Background
People with HIV (PWH) have an increased risk of cardiovascular disease (CVD), however, the underlying mechanisms are not fully understood. Endothelial dysfunction is central to CVD pathology. Endothelial colony-forming cells (ECFC) arise from endothelial progenitors isolated from whole blood and retain the donor’s phenotypic characteristics ex vivo. ECFC are increasingly used to study disease-specific mechanisms in patient cohorts but have been largely overlooked in HIV research. We therefore isolated ECFC from PWH and people without HIV (PWoH) either with or without Pre-Exposure Prophylaxis (PrEP) exposure, to explore differences in ECFC biology.
Methods
Whole blood was obtained from PWH on fully suppressive antiretroviral therapy, PWoH accessing PrEP or not on PrEP attending HIV care or prevention clinics at Chelsea and Westminster Hospital. ECFC were isolated from peripheral blood mononuclear cells (PBMC). DNA and RNA was extracted from ECFC at passage 4. Proviral HIV DNA was measured by qPCR using primers and probes for the integrase gene. mRNA expression of interferon (IFN) signalling (e.g. IFI44, ISG15, MX1), thromboinflammation (e.g. F3, EDN1,VCAM1) and mitochondrial-metabolic (e.g. MTDN1, IDH1) markers was quantified by RT-PCR using SYBR Green.
Results
ECFC were isolated from PWH (PLWH-ECFC; n=8, 75% male, 87.5% white), PWoH accessing PrEP (PrEP-ECFC; n=9, 100% male, 88.9% white) and PWoH not on PrEP (NP-ECFC; n=3, 100% male, 66.7% white). A higher frequency of ECFC colonies were recovered from PLWH compared to people on PrEP when normalised to number of PBMC seeded (0.53 vs 0.13 vs colonies/1×106 PBMC, p=0.0085). In PLWH, proviral HIV DNA was detected in matched PBMC but not in ECFC (1.7 vs 0.0 copies/104 cells, p=0.0002). Elevated IFN, thromboinflammatory, mitochondrial and metabolic associated gene expression was seen in PLWH-ECFC relative to NP- and PrEP-ECFC. IFN gene expression strongly correlated with inflammatory and metabolic genes.
Conclusions
PWH displayed higher ECFC frequencies, although the clinical significance of this is unclear. Despite the absence of proviral HIV DNA, PLWH-ECFC displayed an IFN gene signature which positively correlated with elevated inflammatory and metabolic markers. Collectively, our observations indicate endothelial dysfunction in PLWH-ECFC despite fully suppressive ART, that may be driven by IFN signalling. Further investigation is required to fully define mechanisms of endothelial dysfunction and to identify therapeutic targets in PWH.
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