CROI 2016 Abstract eBook
411 Use of Mass Cytometry to Study Cellular Subsets in CSF and Blood in ART-Treated HIV Shang-Lin Chung ; DianeTrotta; Leah Le; Laurie Andrews; Jennifer Chiarella; Khadir Raddassi; Serena S. Spudich; Brinda Emu Yale Univ Sch of Med, New Haven, CT, USA
Background: HIV leads to immunological abnormalities in the blood and cerebrospinal fluid (CSF) that persist despite effective antiretroviral therapy (ART). Novel methods are needed to comprehensively characterize these changes. Mass cytometry (CyTOF) uses metal-conjugated antibodies to label cells allowing for larger number of markers compared to flow cytometry. We used both CyTOF and flow cytometry to examine cell surface markers in blood and CSF in participants on ART, despite low CSF white blood cell counts (WBC). Methods: Fresh specimens of blood and CSF were analyzed by CyTOF (n=4) and flow cytometry (n=4) from HIV infected participants on ART with plasma viral suppression, and HIV-uninfected individuals (n=3). For CyTOF, 20 markers were analyzed in a single panel (CD3, CD4, CD8, , CD11c, CD14, CD16, CD19, CD27, CD28, CD25, CD38, CD45, CD45RO, CD56, CD123, CD127 , CCR7, HLA-DR, PD-1, cisplatin) to identify T cells (including exhaustion, activation, maturation, regulatory phenotypes), monocytes, NK cells, and dendritic cells. For flow cytometry, two different panels were employed (A) CD3, CD4, CD8, CD27, CD28, CD38, CD45RA, CCR7, HLA-DR, PD- 1; and (B) CD3, CD11c, CD14, CD16, CD19, CD45, CD56, CD123, and HLA-DR. Results: CSF WBC was < 5 cells/mcl except for a single patient with 25 cells/mcl. Total CSF cell count captured on CyTOF ranged between 1981-111443 events (median 5679). Phenotypically-related cell clusters were generated based on a hierarchical algorithm though SPADE (see figure for representative analysis), allowing for evaluation of T and non-T cell subsets. In analysis of CyTOF data, a unique effector memory (EM) subset comprised the largest proportion of CSF CD4+ and CD8+ T cells (CD45RO+CCR7-CD27+CD28+), which was higher than that seen in blood. Data from flow cytometry confirmed that this EM population was higher in CSF compared with blood in CD4+ (65.1% vs 24.9%, p=0.0001) and CD8+ T cells (53.3% vs 20.4%, P=0.0004). This EM subset in CSF had a higher proportion of CD38+/HLA-DR+ (14.1 vs 4.1%, P=0.03 in CD4; 23.3 vs 3.5%, P=0.03 in CD8) and PD-1 (58.3% vs 34.7%, P=0.003) expression as compared to blood by flow. Conclusions: This pilot study has for the first time demonstrated meaningful use of CyTOF to analyze immune cells in CSF. Our results and methodology provide opportunity for future studies to reveal intricate CSF cellular phenotypic patterns of numerous disorders, including neurocognitive disorders associated with treated HIV.
412 CNS Drug Distribution and CSF Inflammation During Suppressive Antiretroviral Therapy Albert Anderson 1 ; Jennifer Iudicello 2 ; Asha R. Kallianpur 3 ; KarenTolentino 2 ; Ron Ellis 2 ; Igor Grant 4 ;Todd Hulgan 5 ; Scott R. Letendre 2 ; for the CHARTER Group 1 Emory Univ, Atlanta, GA, USA; 2 Univ of California San Diego, San Diego, CA, USA; 3 Cleveland Clinic/Lerner Rsr Inst, Cleveland, OH, USA; 4 Univ of California San Diego, La Jolla, CA, USA; 5 Vanderbilt Univ, Nashville, TN, USA Background: Antiretroviral therapy (ART) drugs differ in their distribution into the CNS. Greater estimated ART distribution into the CNS is associated with lower HIV RNA levels in CSF but the relationship with inflammation in the CNS during viral suppression is relatively unknown. The objective of this analysis was to assess this using a training-validation approach with the hypothesis that higher CNS penetration-effectiveness (CPE) values would be associated with lower levels of inflammation-associated biomarkers in CSF.
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