Characterization of multiple P2X receptors in cultured normal human epidermal keratinocytes
Characterization of multiple P2X receptors in cultured normal human epidermal keratinocytes
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DOI：
10.1111/j.0022-202x.2005.23683.x
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发表时间：
2005-04-01
影响因子：
6.5
通讯作者：
Inoue, K
中科院分区：
文献类型：
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作者：
Inoue, K;Denda, M;Inoue, K
ATP门控离子通道(P2X)在人表皮和培养的角质形成细胞中均有表达。本研究的目的是利用全细胞膜片钳技术、逆转录聚合酶链式反应(RT-PCR)和细胞内钙离子浓度的测定，对正常人表皮角质形成细胞(NHEK)中天然的P2X受体进行鉴定。应用三磷酸腺苷产生内向电流，反转电位为0 mV。对ATP的反应表现为两种类型的电流：缓慢脱敏反应和快速脱敏反应。这种缓慢的脱敏反应可被P2X受体拮抗剂-6-偶氮苯基-2‘，5’-二磺酸(PPADS)阻断。我们发现，在分化的细胞中，多个P2X(2)、P2X(3)、P2X(5)和P2X(7)受体亚型的mRNA表达增加。另一方面，在分化的细胞中，G蛋白偶联的P2Y(2)基因表达下调。在分化的细胞中，由α-β-亚甲基三磷酸腺苷(α-β-meATP)和2‘，3’-O-(4-苯甲酰基苯甲酰)三磷酸腺苷(BzATP)引起的[Ca~(2+)](I)升高，而由尿苷5‘-三磷酸(UTP)引起的[Ca~(2+)](I)升高。应用ATP或UVB辐射可增加NHEK中P2X(1)、P2X(2)、P2X(3)和P2X(7)受体的表达。多个P2X受体的表达水平和阳离子内流的变化可能参与了分化的调节和表皮的外部感受器之一。
ATP-gated ion channels (P2X) are expressed in human epidermis and cultured keratinocytes. The aim of this study was to characterize native P2X receptors in normal human epidermal keratinocytes (NHEK) using whole-cell patch clamp technique, RT-PCR, and determination of intracellular Ca2+ concentration ([Ca2+](i)). Application of ATP resulted in an inward current with a reversal potential of 0 mV. Response to ATP showed two types of currents: the slowly desensitizing response and the rapidly desensitizing response. The slowly desensitizing response was blocked by iso-pyridocaphosphate-6-azophenyl-2', 5' disulfonic acid (PPADS), a P2X receptor antagonist. We found that the expression of multiple P2X(2), P2X(3), P2X(5), and P2X(7) receptor subtype mRNA was increased in differentiated cells. On the other hand, the expression of G-protein-coupled P2Y(2) mRNA was downregulated in differentiated cells. Increases in [Ca2+](i) evoked by alpha beta-methylene ATP (alpha beta-meATP) and 2', 3'-O-(4-benzoylbenzoyl) ATP (BzATP) were elevated, whereas elevation of [Ca2+](i) evoked by uridine 5'-triphosphate (UTP) was decreased in differentiated cells. Application of ATP or UVB radiation increased the expression of P2X(1), P2X(2), P2X(3), and P2X(7) receptors in NHEK. Changes in the expression levels and cation influx via multiple P2X receptors might be involved in the regulation of differentiation and one of the epidermal external sensors.