Nuclear Calcium Signals in Immune Responses

免疫反应中的核钙信号

• 批准号: 04454621
• 负责人: NAKANISHI Mamoru
• 金额: $ 3.84万
• 依托单位: Nagoya City University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (B)
• 财政年份: 1992
• 资助国家: 日本
• 起止时间: 1992 至 1993
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-04454621/
• 关键词: Immune Response; Calcium ion; Nuclear signal; Confocal Microscope; T-cell; B-cell; Basophil; Image Analysis; カルシウムイオン; 細胞内情報伝達; サードメッセンジャー; 蛍光性分子ロータ

We have studied receptor-mediated calcium signals in antigen-specific B cells (trinitrophenol-specific B cell clone, TP67.21) and rat basophilic leukemia cells (RBL-2H3) using a confocal fluorescence microscope with an argon ion laser (488nm) and a He-Cd laser (325nm). Confocal fluorescence images of fluo-3-loaded B cells and RBL-2H3 cells, excited by an argon ion laser, became much brighter and more nonhomogeneous than those before antigen stimulation. Time-dependent fluorescence changes in intensities were abrupt and quite similar to the patterns of the intracellular calcium ion concentration [Ca^<2+>]_i observed by a conventional fluorescence microscope using fura-2. From the morphological patterns of the calcium images, the parts of the bright fluorescence seemed to belong to the nucleus in B cells (or RBL-2H3 cells). To confirm the above events we measured the confocal fluorescence images of the nucleus. From the fluorescence images of co-loaded Hoechst 33342 (a DNA-specific fluorescence probe), which excited by a He-Cd laser, the brighter parts of the fluo-3 fluorescence intensities were identified to the nucleus in B cells and RBL-2H3 cells. These nuclear calcium signals were also observed by the addition of thapsigargin, an inhibitor of the intracellular calcium pump. These results suggested the possibility that the increased intranuclear calcium ions may play a nuclear third messenger in B cells and RBL-2H3 cells. Then, we tried to determine whether the increased intranuclear calcium ions are transferred from the cytoplasm or they are released from the nuclear stores. We used Ba^<2+> and Mn^<2+> instead of Ca^<2+>, because Ba^<2+> and Mn^<2+> are known to enter via Ca^<2+> channels but they quench the fluo-3 fluorescence. The results showed that Ba^<2+> and Mn^<2+> entered into the nucleus as well as into the cytoplasm and quenched the fluo-3 fluorescence both in the nucleus and in the cytoplasm. This sug

我们利用共聚焦荧光显微镜，用氩离子激光（488nm）和He-Cd激光（325nm）研究了抗原特异性B细胞（三硝基酚特异性B细胞克隆，TP67.21）和大鼠嗜碱性白血病细胞（RBL-2H3）中受体介导的钙信号。负载fluo-3的B细胞和RBL-2H3细胞在氩离子激光激发下的共聚焦荧光图像比抗原刺激前的图像更亮，更不均匀。荧光强度随时间的变化是突然的，与传统荧光显微镜使用fura-2观察到的细胞内钙离子浓度[Ca^<2+>]_i的模式非常相似。从钙图像的形态模式来看，明亮荧光的部分似乎属于B细胞（或RBL-2H3细胞）的细胞核。为了证实上述事件，我们测量了核的共聚焦荧光图像。通过He-Cd激光激发共负载Hoechst 33342 （dna特异性荧光探针）的荧光图像，确定了B细胞和RBL-2H3细胞中荧光强度较亮的部分为细胞核。这些核钙信号也可以通过添加thapsigargin（一种细胞内钙泵抑制剂）观察到。这些结果提示核内钙离子的增加可能在B细胞和RBL-2H3细胞中发挥核第三信使的作用。然后，我们试图确定增加的核内钙离子是从细胞质转移还是从核库释放。我们使用Ba^<2+>和Mn^<2+>代替Ca^<2+>，因为已知Ba^<2+>和Mn^<2+>通过Ca^<2+>通道进入，但它们猝灭了氟-3荧光。结果表明，Ba^<2+>和Mn^<2+>进入细胞核和细胞质，猝灭了细胞核和细胞质中的氟-3荧光。这见sub

项目成果

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R.Aiyama：“盐酸伊立替康（CPT-11）在水溶液中自缔合的测定。”

T.Furuno: "The effects of an inhibitor ofthe intracelular Ca^<2+> pump on nuclear calcium signals in B cells" Bioimages. 1. 9-12 (1993)

T.Furuno：“细胞内Ca^2泵抑制剂对B细胞核钙信号的影响”生物图像。

T.Furuno: "Confocal fluorescence microscopy for studying the effectsof a tyrosine kinase inhibitor on the nuclear calcium signals in B cells" Bioimages. 1. 175-180 (1993)

T.Furuno：“用于研究酪氨酸激酶抑制剂对 B 细胞核钙信号影响的共聚焦荧光显微镜”Bioimages。

T.Furuno: "A fluorescent molecular rotor probes the kinetic process of degranulation of mast cells." Immunol.Lett.33. 285-288 (1992)

T.Furuno：“荧光分子转子探测肥大细胞脱颗粒的动力学过程。”

Furuno,T. Isoda,R. Nakanishi,M.: "A fluorescence molecular rotor probes the kinetic process of degranulation of mast cells" Immunol.Lett.33. 285-288 (1992)

古野，T.