细胞缝隙连接在丙硫氧嘧啶肝损伤发生中的作用及机制研究

Propylthiouracil (PTU)-induced liver injury is one of the major factors which has limited its drug safety, but the mechanisms are largely unknown. Gap junctions (GJs) are the impartment plasma membrane channels between adjacent cells, which provide a direct way of intercellular communication via the transfer of signaling molecules and substances. Recent researches have found that Drug-induced liver injury (DILI) decreased by inhibition of GJ, suggesting that GJ-mediated signal transmission has a key role in the DILI.Our previous experiments have found that, at the early stage of liver injury, PTU could significantly increase the GJ function in BRL-3A cells. Based on these findings and combined with the progress in recent years, we put up this hypothesise that the engagement of GJs induced by PTU may stimulate transmission of a ‘damage signal’ via GJ channels to neighboring cells and lead to liver injury. To verify this hypothesis, we intend to use chemical and molecular biology methods on BRL-3A,L-O2 cells and BALB / c mice to inhibit the function of GJ. On the above models, we expect to confirmed the role of GJ in PTU-induced liver injury and the related mechanism by means of SEM, western blot, RNAi and HPLC-APCI-MS methods. These studies will contribute to illustrate the effects and mechanism of GJ in PTU-induced liver injury, and provide new strategies and ideas to prevent and cure liver injury caused by PTU in a clinic.

丙硫氧嘧啶（PTU）引起的肝损伤影响其用药安全性，甚至导致患者死亡，但机制未明。细胞缝隙连接（GJ）是相邻细胞之间进行物质交换和信号传递的重要膜通道。新近研究发现干预GJ功能可减弱多种药物的肝毒性，提示经GJ介导的物质和信号传递在药物性肝损伤（DILI）中起重要作用。我们的前期实验发现：肝损伤发生早期，PTU可显著性提高BRL-3A肝细胞的GJ功能。为此，我们推测PTU可通过增强GJ功能，促进其刺激肝细胞产生的损伤信号分子在细胞间传递进而导致肝损伤发生。本课题拟在BRL-3A及L-O2正常肝细胞和BALB/c小鼠上，通过药物和分子生物学技术抑制GJ功能，借助SEM、western blot、RNAi及HPLC-APCI-MS等方法确证上述推测，以明确GJ在PTU肝损伤发生中的作用和相关损伤信号分子的性质及作用机制。研究有助于阐明PTU肝损伤的发生机制，为其临床防治提供新的干预靶点和依据。

丙硫氧嘧啶（Propylthiouracil，PTU）引起的肝损伤影响其用药安全性，但发生机制未明。细胞缝隙连接（gap junction，GJ）是相邻细胞之间形成的一种可以开放和关闭的膜通道结构，它在细胞间的物质交换和信号传递中起重要作用。已有研究提示GJ功能与药物性肝损伤有密切关系。在本研究中，我们首次发现PTU可通过上调连接蛋白32（connexin32，Cx32）的表达增强细胞GJ功能；在细胞及动物模型上，阻断GJ通道可减弱PTU的肝损伤。本研究从GJ介导的“损伤信号”传递角度来探讨PTU肝毒性受GJ功能调控的机制，发现下调Cx32 GJ功能可减少PTU所致肝细胞坏死，降低PTU的胞内摄取率及活性氧（ROS）在细胞间的传递，说明PTU及ROS是可能通过Cx32 GJ传递的“损伤信号”分子。结合PTU自身对GJ功能的上调作用，提示增强GJ功能可促进这些信号分子在胞间的扩散进而导致PTU肝毒性发生。研究成果为揭示GJ在PTU肝损伤发生中的意义提供了资料，有助于阐明PTU肝损伤的发生机制，同时也为临床选择防治药物提供思路。