Discussion
Principal findings
Using novel data and approaches, we offer comprehensive genetic insight into the determinants, cascades, and effects of circulating cytokines in relation to cardiometabolic traits. Our approach generated genetic evidence for effects of obesity measures, hypertension, lipid concentrations, and smoking, on levels of circulating inflammatory cytokines, cytokine cascades underlying systemic responses, and a further three putative therapeutic targets for coronary artery disease (TRAIL, IL1RA, and MCSF).
Obesity, hypertension, and smoking are leading preventable threats to global health.29 Our results suggest that higher body mass index increases multiple mediators of inflammation that affect various processes, including thrombosis (via plasminogen activation inhibitor-1), metabolism (HGF), and endothelial dysfunction (MCP1, TRAIL, ICAM1, and soluble E-selectin). These results for body mass index were corroborated by similar associations for waist circumference and waist to hip ratio. Our analyses also suggested that cigarette smoking contributes to elevating CRP and MCP1 levels, higher systolic blood pressure elevates CRP levels, and low density lipoprotein cholesterol elevates circulating IL6 levels. Taken together, these risk factors seem to increase cardiovascular disease pathogenesis at least partly through inflammatory mediators.30
Comparison with other studies
To better understand cytokine regulatory networks, we also examined the associations of genetically predicted circulating cytokines with levels of other cytokines. Our approach suggests complex relations between circulating cytokines, with VEGF appearing as a master regulator. Indeed, VEGF signalling is already targeted clinically in the treatment of certain cancers and ophthalmic conditions.31 32 Our findings are consistent with an earlier report identifying VEGF as an upstream controller of IL12p70, IL7, IL10, and IL13.14 We replicated and extend these findings to show a wider range of cytokines within this cascade, including drivers of type two immune responses (IL5 and IL13), Jak-STAT cytokine signalling (IFNγ and IL12), and immune modulation (IL10). Although several of these inflammatory cytokines are directly targeted by biological drugs used in routine clinical practice or late stage clinical trials, further research is required to ascertain their potential benefit in the setting of cardiovascular disease. As such, therapeutic targeting of the pathways responsible for the expression or biological signalling of these cytokines might also be possible.
Considering the cardiometabolic outcomes under study, we identified consistent mendelian randomisation and colocalisation evidence for circulating TRAIL increasing coronary artery disease risk; IL1RA and MIF decreasing CRP levels; and IL1RA increasing circulating low density lipoprotein cholesterol, total cholesterol, and glucose concentrations. TRAIL is a ligand involved in initiating apoptosis that has previously been implicated in atherosclerosis.33–35 Our current work extends on this previous work by use of genetic data that supports that higher circulating TRAIL concentrations causally increase coronary artery disease risk, further implicating TRAIL as a therapeutic target. We also found evidence that higher body mass index causally increases circulating TRAIL levels, thus implicating TRAIL as a mediator in the effect of body mass index on coronary artery disease risk. Aligning with earlier work, we showed that variants within the IL1RN gene that increase IL1RA are positively associated with total cholesterol, low density lipoprotein cholesterol, and glucose, but negatively with CRP concentrations,36 which adds support for IL1RA as a therapeutic target in coronary artery disease. Our mendelian randomisation analysis similarly implicated MCSF as a drug target for reducing coronary artery disease risk, triangulating with evidence obtained in studies of mice.37 Although the association between IL1RA and coronary artery disease risk was not robustly supported in our colocalisation analysis, the absence of colocalising signals probably reflects insufficient power rather than genetic confounding. Of relevance, anakinra, which is a recombinant and modified human IL1RA protein, is already used in the treatment of rheumatoid arthritis.38
The mendelian randomisation results using genome-wide selection for instruments provided distinct results to the main results. In particular, genetically proxied circulating soluble E-selectin levels were associated with eight cardiometabolic traits, mostly driven by strong associations in the ABO locus.15 Selection of instruments across the full genome allow for detection of trans-QTL proxies for cytokines. However, this strategy is also likely to include variants that are not specific only to the relevant cytokine, therefore, potentially mis-specifying the exposure and introducing pleiotropic effects violating the mendelian randomisation assumptions.
Strengths and limitations of this study
In our analyses, we aimed to maximise the validity of instruments selected to proxy levels of circulating cytokines by considering only variants that were located at the corresponding gene locus, an established strategy for selecting instruments when investigating drug effects.20 39 Our novel use of both pQTL and eQTL instrument selection criteria offered complementary evidence, and we further used colocalisation analyses to strengthen the evidence for causal effects. However, our work also has limitations. We could not identify suitable genetic proxies for all considered cytokines, and further we may have missed some associations due to insufficient statistical power and the correction imposed for multiple testing. Of note, we could not identify the support for targeting IL6 and MCP1 to reduce cardiovascular disease risk that has been demonstrated in previous mendelian randomisation studies.40 41 The discrepancy for IL6 might be attributable to our approach identifying genetic proxies at the gene for the ligand, rather than its receptor. As a further limitation, mendelian randomisation analyses should not be directly extrapolated to infer the effect of a clinical intervention, as the instruments employed represent the cumulative effect of lifelong genetic predisposition, while a clinical intervention typically represents a discrete event at a particular time point.42 Finally, our genetic data were predominantly obtained from individuals of European ancestry and it is unclear whether these findings will extend to individuals of other ancestries.
Conclusions
By leveraging large scale genetic data, this work offers deeper insight into causal relations between cardiometabolic traits, circulating cytokines, and cardiovascular disease. Our findings replicate previously described therapeutic targets and further identify potential pharmacological opportunities, including perturbation of circulating TRAIL, IL1RA, and MCSF for reducing coronary artery disease risk.