适体技术在肿瘤诊断和治疗中的应用

适体(aptamer)是用配体指数富集法系统演化(SELEX)技术筛选获得的能与多种靶分子特异高效结合的单链或双链寡核苷酸,其制备方便、稳定性好、适用范围广,具有很多蛋白质抗体所不具备的优点.适体技术自出现以来受到广泛重视,在生命科学特别是医学领域发展迅速.本文简要介绍适体技术在肿瘤诊断和治疗中的应用进展.     

BAFF对外周血未成熟B淋巴细胞存活的作用

B淋巴细胞成熟是一个高度的选择过程,需要精细的分化和存活信号.BAFF(TNF家族的B细胞活化因子)能够结合B淋巴细胞,促进BCR介导的细胞增殖.阻断BAFF的作用时外周B细胞数目减少,而在BAFF转基因小鼠中则可检测到高水平的Bcl-2.该研究主要探讨了BAFF在体外对脾脏未成熟移行B细胞(Transitional B lymphocyte)存活的作用.     

DNA去甲基化对急性B淋巴细胞白血病细胞株中DNA甲基转移酶基因及微RNA作用的研究

目的：探索甲基化转移酶抑制剂5-杂氮-2'-脱氧胞苷（5-Aza-CdR）对急性B淋巴细胞白血病（B-ALL）细胞株NALM-6的作用以及对细胞中微RNA（miRNA）表达水平的影响。方法用不同浓度5-Aza-CdR处理NALM-6细胞,采用四甲基偶氮唑盐（MTT）法检测细胞增殖情况,采用荧光定量反转录聚合酶链反应（RT-PCR）检测5-Aza-CdR处理后细胞DNA甲基转移酶（DNMT）基因mRNA表达水平的变化,采用miScript miRNA PCR Array芯片检测去甲基化后细胞中表达量发生改变的miRNA。结果 NALM-6细胞经不同浓度5-Aza-CdR处理不同时间后,细胞生长受抑,最高抑制率达（74.163±0.381）%。5-Aza-CdR作用浓度与DNMT基因mRNA表达水平呈反比,浓度为1000μmol／L的5-Aza-CdR作用细胞72 h后,DNMT-1的相对表达量降至0.453±0.021,DNMT-3L的相对表达量为0.003±0.001, DNMT-3B的相对表达量为0.395±0.019。miScript miRNA PCR Array筛选出3个miRNA（miR-184、miR-23a-3p、miR-34a-5p）与DNA甲基化相关。结论5-Aza-CdR可下调NALM-6细胞中DNMT基因的表达,并对细胞增殖有抑制作用。miR-184、miR-23a-3p和miR-34a-5p在B-ALL的发生、发展中与DNA甲基化相关。     

B淋巴细胞参与肿瘤免疫抑制机制的研究进展

B淋巴细胞作为体液和细胞免疫的重要组成部分,能够通过多种调控方式发挥正性免疫调节作用.然而,越来越多的研究表明,B淋巴细胞亦可通过多种途径参与机体的免疫抑制调节,其中在肿瘤中以免疫抑制作用为主.目前研究多关注于B淋巴细胞特殊亚型-调节性B细胞(regulatory B cells,Bregs)在肿瘤中发挥的免疫抑制性作用,并通过分泌多种细胞因子、调控T细胞的作用以及直接作用于恶性肿瘤细胞等多途径影响肿瘤的发生发展进程.本文就B淋巴细胞在肿瘤免疫抑制机制研究的最新进展作一综述,为B淋巴细胞作为抗肿瘤潜在治疗靶点及治疗策略提供新的思路.     

β—胡萝卜素对人淋巴细胞DNA合成的影响

β—胡萝卜素(β—C)对肿瘤防治作用的研究一直为学者所关注。β—胡萝卜素是植物中含量较高的一种类胡萝卜素,广泛存在于自然界中,没有任何毒副作用并具有食用的安全性。有文献报道,它可以明显地预防紫外线、γ射线及χ射线对机体造成的损伤,对吸烟引起的肺癌具有潜在的预防作用,所以目前对β—胡萝卜素生物学效应的研究日益增多。但其对人体淋巴细胞DNA合成的影响国内外报道尚不多见,本文仅就其在体外对胎儿胸腺和脾脏淋巴细胞的增殖作用进行探讨。     

护骨素和可溶性核因子кB受体活化因子配体在非小细胞肺癌诊断和鉴别诊断中的意义

目的探讨护骨素(OPG)和可溶性核因子кB受体活化因子配体(sRANKL)在非小细胞肺癌(NSCLC)诊断和鉴别诊断中的意义。方法分别选取50例NSCLC患者、25例肺部良性占位患者和25例正常体检者,采用酶联免疫吸附法检测患者中血清OPG和sRANKL的水平。结果NSCLC组的sRANKL水平和sRANKL/OPG比值均高于良性占位组和正常体检组(P<0.05);三组间OPG水平比较,差异无统计学意义(P>0.05)。血清sRANKL和sRANKL/OPG比值诊断NSCLC的最佳截断点分别为>4.20 pmol/L和>0.60,sRANKL诊断的敏感度、特异度和准确性分别为74.0%、84.0%和77.3%,sRANKL/OPG比值诊断的敏感度、特异度和准确性分别为84.0%、88.0%和85.3%。血清sRANKL和sRANKL/OPG比值鉴别诊断肺部良恶性占位病变的最佳截断点分别为>5.24 pmol/L和>0.63,sRANKL鉴别诊断的敏感度、特异度和准确性分别为60.0%、84.0%和68.0%,sRANKL/OPG比值诊断的敏感度、特异度和准确性分别为78.0%、64.0%和73.3%。结论血清sRANKL水平和sRANKL/OPG比值可作为NSCLC的辅助诊断指标。     

体外硫芥对人外周血淋巴细胞DNA的损伤

目的与方法：利用碱性单细胞凝胶电泳（ＳＣＧＥ）技术在体外检测硫芥对人外周血淋巴细胞的损伤情况。结果与结论：结果表明：在硫芥的作用下细胞ＤＮＡ的迁移率和迁移长度增加,且呈显著剂量效应关系。     

铁过量对大鼠淋巴细胞DNA损伤的影响

目的： 探讨不同剂量铁补充对大鼠淋巴细胞DNA损伤的影响。方法：SPF级6~7周龄雄性Wistar大鼠40只,按体质量随机分为4组,每组10只大鼠,即对照组、缺铁组、10倍铁剂量补充组、20倍铁剂量补充组,4组均采用隔日腹腔注射右旋糖酐铁,每次注射0.72 mL,每次注射含Fe2+分别为0.9、0.3、9和18 mg,连续注射6周。4组大鼠自由饮用去离子水,喂饲无铁饲料,于第6周末眼眶取血,使用血清铁试剂盒测定血清铁浓度,采用碱性单细胞凝胶电泳法测定大鼠外周血淋巴细胞DNA损伤状况。结果：缺铁组大鼠血清铁含量为53.54 μmol/L,明显低于正常对照组的77.62 μmol/L(P<0.01),10倍和20倍铁剂量补充组大鼠血清平均铁含量分别达到104.77 μmol/L和205.30 μmol/L,显著高于对照组(P<0.01)。外周血淋巴细胞DNA本底损伤分析显示,缺铁组和正常对照组淋巴细胞DNA损伤总体水平分别为25.30 AU和21.13 AU,两组间差异无统计学意义(P>0.05),10倍和20倍铁剂量补充组DNA自发损伤水平分别为对照组的3.9倍和8.0倍(82.80 AU和169.50 AU),显著高于对照组(P<0.01);然而H2O2与铁过量联合损伤分析显示,大鼠外周血淋巴细胞在10 μmol/L H2O2处理后,缺铁组大鼠DNA氧化损伤水平达到260.40 AU,与对照组(259.00 AU)相比差异无统计学意义(P>0.05),而10倍和20倍铁剂量补充组分别为对照组的1.1倍和1.2倍(293.80 AU和308.88 AU),均明显高于正常对照组(P<0.01)。结论：10倍、20倍铁剂量补充均可提高机体铁的营养状况或增加铁负荷水平;与正常铁摄入水平相比,铁缺乏未见DNA本底及H2O2联合损伤增加,而铁补充过量可引发机体外周血淋巴细胞DNA本底损伤增加及H2O2联合损伤加剧。     

活化的B细胞——肿瘤免疫治疗新方向

肿瘤细胞免疫治疗已取得了丰硕的研究成果,在临床应用中显示出了较好的疗效.其中树突状细胞（DC）研究较多,但由于培养困难而限制了其在临床上的应用.活化的B细胞正好克服了DC这一缺陷,成为肿瘤免疫治疗的新方向.     

壬基酚对牡蛎血淋巴细胞的DNA损伤

【摘要】 背景与目的： 采用单细胞凝胶电泳技术研究壬基酚(nonylphenol, NP)对牡蛎血淋巴细胞DNA的损伤作用。 材料与方法：分别以0(阴性对照)、0.025、0.10、0.25和1.00 mg/L的NP处理牡蛎24 h和48 h,检测NP对血淋巴细胞DNA的损伤作用。 结果： 当NP在0.025 mg/L时可以使牡蛎血淋巴细胞产生轻微的DNA损伤, 染毒24 h后DNA破坏率为6％,48 h后破坏率为20％。随着NP浓度的增加和染毒时间的延长,血淋巴细胞的DNA破坏率相应增加,其中最高浓度的处理组(1.00 mg/L)DNA 损伤最为明显, 染毒24 h后的破坏率为30％,48 h后为60％。 结论： NP对牡蛎血淋巴细胞具有遗传毒性。     

Developing aptamers into tumor diagnostics and therapeutics

Aptamers are small singlestranded nucleic acid molecules that bind a target protein with high affinity and specificity. Due to their stability, low toxicity and immunogenicity, as well as improved safety, aptamers are attractive alternatives to antibody and are therefore suitable for in vivo applications. Aptamers are typically isolated, through a process termed SELEX (systematic evolution of ligands by exponential enrichment), from combinatorial libraries with desired proteins. In the present review, the recent nonconventional aptamer selection process will be discussed together with an overview on the aptamer application in cancer diagnosis and therapy.     

Involvement of p53 in insulin-like growth factor binding protein-3 regulation in the breast cancer cell response to DNA damage

Chemotherapy drugs that induce apoptosis by causing DNA double-strand breaks, upregulate the tumor suppressor p53. This study investigated the regulation of the growth-regulatory protein insulin-like growth factor binding protein-3 (IGFBP-3), a p53 target, by DNA-damaging agents in breast cancer cells. IGFBP-3 was upregulated 1.4- to 13-fold in response to doxorubicin and etoposide in MCF-10A, Hs578T, MCF-7 and T47D cells, which express low to moderate basal levels of IGFBP-3. In contrast, IGFBP-3 was strongly downregulated by these agents in cells with high basal levels of IGFBP-3 (MDA-MB-231, MDA-MB-436 and MDA-MB-468). In MDA-MB-468 cells containing the R273H p53 mutation, reported to display gain-of-function properties, chemotherapy-induced suppression of IGFBP-3 was not reversed by the p53 reactivating drug, PRIMA-1, or by p53 silencing, suggesting that the decrease in IGFBP-3 following DNA damage is not a mutant p53 gain-of-function response. SiRNA-mediated downregulation of endogenous IGFBP-3 modestly attenuated doxorubicin-induced apoptosis in MDA-MB-468 and Hs578T cells. IGFBP-3 downregulation in some breast cancer cell lines in response to DNA-damaging chemotherapy may have clinical implications because suppression of IGFBP-3 may modulate the apoptotic response. These observations provide further evidence that endogenous IGFBP-3 plays a role in breast cancer cell responsiveness to DNA damaging therapy.     

Tumor targeting of vincristine by mBAFF-modified PEG liposomes in B lymphoma cells

Malignant hematologic diseases are highly malignant and refractory to conventional therapies. Ligand-mediated targeting of liposomal anticancer drugs to surface receptors expressed on malignant B cells can be an effective strategy for treating B-cell malignancies. BAFF plays an important role in the maintenance of normal B-cell development and homeostasis and the expression of its receptors is significantly increased in numerous B-cell malignancies. mBAFF (a soluble BAFF mutant with amino acid 217–224 being replaced by two glycine residues) may be used as a competitive inhibitor for BAFF to treat relevant malignant hematologic diseases. It may also hold promise as a novel ligand for targeted anticancer therapy. In this study, we show that liposomes that are sterically stabilized by PEG and surface decorated with mBAFF exhibited strong affinity and specificity to cultured human Raji B lymphoma cells. Vincristine formulated in the targeted liposomes showed significantly higher levels of cytotoxicity towards Raji cells than the nontargeted liposomal drug. Therapeutic experiments in SCID mice implanted with Raji cells showed significantly prolonged survival time with targeted liposomal vincristine compared to either free VCR or vincristine formulated in nontargeted liposomes. These studies suggest the potential of the mBAFF-modified liposomal drugs in targeted therapy of B-cell malignancies.