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目的 了解酪氨酸蛋白激酶信号途径在不同α1肾上腺素受体亚型Ca2 +调控中的作用。方法 用Fura 2荧光探针双波长测定细胞胞浆游Ca2 +浓度 ([Ca2 +]i)的方法 ,在分别转染了α1A、α1B和α1D肾上腺素受体cDNA的HEK2 93细胞 ,观察酪氨酸蛋白激酶抑制剂Genistein和磷脂酶C抑制剂U7312 2 对激动不同α1肾上腺素受体亚型引起的 [Ca2 +]i 变化的影响。结果 预先用U7312 2 (0 1,10 ,5 0 μmol·L-1)与细胞共同孵育 10min ;用Genistein(10 ,10 0 ,2 0 0 μmol·L-1)与细胞共同孵育 1h。在分别转染了α1A、α1B和α1DcDNA的HEK2 93细胞 ,U73 12 2 和Genistein均能浓度依赖性地抑制肾上腺 (10 μmol·L-1)引起的双相 [Ca2 +]i 的升高。在上述细胞 ,U7312 2 (5 0 μmol·L-1)能完全抑制肾上腺素引起的[Ca2 +]i 升高 ;而用最大有效浓度 10 0 μmol·L-1的Genistein只能部分抑制肾上腺素升高 [Ca2 +]i 的作用。结论 在HEK 2 93细胞 ,不同α1肾上腺素受体亚型 (α1A α1B和α1D)激动均能部分通过酪氨酸蛋白激酶信号途径引起Ca2 +释放和Ca2 +内流。α1肾上腺素受体可能通过G蛋白和酪氨酸蛋白激酶两种途径激活磷脂酶C
Objective To investigate the role of tyrosine kinase signaling in the regulation of Ca2 + in different α1 adrenergic receptor subtypes. Methods HEK2 93 cells transfected with α1A, α1B and α1D adrenoceptor cDNAs were respectively treated with Fura 2 fluorescent probe dual-wavelength cytosolic Ca2 + concentration ([Ca2 +] i) Effect of Acid Protein Kinase Inhibitor Genistein and Phospholipase C Inhibitor U7312 2 on [Ca2 +] i Changes Induced by Different α1 Adrenergic Receptors. Results The cells were preincubated with U7312 2 (0 1, 10, 50 μmol·L-1) for 10 min. Cells were incubated with Genistein (10, 10 000 μmol·L-1) for 1 h. In HEK2 93 cells transfected with α1A, α1B and α1D cDNAs, both U73122 and Genistein inhibited the increase of biphasic [Ca2 +] i induced by 10 μmol·L-1 in a concentration-dependent manner. In the above-mentioned cells, U7312 2 (50 μmol·L-1) completely inhibited the increase of [Ca2 +] i induced by epinephrine, whereas Genistein with the maximal effective concentration of 10 μmol·L-1 only partially inhibited the adrenergic Increase the role of [Ca2 +] i. Conclusions In HEK 2 93 cells, the activation of different α1 adrenergic receptor subtypes (α1A α1B and α1D) can both induce Ca2 + release and Ca2 + influx through tyrosine kinase signaling. α1 adrenoceptors may activate phospholipase C through both G-protein and tyrosine protein kinases