METHODS
Details on the materials, methods, definitions of variables and statistical approach may be found in the Supplementary Appendix. Briefly, 905 plasma samples originating from the time of birth (cord blood, n=376), 1 year (n=195) and 8 years (n=334) from the Manchester Asthma and Allergy Study (MAAS) were analysed for their chemokine content (Figure S1). Plasma levels of CCL18 were measured using an in-house DuoSet ELISA kit (R&D Systems) and an in-house multiplex bead assay was setup for analysis of circulating CCL17, CCL22, CXCL10 and CXCL11.
The chemokine levels were thereafter associated to asthma and allergic sensitisation, and CCL18 levels were additionally related to multivariable outcomes. To study whether chemokine levels predicted asthma or allergic sensitisation, binomial logistic regression analyses were performed on natural log transformed chemokine data. Longitudinal analyses were performed using generalised estimating equations (GEE). Population-averaged GEE models were developed to investigate whether the effect of natural log-transformed chemokine levels on the development of asthma or sensitisation changed over time. All models were adjusted for parental atopy, parental smoking and gender. Resulting coefficients represent the increased/decreased odds of the respective outcome per log-unit increase in chemokine levels.
Analyses were conducted in GraphPad Prism 824, IBM SPSS Statistics version 2525, and Stata 15 software.26