Filling the gaps: Towards improved surveillance and monitoring of immunological status relevant to long term co-morbidities in HIV infection

It is approaching 35 years since human immunodeficiency virus (HIV)-1 was first identified in Western Australia (WA) and 20 years since introducing highly active antiretroviral therapy (HAART) transforming HIV management, in the majority of cases, by maintaining ‘undetectable’ HIV-1 RNA and restoring CD4 T cell counts to healthy levels. Despite these advances patients are at increased risk of developing age-associated diseases. This thesis therefore investigates whether expanding laboratory approaches to incorporate aspects of the disease process, that are currently 'invisible' in clinical practice but may be prognostically important or which may inform treatment and prevention strategies at a population level, is warranted. The following aims were addressed; 1) analysing determinants of HIV-1 RNA residual viraemia; 2) investigating monocyte activation during chronic HIV-1 infection and; 3) assessing HIV-1 diversity from large scale sequences throughout WA and Australia, including the collaboration and formation of the Australian Molecular Epidemiology Network (AMEN). The main findings demonstrated HIV-1 residual viraemia was powerfully associated with the pre-treatment level of viraemia even after 10-15 years of starting HAART. Analysis of monocyte activation revealed multiple pathways involved in chronic immune activation responses to HIV-1, by altering CD16+ monocyte expression and elevated levels of sCD14 (previously associated with all-cause mortality), that were not corrected by HAART and therefore prognostically significant. In contrast, levels of other biomarkers (e.g. CXCL10 and sCD163) declined with HAART. Investigations of HIV-1 genetic diversity within the WA cohort (n=1021) and the Australian cohort (n=4873) revealed new challenges for HIV-1 prevention due to significant increases in HIV-1 non-B-subtypes, consistent with an increasing impact of migration, while local transmissions were predominantly HIV-1 B-subtypes. Overall, this thesis provides evidence for possible new approaches that may enhance HIV-1 laboratory practice that could aid clinical management, improve long-term health outcomes for those living with HIV-1 infection and guide effective preventative strategies

Elevated plasma soluble CD14 and skewed CD16+ monocyte distribution persist despite normalisation of soluble CD163 and CXCL10 by effective HIV therapy: a changing paradigm for routine HIV laboratory monitoring?

OBJECTIVE: We investigated plasma and flow cytometric biomarkers of monocyte status that have been associated with prognostic utility in HIV infection and other chronic inflammatory diseases, comparing 81 HIV+ individuals with a range of treatment outcomes to a group of 21 healthy control blood donors. Our aim is to develop and optimise monocyte assays that combine biological relevance, clinical utility, and ease of adoption into routine HIV laboratory practice. DESIGN: Cross-sectional evaluation of concurrent plasma and whole blood samples. METHODS: A flow cytometry protocol was developed comprising single-tube CD45, CD14, CD16, CD64, CD163, CD143 analysis with appropriately matched isotype controls. Plasma levels of soluble CD14 (sCD14), soluble CD163 (sCD163) and CXCL10 were measured by ELISA. RESULTS: HIV status was associated with significantly increased expression of CD64, CD143 and CD163 on CD16+ monocytes, irrespective of the virological response to HIV therapy. Plasma levels of sCD14, sCD163 and CXCL10 were also significantly elevated in association with viremic HIV infection. Plasma sCD163 and CXCL10 levels were restored to healthy control levels by effective antiretroviral therapy while sCD14 levels remained elevated despite virological suppression (p<0.001). CONCLUSIONS: Flow cytometric and plasma biomarkers of monocyte activation indicate an ongoing systemic inflammatory response to HIV infection, characterised by persistent alterations of CD16+ monocyte expression profiles and elevated sCD14 levels, that are not corrected by antiretroviral therapy and likely to be prognostically significant. In contrast, sCD163 and CXCL10 levels declined on antiretroviral therapy, suggesting multiple activation pathways revealed by these biomarkers. Incorporation of these assays into routine clinical care is feasible and warrants further consideration, particularly in light of emerging therapeutic strategies that specifically target innate immune activation in HIV infection

Plasma CXCL10, sCD163 and sCD14 Levels Have Distinct Associations with Antiretroviral Treatment and Cardiovascular Disease Risk Factors - Fig 3

CXCL10, soluble CD14 and soluble CD163 have distinct although overlapping associations with cardiovascular disease risk factors, HIV treatment, ethnicity and age (A) along with different aspects of HIV infection (B).</p

Model 2—Multivariate regression results showing significant associations of plasma biomarker with HIV clinical parameters, CVD risk age, gender, ethnicity and smoking after adjusting for CXCL10, sCD163 and sCD14.

<p>Model 2—Multivariate regression results showing significant associations of plasma biomarker with HIV clinical parameters, CVD risk age, gender, ethnicity and smoking after adjusting for CXCL10, sCD163 and sCD14.</p

sCD14 levels are significantly elevated in HIV patients irrespective of HIV viral load status (A).

<p>In contrast sCD163 levels are only elevated in viremic HIV patients (B) and a similar relationship is seen with CXCL10 levels (C). (Data presented as mean values and 95% confidence intervals; p-values are derived from log transformed data and ANOVA tests were adjusted for multiple comparisons).</p