透過您的圖書館登入
IP:44.204.65.189
  • 學位論文

蓮蓬萃取物對於細菌內毒素誘發肝損傷之影響

The effect of lotus seedpod extracts on lipopolysaccharide-induced liver inflammation in vivo and in vitro

指導教授 : 陳璟賢

摘要


肝臟的保健及肝病的預防是目前所需重視的課題。蓮蓬為傳統中藥材,已有研究顯示蓮蓬富含多酚(polyphone)成分,過去研究發現蓮蓬具有抗氧化、抗老化及抗癌之活性,但對於保肝的應用仍是未知。因此本篇實驗利用動物模式及細胞模式,探討蓮蓬萃取物(lotus seedpod extract;LSE)對於抗肝臟發炎之功效,並分析其分子機制。在動物實驗中,使用ICR小鼠,分別餵食1%與2%的LSE為期4週及8週,並在犧牲前一天由腹腔注射細菌內毒素(lipopolysaccharide ; LPS,20 mg/kgw)誘發小鼠急性肝損傷,犧牲後,取其血液及肝臟組織進行分析。實驗結果顯示,在血液生化數值方面,LPS組的麩胺酸草酸醋酸轉胺酶(glutamate oxaloacetate transaminase;GOT)、麩胺酸丙酮酸轉胺酶(glutamate pyruvate transaminase;GPT)、血液尿素氮(blood urea nitrogen;BUN)及三酸甘油脂(triglycerides;TG)的表現量顯著高於控制組,而餵食2% LSE的組別其表現量和LPS組相比下有顯著降低的趨勢。在發炎因子的測定當中,LPS組之TNF-a及IL-6均顯著上升,而餵食LSE的組別均有下降的趨勢。在肝臟氧化壓力測定方面,注射LPS後其氧化壓力上升,餵食2% LSE的組別有顯著降低的趨勢。而肝臟抗氧化酵素測定方面,在感染LPS後,小鼠肝臟中的麩胱苷肽過氧化酶(glutathione peroxidase;GPx)、麩胱苷肽(glutathione;GSH)及超氧歧化酶(superoxide dismutase;SOD)的活性均下降,而補充LSE的組別其活性均有上升的趨勢。以西方墨點法分析發炎相關蛋白,結果顯示LSE能降低因LPS誘發發炎相關蛋白COX-2 (cyclooxygenase-2)、iNOS (inducible nitric oxide synthase)及NF-κB (nuclear factor kappa B)的表現。上述結果顯示,平日攝取蓮蓬萃取物能夠預防肝臟急性發炎的發生。在細胞實驗中,以LPS誘發HepG2細胞產生發炎反應,並給予不同劑量LSE ( 1、2.5、5 µg/ml)或EGC 2 µM,結果顯示LSE及EGC能降低細胞IL-6的表現量。以西方墨點法分析,結果顯示LSE透過能抑制TLR4 (Toll-like receptor 4)路徑降低IκB-α之磷酸化,進而抑制NF-κB的轉錄活性,導致發炎相關因子的表現量下降。綜合以上研究結果,未來蓮蓬萃取物可能有潛力應用於保肝產品的開發。

並列摘要


Liver protection and hepatopathy prevention are very important issues in the world. Lotus seedpod, rich in polyphenol, is a traditional herbal and has been shown to possess antioxidant, anti-aging and anti-cancer activities. However, the effect of lotus seedpod on liver protection is still unknown. This study was to investigate the inhibitory effect and molecular mechanism of lotus seedpod extract (LSE) on liver inflammation and injury in vitro and in vivo. ICR mice were treated with 20 mg/kgw of lipopolysaccharide (LPS), an inducer of acute liver injury, or LPS combined LSE (1% or 2%) administration for 4 weeks and 8 weeks. The results showed that serum levels of glutamate oxaloacetate transaminase (GOT), glutamate pyruvate transaminase (GPT), blood urea nitrogen (BUN) and triglycerides (TG) in LPS-induced mice were increased as compared with control group. 2% LSE-treated group could decrease the GOT, GPT, TG and BUN levels when compared with the LPS alone group. In inflammatory cytokines analysis, we found that tumor necrosis factor (TNF)-alpha and interleukin (IL)-6 levels in plasma were significant increased after LPS injection, while both intake 1% and 2% of LSE could reduce serum levels of the cytokines. In determination of oxidative stress in liver, LPS could increase thiobarbituric acid relative substances (TBARS) level, while LSE treatments could significantly decrease oxidative stress level in mice. In measurement of antioxidant enzyme activity, LPS also decrease glutathione peroxidase (GPx) activity, glutathione (GSH) level and superoxide dismutase (SOD) activity in liver, while LSE treatments could increase these enzyme activities. Western blotting demonstrated that LSE inhibited the expression of cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS) and nuclear factor kappa B (NF-κB) in liver. These results indicated that LSE exhibited protective effects against LPS-induced acute liver injury in mice. In vitro, the data show that the level of IL-6 in HepG2 cells after treatment LPS was increased, and non-cytotoxic dose of LSE would reduce the level of IL-6. Western blotting data showed that LSE inhibited the expressions of Toll-like receptor 4 (TLR4) and MyD88. Phosphorylation of IκB-α by LPS-induced was efficiently inhibited by LSE treatment. LSE also suppressed the nuclear levels of NF-κB. Further, the protein level of COX-2 and iNOS were also decreased in LSE treatment group. In conclusion, LSE exhibited potent protective effects against LPS-induced liver injury in mice and anti-inflammatory signaling pathway in HepG2 cells. Thus, LSE might be used as a potential product for liver protection.

參考文獻


102. 柯志坤,蘑菇對於細菌內毒素誘發氧化壓力動物模式抗氧化能力之效用。中山醫學大學,台中市,2007。
4. Gregory SH, Wing EJ, Danowski KL, van Rooijen N, Dyer KF, Tweardy DJ. IL-6 produced by Kupffer cells induces STAT protein activation in hepatocytes early during the course of systemic listerial infections. J Immunol. 1998, 160: 6056-61.
1. Tacke F, Luedde T, Trautwein C. Inflammatory pathways in liver homeostasis and liver injury. Clin Rev Allergy Immunol. 2009, 36:4-12.
2. Bilzer M, Roggel F, Gerbes AL. Role of Kupffer cells in host defense and liver disease. Liver Int. 2006, 26: 1175-86.
3. Gregory SH, Wing EJ. Neutrophil-Kupffer cell interaction: a critical component of host defenses to systemic bacterial infections. J Leukoc Biol. 2002, 72: 239-48.

被引用紀錄


賴彥勳(2017)。蓮蓬萃取物應用於保肝護腎之研究〔碩士論文,中山醫學大學〕。華藝線上圖書館。https://www.airitilibrary.com/Article/Detail?DocID=U0003-2308201712520400

延伸閱讀