Supplementary Material for: Circulating Plasmacytoid Dendritic Cells are Numerically and Functionally Deficient in Trauma Patients

Introduction: Dendritic cells (DCs) are crucial in immune responses but their roles in trauma patients remain poorly understood. This study aimed to investigate the frequencies and functions of plasmacytoid DCs (pDCs) and conventional DCs (cDCs) in trauma patients. Methods: This study included 25 trauma patients and 40 healthy controls. Using flow cytometry, the frequencies, cytokine production (IFN-α, TNF-α, IL-12), and expression of co-stimulatory/inhibitory (CD86/CD274) markers were analyzed. The relationship between CD274+ pDCs and the Injury Severity Score (ISS) was also evaluated. Additionally, in vitro experiments were conducted using proinflammatory cytokine stimulation and blocking antibodies to assess their effects on DC activation. Results: Trauma patients showed decreased percentages and absolute numbers of pDCs, while cDC levels remained unchanged. Cytokine production was impaired, with reduced IFN-α and TNF-α from pDCs and IL-12 from cDCs. CD86+ and CD274+ expression were increased in pDCs but decreased in cDCs, with CD274+ pDCs positively correlating with ISS scores. In vitro studies revealed that proinflammatory cytokines enhanced activation markers, peaking at 72 hours; this observation was reversed by blocking antibodies. Conclusion: This study demonstrates that trauma induces both numerical and functional deficiencies in circulating pDCs despite increased activation marker expression. The correlation between CD274+ pDCs and trauma severity, along with the temporal dynamics of cytokine-induced activation, suggests a complex role for pDCs in post-traumatic immune responses and provides new insights for developing targeted therapies for trauma-related immune complications.