Lu Hai Ying, Zhang Guo Qiang, Li Ji Cheng
Institute of Cell Biology, Zhejiang University, Hangzhou 310031.
Fen Zi Xi Bao Sheng Wu Xue Bao. 2006 Jun;39(3):249-57.
The aim of this study was to examine the microsatellite instability (MSI) and loss of heterozygosity (LOH) of locus D17S396 on chromosome 17 and their influence on the expression of nm23H1 in gallbladder tumors, which may provide experimental basis for the tumor occurrence and metastasis. Techniques such as DNA extraction from formalin-fixed paraffin-embedded tissues, polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP), ordinary silver stain were used to study MSI and LOH of locus D17S396. Envision immunohistochemistry and Leica-Qwin computer imaging techniques were used to assess the expression of gene nm23H1. In our experiment, the frequency of genetic instability of malignant gallbladder tumors was 42.55%, which was higher than that of gallbladder adenomas, while there were no genetic instability occurred in chronic cholecystitis tissue. The frequency of LOH seemed higher with the deteriorism of gallbladder tumor. Among 47 gallbladder carcinomas, the frequency of LOH and MSI were different between different differentiation cases (P < 0.05), and the frequency of LOH in liver and lymph node metastasis cases was significantly higher than those without metastasis (P < 0.01). Moreover, the frequency of LOH was higher in stage Nevin IV and V when compared with stage I, II and III. However, the frequency of MSI showed contrary correlation with some clinicopathologic characteristics. The expression of nm23H1 in gallbladder carcinoma, gallbladder adenoma and chronic cholecystitis tissue were different (P < 0.05). The case with lymph node metastasis showed significantly lower nm23H1 expression than those without lymph node metastasis (P < 0.01). Nevin stage IV and V also exhibited lower nm23H1 expression levels compared with stage I, II and Ill. Furthermore, there was no difference in nm23H1 protein expression intensity analyzed by computer imaging techniques. In gallbladder carcinomas, the positive frequency of nm23H1 protein in LOH positive group was lower than that of LOH negative group (P < 0.05). The results indicated that the genetic instability of nm23H1 gene might be implicated in pathogenesis and progression of gallbladder tumor. Both MSI and LOH of nm23H1 gene controlled the development of gallbladder tumor independently in different paths. MSI may be an early stage molecule marker of gallbladder carcinoma. LOH may be molecule marker for the deteriorism of gallbladder tissue, which could inhibit the expression of nm23H1 in local tissue of gallbladder carcinoma and endow it with high aggressive and poor prognosis. Increasing the amount of nm23H1 protein expression could effectively restrain gallbladder carcinoma metastasis and improve prognosis of patients.
本研究旨在检测17号染色体上D17S396位点的微卫星不稳定性(MSI)和杂合性缺失(LOH)及其对胆囊肿瘤中nm23H1表达的影响,为肿瘤的发生和转移提供实验依据。采用从福尔马林固定石蜡包埋组织中提取DNA、聚合酶链反应-单链构象多态性(PCR-SSCP)、普通银染等技术研究D17S396位点的MSI和LOH。采用Envision免疫组化和Leica-Qwin计算机成像技术评估基因nm23H1的表达。在我们的实验中,恶性胆囊肿瘤的基因不稳定频率为42.55%,高于胆囊腺瘤,而慢性胆囊炎组织未发生基因不稳定。随着胆囊肿瘤恶化,LOH频率似乎更高。在47例胆囊癌中,不同分化程度病例的LOH和MSI频率不同(P<0.05),肝转移和淋巴结转移病例的LOH频率显著高于无转移病例(P<0.01)。此外,与Nevin I、II和III期相比,IV和V期的LOH频率更高。然而,MSI频率与一些临床病理特征呈相反的相关性。nm23H1在胆囊癌、胆囊腺瘤和慢性胆囊炎组织中的表达不同(P<0.05)。有淋巴结转移的病例nm23H1表达明显低于无淋巴结转移的病例(P<0.01)。与Nevin I、II和III期相比,IV和V期的nm23H1表达水平也较低。此外,通过计算机成像技术分析nm23H1蛋白表达强度无差异。在胆囊癌中,LOH阳性组nm23H1蛋白的阳性频率低于LOH阴性组(P<0.05)。结果表明,nm23H1基因的遗传不稳定可能与胆囊肿瘤的发生和进展有关。nm23H1基因的MSI和LOH均通过不同途径独立控制胆囊肿瘤的发展。MSI可能是胆囊癌的早期分子标志物。LOH可能是胆囊组织恶化的分子标志物,它可抑制胆囊癌局部组织中nm23H1的表达,使其具有高侵袭性和预后不良。增加nm23H1蛋白表达量可有效抑制胆囊癌转移,改善患者预后。