蒲葵醇類萃取物抑制C型肝炎病毒複製與降低病毒誘導性發炎反應之研究

C型肝炎病毒的感染，造成肝臟長期處於慢性發炎狀態，易導致肝纖維化、肝硬化甚至提高罹患肝癌的風險，因而，C型肝炎疾病成為全球公共衛生問題的隱憂。目前尚未研發出預防性C型肝炎疫苗，且現今治療性藥物並非適合所有的病患，卻帶來其他併發的副作用以及產生抗藥性反應，因此，研究開拓新穎的抗病毒藥物與療法是一個十分重要的目標。本實驗研究方向為探討蒲葵醇類萃取物(Methanol extract of the fruits of Livistona chinensis; LCMF )抑制C型肝炎病毒複製機轉與降低病毒性所誘導的發炎反應。根據我們實驗數據發現，使用兩種檢測抗C型肝炎病毒系統，在無藥物毒性反應下於C型肝炎複製子細胞株與病毒感染模式其EC50 (Effective concentration)分別為33.2與36.5 μg/ml，並探討LCMF其抑制病毒複製的機制，是藉由銳減細胞中環氧合酶-2 (Cyclooxygenase -2；COX-2 )基因過度表現，而減弱病毒複製能力。更進一步觀察，LCMF能阻斷位於COX-2基因上游的調節因子－核轉錄因子(nuclear transcription factor kappa B；NF-κB)進入細胞核中調節基因轉錄，並且LCMF抑制MAPK (Mitogen-activated protein kinases)中Jnk傳訊因子活化。另一方面，實驗結果顯示，LCMF可降低C型肝炎病毒刺激所產生的發炎激素分泌，包含TNF-α (Tumor necrosis factor -α)、IL-1β (Interleukin-1β)、iNOS (inducible NO synthase)，代表其具有抗病毒性誘導的發炎反應。最後，評估LCMF與藥物合併治療效果，其有助於提高干擾素INF-α或其他病毒抑制劑Telaprevir以及Sofosbuvir 之抗C型肝炎病毒複製效用。總結而言，LCMF是透過NF-κB -COX-2途徑抑制C型肝炎病毒複製，且具抗發炎作用之能力，其可能成為一種有潛力待研究開發的藥物或療法。

關鍵字

蒲葵萃取物； C型肝炎病毒；環氧合酶-2 ；核轉錄因子；病毒誘導性發炎反應

並列摘要

Hepatitis C virus (HCV) infection cause formation of chronic inflammation in liver over a long period and result in hepatic fibrosis, cirrhosis even hepatocellular carcinoma (HCC). Current therapies are limited due to serious adverse effects and drug resistance. Development of innovative antiviral therapies for HCV infection represents an important goal of public health. In this study, we attempted to investigate the mechanism of methanol extracts from the fruits of Livistona Chinesis, termed as LCMF, against viral replication and its anti-inflammatory effect on HCV-infected hepatocyte. We found that LCMF exhibited the inhibtion of HCV replication with an EC50 value (effective concentration) of 33.2 and 36.5 μg/ml without cytotoxicity, in HCV replicon cell line and infectious system, respectively. Next, our results demonstrated that LCMF suppressed HCV replication through down-regulation of the COX-2 (Cyclooxygenase -2) gene overexpression in HCV replicon cells. Furthermore, LCMF can prevent the translocation of free NF-κB into nucleus and for activation of transcription of COX-2 gene expression. In addition, LCMF also cause the inhibitory effect on mitogen-activated protein kinase (MAPK) signaling pathway in HCV replicon cell line. The results showed that phospho-JNK protein levels were reduced by LCMF treatment in a time-dependent manner. On the other hand, it has been shown that LCMF can reduce the production of HCV-stimulated proinflammatory cytokines, such as TNF-α ( tumor necrosis factor-α), IL-1β(interleukin-1β), and iNOS ( inducible nitric oxide synthase). Finally, we comfirmed that the combination of LCMF with interferon-α or other viral inhibitor, such as Telaprevir and Sofosbuvir, have synergistic effects against HCV replication. In summary, LCMF can inhibit the HCV replication via down-regualtion of COX-2 expression and attenuate HCV-induced inflammation. These ﬁndings indicated that LCMF could be as a poteintial supplement against HCV infection.