Exposure to 4,4′‐Methylene Diphenyl Diisocyanate Upregulates Chemokines CCL2, CCL3, CCL5, and IL‐8 Transcription Through Activation of Calcineurin/NFAT Signaling in Macrophages

Abstract

Exposure to the most widely used monomericdiisocyanate, 4,4′‐methylene diphenyl diisocyanate (MDI), in the occupational setting may lead to the development of occupational asthma (OA). Currently, the underlying molecular mechanism(s) by which MDI induces OA remain an active area of research. Airway macrophage dysfunctions such as decreased phagocytosis and efferocytosis as well as increased production of reactive oxygen species (ROS), nitric oxide(NO) and immune mediators, etc., play important roles in asthma pathogenesis. Previously, we demonstrated that the downregulation of microRNA (miR)‐206‐3p and miR‐381‐3p mediated activation of the Calcineurin/nuclear factors of activated T‐cells (NFATs) signaling pathway which induced endogenous inducible nitric oxide synthase (iNOS) transcription in macrophages and bronchoalveolarlavage cells (BALCs) after acute MDI exposure. However, whether MDI may affect expression of other asthma‐associated immune mediators in macrophages/BALCs and the underlying molecular mechanisms driving these effects are currently unknown. We hypothesize that MDI exposure induces several asthma‐associated mediators through Calcineurin/NFAT signaling activation in macrophages. We determined the expression of macrophage‐secreted, asthma‐associated mediators including cytokines (IL1‐β, TNF‐α, and IL‐6); chemokines (CCL2, CCL3, CCL5, CCL11, and IL‐8), and growth factors (GM‐CSF and TGF‐β) from both MDI‐exposed murine BALCs and MDI‐glutathione(GSH) conjugate‐treated differentiated THP‐1 macrophages using RT‐qPCR. Both in vivo murine MDI aerosol and in vitro MDI‐GSH exposures result in the upregulation of endogenous IL1‐β, TNF‐α, IL‐6, CCL2, CCL3, CCL5, and TGF‐β. Treatment of THP‐1 macrophages with 1 μM Tacrolimus (FK506), a specific Calcineurin/NFAT signaling inhibitor, blocked the induction of CCL2, CCL3, CCL5, and IL‐8 expression by MDI‐GSH conjugates but not IL1‐β, TNF‐α, IL‐6, and TGF‐β. These results indicate that Calcineurin/NFAT signaling activation plays an important role for CCL2, CCL3, CCL5, and IL‐8 transcriptional upregulation following MDI exposure in macrophage. These results suggest MDI exposure may result in recrutement of many different cell types such as eosinophils, T‐cells, monocytes, macrophages, etc., into the airway by induction of chemokines transcription via Calcineurin/NFAT signaling activation in the macrophages/BALCs.Support or Funding InformationThis work was supported by intramural funds from the National Institute for Occupational Safety and Health, Centers for Disease Control and Prevention.