本文關鍵詞：Na-PCP誘導L-02肝細胞氧化應激和線粒體損傷的實驗研究　出處：《中南大學》2008年碩士論文　論文類型：學位論文

  更多相關文章： 五氯酚鈉(Na-PCP) L-02肝細胞 氧化應激 線粒體損傷

【摘要】： 目的: 在體外試驗系統(tǒng)(in vitro test),研究有機氯農(nóng)藥-五氯酚鈉(Pentachlorophenol Sodium,Na-PCP)對L-02肝細胞氧化應激和線粒體損傷,為進一步了解五氯酚鈉的毒作用機制奠定實驗基礎。 方法: 以L-02肝細胞為受試細胞,通過MTT試驗檢測不同濃度Na-PCP對L-02肝細胞存活率的影響,選擇細胞存活率為20%～90%的濃度進行后續(xù)實驗。設置1個對照組和5個Na-PCP處理組,即12.812mg/L、19.218mg/L、25.624mg/L、32.030mg/L和38.436mg/L,處理時間為24h,采用化學比色法檢測Na-PCP對L-02肝細胞超氧化物岐化酶(superoxide dismutase,SOD)活力、谷胱甘肽(reduced glutathione,GSH)和丙二醛(malondialdehyde,MDA)含量的影響;采用熒光分光光度法檢測Na-PCP對L-02肝細胞線粒體膜電位(△Ψm)和線粒體通透性轉運孔(permeability transition pore,PTP)的影響。設置1個對照組和9.609mg/L、12.812mg/L、19.218mg/L三個處理組,處理時間為24h,采用Western Blotting法檢測Na-PCP對肝細胞p53、細胞色素C和凋亡誘導因子(apoptosis-inducing factor,AIF)的蛋白水平的影響。 結果: 1.在9.609mg/L～38.436mg/L濃度范圍內,Na-PCP能明顯引起L-02肝細胞存活率的降低,且存在著濃度.反應關系(r=-0.978,P＜0.01)。 2.與對照組相比,12.812mg/L～38.436mg/L Na-PCP均可導致L-02肝細胞內MDA含量增加(r=0.956,P＜0.01),SOD活性下降(r=-0.900,P＜0.01),GSH含量減少(r=-0.975,P＜0.01)。 3.與對照組比較,在12.812mg/L～38.436mg/L濃度范圍內,隨著Na-PCP染毒濃度的增加,線粒體PTP開放度明顯增大(P＜0.01),且PTP開放度與濃度之間呈現(xiàn)明顯的濃度-反應關系(r=0.997,P＜0.01)。 4.與對照組比較,在12.812mg/L～38.436mg/L濃度范圍內,Na-PCP導致熒光吸光度值升高,線粒體膜電位明顯降低(P＜0.01),且膜電位的下降呈濃度依賴性(r=0.795,P＜0.01)。 5.經(jīng)9.609mg/L、12.812mg/L和19.218mg/L Na-PCP處理L-02肝細胞,各組細胞p53、細胞色素C和AIF的蛋白水平明顯高于對照組細胞(P＜0.05)。 結論: Na-PCP在9.609mg/L～38.436mg/L的濃度范圍內,能引起L-02肝細胞存活率下降,誘發(fā)氧化損傷,表現(xiàn)為SOD酶活力和GSH含量下降,MDA含量上升。Na-PCP導致肝細胞線粒體膜電位崩潰和通透性轉運孔開放度增大,并使得細胞色素C和AIF從線粒體釋放增多。Na-PCP導致p53蛋白水平增高。
[Abstract]:Objective:
In vitro test system (in vitro test), organochlorine pesticides (Pentachlorophenol - Sodium, Na-PCP of sodium pentachlorophenate) on L-02 hepatocyte oxidative stress and mitochondrial damage, provide an experimental basis for understanding the toxicity mechanism of pentachlorophenol.
Method:
In L-02 liver cells as test cells, the influence of the survival rate of L-02 cells was detected by MTT with different concentration of Na-PCP, the cell survival rate was 20% ~ 90% concentration for subsequent experiments. 1 control group and 5 Na-PCP group, 12.812mg/L, 19.218mg/L, 25.624mg/L, 32.030mg/L and 38.436mg/L, processing time 24h, detection of Na-PCP in L-02 cell superoxide dismutase by chemical colorimetric method (superoxide, dismutase, SOD) activity, glutathione (reduced glutathione GSH) and malondialdehyde (malondialdehyde, MDA) content influence; detection of Na-PCP by fluorescence spectrophotometry of L-02 hepatocyte mitochondria membrane potential (lpli m) and mitochondrial permeability transition pore (permeability transition, pore, PTP). The effects of 1 control group and 9.609mg/L, 12.812mg/L, 19.218mg/L three groups, treatment time is 24h, using Western Blotting method. The effect of Na-PCP on the protein level of p53, cytochrome C and apoptosis inducible factor (apoptosis-inducing factor, AIF) in liver cells.
Result:
1. in the range of 9.609mg/L ~ 38.436mg/L concentration, Na-PCP could significantly reduce the survival rate of L-02 hepatocytes, and there was a concentration. The reaction (r=-0.978, P < 0.01).
2., compared with the control group, 12.812mg/L to 38.436mg/L Na-PCP could increase the MDA content in L-02 cells (r=0.956, P < 0.01), SOD activity decreased (r=-0.900, P < 0.01), and the content of GSH decreased (r=-0.900, < 0.01).
3. compared with the control group, in the concentration range of 12.812mg/L to 38.436mg/L, with the increase of Na-PCP concentration, the mitochondrial PTP openness increased significantly (P < 0.01), and there was an obvious concentration response relationship between PTP openness and concentration (r=0.997, P < 0.01).
4. compared with the control group, in the concentration range of 12.812mg/L to 38.436mg/L, Na-PCP led to an increase in fluorescence absorbance and a decrease in mitochondrial membrane potential (P < 0.01), and the decrease of membrane potential was in a concentration dependent manner (r=0.795, P < 0.01).
5., 9.609mg/L, 12.812mg/L and 19.218mg/L Na-PCP were used to treat L-02 hepatocytes. The protein levels of p53, cytochrome C and AIF in each group were significantly higher than those in control group (P < 0.05).
Conclusion:
The concentration of Na-PCP in the range of 9.609mg/L to 38.436mg/L, L-02 can cause liver cell survival rate decreased, the oxidative damage induced by decreased SOD activity and GSH content, MDA content increased.Na-PCP induced hepatic mitochondrial membrane potential collapse and permeability transition pore opening degree increases, and the cytochrome C release from mitochondria increased.Na-PCP and AIF lead to increased p53 protein levels.

【學位授予單位】：中南大學
【學位級別】：碩士
【學位授予年份】：2008
【分類號】：R363

【參考文獻】

相關期刊論文 前10條

1 宋瑞霞;阮鴻潔;景欣月;劉征濤;;五氯酚鈉的細胞遺傳毒性研究[J];癌變.畸變.突變;2007年05期

2 呂華東,黃心宜,林玉珍,王謀鳳,林仲;環(huán)境中五氯酚污染監(jiān)測與人體攝入蓄積情況[J];福建環(huán)境;1996年01期

3 徐蘭琴;陶濤;鄧穗德;呂嘉春;冉丕鑫;;醛酮類物質染毒對小鼠肺臟抗氧化能力的影響[J];廣州醫(yī)學院學報;2007年03期

4 包志成,王克歐,康君行,趙立文;五氯酚及其鈉鹽中氯代二惡英類分析[J];環(huán)境化學;1995年04期

5 張民,王s，

本文編號：1417703