Multiplex Analysis of Cytokines and Chemokines in Persons Aging With or Without HIV

Abstract

People with HIV (PWH) on combination antiretroviral therapy (cART) are living longer lives due to modern cART advances and increased routine medical care. The full landscape of aging with HIV is unclear; given that HIV emerged relatively recently in human history and initially had a high mortality rate, there has not been a substantially aged population to evaluate. In this study, we set out to perform high-throughput plasma analyte profiling by multiplex analysis, focusing on various T helper (Th)-related cytokines, chemokines, and proinflammatory and anti-inflammatory cytokines. The primary goals being to provide reference ranges of these analytes for aging PWH cohorts, as well as testing the utility of high-throughput multiplex plasma assays. The cohort used in this study comprised age-matched healthy donors (32.6-73.5 years of age), PWH on cART (26.7-60.2 years of age), and viremic PWH (27.5-59.4 years of age). The patients in each group were then stratified across the age span to examine age-related impacts of these plasma biomarkers. Our results largely indicate feasibility of plasma analyte monitoring by multiplex and demonstrate a high degree of person-to-person variability regardless of age and HIV status. Nonetheless, we find multiple associations with age, duration of known infection, and viral load, all of which appear to be driven by either prolonged HIV disease progression or long-term use of cART.

Keywords: aging; inflammation; soluble mediators.

FIG. 1.

Plasma concentrations of Th1/Th2 analytes. Violin plots of Th1/Th2 analyte concentrations among HD, cART, and viremic individuals younger than vs older than 45 years. (A) GM-CSF; (B) IFN-γ; (C) IL-1β; (D) IL-2; (E) IL-4; (F) IL-5; (G) IL-6; (H) IL-8; (I) IL-12p70; (J) IL-13; (K) IL-18; (L) TNF-α. cART, combination antiretroviral therapy; GM-CSF, granulocyte macrophage colony-stimulating factor; HD, healthy donors; IFN-γ, interferon-gamma; IL, interleukin; TNF-α, tumor necrosis factor-alpha.

FIG. 2.

Plasma concentrations of Th9/Th17/Th22/Treg analytes. Violin plots of Th9/Th17/Th22/Treg analyte concentrations among HD, persons on cART, and viremic PWH younger than vs older than 45 years. (A) IL-9; (B) IL-10; (C) IL-17; (D) IL-21; (E) IL-22; (F) IL-23; (G) IL-27. PWH, people with HIV.

FIG. 3.

Plasma concentrations of inflammatory cytokines. Violin plots of inflammatory cytokine concentrations among HD, persons on cART, and viremic PWH younger than vs older than 45 years. (A) IFN-α; (B) IL-1α; (C) IL-1RA; (D) IL-7; (E) IL-15; (F) IL-31; (G) TNF-β. TNF-β, tumor necrosis factor-beta.

FIG. 4.

Plasma concentrations of chemokines. Violin plots of chemokines concentrations among HD, person on cART, and viremic PWH younger than vs older than 45 years. (A) CCL11*; (B) CXCL1; (C) CXCL10; (D) CCL2; (E) CCL3; (F) CCL4; (G) CCL5; (H) SDF-1α. *CCL11 concentrations were higher among cART older than 45 years group compared to viremic younger than 45 years group (p = .0336).

FIG. 5.

Luminex output. Heatmap plotted as individual samples showing analyte expressions per group: HD, PWH on cART, and viremic PWH. Each of these three groups was then divided into age groups, younger than 45 and older than 45 years. Colorimetric scale displays z-score values for that sample. Mean analyte expression was calculated per row and z-scores were calculated per sample. Color images are available online.

FIG. 6.

PCA output. PCA reduction using no group labels and all analytes. (A) PCA plot with color of points denoting group. Filled circles represent samples older than 45 years and empty circle represents samples younger than 45 years. (B) Analyte loadings from PCA analysis. Arrows denote magnitude and direction of loading for each principal component. PCA, principal component analysis. Color images are available online.