通過線粒體DNA D-Loop區序列分析鑒別多結節性肝細胞性肝癌細胞克隆起源_《中國普外基礎與臨床雜志》

作者：

李仕來 ¹ , 黎樂群 ² , 彭濤 ² , 蘇銘 ²

1.廣西醫科大學第一附屬醫院急診科（南寧530021）;;
2.廣西醫科大學第一附屬醫院肝膽外科（南寧530021）;

關鍵詞：

線粒體DNA D-Loop區多結節性肝癌克隆起源

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目的探討通過線粒體 DNA (mtDNA) D-Loop 區序列分析鑒別多結節性肝細胞性肝癌（肝癌）細胞克隆起源的可行性。方法實驗組為多結節肝癌組，標本來自廣西醫科大學第一附屬醫院肝膽外科2004年4月至2007年8月期間連續收治的42例共112個結節性HCC行根治性手術切除的腫瘤組織。對照組來自同期單結節HCC手術切除組織共20例40個樣本，分2個亞組，對照組Ⅰ為16例單結節肝癌的2塊不相鄰的癌組織；對照組Ⅱ為4例伴有肉眼門靜脈癌栓的肝癌患者，取癌組織和癌栓各1塊；正常對照組來自同期肝移植供肝或肝外傷切除的肝組織共5例。用PCR結合直接測序的方法檢測各樣本組織mtDNA D-Loop 區的序列，并分析各例癌結節中序列異同情況。結果在42例實驗組標本中，共有131個位點發生堿基變異，其中15個位點為點突變，9個位點發生插入，16個位點發生缺失，共出現98個多態性變化，位點總變異率為11.7%（131/1 122，1 122為mtDNA D-Loop 區全長）；實驗組中有20例各結節的mtDNA D-Loop 區序列存在差異，可能為多中心（MO）起源，22例各結節的mtDNA D-Loop 區序列完全相同，可能為單中心（即肝內轉移，IM）起源。對照組20例中各樣本的mtDNA D-Loop 區序列完全相同，為相同細胞克隆起源。正常對照組中共有14個位點發生堿基變異，均為多態性，其中NT 479 A gt;G為新的多態性。結論 mtDNA D-Loop區序列具有較高的變異率，應用PCR結合直接測序的方法檢測該區序列并比較各個癌結節的DNA序列異同，可以快速、簡單、有效地為區分IM和MO起源提供參考。

引用本文： 李仕來,黎樂群,彭濤,蘇銘. 通過線粒體DNA D-Loop區序列分析鑒別多結節性肝細胞性肝癌細胞克隆起源. 中國普外基礎與臨床雜志, 2010, 17(6): 567-572. doi: 復制

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13.	Nowell PC. The clonal evolution of tumor cell populations [J]. Science, 1976; 194(4260): 23-28.
14.	Calabrese P, Tavaré S, Shibata D. Pretumor progression: clonal evolution of human stem cell populations [J]. Am J Pathol, 2004; 164(4): 1337-1346.
15.	Tang ZY, Ye SL, Liu YK, et al. A decade’s studies on metastasis of hepatocellular carcinoma [J]. J Cancer Res Clin Oncol, 2004; 130(4): 187-196.
16.	Copeland WC, Wachsman JT, Johnson FM, et al. Mitochondrial DNA alterations in cancer [J]. Cancer Invest, 2002; 20(4): 557-569.
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26.	Okochi O, Hibi K, Uemura T, et al. Detection of mitochondrial DNA alterations in the serum of hepatocellular carcinoma patients [J]. Clin Cancer Res, 2002; 8(9): 2875-2878.

1. Yamamoto T, Kajino K, Kudo M, et al. Determination of the clonal origin of multiple human hepatocellular carcinomas by cloning and polymerase chain reaction of the integrated of hepatitis B virus DNA [J]. Hepatology, 1999; 29(5): 1446-1452.
2. Liver Cancer Study Group of Japan. Classification of primary liver cancer [S]. Frist English Edition, Kanehara Shuppan, Tokyo， 1997： 1-22.
3. Matsumoto Y, Fuji H, Matsuda M, et al. Multicentric occurrence of hepatocellular carcinoma: diagnosis and clinical significance [J]. J Hepatobiliary Pancreat Surg, 2001; 8(5): 435-440.
4. Esumi M, Aritaka T, Arri M, et al. Clonal origin of human hepatoma detemined by integration of hepatitis B virus DNA [J]. Cancer Res, 1986; 46(11): 5767-5771.
5. Chen PJ, Chen DS, Lai MY, et al. Clonal origin of recurrent hepatocellular carcinomas [J]. Gastroenterology, 1989; 96(2 Pt 1): 527-529.
6. Hsu HC, Chiou TJ, Chen JY, et al. Clonality and Clonal evolution of hepatocellular carcinoma with multiple nodules [J]. Hepatology, 1991; 13(5): 923-928.
7. Oda T, Tsuda H, Scarpa A, et al. Mutation pattern of the p53 gene as a diagnostic marker for multiple hepatocellular carcinoma [J]. Cancer Res, 1992; 52(13): 3674-3678.
8. Peng SY, Lai PL, Chu JS, et al. Expression and hypomethylation of alpha-fetoprotein gene in unicentric and multicentric human hepatocellular carcinomas [J]. Hepatology, 1993; 17(1): 35-41.
9. Chen YJ, Yeh SH, Chen JT, et al. Chromosomal changes and clonality relationship between primary and recurrent hepatocellular carcinoma [J]. Gastroenterology, 2000; 119(2): 431-440.
10. Okamoto M, Utsunomiya T, Wakiyama S, et al. Specific gene-expression profiles of noncancerous liver tissue predict the risk for multicentric occurrence of hepatocellular carcinoma in hepatitis C virus-positive patients [J]. Ann Surg Oncol, 2006; 13(7): 947-954.
11. Yamada S, Nomoto S, Fujii T, et al. Correlation between copy number of mitochondrial DNA and clinico-pathologic parameters of hepatocellular carcinoma [J]. Eur J Surg Oncol, 2006; 32(3): 303-307.
12. 胡義德, 錢桂生, 李淑平, 等. 人癌細胞線粒體DNA控制區序列特征分析 [J]. 遺傳, 1999; 21(4): 1-6.
13. Nowell PC. The clonal evolution of tumor cell populations [J]. Science, 1976; 194(4260): 23-28.
14. Calabrese P, Tavaré S, Shibata D. Pretumor progression: clonal evolution of human stem cell populations [J]. Am J Pathol, 2004; 164(4): 1337-1346.
15. Tang ZY, Ye SL, Liu YK, et al. A decade’s studies on metastasis of hepatocellular carcinoma [J]. J Cancer Res Clin Oncol, 2004; 130(4): 187-196.
16. Copeland WC, Wachsman JT, Johnson FM, et al. Mitochondrial DNA alterations in cancer [J]. Cancer Invest, 2002; 20(4): 557-569.
17. Klaunig JE, Kamendulis LM. The role of oxidative stress in carcinogenesis [J]. Annu Rev Pharmacol Toxicol, 2004; 44: 239-267.
18. Lièvre A, Blons H, Houllier AM, et al. Clinicopathological significance of mitochondrial D-Loop mutations in head and neck carcinoma [J]. Br J Cancer, 2006; 94(5): 692-697.
19. Han CB, Li F, Zhao YJ, et al. Variations of mitochondrial D-Loop region plus downstream gene 12S rRNA-Trna(phe) and gastric carcinomas [J]. World J Gastroenterol, 2003; 9(9): 1925-1929.
20. Yamamoto H, Tanaka M, Katayama M, et al. Significant existence of deleted mitochondrial DNA in cirrhotic liver surrounding hepatic tumor [J]. Biochem Biophys Res Commun, 1992; 182(2): 913-920.
21. Nishikawa M, Nishiguchi S, Shiomi S, et al. Somatic mutation of mitochondrial DNA in cancerous and noncancerous liver tissue in individuals with hepatocellular carcinoma [J]. Cancer Res, 2001; 61(5): 1843-1845.
22. Lee HC, Li SH, Lin JC, et al. Somatic mutations in the D-Loop and decrease in the copy number of mitochondrial DNA in human hepatocellular carcinoma [J]. Mutat Res, 2004; 547(1-2): 71-78.
23. Ikeda K, Arase Y, Kobayashi M, et al. Significance of multicentric cancer recurrence after potentially curative ablation of hepatocellular carcinoma: a longterm cohort study of 892 patients with viral cirrhosis [J]. J Gastroenterol, 2003; 38(9): 865-876.
24. Utsunomiya I. Pathological study on multicentric occurrence of hepatocellular carcinoma [J]. Kurume Med J, 2005; 52(4): 133-138.
25. Fliss MS, Usade1 H, Caballero OL, et al. Facile detection of mitochondrial DNA mutations in tumors and bodily fluids [J]. Science, 2000; 287(5460): 2017-2019.
26. Okochi O, Hibi K, Uemura T, et al. Detection of mitochondrial DNA alterations in the serum of hepatocellular carcinoma patients [J]. Clin Cancer Res, 2002; 8(9): 2875-2878.

《中國普外基礎與臨床雜志》

通過線粒體DNA D-Loop區序列分析鑒別多結節性肝細胞性肝癌細胞克隆起源

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《中國普外基礎與臨床雜志》

通過線粒體DNA D-Loop區序列分析鑒別多結節性肝細胞性肝癌細胞克隆起源

摘要 全文 圖表 視頻 參考文獻 施引文獻 補充材料

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