Highly multiplexed cytokine analysis of bronchoalveolar lavage and plasma reveals age-related dynamics and correlates of inflammation in children

ABSTRACT Despite the central role of cytokines in mediating inflammation that underlies a range of childhood diseases, cytokine testing remains primarily limited to research settings and surrogate markers of inflammation are often used to inform clinical diagnostic and treatment decisions. There are currently no reference ranges available for cytokines in healthy children, either systemically (in blood) or at sites of disease (such as the lung). In our study, we aimed to develop an openly accessible reference of cytokines in the airways and blood of healthy children spanning 1 to 16 years of age. We examined how cytokine concentration changes during childhood and assessed whether a core set of cytokine markers could be used to indirectly evaluate the response of a broad spectrum of inflammatory analytes. To develop our reference, a total of 78 unique analytes were measured in cell-free bronchoalveolar lavage (BAL) and plasma from 78 children. We showed that age profoundly impacts soluble immune analyte concentration in both sample types and identified a highly correlative core set of 10 analytes in BAL and 11 analytes in plasma capable of indirectly evaluating the response of up to 34 inflammatory mediators. This study addresses an urgent need to develop reference ranges for cytokines in healthy children to aid in diagnosis of disease, to determine eligibility for, and to monitor the effects of, cytokine-targeted monoclonal antibody therapy. Competing Interest Statement The authors have declared no competing interest. Funding Statement This work was funded by Chan Zuckerberg Initiative Single-Cell Biology Grants (2021-237883, 2020-217531). We thank the children and parents who participated in our studies, without whom this work would not exist. Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: The studies involving human participants were reviewed and approved by Royal Childrens Hospital Human Research Ethics Committee (HREC 25054 and 88144). Written informed consent to participate in the studies was provided by the participants legal guardian/next of kin. I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes Data Availability All data produced in the present work are contained in the manuscript