Georas, S; Williams, MA; Little, EL; Stewart, JC; Emo, J; Bauer, SM; Frasier, LM; Chalupa, DC; Vora, R; Pietropaoli, AP; Utell, MJ; Frampton, MW
Introduction: Exposure to ultrafine particulate matter (UFP) may worsen asthma, although the mechanisms involved are uncertain. We tested the hypothesis that UFP activates dendritic cells (DC), key sensors of the innate immune system. DC are heterogeneous and derive from circulating CD14+ myeloid precursors. Circulating myeloid DC (mDC) and plasmacytoid DC (pDC) can be detected using multi-color flow cytometry.
Methods: We recruited 10 adult human asthmatic subjects (n=5 with the “null” polymorphism for glutathione-S-transferase M1) and exposed them for 2 hours at rest to concentrated ambient UFP (mean 2.96 x10^5 particles/cm^3, count median diameter 90.9 nm) or filtered air using a double-blinded cross-over design. Phlebotomy, spirometry, and airway production of nitric oxide (NO) were performed at intervals up to 24 hours after exposure. The numbers and activation state of circulating CD14+ monocytes and DC subsets were analyzed using flow cytometry 3 hours after exposure. DC subsets were defined as lineage negative (lin-) cells not expressing CD3, CD16, CD19, CD32, or 7AAD, and separated into mDC1, mDC2 and pDC subsets with a panel of specific surface markers. CD14+ cells obtained 3 hours after exposure were differentiated into DC in vitro using standard techniques, and restimulated with CD40-ligand or lipopolysaccharide for 48 hours. We monitored cell activation using anti-CD40, CD80, CD86, and MHCII antibodies.
Results: Exposure to concentrated UFP was well-tolerated, and did not alter the FEV1 or airway NO kinetics. Expression of CD40 was significantly reduced on CD14+ monocytes 3 hours after UFP exposure compared to filtered air (p=0.03). The numbers of lin- cells decreased significantly 3 hours after UFP but not filtered air (p=0.02), but this was not reflected by changes in the frequencies or activation of mDC1, mDC2, or pDC subsets, although expression of CD40 tended to be lower on mDC1. Inducible expression of CD40 also tended to be lower on DC differentiated from CD14+ precursors obtained 3 hours after UFP exposure compared to filtered air. The number of blood eosinophils increased 24 hours after UFP exposure relative to air (p=0.02). None of these results were significantly affected by GSTM1 genotype.
Conclusion: Exposure to UFP results in alterations in circulating eosinophils, monocytes, myeloid DC and probably DC precursors in asthmatic subjects. Future studies are needed to determine whether this reflects recruitment of DC subsets out of the circulation or direct activation in vivo. Myeloid DC may be a previously unsuspected target of inhaled UFP in human subjects.
