Analysis of Immunoglobulin E Antibody Production in the Human Cell Line by Polycyclic Aromatic Hydrocarbon Treatments: Considerations of Culture Conditions and Dimethyl Sulfoxide Cytotoxicity

To determine allergic reactions of polycyclic aromatic hydrocarbons (PAHs) in humans, culture conditions of an in vitro bioassay system were investigated to evaluate immunoglobulin E (IgE) antibody production using the human myeloma cell line (U266 cells) by PAH treatments.In the preliminary experiment, the cell count of U266 was observed under a microscope at 12, 24, 48, and 72 h of incubation.As a result, many dead cells were observed in the 72 h culture.Accordingly, changes within 48 h were examined in detail.The cell viability reached a plateau in 24 h culture.The IgE antibody production gradually increased during the 48-h incubation period and was higher at 48 h than at 24 h.Therefore, the incubation time was determined to be 48 h.In cell culture experiments, liposoluble materials were dissolved in dimethyl sulfoxide (DMSO).Next, the DMSO cytotoxicity was examined.The results demonstrated that 0.5% and 1% of DMSO were toxic; however, 0.1% and 0.05% were nontoxic.Furthermore, benzopyrene and pyrene (each 10 -8 M) were dissolved in 0.5% and 0.1% DMSO, respectively.We investigated the effects of cytotoxicity and IgE production on U266 cells by adding PAH dissolved in 0.5% or 0.1% of DMSO.Synergistic effects of PAHs (benzo[a]pyrene and pyrene) and 0.5% of DMSO were detected.Therefore, using DMSO concentrations of <0.1% was recommended to study the effects of liposoluble substances on cultured cells.