DISCUSSION
Minimally invasive biomarkers, such as serum proteins, are widely sought after for their value in defining disease states and outcomes. In adolescent and adult AD patients, a high total serum IgE level was a predictive factor for poor disease prognosis [19]. Interestingly, in these pediatric patients, many inflammatory serum biomarkers demonstrated stronger correlation to disease severity better than IgE alone [20]. Thus, the focus of our study was to highlight key biomarkers in early AD that correlate with disease and potentially predict the persistence or resolution of disease.
Multiple analytes correlated strongly with pediatric AD severity and, most significantly, known disease markers, IL-13, MCP-4, and TARC, correlated with SCORAD across all time points from infancy to 5 years of age, recapitulating the importance of type 2 immune response early in AD pathogenesis. Correlations with chemokines and monocyte chemoattractant proteins underscores the role of eosinophils and monocytes in early AD, while TARC and IL-13 are both mediators of Th2 inflammation and implicated in pediatric and adult AD, emphasizing that these analytes could represent critical nodes for better understanding AD heterogeneity and disease progression [7, 21, 22]. Although standard of care treatments lead to a decreased concentration of these biomarkers, the observations and changes observed in our study demonstrate the strong relationship these markers have with the disease pathology in pediatric patients. Importantly, this study included ultrasensitive measurements of numerous cytokines for which quantitative assessment has only recently been possible, including IL-13, which is implicated as a driver of AD based on clinical responses to lebrikizumab and dupilumab, further implicating IL-13 in disease severity even in infancy [5, 23, 24].
Analytes described as drivers of adult AD, such as IL-22 and IL-17A [7, 18], did not possess strong correlations to disease severity within these pediatric AD samples. This is likely due to the slow increase in serum concentration over time, limiting the role of IL-22 in pediatric AD as seen in previous reports [7]. Though levels of IL-17A were not correlated to SCORAD and present in very low levels, as seen in previous pediatric studies [25], Th17-related fibroblast markers, CCL19 and CCL20, were observed at various timepoints throughout our study and CCL20 positively correlated with Th2 circulating cell populations at Y1 pointing toward an intriguing immunological overlap of inflammatory fibroblasts and T cells. Pathway analysis of proteins upregulated in Y5 samples reveals the emergence of innate pathways over time and multiple gene ontology results indicate that dendritic cells (DCs) may increase with age. Increased conventional dendritic cells were previously described as being protective for wheezing in these pediatric eczema patients [10], and stimulation of these conventional DCs isolated from atopic infants produced more interferon and IL-10 than those derived from non-atopic infants at initial visit and the one year follow-up visit [26]. Mounting evidence demonstrates this appearance of symptoms known as the atopic march tracks with early disruption of immune development and can have lasting effect into adulthood [27]. This link between increased CCL19 and DCs further indicates a role in atopic march from AD to asthma for CCR7, the receptor for CCL19, which can coordinate T cell, B cell, and DC responses across pathologies [28].
Matrix remodeling proteins (MMP-10) and chemoattractant molecules (MCP-3, MCP-2, CXCL5) provide further insights into the complexity of disease mechanisms significantly associated with the progression of the pediatric form of AD. The connections of these markers to disease severity contextualize a shift toward Th1 immunity and myeloid involvement developing throughout the course of the pediatric AD patient journey concomitant with dominant Th2 responses throughout. The identification of CXCL5 and IL-5 as important disease severity predictive biomarkers for pediatric AD highlights the innate allergic response early in disease progression. The high levels of sCD40L that we observed in pediatric patients aligned with prior published observations that this protein is elevated in the blood of pediatric patients [6]. Additionally, sCD40L concentrations correlated throughout the study with CD4+Th2 cells which might link to the role of CD4+CD40L+ cells in the priming of other immune cell types including CD14+ monocytes and CD8+ T cells [29]. This work aids our contextualization of a shift toward Th1 immunity and myeloid involvement developing throughout the course of the pediatric AD patient journey concomitant with Th2 responses presenting initially and persisting throughout the course of disease.
The previously reported circulating protein profiles of pediatric AD have provided robust assessment of correlation analysis at a single timepoint [6]. Our study builds upon these important observations via the monitoring of circulating biomarkers over a development period of four years adding value to the concept that objective biomarkers could be used to objectively assess disease severity, as has been demonstrated in adult AD [30]. We observed subsets of inflammatory proteins that correlated with each other in a manner consistent with early disease skewing toward type 2 immune responses. Importantly, forward selection modeling identified 18 analytes, such as IL-18, IL-13, and MMP-10, in infant serum that could be used to predict AD severity four years later; therefore, these analytes might represent critical nodes to better understand AD heterogeneity and disease progression. The measurement of markers identified in Figure 6 should provide guidance for therapeutic intervention strategies at early stages of life for pediatric patients suffering from AD. This model will likely have predictive value for AD persistence past infancy and will be useful in further defining the pathogenesis of atopic disease allowing us to begin to see ever branching nodes delineating the complex underlying immune regulation. This, in turn, may guide therapeutic intervention and the prognostic value of such serum protein measurement, outlining the need for early treatment options in pediatric patients.
Our data highlights additional questions for the field regarding the progression of AD as children age through adolescence into adulthood. We did not observe elevation of IL-4 in the serum of these patients, which could be attributed to the technical limitations of assays available at time of testing, considering that IL-13 was measured on an ultra-sensitive platform. We also note that though MCP-4 and TARC were moderately correlated with disease severity, they did not factor into the predictive modeling of disease progression possibly due to these proteins being more broadly involved in atopic mechanisms beyond AD [31]. Future studies with focused outcomes on other atopy measures can provide predictive insight into mechanisms of wheezing or lung function. Correlations to SCORAD allow us to minimize potential error that is introduced in a longitudinal study, especially in pediatrics as maturation and growth are heavily involved in immune system development. This cohort and data included in this manuscript enhance our knowledge regarding the systemic presentation of AD in children and potential biomarker profiles which aid in predication of disease persistence or resolution.