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摘要:
目的 建立EBV潜伏感染的套细胞淋巴瘤模型，为淋巴瘤预防与治疗的研究奠定基础。 方法 利用新霉素耐性的重组遗传基因EBV（NeoR EBV）和B95-8 EBV对套细胞淋巴瘤细胞株SP49、SP53和Jeko-1进行感染，选出EBV阳性的MCL细胞株，采用免疫荧光法和Southern blot进行鉴定，采用Western blot和流式细胞仪检测EBV阳性的MCL细胞株的免疫表型及生物学活性。 结果 成功获得SP49/EBV、SP53/EBV细胞株，均表达EBER1、EBNA2和LMP1，其免疫表型只有CD38表达增强，两细胞株均具有低增殖功能，凋亡抵抗性，成瘤能力下降等特点。 结论 本研究建立的SP49/EBV和SP53/EBV细胞株，具有体内EBV潜伏感染的静息B细胞低增殖活性的特征，可成为研究体内EBV潜伏感染的良好模型。 Abstract:Objective To establish a mantle cell lymphoma (MCL) model latently infected by EBV for researches of prevention and treatment of lymphoma. Methods Neomycin resistant recombinant EBV (NeoR EBV) and B95-8 EBV genes were used to infect B cell lines of SP49, SP53 and Jeko-1. EBV-positive MCL cell lines were selected through immunofluorescence and Southern blot. The immune phenotypes and biological activities of EBV-positive MCL cell lines were detected by Western blot and flow cytometry. Results SP49/EBV and SP53/EBV cell lines expressing EBER1, EBNA2 and LMP1 were successfully established, with only enhanced CD38 immune phenotype. The abilities of proliferation, apoptotic resistance as well as tumor forming decreased in both cell lines. Conclusion SP49/EBV and SP53/EBV cell lines established in this study were characterized with lower abilities of proliferation similar to resting B cells latently infected by EBV in vivo. Therefore the above cell lines may be good models of research for latent EBV infection in vivo. -
Key words:
- EB virus /
- Latent infection /
- Resting B cell /
- Mantle cell lymphoma
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图 1 蛋白印迹法检测Bcl-2、P53和CyclinD1蛋白
Figure 1. 1 Detection of Bcl-2, P53 protein and CyclinD1 using Western blot EBNA2×1 000;LMP-1×1 000;EBER1×400
图 2 SP53和SP53/EBV细胞株的EBNA2、LMP-1蛋白荧光像及EBERs原位杂交像
Figure 2. Immunofluorescent staining of SP53 and SP53/EBV in EBNA2 and LMP-1 cell lines and EBERs in situ hybridization. (Original EBNA2× 1000; LMP-1×1000; EBER1×400)
图 3 DNA印迹法检出EB病毒DNA
用BamHI（a）或BamHI/Bgl II（b）消化DNA，电泳后用BamHI-W fragememt作为探针，检出EBV DNA断片。1：SP50B，2：Akata-EBV，3：SP49，4：EBV-carrying SP49，5：SP-53，6，7：EBV-carrying SP53。6和7表示不同时期感染EB病毒的SP53细胞株
Figure 3. Southern blot detection of EB virus DNA
图 4 SP53/EBV裸鼠淋巴瘤实验模型肿瘤组织（箭头）HE染色× 100 CD5免疫组化照片×100
Figure 4. SP53/EBV experimental models of lymphoma in nude mice (arrow), H&E staining×100, CD5 IHC×100
表 1 使用的抗体
Table 1. The antibodies applied in the stady
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