T-lymphocyte Phenotypic and Mitochondrial Parameters as Markers of Incomplete Immune Restoration in People Living with HIV+ on Long-Term cART

Background/Objectives: Restored CD4 absolute counts (CD4AC) and CD4/CD8 ratio in the settings of continuous antiretroviral treatment (ART) do not exclude a low-level immune activation associated with HIV reservoirs, microbial translocation, or the side-effects of ART itself, that accelerates ageing of people living with HIV (PLHIV). To delineate biomarkers of incomplete immune restoration in PLHIV on successful ART, we evaluated T-lymphocyte mitochondrial parameters in relation to phenotypic markers of immune exhaustion and senescence. Methods: PLHIV with sustained viral supression, CD4AC >500 and CD4/CD8 ratio >0.9 on ART (n=39) were compared to age-matched ART-naïve donors (n=27) and HIV(-) healthy controls (HC, n=35). CD4 and CD8 differentiation and effector subsets (CCR7/CD45RA and CD27/CD28), activation, exhaustion, and senescence markers (CD38, CD39 Treg, CD57, TIGIT, PD-1) were determined by flow cytometry. Mitochondrial mass (MM) and membrane potential (MMP) of CD8 and CD4 T cells were evaluated with MitoTracker Green and Red flow cytometry dyes. Results: ART+PLHIV differed from HC by increased CD4 TEMRA (5.3 (2.1-8.8) vs. 3.2 (1.6-4.4), p<0.05), persistent TIGIT+CD57-CD27+CD28- CD8+ subset (53.9 (45.5-68.9) vs. 40.1 (26.7-58.5), p<0.05), and expanding preapoptotic TIGIT-CD57+CD8+ effectors (9.2 (4.3-21.8) vs. 3.0 (1.5-7.3), p<0.01) in correlation with increased CD8+ MMP (2527 (1675-4080) vs.1477 (1280-1691), p<0.01). These aberrations were independent of age, time to ART, ART duration, and were combined with an increasing CD4 T cell MMP and MM. Conclusions: In spite of recovered CD4AC and CD4/CD8, the increased CD8+ MMP combined with elevated markers of exhaustion and senescence in ART+PLHIV, signals a malfunction of the CD8 effector pool, that may compromise viral reservoirs latency.