Abstract
Mast cells, in addition to endocrine cells and neurons, are typical secretory cells. Their function in allergic inflammation is to secrete inflammatory mediators from secretory vesicles. Intracellular synthesized inflammatory mediators are transported by vesicular monoamine transporters (VMATs) to vesicles where they are stored. After stimulation, the contents of the secretory vesicles are released via exocytosis. This study established a high throughput imaging screening system to monitor the functions of secretory vesicles in mast cells, including molecular uptake via VMAT2 and the exocytotic process, by using a novel fluorescent probe, FFN206, which was developed as a VMAT2 substrate. After loading with FFN206, the rapid uptake of FFN206 was observed and secretory vesicles in mouse bone marrow derived mast cells and a cultured mast cell line were clearly visualized. FFN206 uptake by secretory vesicles was time-dependent and was blocked by reserpine. Furthermore, exocytotic trafficking was monitored dynamically by real-time high-throughput fluorescence quantitation. In the present study, we verified the application of FFN206 for the monitoring of functional vesicles. This high-throughput screening system may benefit instinctive drug evaluation.
Highlights
Mast cells, in addition to exocrine cells, endocrine cells and neurons, are typical secretory cells
Secretory vesicles with different sizes were observed in primary differentiated mast cells derived from +/+, bg/+, and bg/bg mice. bg/bg mice are used as a Chediak-Higashi syndrome
The images of primary differentiated bone marrow-derived mast cells (BMMC) and cultured RBL-2H3 cells suggested that labeling using FFN206 is a useful technique for the morphological and functional studies of living mast cells
Summary
In addition to exocrine cells, endocrine cells and neurons, are typical secretory cells. They are involved in the innate and adaptive immune systems and their roles in allergic and anaphylactic reactions are well characterized [1, 2]. Vesicles in mast cells take up and store mediators such as biological amines, peptidoglycan, chymase and tryptase [3]. The granule content is important in determining the fate of granule formation and maturation [4, 5]. The characteristics of vesicle contents and cell activity in mast cells have been widely studied to clarify the mechanisms underlying their pathological role.
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