Molecular Biology Of Mast Cell Growth And Differentiation

人类肥大细胞起源于多能祖细胞，并作为未成熟细胞从骨髓迁移到组织部位，包括肺和胃肠道。在那里，这些前体成熟并参与先天和获得性免疫反应，产生细胞因子和其他炎症介质。因此，肥大细胞的生长和发育可能发生在与外部环境接触的组织中，肥大细胞在发育过程中可能暴露于细菌产物中，这可能对其随后的行为产生影响。与这一观点一致的观察结果是，肥大细胞在体外和体内均表达toll样受体（TLR） 1-7和9；暴露于细菌产物如内毒素或肽聚糖会导致细胞因子的表达和释放。然而，在一个相关的问题中，我们探讨了细菌衍生产品是否会改变HuMC的生长和发育。

Human mast cells originate from pluripotential progenitor cells and migrate as immature cells from the bone marrow to tissue sites including the lung and gastrointestinal tract. There these precursors mature and participate in both innate and acquired immune responses with production of cytokines and other inflammatory mediators. Mast cell growth and development thus may occur in a tissue that interfaces with the external environment, potentially exposing mast cells during their development to bacterial products which could have an impact on their subsequent behavior. Consistent with this idea is the observation that mast cells are known to express Toll-like receptors (TLR) 1-7, and 9 both in vitro and in vivo; and exposure to such bacterial products as endotoxin or peptidoglycan leads to expression and release of cytokines. However, and in a related question, we explored whether bacteria-derived products alter the growth and development of HuMC. We thus performed long and short-term cultures to which we added LPS or PGN. We followed specific mast cell characteristics including growth; surface FcepsilonRI and CD117 expression; degranulation, LTC4 and PGD2 release; protease expression and composition, and cytokine release. Over 6 wks of culture, LPS had minimal effect on HuMC numbers but increased CD117, tryptase and chymase expression. PGN inhibited HuMC development. For mature mast cells, LPS in the presence of rhSCF (10 ng/ml) increased CD117, tryptase, chymase and carboxypeptidase expression, primarily in CD117low HuMC. LPS decreased FcepsilonRI expression and granule release; but had no effect on LTC4 and PGD2 production. PGN reduced HuMC numbers; and CD117 and tryptase expression. IL-1 and IL-6 (in addition to IL-8 and IL-12) were detected in short-term culture supernatants of LPS treated cells, and reproduced the increases in CD117, tryptase, chymase, and carboxypeptidase expression observed in the presence of LPS. Comparative studies with mouse bone marrow-derived mast cells from wild type, but not TLR4 knockout mice, showed increases in mRNA of mouse mast cell chymases MMCP-1, MMCP-2 and MMCP-4. We thus concluded PGN inhibits human mst cell growth, while LPS exerts its primary effects on mature mast cells by altering cytokine production and protease composition, particularly at low concentrations of SCF. These data thus demonstrate the ability of bacterial products to alter human mast cell mediator production, granular content, and number which may be particularly relevant at mucosal sites where mast cells are exposed to these products. In a question relevant to the evaluation of allergic sensitization, we are examining the ability of a component of shrimp, a major food allergen, to directly activate human mast cells. It is known that shrimp extracts, used as skin testing reagents, cannot be used at higher concentrations, as this leads to non-specific reactions; that is, these reactions occur in all individuals and thus do not correlate with clinical sensitivity. To identify the component of shrimp that directly activates mast cells, we fractionated shrimp extracts by size using column chromatography (S-300) and screened fractions for direct mast cell degranulating activity using the LAD2 human mast cell line. Active fractions were sequenced. Tropomyosin (Pen a 1) was identified as the active material. Recombinant Pen a 1 (rPen a 1) was used in degranulation experiments and did directly degranulate mast cells. In vitro degranulation experiments were then undertaken. Pertussis toxin and inhibitors of PI3K, PLCgamma, PKC and Src did not prevent degranulation. rPen a 1 did increase cytosolic calcium concentrations by promoting liberation from intracellular stores and calcium entry. These observations appear to explain how shrimp extracts at higher concentrations lead to non-specific skin test reactivity. In vivo experiments in an animal model are underway.