Patho-immune Mechanisms of Hypertension in HIV: a Systematic and Thematic Review

Purpose of Review To systematically review recent findings on the role of immune cell activation in the pathogenesis of hypertension in people living with HIV (PLWH) and compare studies from Sub-Saharan Africa with what is reported in the USA and European literature according to guidelines of the Preferred Reporting Items for Systematic Reviews and Meta-Analyses. Recent Findings PLWH have an increased risk for development of hypertension and cardiovascular disease. Chronic immune activation contributes to hypertension but the inflammatory milieu that predisposes PLWH to hypertension is poorly understood. We identified 45 relevant studies from 13 unique African countries. The prevalence of hypertension in PLWH on antiretroviral therapy (ART) and the ART-naive PLWH ranged from 6 to 50% and 2 to 41%, respectively. Interleukin (IL)-17A, interferon (IFN)-γ, and higher CD4+ T cell counts were associated with hypertension in ART-treated participants. Summary Targeting adaptive immune activation could provide improved care for hypertensive PLWH. Further research is needed to characterize the inflammatory milieu contributing to hypertension in PLWH especially in African populations where the global burden of HIV is the highest.


Introduction
The introduction of antiretroviral therapy (ART) has improved survival among people living with HIV (PLWH), but this success is accompanied by a high burden of cardiovascular disease [1][2][3]. Hypertension is a major risk factor to cardiovascular disease and its prevalence is higher in PLWH despite viral suppression by ART [4]. The mechanisms contributing to hypertension and cardiovascular disease in PLWH are not fully understood.
A heightened systemic inflammatory process including activation of the innate and adaptive immune systems contributes to the development of hypertension in the general population and in experimental animal studies [5][6][7]. However, it is unknown which innate and adaptive immune factors are most closely linked to hypertension in PLWH. At present, much of our understanding of the role of immune cell activation in the pathogenesis of hypertension comes from experimental animal and human studies in the HIV-negative context, but similar inflammatory biomarkers implicated in these HIVnegative studies have also been associated with hypertension in HIV infection. In this review, we present the current understanding on HIV, hypertension, and immune activation and how these may be related and highlight gaps and the need for further studies. We review studies conducted in Sub-Saharan Africa where the global HIV infection burden is the highest. We also performed a comparative analysis contrasting studies from Sub-Saharan Africa with what is reported in the USA and European literature.
The aim of our current study was to estimate the prevalence of hypertension in PLWH from African studies, identify inflammatory markers or markers of immune activation associated with hypertension in antiretroviral (ART)-treated HIVpositive individuals, and to explore the possible mechanistic interaction between immune activation, hypertension, and HIV in cohorts from Sub-Saharan Africa and contrast with European and US cohorts.

Methods
We followed guidelines of the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) [8,9].
Eligibility Criteria for Studies Included in the Review We considered studies on HIV-positive human subjects with or without the HIV-negative population as controls. For exposure and outcome variables, we considered HIV infection as the exposure and our outcome measures were hypertension or blood pressure, inflammation, and/or immune activation. Only published articles in peer-reviewed journals were considered. Articles in languages other than English were not considered. We only included studies from Sub-Saharan African, USA, and European HIV-positive populations that reported on hypertension and HIV excluding studies that did not report HIV at all. We also excluded studies reporting opportunistic infections and other coinfections to reduce bias when deducting on the contribution of HIV on hypertension. In case of duplicate publications, the article with more complete data was included. Extracted information focused on study design, population, sample size, country where study was conducted, and details of findings with conclusions. The source of research articles was derived from Pubmed, Medline, google scholar, and google search. Dates of coverage were not specified to filter studies addressing hypertension and inflammation or immune activation in HIV owing to the scarcity of published studies. Other articles were identified from reference lists of related studies from the included study. Search terms that were used in databases employed the use of Boolean operators "AND," "OR," and "NOT" to refine our search by combining or limiting terms. The key terms used to search for articles were "hypertension," "blood pressure," "immune activation," "inflammation," "Africa," and "HIV." Studies were screened based on abstract information that reported HIV or/and inflammation or immune activation and hypertension or blood pressure. A full article was then obtained and screened to ensure eligibility. The PRISMA 2009 Flow Diagrams for article identification, screening, eligibility, and inclusion in Sub-Saharan African and the Western populations are shown in Supplementary Figures 1 and 2 found at the f o l l o w i n g l i n k : h t t p s : / / f i g s h a r e . c o m / s / 38ff86137d00153423ac. For data extraction, we used a data collection form that identifies the author, study, population, sample size, methods, findings, and limitations.
Risk of bias was assessed by two reviewers. For the criteria used to assess internal validity of included studies, we made use of the Cochrane Risk of Bias tool, paying attention especially to parts such as completeness of outcome data, selective outcome reporting, reporting bias, and any other bias that may affect the exposure-outcome relationship. For any disagreements between the reviewers over the risk of bias in particular studies, this was resolved by discussion, with involvement of a third review author where necessary.

Data Synthesis
We did not conduct a meta-analysis. We only conducted a systematic thematic review due to the qualitative nature of our study and due to excessive heterogeneity of population, outcome, or methodology. To assist with the result and discussion write-ups, we constructed summary tables and provided a narrative synthesis (thematic or content analysis). This includes investigation of the similarities and the differences between the findings of different studies, as well as exploration of patterns in the data. Reasons for similarities and differences in the findings were also explored systematically. Studies were grouped based on reported parameters. We also considered how the results of studies might be affected by factors such as methodological differences, variable characteristics of the populations studied, or interventions investigated. Conceptual models were used to explore relationships and patterns from study findings and we related the findings to existing concepts from animal models to generate hypotheses and the need for further studies.

Results
Characteristics of the Studies We performed a systematic review of studies on HIV, hypertension, and immunity from countries in Sub-Saharan Africa, USA, and Europe. For Sub-Saharan Africa studies shown in Table 1, we included 45 studies from 13 countries. Of the 45 studies, 27 were crosssectional, 15 were prospective, and three were retrospective. Most of the studies (35) reported on both hypertension and HIV. The prevalence of hypertension was highest in the HIVpositive group on ART (ranging from 6.4 to 50.2%) followed by HIV-negative controls (13.7 to 44%). ART-naive patients recorded the lowest prevalence ranging from 2 to 41% by group comparisons. The prevalence differences of groups within studies were highest between ART treated and ARTnaive HIV-positive groups (35.7) followed by ART treated and HIV-negative group (18.7) with the least difference in magnitude between HIV negative and ART-naive groups.
When we included USA and European studies, shown in Supplementary Table 1 found at the following link: https:// figshare.com/s/38ff86137d00153423ac, the global prevalence of hypertension in PLWH ranged from 4 to 57% (van Ziest et al. 2017) [10]. Hypertension was more prevalent in HIV uninfected (71%) versus HIV infected (57%) in a large longitudinal study by Armah et al. [11]. HIV-infected veterans with HIV-1 RNA ≥ 500 copies/mL or CD4 count < 200 cells/ μL had a significantly higher prevalence of elevated IL-6 and D-dimer after adjusting for comorbidities and had significantly higher prevalence of elevated sCD14 compared to uninfected veterans. Manner et al. [12] reported prevalence of 35% hypertension in HIV-infected individuals and the prevalence did not change during the follow-up time (3.4 ± 0.8 years). CD4 T cell count < 50 cells/μL and increased duration of ART were independent predictors of sustained hypertension throughout the study period. Older age, male gender, BMI > 25 kg/m 2 , and baseline CD4 cell count ≥ 200 cells/μL were also independent predictors of sustained hypertension. Markers of microbial translocation predicted hypertension in HIV-infected individuals. Manner et al. found that both LPS and sCD14 independently predicted subsequent blood pressure levels after adjustment for age and gender [12]. These results suggest that ART may act as a contributing factor to inflammation and the increased prevalence of hypertension in the PLWH.
Higher Levels of IL-17A, IFN-γ, and CD4 + T Cells Are Associated with Hypertension in HIV We found that among studies reporting on both inflammation, immunity, and hypertension in HIV (Table 2), higher levels of IL-17A, IFN-γ, [13••], and CD4 + T cell count (Peck et al.) [4] were significantly associated with hypertension in ART treated HIV-positive individuals. The cross-section study by Chepchirchir et al. reported that females were more likely to have higher IL-17A levels than males and IL-17A was affected by BMI but not stress levels, ART, World Health Organization (WHO) stage, and CD4 + count [13••]. However, inflammatory cytokines IL-2, IL-6, IL-8, tumor necrosis factor alpha (TNF-α), and antiinflammatory cytokines IL-4 and IL-10 were not associated with hypertension. The cross-section study by Peck et    Prevalence of Hypertension Is Higher in ART-Treated PLWH As shown in Table 3, hypertension prevalence was mostly higher in men than women [15][16][17] and higher among ART treated versus ART naive (28.7% vs 5.3%; 17% vs 2%; 30 vs 21.9% and 38% vs 19%, respectively) [17,18]. However, one study [22] reported contradictory findings (ART vs ART naive, 12.3 vs 19%). In another study, [17] reported higher prevalence of hypertension in ART treated group (28.7%) compared to the HIV-negative controls (16.3%) but two studies [22,25]  ART Treatment in PLWH Is Associated with Higher Inflammation As shown in Table 4, we found that PLWH on ART had higher inflammatory markers including IL-6, CRP, intracellular adhesion molecule 1 (ICAM-1), and vascular adhesion molecule 1 (VCAM-1) compared with HIV-negative individuals (Fourie et al.) [55]. Intermediate monocytes (CD14 + 16 + ) increased with viremia in immunecompromised patients and microbial translocation was a major force driving chronic systemic inflammation in HIVpositive individuals on ART.

Discussion
In the current study, we found that the prevalence of hypertension in PLWH in Sub-Saharan African countries ranged from 2.0 to 50.2% with most cases among those receiving ART. Besides traditional risk factors and the effect of ART on blood pressure, IL-17A, IFN-γ, and CD4 + T cells were among the inflammatory parameters associated with hypertension in ART treated PLWH [13••, 56]. The mechanism of interaction between immune activation or inflammation and hypertension in HIV remains elusive and warrants further studies. Our findings and conceptual hypothesis of how immune activation may contribute to hypertension in HIV is shown in Fig. 1. We propose that HIV viral protein and ART interacts with components of the immune system to synergistically induce kidney damage, vascular dysfunction, alterations in sympathetic nervous outflow and lead to hypertension. This process is exacerbated by presence of traditional risk factors exacerbates including obesity, excess dietary salt intake, smoking, and genetic predisposition.  [58]. However, in our study, we further segregated prevalence by HIV infection and ART treatment and report magnitude of differences between the groups (Table 1). Higher hypertension prevalence was reported in ART treated PLWH compared with ART-naive participants except in the study by Ogunmola et al. who reported the opposite [22]. This      Immune mechanism or inflammation as it relates to hypertension was not addressed  Immune mechanism or inflammation as it relates to hypertension was not addressed contradictory finding is likely due to differential effects of specific ART regimens and other traditional risk factors differing between study populations. The effect of specific regimens on blood pressure has not yet been well established except for the low to moderate increase attributed to nonnucleoside reverse transcriptase inhibitors (NNRTI's) and protease inhibitors (PIs) [59,60]. Prior studies have shown that patients become hypertensive in most cases at least after 2 years of ART and systolic pressure increases further after 5 years of ART [60], and that there is no association between HIV status, ART, and hypertension following short-term follow-up of less than 2 years [22,61].

Markers of Immune Activation or Inflammation Associated with Hypertension in HIV
Traditional risk factors associated with hypertension in HIV such as older age, male gender, African-American, higher BMI, central obesity, previous CV events, chronic kidney disease, family history of hypertension and CVD, diabetes, and dyslipidemia have been well studied [57••, 58]. It is also well established that HIV infection and exposure to ART (more than 2 years), through metabolic disturbances and endothelial dysfunction, might have an additional role in the development of hypertension in HIV patients [57••, 58]. However, little is known about the contribution of the innate and adaptive immune system in the development or propagation of hypertension in HIV. In our systematic review, we found that an improved immune status as determined by higher CD4 T cells was associated with higher hypertension prevalence [17]. However, one study be Okello et al. found that CD4 T cell count of less than 100 was associated with incident hypertension [61]. Martin-Iguacel et al. and Nguyen et al. also reported similar findings [57••, 58]. We found that T cell-derived cytokines IL-17A and IFN-γ were associated with hypertension in ART treated PLWH as reported by Chepchirchir et al. [13••]. These cytokines have been reported to contribute to the genesis of hypertension in the HIV-negative general population and experimental animal studies [6]. Further studies are needed to ascertain the contribution of CD4 T cells and their cytokines IL-17 and IFN-γ in concert with existing traditional risk factors on blood pressure elevation in HIV.

Immune Activation, Inflammation, and Hypertension
The role of innate and adaptive immunity including specific cell types and cytokines in the development and maintenance of hypertension has been extensively studied in humans and animal models elsewhere [6, 7, 62•, 63-65]. Using multiple experimental animal models, studies have shown that hypertensive stimuli including angiotensin II, aldosterone, endothelin-1, and salt induce activation of immune cells, which infiltrate the vasculature and the kidneys, and release cytokines that induce increased salt and water retention leading to hypertension [6]. This process is mediated in part by increases oxidative stress leading to oxidation of fatty acids and formation of isolevuglandins (IsoLGs) in antigenpresenting cells. IsoLGs activate these cells to produce proinflammatory cytokines IL-6, IL-1β, and IL-23, express costimulatory proteins CD80 and CD86, and activate T cells to produce pro-hypertensive cytokines IFN-γ and IL-17A [7]. In humans, plasma isoprostanes, which are produced in concert with IsoLGs, are elevated in hypertension and markedly elevated in patients with resistant hypertension, and IsoLGs are markedly elevated in circulating monocytes of hypertensive patients [7]. IL-17A and IFN-γ have been implicated in the genesis and maintenance of hypertension due in part to their direct effect in causing endothelial dysfunction and renal damage [6,66]. Thus, similar immuno-pathophysiological mechanisms underlying hypertension in the general population may also contribute to elevated blood pressure in PLWH.

Potential Mechanisms Leading to Inflammation in HIV
The pathophysiology of HIV-related hypertension seems to emerge from three factors: HIV-related inflammation, HIVrelated proteins, and genetic predisposition [59]. It is believed that the HIV viral proteins (negative factor (Nef), transcription proteins (Tat) and glycoprotein 120 (gp-120)) induce hypertension through vascular oxidative stress, smooth myocyte proliferation and migration, and endothelial dysfunction especially in patients with high HIV viral load [59]. Tat, a transactivator protein for HIV replication, is known to be secreted extracellularly by infected cells and has been shown to activate endothelial cells by increasing expression of endothelial-leukocyte adhesion molecules such as intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and E-selectin which induce initial binding of leukocytes to the blood vessel wall [67]. The levels of soluble ICAM-1 concentration correlate with HIV disease as well as reduction in CD4 count. Tat also induces IL-6 production which increases endothelial permeability [67]. Tat can also suppress the bone morphogenic protein receptor 2 (BMPR-2) responsible for regulating endothelial cell proliferation and survival. This results in increased vascular smooth muscle proliferation and activation of endothelial cells leading to pulmonary arterial hypertension (PAH) [68]. In HIV-positive patients, Tat has been shown to increase the transcription of IL-17 and secretion by T cells causing a pro-inflammatory milieu and has been associated with a devastating immune reconstitution inflammatory syndrome (IRIS) in the brain [69]. The HIV Gp120 is found on the surface of the HIV envelope and can also be found in circulation from viral turn over [70]. Gp120 stimulates monocytes and macrophages to release pro-inflammatory cytokines, promotes an increase in markers of apoptosis, and stimulates the secretion of endothelin-1, a potent vasoconstrictor [68]. It induces endothelial apoptosis by interacting with CXCR4 also expressed on endothelial cells [70]. Gp120 and TNF-α synergistically decrease eNOS and NO in human coronary artery endothelial cells [70]. DCs, the most potent antigen-presenting cells of the innate immune system, have been shown to interact with components of HIV-1 both intrinsically (following fusion of cellular and viral membrane) and extrinsically (prior to being infected) through pathogen recognition receptors (PRRs) resulting in activation of the adaptive immune system [71]. Through their CD28/B7 ligands CD80 and CD86 (also expressed on monocytes and B cells), DCs provide a costimulatory signal necessary for T cell activation and survival. In hypertension, there is an increase in B7 ligand expression leading to T cell activation [5,72]. An important element in the activation of T cells by DCs and other cells in hypertension is antigen presentation. It is unclear what antigens are presented to T cells in HIV and it is not known if IsoLG-adducted peptides might play a role in hypertension associated with HIV.
Monocyte activation has an important role in HIV-infected persons on ART. Monocytes are chronically activated during HIV infection and it is now evident that inflammatory mediators produced by monocytes (especially IL-6), independent of T cell activation, also predict serious non-AIDS events (SNAEs) in virologically suppressed HIV-infected persons treated with ART [73]. IL-6 production was higher in monocytes than other cells and was associated with increased odds of SNAE and mortality but not the percentage of activated CD4 and CD8 T cells (those expressing CD38 or CD38 and HLA-DR) [73]. Infected monocytes, especially intermediate monocytes expressing CD14 + and CD16 + , produce more proinflammatory cytokines as compared to their counterpart. They adhere to endothelial cells and transmigrate into the sub endothelial area where their pro-inflammatory activity is increased. It is evident that in HIV, monocytes and macrophages have a reduced phagocytic activity and they age prematurely [70]. Monocytes/macrophages have been implicated in the pathogenesis of hypertension in the general population and experimental animal models [5].
Several mechanisms of immune cell activation have been reported in HIV. First, there is direct binding of the HIVenvelope protein gp120/160 to CD4 and/or C-C chemokine receptor type Fig. 1 Conceptual schematic of the effect of HIV infection and treatment can activate the immune system leading to hypertension. Viral proteins and/or antiretroviral therapy (ART) activates antigen presenting and T cells which infiltrate the vasculature and the kidneys and release cytokines IL-6, IL-17A, and IFN-γ which promote vascular dysfunction, retention of sodium, and water, leading to hypertension IL-6 production by monocytes and macrophages is increased following HIV infection and is a valuable prognostic marker for disease progression [77,78]. Soluble CD14 (sCD14) produced from monocytes and macrophages is also elevated in HIV-infected persons regardless of treatment status and is an independent predictor of mortality [73]. Anzinger et al. reported that increased microbial translocation from the gut resulting from depletion of CD4 + T cells in acute HIV infection results in monocyte activation and higher sCD14 [73]. The expression of programmed death-1 (PD-1), a member of the B7:28 inhibitory molecules, is rapidly increased on CD4 + and CD8 + T cells, B cells, natural killer cells, and monocytes as they interact with PD-L1, its ligand on antigenpresenting cells such that engagement on virus-infected cells leads to impaired generation of effector T cell responses [74]. Other markers which have been associated with immune activation in HIV include increased expression of CD28 − , CD57 + and reduced expression of CD127 on CD4 + and CD8 + T cells [79,80].
In our analysis, we found HIV infection alone is associated with an inflammatory milieu involving higher levels of IL-6, CRP, VCAM-1 ICAM-1, and intermediate monocytes (CD14 + 16 + ) among studies conducted in Sub-Saharan African cohorts [17,55]. There is a generalized systemic immune activation following HIV infection which subsides but still persists at low-grade or subclinical following initiation of ART involving activation of both CD4 + and CD8 + T cells expressing high levels of CD38 [74]. Other T cell activation markers such as human leukocyte antigen-antigen D related (HLA-DR) as well as increases in pro-inflammatory cytokines tumor necrosis factor alpha (TNF-α), interleukin 6 (IL-6), and interleukin 1 beta (IL-1β) have been reported in PLWH in the western countries [74]. Increased expression of CD38 alone or in concert with HLA-DR + in T cells is a marker of disease progression and mortality in some cases correlating directly with HIV viral load and indirectly with CD4 + count [74]. This continuous immune activation and inflammation has been proposed to contribute to endothelial dysfunction, hypertension, and cardiovascular disease [81].

Limitations, Conclusion, and Future Perspectives
Almost all studies except one [13••] were not designed to compare blood pressure or hypertension and inflammation in PLWH as the primary outcomes. There is need for additional prospective and mechanistic studies to establish the relationship between inflammation and hypertension in HIV infection, and ART. Most studies addressing the mechanistic role of inflammation in the genesis and progression of hypertension have been conducted in murine models and humans in the HIV-negative context. The inflammatory milieu in HIV infection is obviously complex and very different from the general population and further studies are needed to determine the specific contribution of the various attributes in HIV including the viral proteins and ART on inducing inflammation associated with hypertension.