| Traditional Chinese medicine was formulated in pill, powder, ointment and Dan. TCM injections (Traditional Chinese Medicine Injection, TCMI) is a new dosage form of the modernization process. Because of its easy absorption, rapid onset, and convenient features, it open the market quickly and achieved good clinical efficacy. However, with the increase in the use of traditional Chinese medicine injections, there are more and more adverse events, caused widespread concern in the industry. Most of adverse reactions caused by TCMI are allergic reactions or anaphylactoid reactions,. Anaphylactoid Reactionis due to the non-IgE-mediated mast cell and basophil degranulation, releasing of inflammatory mediators. Anaphylactoid reactions and anaphylaxis symptoms are mostly the same and difficult to differentiate in clinical. The method of detecting the allergies caused by TCMI is animal experiment in the current Chinese pharmacopoeia;which is not specifically for Anaphylactoid Reaction evaluation. Though most TCMI have had undergone strict preclinical evaluation, its still cause allergy frequently, which indicate the method is not sensitive enough. Thus, it’s necessary to optimize the current method of detecting the allergies and anaphylactoid reaction caused by TCMI. In this study, we intend to establish an allergy evaluation model for evaluate the potential of TCMI anaphylactoid reaction with RBL-2H3cells and rat peritoneal mast cells.1Establishment of research methodIn this study, rat peritoneal mast cells and RBL-2H3cells were used as mast cells and basophils cell model, The method of separate and extract rat peritoneal mast cells was established by our laboratory. The mast cells were identified, and the activity as well as the stability were studied in vitro. The extraction method is simple, reliable, and the purity and the activity of mast cells were consistent with experimental requirements. RBL-2H3cells were cultured, and the degranulated cells were tested with β-hexosaminidase and histamine in supernatant. In this study, we also compared the degranulation character of the two cells, and optimize the fluorescence detection methods of histamine.2Using the cell model to evaluating the Anaphylactoid reaction induced by traditional Chinese medicine injectionShengmai injection was choosed as sample of TCMI because of its the clinical adverse reactions. the degranulation of RBL-2H3cells and rat peritoneal mast cell were studied with Shengmai injection. the cytotoxic of Shengmai injections were evaluated by the MTT assay and flow cytometryConclusion1Optimized and established an anaphylactoid evaluation model with rat peritoneal mast cells and RBL-2H3cells. The optimized of fluorescence detection methods of histamine were found:the condensation time in3min, the reaction was terminated after15-30min, which can be the best condition to detect fluorescence. Study of particle properties of rat peritoneal mast cells and RBL-2H3cells found that the content of histamine in mast cells is rich and has a high stimulating released rate. Anaphylactoid reaction should be use histamine as detect targets; and RBL-2H3cells are rich in β-hexosaminidase enzyme, and has a high stimulating release rate, Anaphylactoid reaction should adopt the β-hexosaminidase as detect targets.2Using this method to evaluate anaphylactoid reaction caused by shengmai injections. The results showed that at the concentration of12.5-100μL/mL, Shengmai injection can increase the release rate of histamine in mast cells (P<0.05), at the concentration of25-100μL/mL can cause β-amino glucosidase of RBL-2H3cells release rate increased (P<0.05). Both cells did not show cytotoxicity at the above dosage ranges. The results suggest that Shengmai injection can induced anaphylactoid which may be one of its clinical adverse reaction mechanism. |
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