Methylenediphenyl diisocyanate binds membrane and intracellular proteins of intact THP-1 cells.
Mhike-M; Hettick-JM; Law-BF; Bledsoe-TA; Chipinda-I; Lemons-AR; Green-BJ; Nayak-AP; Simoyi-R; Beezhold-D; Siegel-PD
Toxicologist 2014 Mar; 138(1):537
Diisocyanates (dNCO) are used in the production of polyurethane products and can cause occupational asthma. Although dNCO asthma displays the same pathological hallmarks as other allergic asthmas, dNCO-specific IgE can only be detected within a small subset of dNCO asthmatics. Because of this low prevalence, we propose that methylenediphenyl diisocyanate (MDI) may adduct to cell membrane and intracellular proteins, contributing to the development of non-IgE asthma. In this study, human monocytic leukemia (THP-1) cells were exposed to varying concentrations of MDI. After exposure, cells were processed into soluble and membrane fractions, dialyzed, and analyzed for extent of MDI protein binding, following acid hydrolysis, by quantification of the MDI hydrolysis product, methylenedianiline (MDA). Proteins were electrophoresed on sodium dodecyl sulfate polyacrylamide gels and protein bands were excised, hydrolyzed and analyzed for MDA. Proteins from the bands were identified using proteomic mass spectrometry. Protein bound MDI was detected in a dose-dependent manner and quantified from the membrane and soluble fractions of MDI exposed THP-1 cells. Eleven bands from the soluble fraction and 2 bands from the membrane fraction were identified as containing MDI bound proteins. The extent of MDI intracellular protein binding was not affected by cytochalasin D, a chemical which binds actin filaments and inhibits active uptake into cells. Proteins identified in the soluble fraction were all of intracellular origin suggesting that reactive MDI can cross the cell membrane and subsequently haptenate intracellular proteins. The results of the present study support the potential involvement of dNCO haptenated membrane and intracellular proteins in development of asthma.
Bronchial-asthma; Allergic-reactions; Allergens; Allergies; In-vitro-study; Proteins; Protein-biochemistry; Cellular-function; Cellular-reactions; Respiratory-system-disorders; Pulmonary-system-disorders; Pulmonary-function
The Toxicologist. Society of Toxicology 53rd Annual Meeting and ToxExpo, March 23-27, 2014, Phonex, Arizona