Regulatory system of human mast cell production

利用干细胞因子(SCF)选择性地从CD34+脐带血细胞中分离培养肥大细胞，研究IL-6对人肥大细胞生长和特性的影响。在含有肥大细胞的培养液中加入IL-6可显著减少SCF在液体培养中生长的子代数量。根据克隆细胞培养实验的结果，这种IL-6介导的肥大细胞生长抑制可能部分是由于前体水平的抑制。此外，流式细胞仪分析显示，在SCF+IL-6的存在下培养的肥大细胞c-kit表达降低。培养的肥大细胞暴露于SCF+IL-6后，与单独使用SCF相比，肥大细胞的大小、凝乳酶阳性细胞的频率和细胞内组胺水平均显著增加。流式细胞仪分析显示，…上IL-6受体(IL-6R)和gp130的表达水平很低，但有显著意义加入抗IL-6R抗体或抗gp130抗体均可阻断IL-6的生物学功能。尽管在SCF刺激下，IL-4对培养的肥大细胞的作用与IL-6相似，但对比实验结果表明，这两种细胞因子的调节机制不同。综上所述，目前的研究结果表明，IL-6通过IL-6R-gp130系统直接调节依赖干细胞因子的人肥大细胞的发育。接下来，我们使用无血清培养系统研究了维甲酸对人肥大细胞发育的影响。以10周培养的脐血CD34+细胞来源的肥大细胞为靶细胞，全反式维甲酸(ATRA)和9-顺式维甲酸(9-cis RA)均以剂量依赖的方式抑制SCF刺激的子代生长。根据单一的CD34^+c-kit^+脐带血细胞培养研究，肥大细胞发育的早期阶段似乎对RA不那么敏感。适宜浓度的RAS还可降低肥大细胞组胺浓度(SCF组为3.00±0.47pg/cell，SCF+ATRA组为1.44±0.18pg/cell，SCF+9-cis RA组为1.41±0.10pg/cell)。结果显示，培养10周的肥大细胞表达RARα、RARβ、RXRα和RXRβ。在10^lt；-7&gt；M中加入RAR选择性激动剂减少了在SCF中生长的肥大细胞的数量，而在10^lt；-8&gt；M中加入RXR选择性激动剂则无效。在RAR亚型选择性维甲酸中，只有RARα激动剂在抑制肥大细胞生长方面与10^lt；-7&gt；M的维甲酸相当。相反，过量浓度的RARα拮抗剂的加入深刻地抵消了维甲酸介导的抑制效应。这些结果表明，RA通过一种特定的受体抑制人肥大细胞前体细胞依赖于SCF的分化。较少

We attempted to clarify the effects of IL-6 on the growth and properties of human mast cells using cultured mast cells selectively generated by stem cell factor (SCF) from CD34^+ cord blood cells. The addition of IL-6 to cultures containing mast cells resulted in a substantial reduction of the number of progenies grown by SCF in the liquid culture. This IL-6-mediated inhibition of mast cell growth may be due in part to the suppression at the precursor level, according to the results of a clonal cell culture assay. Moreover, a flow cytometric analysis showed that the cultured mast cells grown in the presence of SCF+IL-6 had decreased c-kit expression. The exposure of cultured mast cells to SCF+IL-6 also caused substantial increases in the cell size, frequency of chymase-positive cells and intracellular histamine level compared with the values obtained with SCF alone. The flow cytometric analysis revealed low but significant levels of expression of IL-6 receptor (IL-6R) and gp130 on the … More cultured mast cells grown with SCF.The addition of either anti-IL-6R antibody or anti-gp130 antibody abrogated the biological functions of IL-6. Although IL-4 exerted an effect similar to that of IL-6 on the cultured mast cells under stimulation with SCF, the results of comparative experiments suggest that the two cytokines use different regulatory mechanisms. Taken together, the present findings suggest that IL-6 modulates SCF-dependent human mast cell development directly via an IL-6R-gp130 system.Next, we examined the effects of retinoids on the human mast cell development using a serum-deprived culture system. When 10-week cultured mast cells derived from CD34^+ cord blood cells were used as target cells, both all-trans retinoic acid (ATRA) and 9-cis RA inhibited the progeny generation under stimulation with SCF in a dose-dependent manner. The early steps in mast cell development appear to be less sensitive to RA according to the single CD34^+c-kit^+ cord blood cell culture study. The optimal concentration of RAs also reduced the histamine concentration in the cultured mast cells (3.00±0.47 pg/cell in SCF alone, 1.44±0.18 pg/cell in SCF+ATRA, and 1.41±0.10 pg/cell in SCF+9-cis RA). RT-PCR analyses showed the expression of RARα, RARβ, RXRα and RXRβ mRNA in 10-week cultured mast cells. The addition of an RAR-selective agonist at 10^<-10> M to 10^<-7> M decreased the number of mast cells grown in SCF, whereas an RXR-selective agonist at up to 10^<-8> M was inactive. Among RAR subtype selective retinoids used at 10^<-9> M to 10^<-7> M, only the RARα agonist was equivalent to ATRA at 10^<-7> M in its ability to inhibit mast cell growth. Conversely, the addition of excess concentrations of a RARα antagonist profoundly counteracted the retinoid-mediated suppressive effects. These results suggest that RA inhibits SCF-dependent differentiation of human mast cell progenitors through a specific receptor. Less