An HA-1-DC nucleic-acid vaccine was constructed to induce anti-leukemia effect after hematopoietic stem cell transplantation (HSCT). DCs were generated from HSCT donors in vitro, and its immunologic activity was assay...An HA-1-DC nucleic-acid vaccine was constructed to induce anti-leukemia effect after hematopoietic stem cell transplantation (HSCT). DCs were generated from HSCT donors in vitro, and its immunologic activity was assayed by using flow cytometry and mixed lymphocytes reaction. HA-1 gene was electroporated into the cultured DCs to construct a DC nucleic-acid vaccine. After transfection for 48 h, the expression of HA-1 protein could be detected by using Western blot. The DCs were cultured with syngenic lymphocytes to induce specific cytotoxic T lymphocytes (CTLs). The cytoxicity of the CTLs was detected by LDH assay. The results showed that The DCs derived from peripheral blood monocytes (PBMCs) expressed the phenotype of DCs, and were effective in stimulating proliferation of the allogenic lymphocytes. After electroporating for 48-h, HA-1 protein was detected by using Western blot. The cytotoxity of inducing CTLs was higher than the control group. It was suggested that minor histocompatibility antigen HA-1 could be considered as a target of immunotherapy against leukemia after HSCT.展开更多
核酶和脱氧核酶因其催化活性高度依赖金属离子,限制了其在核酸药物领域的应用,因此开发不依赖金属离子的修饰核酶成为研究热点。本研究测试了野生型Tgo DNA聚合酶同时识别两种糖环修饰的2'-氟代阿拉伯糖核苷酸和两种碱基修饰的脱氧...核酶和脱氧核酶因其催化活性高度依赖金属离子,限制了其在核酸药物领域的应用,因此开发不依赖金属离子的修饰核酶成为研究热点。本研究测试了野生型Tgo DNA聚合酶同时识别两种糖环修饰的2'-氟代阿拉伯糖核苷酸和两种碱基修饰的脱氧核糖核苷酸的活性和忠实性,构建了含有这4种修饰核苷酸的修饰文库,利用指数富集配体的系统进化(systematic evolution of ligands by exponential enrichment,SELEX)技术筛选出具有RNA切割活性的修饰核酶。与传统方法不同,本研究以修饰文库为模板直接进行PCR扩增,减少逆转录反应步骤,简化了筛选流程。本研究以全RNA底物为靶标进行筛选,通过对测序结果的同源性分析和二级结构预测,验证了富集序列中拷贝数前20序列的切割活性。结果表明,修饰核酶R4-11具有RNA切割活性,并鉴定了其选择性切割位于rC-rA之间的磷酸二酯键。本研究为修饰核酶的筛选提供了模式和平台,并为沉默疾病相关mRNA的靶向治疗提供了新型工具。展开更多
背景:miRNA作为重要的基因转录后调控因子,在骨质疏松症的发生和发展过程中发挥着关键作用。通过对miRNA调节骨质疏松症生物学的深入探究,其潜在的疗愈机制得以揭示,此领域已成为当前研究的热门焦点。目的:探讨miRNA在骨质疏松症发生中...背景:miRNA作为重要的基因转录后调控因子,在骨质疏松症的发生和发展过程中发挥着关键作用。通过对miRNA调节骨质疏松症生物学的深入探究,其潜在的疗愈机制得以揭示,此领域已成为当前研究的热门焦点。目的:探讨miRNA在骨质疏松症发生中的调控作用及其分子机制,并对以miRNA为靶点的骨质疏松症治疗策略所遭遇的关键问题及其解决方案进行综述。方法:以“miRNA,osteoporosis,angiogenesis,osteogenesis,genetherapy,drugdelivery”为英文检索词,以“miRNA,骨质疏松,基因治疗,核酸药物,递送载体”为中文检索词,检索PubMed、Web of Science数据库和中国知网2025年3月以前发表的文献。通过阅读文题和摘要进行初步筛选,排除相关性差、信息陈旧或观点重复且缺乏权威性的文献,最后纳入138篇文献进行综述。结果与结论:①miRNA是一种高效的、应用范围广泛、可精准调控细胞活动的非编码RNA,在调控骨细胞功能与骨血管生成方面展现出显著的优势,在骨质疏松症的治疗中具有潜在价值;②尽管基于miRNA的靶向治疗药物在其他疾病领域已进入临床前研究阶段,但在临床转化过程中,仍面临核酸体内稳定性不足及脱靶效应的挑战;③针对miRNA疗法所面临的挑战,研究者们提出了多种应对策略,包括精准定位miRNA的靶基因降低脱靶效应;化学修饰提高核酸药物在体内的稳定性;降低核酸生产成本推进研究;利用病毒载体、外泌体和各类生物材料优化核酸药物的递送途径;④科技的进步在提升核酸药物载体性能上持续创新,未来终将达到精确而高效的药物递送及靶向治疗效果。展开更多
文摘An HA-1-DC nucleic-acid vaccine was constructed to induce anti-leukemia effect after hematopoietic stem cell transplantation (HSCT). DCs were generated from HSCT donors in vitro, and its immunologic activity was assayed by using flow cytometry and mixed lymphocytes reaction. HA-1 gene was electroporated into the cultured DCs to construct a DC nucleic-acid vaccine. After transfection for 48 h, the expression of HA-1 protein could be detected by using Western blot. The DCs were cultured with syngenic lymphocytes to induce specific cytotoxic T lymphocytes (CTLs). The cytoxicity of the CTLs was detected by LDH assay. The results showed that The DCs derived from peripheral blood monocytes (PBMCs) expressed the phenotype of DCs, and were effective in stimulating proliferation of the allogenic lymphocytes. After electroporating for 48-h, HA-1 protein was detected by using Western blot. The cytotoxity of inducing CTLs was higher than the control group. It was suggested that minor histocompatibility antigen HA-1 could be considered as a target of immunotherapy against leukemia after HSCT.
文摘核酶和脱氧核酶因其催化活性高度依赖金属离子,限制了其在核酸药物领域的应用,因此开发不依赖金属离子的修饰核酶成为研究热点。本研究测试了野生型Tgo DNA聚合酶同时识别两种糖环修饰的2'-氟代阿拉伯糖核苷酸和两种碱基修饰的脱氧核糖核苷酸的活性和忠实性,构建了含有这4种修饰核苷酸的修饰文库,利用指数富集配体的系统进化(systematic evolution of ligands by exponential enrichment,SELEX)技术筛选出具有RNA切割活性的修饰核酶。与传统方法不同,本研究以修饰文库为模板直接进行PCR扩增,减少逆转录反应步骤,简化了筛选流程。本研究以全RNA底物为靶标进行筛选,通过对测序结果的同源性分析和二级结构预测,验证了富集序列中拷贝数前20序列的切割活性。结果表明,修饰核酶R4-11具有RNA切割活性,并鉴定了其选择性切割位于rC-rA之间的磷酸二酯键。本研究为修饰核酶的筛选提供了模式和平台,并为沉默疾病相关mRNA的靶向治疗提供了新型工具。
文摘背景:miRNA作为重要的基因转录后调控因子,在骨质疏松症的发生和发展过程中发挥着关键作用。通过对miRNA调节骨质疏松症生物学的深入探究,其潜在的疗愈机制得以揭示,此领域已成为当前研究的热门焦点。目的:探讨miRNA在骨质疏松症发生中的调控作用及其分子机制,并对以miRNA为靶点的骨质疏松症治疗策略所遭遇的关键问题及其解决方案进行综述。方法:以“miRNA,osteoporosis,angiogenesis,osteogenesis,genetherapy,drugdelivery”为英文检索词,以“miRNA,骨质疏松,基因治疗,核酸药物,递送载体”为中文检索词,检索PubMed、Web of Science数据库和中国知网2025年3月以前发表的文献。通过阅读文题和摘要进行初步筛选,排除相关性差、信息陈旧或观点重复且缺乏权威性的文献,最后纳入138篇文献进行综述。结果与结论:①miRNA是一种高效的、应用范围广泛、可精准调控细胞活动的非编码RNA,在调控骨细胞功能与骨血管生成方面展现出显著的优势,在骨质疏松症的治疗中具有潜在价值;②尽管基于miRNA的靶向治疗药物在其他疾病领域已进入临床前研究阶段,但在临床转化过程中,仍面临核酸体内稳定性不足及脱靶效应的挑战;③针对miRNA疗法所面临的挑战,研究者们提出了多种应对策略,包括精准定位miRNA的靶基因降低脱靶效应;化学修饰提高核酸药物在体内的稳定性;降低核酸生产成本推进研究;利用病毒载体、外泌体和各类生物材料优化核酸药物的递送途径;④科技的进步在提升核酸药物载体性能上持续创新,未来终将达到精确而高效的药物递送及靶向治疗效果。
