眼附属器黏膜相关淋巴组织淋巴瘤的Bcl-10基因突变研究

目的探讨Bcl-10基因在我国人群眼附属器黏膜相关淋巴组织（MALT）淋巴瘤、不典型淋巴组织增生和淋巴组织反应性增生中的表达和新突变。方法收集第二军医大学附属长征医院眼科患者新鲜冰冻组织标本31例,其中眼附属器MALT淋巴瘤23例,不典型淋巴组织增生4例,淋巴组织反应性增生4例。采用分子生物学方法获得Bcl-10基因,以双脱氧Sanger法行DNA测序,Blast比对分析后,得到突变碱基。同时行免疫组织化学检测和免疫荧光定位,用激光共焦显微镜检测Bcl-10和NF—κB的共定位情况。结果31例中,检出14例眼附属器MALT淋巴瘤有Bcl-10基因表达,其中10例发现新的基因突变。4例不典型淋巴组织增生和4例淋巴组织反应性增生中,各有1例发生新突变。免疫组织化学检测发现异常的Bcl-10基因在14例MALT淋巴瘤中均有表达（60．8％）,其中中等强度核表达为6例,弱到中等强度胞质表达为8例。2例不典型增生标本胞质中有异常Bcl-10表达,1例反应性增生标本胞质中亦见异常Bcl-10表达。免疫荧光定位发现NF—κB的反应因子Iκα在20例胞质中弥漫性表达,其中Bcl-10和Iκα共表达有14例。结论Bcl-10基因在我国人群眼附属器MALT淋巴瘤中有新的突变形式,检测到的基因突变分布与病理诊断一致。基因检测的灵敏度高于病理诊断,在尚无形态学改变及其他可供鉴别的指标下,能判断出病变阶段和性质改变,可作为早期诊断的灵敏指标之一。（中华眼科杂志,2007,43：1010—1016）

Significance of detection of Bcl-10 novel mutation in ocular adnexal mucosa-associated lymphoid tissue lymphoma

OBJECTIVE: To examine the expression of Bc1-10 gene in mucosa-associated lymphoid tissue (MALT) lymphomas, atypical lymphatic hyperplasia and reactive lymphatic hyperplasia; as well as to discover novel mutations and their role in clinical diagnosis and pathogenesis of ocular adnexal lymphatic disorders. METHODS: Thirty-one specimens of ocular adnexal lymphoma were obtained from the department of ophthalmology of Shanghai Changzheng hospital, second military medical university during surgery and were preserved in liquid nitrogen immediately after harvesting. The expression of Bc1-10 gene in ocular adnexal specimens was examined by molecular biological methods. The DNA sequences were analyzed by Sanger method and the results were compared with those of the Gene Bank blast to identify novel mutations. The protein expression of mutant Bc1-10 and NF-kappaB in these specimens were detected by immunohistological method and immunofluorescence. Confocal microscope was used to observe the colocalization of Bc1-10 and NF-kappaB expressions. RESULTS: Of the 31 specimens, 14 were positive for Bc1-10, of which 10 showed novel mutations of Bc1-10. One of the four atypical lymphatic hyperplasia specimens was positive for Bc1-10, this mutation was reported by others previously. One of the four reactive lymphatic hyperplasia specimens was positive for Bc1-10. Fourteen (60.8%) of the 23 MALT lymphoma specimens were positive for aberration of Bc1-10, including 6 cases with nuclear expression at moderate intensity and 8 cases with expression in the cytoplasm at weak to moderate intensities. Mutations of Bcl-10 were also expressed in the cytoplasm at moderate intensity in two cases with atypical lymphatic hyperplasia and expressed at moderate intensity in one case with reactive lymphatic hyperplasia. I kappa alpha was expressed diffusely in the cytoplasm in 20 cases, among which co-expression of I kappa alpha and Bcl-10 was present in 14 cases. CONCLUSIONS: Novel mutation of Bc1-10 gene in ocular adnexal MALT lymphoma is detected in Chinese patients. Detection of genetic mutation is consistent with and more sensitive than the pathological diagnosis. It can be used to identify the stage and the quality of the disease even no morphological changes are present for discrimination. It can be used as a sensitive index for early diagnosis.