Frailty in Middle-Aged and Older People With HIV: A Prospective Cohort Study. J Acquir Immune Defic Syndr. 2021;88 (5) :518-527.Abstract
BACKGROUND: People with HIV (PWH) have increased frailty risk at younger ages compared with the general population. Multimorbidity is associated with frailty, yet effects of specific comorbidities on transition to frailty in PWH are unknown. SETTING: Prospective study of 219 PWH age 45 years or older in the National NeuroAIDS Tissue Consortium. METHODS: Frailty status was categorized using Fried frailty phenotype criteria. Comorbidities [bone disease, cardiovascular disease, cerebrovascular disease, liver disease, renal disease, diabetes, chronic obstructive pulmonary disease (COPD), hypertension, obesity, cancers, neuropsychiatric conditions] were assessed from longitudinal data. Associations between baseline comorbidities and transition to frailty within 30 months were analyzed using Kaplan-Meier and Cox regression models. Grip strength was assessed using mixed-effects models. RESULTS: At baseline, the median age was 61 years, 73% were male 98% were on antiretroviral therapy, 29% had ≥3 comorbidities, 27% were robust, and 73% were pre-frail. Cerebrovascular disease, diabetes, and COPD were independent predictors of transition to frailty within 30 months in models adjusted for age, sex, and multimorbidity (≥3 additional comorbidities) [hazard ratios (95% confidence intervals) 2.52 (1.29 to 4.93), 2.31 (1.12 to 4.76), and 1.82 (0.95 to 3.48), respectively]. Furthermore, cerebrovascular disease, diabetes, COPD, or liver disease co-occurring with multimorbidity was associated with substantially increased frailty hazards compared with multimorbidity alone (hazard ratios 4.75-7.46). Cerebrovascular disease was associated with decreased baseline grip strength (P = 0.0001), whereas multimorbidity, diabetes, and COPD were associated with declining grip strength (P < 0.10). CONCLUSIONS: In older PWH, cerebrovascular disease, diabetes, COPD, or liver disease co-occurring with multimorbidity is associated with substantially increased risk of becoming frail within 30 months. Interventions targeting these comorbidities may ameliorate frailty and age-related functional decline in PWH.
Aldersley J, Lorenz DR, Mouw KW, D'Andrea AD, Gabuzda D. Genomic Landscape of Primary and Recurrent Anal Squamous Cell Carcinomas in Relation to HPV Integration, Copy-Number Variation, and DNA Damage Response Genes. Mol Cancer Res. 2021;19 (8) :1308-1321.Abstract
The incidence of anal squamous cell carcinoma (ASCC) has been increasing, particularly in populations with HIV. Human papillomavirus (HPV) is the causal factor in 85% to 90% of ASCCs, but few studies evaluated HPV genotypes and integrations in relation to genomic alterations in ASCC. Using whole-exome sequence data for primary (n = 56) and recurrent (n = 31) ASCC from 72 patients, we detected HPV DNA in 87.5% of ASCC, of which HPV-16, HPV-18, and HPV-6 were detected in 56%, 22%, and 33% of HIV-positive (n = 9) compared with 83%, 3.2%, and 1.6% of HIV-negative cases (n = 63), respectively. Recurrent copy-number variations (CNV) involving genes with documented roles in cancer included amplification of PI3KCA and deletion of APC in primary and recurrent tumors; amplifications of CCND1, MYC, and NOTCH1 and deletions of BRCA2 and RB1 in primary tumors; and deletions of ATR, FANCD2, and FHIT in recurrent tumors. DNA damage response genes were enriched among recurrently deleted genes in recurrent ASCCs (P = 0.001). HPV integrations were detected in 29 of 76 (38%) ASCCs and were more frequent in stage III-IV versus stage I-II tumors. HPV integrations were detected near MYC and CCND1 amplifications and recurrent targets included NFI and MUC genes. These results suggest HPV genotypes in ASCC differ by HIV status, HPV integration is associated with ASCC progression, and DNA damage response genes are commonly disrupted in recurrent ASCCs. IMPLICATIONS: These data provide the largest whole-exome sequencing study of the ASCC genomic landscape to date and identify HPV genotypes, integrations, and recurrent CNVs in primary or recurrent ASCCs.
Lorenz DR, Mukerji SS, Misra V, Uno H, Gelman BB, Moore DJ, Singer EJ, Morgello S, Gabuzda D. Multimorbidity networks associated with frailty among middle aged and older people with HIV. AIDS. 2021.Abstract
OBJECTIVE: People with human immunodeficiency virus (HIV) (PWH) have increased prevalence of multimorbidity and frailty at younger ages compared to the general population. This study investigated individual and combinatorial effects of neuropsychiatric and medical comorbidities as predictors of frailty in PWH. DESIGN: Analysis of data from the National NeuroAIDS Tissue Consortium, a longitudinal observational cohort. METHODS: 524 PWH over age 40 were classified using Fried's Frailty criteria. Twelve comorbidities were documented from longitudinal data and associations between individual and co-occurring comorbidities with frailty were assessed using weighted network and logistic regression analyses. RESULTS: At frailty assessment between 2015-2020, median age was 61 years, 76% were male, 94% were on ART, 73% had two or more comorbidities, 24% were frail, and 52% were prefrail. Among individual comorbidities, highest odds of frailty were in participants with depressive symptoms (adjusted odds ratio [aOR], 95% confidence interval [CI] 3.48 [2.22-5.46]]), followed by bone disease and COPD (2.47 [1.28-4.72] and 2.13 [1.36-3.34], respectively). Among co-occurring comorbidities, highest odds of frailty were in participants having depressive symptoms with diabetes, hypertension, or obesity (aORs [95% CIs] 5.29 [2.32-12.08], 5.21 [2.65-10.40], 4.85 [2.39-9.95], respectively), cognitive impairment with diabetes or renal disease, (2.81 [1.38-5.68] and 2.53 [1.26-5.03], respectively), renal disease with cardiovascular disease (2.81 [1.32-6.01]), and diabetes with obesity (2.76 [1.39-5.45]). CONCLUSIONS: Co-occurrence of depressive symptoms, cognitive impairment, diabetes, or renal disease with other medical conditions substantially increases odds of frailty in older PWH. Identifying and treating these comorbidities may help to reduce functional decline with aging in PWH.
Lorenz DR, Misra V, Chettimada S, Uno H, Wang L, Blount BC, De Jesús VR, Gelman BB, Morgello S, Wolinsky SM, et al. Acrolein and other toxicant exposures in relation to cardiovascular disease among marijuana and tobacco smokers in a longitudinal cohort of HIV-positive and negative adults. EClinicalMedicine. 2021;31 :100697.Abstract
Background: Marijuana smoke contains some of the same toxicants present in tobacco smoke. Marijuana smoking is prevalent among HIV+ individuals, but few studies have characterized smoke-related toxicants or associated health outcomes in exclusive marijuana users. Methods: This longitudinal study included 245 participants over age 40 (76% HIV+). 33 plasma and 28 urine metabolites of nicotine, ∆-9-trans-tetrahydrocannabinol, polycyclic aromatic hydrocarbons, and volatile organic compounds were assayed by liquid or gas chromatography/mass spectrometry. Exposures and health outcomes were assessed from surveys and medical records. Findings: At baseline, 18% of participants were marijuana-only smokers, 20% tobacco-only smokers, and 24% dual marijuana-tobacco smokers (median (IQR) age 53 (47-60) years, 78% male, 54% white race). Marijuana smoking was independently associated with elevated plasma naphthalenes, 2-hydroxyfluorene sulfate, 4-vinylphenol sulfate, and o-cresol sulfate 0·05) and urine acrylonitrile and acrylamide metabolites (0·05), but levels were lower than those associated with tobacco smoking. Acrolein metabolite N-Acetyl-S-(3-hydroxypropyl)-l-cysteine (3HPMA) was significantly elevated in plasma and urine in tobacco-only and dual but not marijuana-only smokers, and correlated with nicotine metabolites (0·05). The highest tertile of 3HPMA was associated with increased cardiovascular disease diagnoses independent of tobacco smoking, traditional risk factors, and HIV status (odds ratio [95% CI] 3·34 [1·31-8·57];   0·012). Interpretation: Smoke-related toxicants, including acrylonitrile and acrylamide metabolites, are detectable in exclusive marijuana smokers, but exposures are lower compared with tobacco or dual smokers. Acrolein exposure is increased by tobacco smoking but not exclusive marijuana smoking in HIV+ and HIV- adults, and contributes to cardiovascular disease in tobacco smokers. Funding: U.S. NIH.
Pahwa S, Deeks S, Zou S, Tomitch N, Miller-Novak L, Caler E, Justice A, Sacktor N, Gabuzda D, Hunt PW, et al. NIH Workshop on HIV-Associated Comorbidities, Coinfections, and Complications: Summary and Recommendation for Future Research. J Acquir Immune Defic Syndr. 2021;86 (1) :11-18.Abstract
BACKGROUND: With potent antiretroviral therapy and simplified regimens, people living with HIV (PWH) are achieving near-normal lifespans but not necessarily a normal health span or healthy aging. PWH have a higher than expected risk of developing a number of non-AIDS comorbidities, coinfections, and complications (CCC), often against a background of stigma, poverty, and isolation. SETTING: To gain a better understanding of research needs for HIV-associated CCC, the NIH convened a 2-day workshop (HIV-associated CCC, or HIV ACTION). METHODS: A cross-institute NIH planning committee identified 6 key research areas: epidemiology and population research, pathogenesis and basic science research, clinical research, implementation science research, syndemics research and international research in low and middle income countries. Investigators were selected to lead working groups (WGs) to assess the state-of-the-art and identify 3-5 priority areas in each field before the workshop. A 2-day program at the NIH was developed which included presentations by invited experts and WG members. RESULTS: Over 400 participants attended the workshop. After general and individual WG discussions, the most pressing gaps, questions, or proposed action items were identified. Priority lists of pressing research issues were presented by cochairs of each WG. A detailed report is posted at the NHLBI website. This article reports the streamlined priority list and a summary of WG discussions to inform investigators of current priorities in the field. CONCLUSION: Collaborative efforts of many disciplines are needed to improve the health and wellbeing of PWH. Several common themes emerged across WG representing potential priorities for investigators and recommendations for the NIH.
2020
Chettimada S, Lorenz DR, Misra V, Wolinsky SM, Gabuzda D. Small RNA sequencing of extracellular vesicles identifies circulating miRNAs related to inflammation and oxidative stress in HIV patients. BMC Immunol. 2020;21 :57.Abstract

BACKGROUND: Extracellular vesicles (EVs) are nano-sized particles secreted by most cells. EVs carry nucleic acids that hold promise as potential biomarkers in various diseases. Human immunodeficiency virus type 1 (HIV) infects CD4+ T cells and induces immune dysfunction, inflammation, and EV secretion, but little is known about EV small RNA cargo in relation to immune dysregulation in HIV-infected individuals. Here, we characterize small RNA carried by circulating EVs in HIV-positive subjects on antiretroviral therapy (ART) relative to uninfected controls by next-generation RNA sequencing. RESULTS: Plasma EVs isolated from HIV-positive and HIV-negative subjects in test (n = 24) and validation (n = 16) cohorts were characterized by electron microscopy, nanoparticle tracking analysis, and immunoblotting for exosome markers. EVs were more abundant in plasma from HIV-positive compared to HIV-negative subjects. Small RNA sequencing of plasma EVs in the test cohort identified diverse small RNA species including miRNA, piRNA, snRNA, snoRNA, tRNA, and rRNA, with miRNA being the most abundant. A total of 351 different miRNAs were detected in plasma EVs, with the top 50 miRNAs accounting for 90% of all miRNA reads. miR-26a-5p was the most abundant miRNA, followed by miR-21-5p and miR-148-3p. qRT-PCR analysis showed that six miRNAs (miR-10a-5p, - 21-5p, -27b-3p, - 122-5p, -146a-5p, - 423-5p) were significantly increased in plasma EVs from HIV-positive compared to HIV-negative subjects in the validation cohort. Furthermore, miR-21-5p, -27b-3p, -146a-5p, and - 423-5p correlated positively with metabolite markers of oxidative stress and negatively with anti-inflammatory polyunsaturated fatty acids. Over-representation and pathway enrichment analyses of miRNAs and their target genes predicted functional association with oxidative stress responses, interferon gamma signaling, Toll-like receptor signaling, TGF beta signaling, and Notch signaling. CONCLUSIONS: HIV-positive individuals on ART have increased abundance of circulating EVs carrying diverse small RNAs, with miRNAs being the most abundant. Several miRNAs associated with inflammation and oxidative stress are increased in circulating EVs of HIV-positive individuals, representing potential biomarkers of targetable pathways that contribute to disease pathogenesis.

PubMed

DOI

Zhang S, Nguyen HT, Ding H, Wang J, Zou S, Liu L, Guha D, Gabuzda D, Ho DD, Kappes JC, et al. Dual pathways of human immunodeficiency virus (HIV-1) envelope glycoprotein trafficking modulate the selective exclusion of uncleaved oligomers from virions. J Virol. 2020.Abstract

The human immunodeficiency virus (HIV-1) envelope glycoprotein (Env) trimer is transported through the secretory pathway to the infected cell surface and onto virion particles. In the Golgi, the gp160 Env precursor is modified by complex sugars and proteolytically cleaved to produce the mature functional Env trimer, which resists antibody neutralization. We observed mostly uncleaved gp160 and smaller amounts of cleaved gp120 and gp41 Envs on the surface of HIV-1-infected or Env-expressing cells; however, cleaved Envs were relatively enriched in virions and virus-like particles (VLPs). This relative enrichment of cleaved Env in VLPs was observed for wild-type Envs, for Envs lacking the cytoplasmic tail and for CD4-independent, conformationally flexible Envs. On the cell surface, we identified three distinct populations of Envs: 1) the cleaved Env was transported through the Golgi, was modified by complex glycans, formed trimers that crosslinked efficiently, and was recognized by broadly neutralizing antibodies; 2) a small fraction of Env modified by complex carbohydrates escaped cleavage in the Golgi; and 3) the larger population of uncleaved Env lacked complex carbohydrates, crosslinked into diverse oligomeric forms, and was recognized by poorly neutralizing antibodies. This last group of more "open" Env oligomers reached the cell surface in the presence of Brefeldin A, apparently bypassing the Golgi apparatus. Relative to Envs transported through the Golgi, these uncleaved Envs were counterselected for virion incorporation. By employing two pathways for Env transport to the surface of infected cells, HIV-1 can misdirect host antibody responses towards conformationally flexible, uncleaved Env without compromising virus infectivity.IMPORTANCE The envelope glycoprotein (Env) trimers on the surface of human immunodeficiency virus (HIV-1) mediate the entry of the virus into host cells and serve as targets for neutralizing antibodies. The cleaved, functional Env is incorporated into virus particles from the surface of the infected cell. We found that an uncleaved form of Env is transported to the cell surface by an unconventional route, but this non-functional Env is mostly excluded from the virus. Thus, only one of the pathways by which Env is transported to the surface of infected cells results in efficient incorporation into virus particles, potentially allowing the uncleaved Env to act as a decoy to the host immune system without compromising virus infectivity.

PubMed

DOI

Gabuzda D, McArthur JC, Letendre SL. Efficacy and safety of two-drug regimens for treatment of HIV in the central nervous system. Aids. 2020;34 :1975-1977.Abstract
Gabuzda D, Jamieson BD, Collman RG, Lederman MM, Burdo TH, Deeks SG, Dittmer DP, Fox HS, Funderburg NT, Pahwa SG, et al. Pathogenesis of Aging and Age-related Comorbidities in People with HIV: Highlights from the HIV ACTION Workshop. Pathog Immun. 2020;5 :143-174.Abstract

People with HIV (PWH) experience accentuated biological aging, as defined by markers of inflammation, immune dysfunction, and the epigenetic clock. They also have an elevated risk of multiple age-associated comorbidities. To discuss current knowledge, research gaps, and priorities in aging and age-related comorbidities in treated HIV infection, the NIH program staff organized a workshop held in Bethesda, Maryland in September 2019. This review article describes highlights of discussions led by the Pathogenesis/Basic Science Research working group that focused on three high priority topics: immunopathogenesis; the microbiome/virome; and aging and senescence. We summarize knowledge in these fields and describe key questions for research on the pathogenesis of aging and age-related comorbidities in PWH. Understanding the drivers and mechanisms underlying accentuated biological aging is a high priority that will help identify potential therapeutic targets to improve healthspan in older PWH.

PubMed

DOI

Mukerji SS, Misra V, Lorenz DR, Chettimada S, Keller K, Letendre S, Ellis RJ, Morgello S, Parker RA, Gabuzda D. Low Neuroactive Steroids Identifies a Biological Subtype of Depression in Adults with Human Immunodeficiency Virus on Suppressive Antiretroviral Therapy. J Infect Dis. 2020.Abstract
BACKGROUND: The prevalence and mortality risk of depression in people with Human Immunodeficiency Virus (HIV)-infection (PWH) on antiretroviral therapy (ART) is higher than in the general population, yet biomarkers for therapeutic targeting are unknown. Here, we aimed to identify plasma metabolites associated with depressive symptoms in PWH on ART. METHODS: This is a prospective study of ART-treated HIV-infected adults with or without depressive symptoms assessed using longitudinal Beck Depression Inventory scores. Plasma metabolite profiling was performed in two independent cohorts (total n=99) using liquid and gas chromatography and tandem mass spectrometry. RESULTS: Participants with depressive symptoms had lower neuroactive steroids (dehydroepiandrosterone sulfate (DHEA-S), androstenediols, pregnenolone sulfate) compared to those without depressive symptoms. Cortisol/DHEA-S ratio, an indicator of hypothalamic-pituitary-adrenal axis imbalance, was associated with depressive symptoms (p<0.01) due to low DHEA-S, while cortisol was similar between groups. The odds of having depressive symptoms increased with higher cortisol/DHEA-S ratios [odds 2.5 per z-score, 95% confidence interval 1.3-4.7], independent of age and gender. Kynurenine to tryptophan ratio showed no significant associations. CONCLUSIONS: These findings suggest that altered neuroactive steroid metabolism may contribute to the pathophysiology of depression in ART-treated HIV-infected adults, representing a potential biological pathway for therapeutic targeting.
2019
Guha D, Lorenz DR, Misra V, Chettimada S, Morgello S, Gabuzda D. Proteomic analysis of cerebrospinal fluid extracellular vesicles reveals synaptic injury, inflammation, and stress response markers in HIV patients with cognitive impairment. J Neuroinflammation. 2019;16 (1) :254.Abstract
BACKGROUND: Extracellular vesicles (EVs) are nano-sized particles present in most body fluids including cerebrospinal fluid (CSF). Little is known about CSF EV proteins in HIV+ individuals. Here, we characterize the CSF EV proteome in HIV+ subjects and its relationship to neuroinflammation, stress responses, and HIV-associated neurocognitive disorders (HAND). METHODS: CSF EVs isolated from 20 HIV+ subjects with (n = 10) or without (n = 10) cognitive impairment were characterized by electron microscopy, nanoparticle tracking analysis, immunoblotting, and untargeted LC/MS/MS mass spectrometry. Functional annotation was performed by gene ontology (GO) mapping and expression annotation using Biobase Transfac and PANTHER software. Cultured astrocytic U87 cells were treated with hydrogen peroxide for 4 h to induce oxidative stress and EVs isolated by ultracentrifugation. Selected markers of astrocytes (GFAP, GLUL), inflammation (CRP), and stress responses (PRDX2, PARK7, HSP70) were evaluated in EVs released by U87 cells following induction of oxidative stress and in CSF EVs from HIV+ patients by immunoblotting. RESULTS: Mass spectrometry identified 2727 and 1626 proteins in EV fractions and EV-depleted CSF samples, respectively. CSF EV fractions were enriched with exosomal markers including Alix, syntenin, tetraspanins, and heat-shock proteins and a subset of neuronal, astrocyte, oligodendrocyte, and choroid plexus markers, in comparison to EV-depleted CSF. Proteins related to synapses, immune/inflammatory responses, stress responses, metabolic processes, mitochondrial functions, and blood-brain barrier were also identified in CSF EV fractions by GO mapping. HAND subjects had higher abundance of CSF EVs and proteins mapping to GO terms for synapses, glial cells, inflammation, and stress responses compared to those without HAND. GFAP, GLUL, CRP, PRDX2, PARK7, and HSP70 were confirmed by immunoblotting of CSF EVs from subjects with HAND and were also detected in EVs released by U87 cells under oxidative stress. CONCLUSIONS: These findings suggest that CSF EVs derived from neurons, glial cells, and choroid plexus carry synaptic, immune/inflammation-related, and stress response proteins in HIV+ individuals with cognitive impairment, representing a valuable source for biomarker discovery.
Lorenz DR, Misra V, Gabuzda D. Transcriptomic analysis of monocytes from HIV-positive men on antiretroviral therapy reveals effects of tobacco smoking on interferon and stress response systems associated with depressive symptoms. Hum Genomics. 2019;13 (1) :59.Abstract
BACKGROUND: Tobacco smoking induces immunomodulatory and pro-inflammatory effects associated with transcriptome changes in monocytes and other immune cell types. While smoking is prevalent in HIV-infected (HIV+) individuals, few studies have investigated its effects on gene expression in this population. Here, we report whole-transcriptome analyses of 125 peripheral blood monocyte samples from ART-treated HIV+ and uninfected (HIV-) men enrolled in the Multicenter AIDS Cohort Study (MACS) (n = 25 HIV+ smokers, n = 60 HIV+ non-smokers, n = 40 HIV- non-smoking controls). Gene expression profiling was performed using Illumina HumanHT-12 Expression BeadChip microarrays. Differential expression analysis was performed with weighted linear regression models using the R limma package, followed by functional enrichment and Ingenuity Pathway analyses. RESULTS: A total of 286 genes were differentially expressed in monocytes from HIV+ smokers compared with HIV- non-smokers; upregulated genes (n = 180) were enriched for immune and interferon response, chemical/stress response, mitochondria, and extracellular vesicle gene ontology (GO) terms. Expression of genes related to immune/interferon responses (AIM2, FCGR1A-B, IFI16, SP100), stress/chemical responses (APAF1, HSPD1, KLF4), and mitochondrial function (CISD1, MTHFD2, SQOR) was upregulated in HIV+ non-smokers and further increased in HIV+ smokers. Gene expression changes associated with smoking in previous studies of human monocytes were also observed (SASH1, STAB1, PID1, MMP25). Depressive symptoms (CES-D scores ≥ 16) were more prevalent in HIV+ tobacco smokers compared with HIV+ and HIV- non-smokers (50% vs. 26% and 13%, respectively; p = 0.007), and upregulation of immune/interferon response genes, including IFI35, IFNAR1, OAS1-2, STAT1, and SP100, was associated with depressive symptoms in logistic regression models adjusted for HIV status and smoking (p < 0.05). Network models linked the Stat1-mediated interferon pathway to transcriptional regulator Klf4 and smoking-associated toll-like receptor scaffolding protein Sash1, suggesting inter-relationships between smoking-associated genes, control of monocyte differentiation, and interferon-mediated inflammatory responses. CONCLUSIONS: This study characterizes immune, interferon, stress response, and mitochondrial-associated gene expression changes in monocytes from HIV+ tobacco smokers, and identifies augmented interferon and stress responses associated with depressive symptoms. These findings help to explain complex interrelationships between pro-inflammatory effects of HIV and smoking, and their combined impact on comorbidities prevalent in HIV+ individuals.
Solomon IH, Chettimada S, Misra V, Lorenz DR, Gorelick RJ, Gelman BB, Morgello S, Gabuzda D. White Matter Abnormalities Linked to Interferon, Stress Response, and Energy Metabolism Gene Expression Changes in Older HIV-Positive Patients on Antiretroviral Therapy. Mol Neurobiol. 2019.Abstract
Neurocognitive impairment (NCI) remains a significant cause of morbidity in human immunodeficiency virus (HIV)-positive individuals despite highly active antiretroviral therapy (HAART). White matter abnormalities have emerged as a key component of age-related neurodegeneration, and accumulating evidence suggests they play a role in HIV-associated neurocognitive disorders. Viral persistence in the brain induces chronic inflammation associated with lymphocytic infiltration, microglial proliferation, myelin loss, and cerebrovascular lesions. In this study, gene expression profiling was performed on frontal white matter from 34 older HIV+ individuals on HAART (18 with NCI) and 24 HIV-negative controls. We used the NanoString nCounter platform to evaluate 933 probes targeting inflammation, interferon and stress responses, energy metabolism, and central nervous system-related genes. Viral loads were measured using single-copy assays. Compared to HIV- controls, HIV+ individuals exhibited increased expression of genes related to interferon, MHC-1, and stress responses, myeloid cells, and T cells and decreased expression of genes associated with oligodendrocytes and energy metabolism in white matter. These findings correlated with increased white matter inflammation and myelin pallor, suggesting interferon (IRFs, IFITM1, ISG15, MX1, OAS3) and stress response (ATF4, XBP1, CHOP, CASP1, WARS) gene expression changes are associated with decreased energy metabolism (SREBF1, SREBF2, PARK2, TXNIP) and oligodendrocyte myelin production (MAG, MOG), leading to white matter dysfunction. Machine learning identified a 15-gene signature predictive of HIV status that was validated in an independent cohort. No specific gene expression patterns were associated with NCI. These findings suggest therapies that decrease chronic inflammation while protecting mitochondrial function may help to preserve white matter integrity in older HIV+ individuals.
Lorenz DR, Uno H, Wolinsky SM, Gabuzda D. Effect of Marijuana Smoking on Pulmonary Disease in HIV-Infected and Uninfected Men: A Longitudinal Cohort Study. EClinicalMedicine. 2019.Abstract
Background
Lung disease is a common comorbidity in people with HIV/AIDS, independent of smoking status. The effects of marijuana smoking on risk of lung disease in HIV-infected individuals are unclear.
Methods
In this prospective cohort study, we quantified lung disease risk among men enrolled in the Multicenter AIDS Cohort Study (MACS), a long-term observational cohort of HIV-infected and uninfected men who have sex with men. Eligible participants were aged ≥30 years with self-reported marijuana and tobacco smoking data from biannual study visits between 1996 and 2014. Pulmonary diagnoses were obtained from self-report and medical records. Analyses were performed using Cox models and Generalized Estimating Equations adjusted for tobacco smoking, CD4 T cell count, and other risk factors.
Findings
1630 incident pulmonary diagnoses were reported among 1352 HIV-seropositive and 1352 HIV-seronegative eligible participants matched for race and baseline age (53,794 total person-visits, median follow-up 10.5 years). 27% of HIV-infected participants reported daily or weekly marijuana smoking for one or more years in follow-up, compared to 18% of uninfected participants (median 4·0 and 4·5 years daily/weekly use, respectively). HIV-infected participants had an increased likelihood of infectious or non-infectious pulmonary diagnoses compared to uninfected participants (33·2% vs. 21·5%, and 20·6% vs. 17·2%, respectively). Among HIV-infected participants, recent marijuana smoking was associated with increased risk of infectious pulmonary diagnoses and chronic bronchitis independent of tobacco smoking and other risk factors for lung disease (hazard ratio [95% confidence interval] 1·43 [1·09–1·86], and 1·54 [1·11–2·13], respectively); these risks were additive in participants smoking both substances. There was no association between marijuana smoking and pulmonary diagnoses in HIV-uninfected participants.
Interpretation
In this longitudinal study, long-term marijuana smoking was associated with lung disease independent of tobacco smoking and other risk factors in HIV-infected individuals. These findings could be used to reduce modifiable risks of lung disease in high-risk populations.
Guha D, Mukerji SS, Chettimada S, Misra V, Lorenz DR, Morgello S, Gabuzda D. CSF extracellular vesicles and neurofilament light protein as biomarkers of CNS injury in HIV-infected patients on antiretroviral therapy. AIDS. 2019.Abstract
OBJECTIVE: The relationship of cerebrospinal fluid (CSF) extracellular vesicles (EVs) to neurocognitive impairment (NCI) in HIV-infected individuals is unclear. Here, we characterize CSF EVs and their association with CNS injury-related biomarkers (neurofilament light [NFL], S100B, neopterin) and NCI in HIV+ subjects on combination antiretroviral therapy (cART). DESIGN: Cross-sectional and longitudinal study of CSF samples from HIV+ subjects on cART. METHODS: NFL, S100B, and neopterin were measured by ELISA in 190 CSF samples from 112 subjects (67 HIV+ and 45 HIV-). CSF EVs were isolated and characterized by electron microscopy, nanoparticle tracking analysis, immunoblotting for exosome markers (CD9, CD63, CD81, FLOT-1), and ELISA for HLA-DR. RESULTS: HIV+ subjects had median age 52 years, 67% with suppressed plasma viral load (< 50 copies/ml), median CD4 nadir 66 cells/μl and CD4 count 313 cells/μl. CSF NFL, S100B, and neopterin levels were higher in HIV+ vs. HIV- subjects, and nonsuppressed vs. suppressed HIV+ subjects. While CSF NFL and S100B levels were higher in NCI vs. unimpaired HIV+ subjects (p < 0.05), only NFL was associated with NCI in adjusted models (p < 0.05). CSF EVs were increased in HIV+ vs. HIV- subjects, and NCI vs. unimpaired HIV+ subjects (p < 0.001), and correlated positively with NFL (p < 0.001). HLA-DR was enriched in CSF EVs from HIV+ subjects with NCI (p < 0.05), suggesting myeloid cells are a potential source of CSF EVs during HIV infection. CONCLUSIONS: Increased CSF EVs correlate with neuronal injury biomarker NFL in cART-treated HIV+ individuals with neurocognitive impairment, suggesting potential applications as novel biomarkers of CNS injury.This is an open-access article distributed under the terms of the Creative Commons Attribution-Non Commercial-No Derivatives License 4.0 (CCBY-NC-ND), where it is permissible to download and share the work provided it is properly cited. The work cannot be changed in any way or used commercially without permission from the journal. http://creativecommons.org/licenses/by-nc-nd/4.0.
2018
Aldersley J, Lorenz DR, Misra V, Uno H, Gabuzda D. Increased risk of anal squamous cell carcinoma in HIV-positive men with prior HBV infection. AIDS. 2018.Abstract
OBJECTIVE(S): HIV-positive individuals have elevated rates of anal squamous cell carcinoma (SCC), and sexually transmitted infections with its causative agent, high-risk human papillomavirus, and other oncoviruses including hepatitis B virus (HBV). HBV infection can cause liver cancer, and has been associated with increased risk of some extra-hepatic cancers including biliary tract cancer, pancreatic cancer, and non-Hodgkin lymphoma. Whether HBV is associated with anal SCC risk is unknown. DESIGN: Prospective study of anal SCC risk in HIV-positive and -negative men who have sex with men (MSM) in the Multicenter AIDS Cohort Study from 1984-2014. METHODS: Poisson regression models were used to examine the association between past or current HBV infection (positive tests for HBV core antibodies, surface antigen, and/or DNA) and anal SCC risk. RESULTS: We observed 53 cases of anal SCC among 5,298 participants with 79,334 person-years follow-up. Among HIV-positive men, past or current HBV infection was associated with anal SCC risk in models adjusted for age, CD4+ cell counts, HAART use, and other risk factors (incidence rate ratio [IRR], 95% confidence interval [CI] 3.15, 1.27-7.82). Additional risk factors included immunological parameters one and six years prior to diagnosis (IRR, 95% CI 2.45, 1.31-4.58 and 2.44, 1.3-4.59 for CD4+ counts <500 cells/μl; 2.43, 1.34-4.42 and 2.77, 1.5-5.11 for CD4:CD8 ratios <0.5, respectively). Among HIV-negative men, IRR for prior HBV and anal SCC risk was similar, but not significant due to small number of cases. CONCLUSIONS: HIV-positive MSM with prior HBV infection have increased anal SCC risk. This population may benefit from screening.This is an open access article distributed under the terms of the Creative Commons Attribution-Non Commercial-No Derivatives License 4.0 (CCBY-NC-ND), where it is permissible to download and share the work provided it is properly cited. The work cannot be changed in any way or used commercially without permission from the journal. http://creativecommons.org/licenses/by-nc-nd/4.0.
Dutta A, Uno H, Lorenz DR, Wolinsky SM, Gabuzda D. Low T-cell subsets prior to development of virus-associated cancer in HIV-seronegative men who have sex with men. Cancer Causes Control. 2018.Abstract
Immunological parameters that influence susceptibility to virus-associated cancers in HIV-seronegative individuals are unclear. We conducted a case-control cohort study of immunological parameters associated with development of incident virus-associated cancers among 532 HIV-seronegative men who have sex with men (MSM) enrolled in the Multicenter AIDS Cohort Study (MACS) with median (IQR) 21 (8-26) years of follow-up. Thirty-two incident virus-associated cancers (anal cancer, non-Hodgkin lymphoma, liver cancer, other cancers with etiologies linked to human papillomavirus, Epstein-Barr virus, hepatitis B virus, or human herpesvirus-8) were identified among 3,408 HIV-seronegative men in the MACS during 1984-2010. Cases were matched for demographics, smoking, and follow-up to 500 controls without cancer. Mixed-effects and Cox regression models were used to examine associations between nadir or recent CD4, CD8, and white blood cell (WBC) counts or CD4:CD8 ratios and subsequent diagnosis of virus-associated cancers. Men with incident virus-associated cancers had lower CD4 and WBC counts over a 6-year window prior to diagnosis compared to men without cancer (p = 0.001 and 0.03, respectively). Low CD4 cell count and nadir, CD4 count-nadir differential, and CD4:CD8 ratio nadir were associated with increased 2-year risk of incident virus-associated cancers in models adjusted for demographics and smoking (hazard ratios 1.2-1.3 per 100 or 0.1 unit decrease, respectively; p < 0.01). Other associated factors included heavy smoking and past or current hepatitis B virus infection. These findings show that low CD4 cell counts, CD4 nadir, and CD4:CD8 cell ratios are independent predictors for subsequent risk of virus-associated cancers in HIV-seronegative MSM.
Chettimada S, Lorenz DR, Misra V, Dillon ST, Reeves KR, Manickam C, Morgello S, Kirk GD, Mehta SH, Gabuzda D. Exosome markers associated with immune activation and oxidative stress in HIV patients on antiretroviral therapy. Sci Rep. 2018;8 (1) :7227.Abstract
Exosomes are nanovesicles released from most cell types including immune cells. Prior studies suggest exosomes play a role in HIV pathogenesis, but little is known about exosome cargo in relation to immune responses and oxidative stress. Here, we characterize plasma exosomes in HIV patients and their relationship to immunological and oxidative stress markers. Plasma exosome fractions were isolated from HIV-positive subjects on ART with suppressed viral load and HIV-negative controls. Exosomes were characterized by electron microscopy, nanoparticle tracking, immunoblotting, and LC-MS/MS proteomics. Plasma exosomes were increased in HIV-positive subjects compared to controls, and correlated with increased oxidative stress markers (cystine, oxidized cys-gly) and decreased PUFA (DHA, EPA, DPA). Untargeted proteomics detected markers of exosomes (CD9, CD63, CD81), immune activation (CD14, CRP, HLA-A, HLA-B), oxidative stress (CAT, PRDX1, PRDX2, TXN), and Notch4 in plasma exosomes. Exosomal Notch4 was increased in HIV-positive subjects versus controls and correlated with immune activation markers. Treatment of THP-1 monocytic cells with patient-derived exosomes induced expression of genes related to interferon responses and immune activation. These results suggest that exosomes in ART-treated HIV patients carry proteins related to immune activation and oxidative stress, have immunomodulatory effects on myeloid cells, and may have pro-inflammatory and redox effects during pathogenesis.
Mukerji SS, Misra V, Lorenz DR, Uno H, Morgello S, Franklin D, Ellis RJ, Letendre S, Gabuzda D. Impact of Antiretroviral Regimens on CSF Viral Escape in a Prospective Multicohort Study of ART-Experienced HIV-1 Infected Adults in the United States. Clin Infect Dis. 2018.Abstract
Background: Cerebrospinal fluid (CSF) viral escape occurs in 4-20% of HIV-infected adults, yet the impact of antiretroviral therapy (ART) on CSF escape is unclear. Methods: Prospective study of 1063 participants with baseline plasma viral load (VL) ≤400 copies/ml between 2005-2016. Odds ratio for ART regimens (PI with nucleoside reverse transcriptase inhibitor [PI+NRTI] versus other ART) and CSF escape was estimated using mixed-effects models. Drug resistance mutation frequencies were calculated. Results: Baseline mean age was 46, median plasma VL, CD4 nadir, and CD4 count were 50 copies/mL, 88 cells/μL, and 424 cells/μL, respectively; 48% on PI+NRTI, 33% on non-NRTI, and 6% on integrase inhibitors. During median follow-up of 4.4 years, CSF escape occurred in 77 participants (7.2%). PI+NRTI use was an independent predictor of CSF escape (OR 3.1 [95% CI 1.8-5.0]) in adjusted analyses and models restricted to plasma VL ≤50 copies/ml (p<0.001). Regimens containing atazanavir (ATV) were a stronger predictor of CSF viral escape than non-ATV PI+NRTI regimens. Plasma and CSF M184V/I combined with thymidine-analog mutations were more frequent in CSF escape versus no escape (23% vs. 2.3%). Genotypic susceptibility score-adjusted CNS penetration-effectiveness (CPE) values were calculated for CSF escape with M184V/I mutations (n=34). Adjusted CPE values were low (<5) for CSF and plasma in 27 (79%) and 13 (38%), respectively, indicating suboptimal CNS drug availability. Conclusions: PI+NRTI regimens are independent predictors of CSF escape in HIV-infected adults. Reduced CNS ART bioavailability may predispose to CSF escape in patients with M184V/I mutations. Optimizing ART regimens may reduce risk of CSF escape.
Gumber S, Amancha PK, Yen P-J, Villinger F, Gabuzda D, Byrareddy SN. In vivo characterization of macrophage-tropic simian immunodeficiency virus molecular clones in rhesus macaques. J Neurovirol. 2018;24 (4) :411-419.Abstract
Macrophages are a major target of HIV/SIV infection and play an important role in pathogenesis by serving as viral reservoirs in the central nervous system. Previously, a unique early SIVmac251 envelope (Env) variant, deSIV147 was cloned from blood of a rhesus macaque with rapid disease progression and SIV-associated encephalitis. Here, we show that infectious molecular clone deSIV147 caused systemic infection in rhesus macaques following intravenous or intrarectal exposure. Next, we inoculated deSIV147 into macaques depleted of CD4+ T cells and found that animals were SIV-positive, with high plasma and CSF viral loads. These macaques also showed SIVp17-positive macrophages in brain, lymph nodes, colon, lung, and liver. Furthermore, accumulation of perivascular macrophages, multinucleated giant cells, and microgliosis was detected. These findings suggest that the neurotropic deSIV147 clone will be useful to study macrophage infection in HIV/SIV-associated neurocognitive disorders, gain insights into myeloid cell reservoirs in brain and other anatomical sites, as well as test strategies for eradication.

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