CD14是一種表現在單核球細胞膜上的分化抗原，已知是LPS的接受器之一。由過去的研究顯示CD14啟動子-159T的多型性與其基因的表現有關，不論是在PBMC上mCD14以及血清中的sCD14其表現量均是TT型的族群大於CC型，而CD14-159T與新生兒膽道閉鎖疾病的發生率也有相關性。已有文獻證實CD14的表現量，在膽道結紮的動物模式以及膽道閉鎖病人的肝臟組織切片中都有上升的現象，過去被認為可能是單核細胞被活化所引起，但是CD14基因詳細調控機轉仍未被釐清。但也有報告證實人類的肝細胞也會合成CD14分子，屬於一種發炎急性期反應蛋白質，而我們自己的研究結果，也證明膽道閉鎖病患其肝細胞中CD14會明顯增加。因此本研究的目標著重在探討CD14在肝細胞中的表現及在膽汁鬱積的情形之調控機轉。我們首先構築CD14啟動子報導基因質體，以分析報導基因的方式來探討CD14在肝細胞的調控機轉。結果顯示CD14在肝細胞的表現，其主要調節的活性的重要區域在啟動子-300至-232之間，而在-376至-358之間的GATA-1結合位，則是一個抑制活性的區域。我們實驗室先前的研究發現臨床膽道閉鎖病人的血漿中sCD14濃度上升，而且其肝臟中的CD14 mRNA表現量明顯增加，由於膽汁鬱積會造成肝細胞內膽鹽堆積，而膽鹽的刺激會透過活化PKC的機制，促使肝細胞進行細胞凋亡而造成肝臟損傷。所以我們直接利用PMA(PKC agonist)來釐清PKC活化對CD14表現的影響。結果顯示在Huh-7細胞株，CD14啟動子的活性能被PMA誘導，且能被RO318220 (PKC inhibitor)所抑制。利用PMA刺激肝細胞三小時後，CD14 mRNA表現量也呈現明顯上升。綜合以上研究，我們證實CD14在肝細胞中具有高度的轉錄活性，而在膽汁鬱積的過程中，其活性上升的機制，可能和PKC的訊息傳導路徑有關。

CD14 is a pattern recognition receptor that plays a central role in innate immunity through recognition of lipopolysaccarides. Recently, our group has showed CD14 -159T promoter polymorphism was related with the gene expression. Homozygous carriers of the T allele have a significant increase in mCD14 on monocytes and sCD14 in sera. In our previous study, The CD14 -159T promoter polymorphism was also associated with the development biliary atresia (BA) and idiopathic neonatal cholestasis. Although increased expression of CD14 in liver specimens of BA patients have been reported, the exact mechanism of over-expression of CD14 in cholestasis is yet to be elucidated. Human hepatocytes were demonstrated to produce CD14 as an acute phase protein, whereas little literature documented the regulation mechanisms of CD14 gene expression in liver. Therefore, the specific goal of this study is to investigate the regulation mechanism of CD14 gene expression in Huh-7 cells. Preliminary data of our laboratory, significantly higher hepatic CD14 mRNA expression and plasma sCD14 levels in BA patients were demonstrated. It implies that CD14 might be involved in pathogenesis and progression of BA. The basal activity of CD14 expression in hepatocytes was investigated by promoter report assays. Based on the data of serial deletion analysis, the hepatocyte-specific regulation region of CD14 promoter is localized in -300/-232 segment, and the GATA-1 binding element within -376/-358 was demonstrated to act as a repressor. It was reported that accumulated bile salts may cause liver damage and induce the apoptosis of hepatocytes via the activation of protein kinase C (PKC). Our results also showed the activity of CD14 promoter and the levels of CD14 mRNA can be activated by PMA (PKC agonist) and inhibited by RO318220 (PKC inhibitor). It implies that PKC signal pathway may involve in CD14 production in hepatocytes during cholestasis. In conclusion, this study demonstrated CD14 was highly expressed in hapatocytes and might be up-regulated in cholestasis via PKC activation.

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