Development of slow release reagent of NFkB decoy oligodeoxynucleotides for the treatment of rheumatic arthritis

我们试图延长NFkB oligodeoxynudeotid.es的释放而不降解。首先，我们测试了使用去端肽胶原将FITC标记的寡核苷酸引入培养细胞的效率。转移至HeLa、BHK-21和人舌癌细胞系SAS后48小时未检测到荧光。然后，我们开发了HVJ（日本血凝病毒;仙台病毒）包膜载体，该载体可以引入合成的寡核苷酸、蛋白质、肽和化学药物以及基因。通过温和去污剂处理和离心将FITC标记的寡核苷酸掺入HVJ包膜载体中。将上述培养细胞与HVJ包膜载体温育10分钟后，在几乎所有细胞核中观察到荧光。当使用HVJ包膜载体将NFkB诱饵寡核苷酸引入癌细胞如SAS和HeLa细胞中时，通过抑制NFkB诱导的凋亡而增强辐射后的细胞凋亡。 关于我们 基因表达。当使用HVJ包膜载体将FITC标记的NFkB诱饵寡核苷酸引入食蟹猴的关节间隙时，在滑膜和关节软骨处均检测到荧光。接下来，用各种聚合物包埋HVJ包膜载体。首先，在没有聚合物的情况下，HVJ包膜载体通过静脉内注射到达脾脏，并且在小鼠脾脏的边缘区检测到FITC-寡核苷酸。然后用硫酸鱼精蛋白修饰含荧光素酶基因的载体，并将复合物注射到小鼠尾静脉。仅在肺中检测到基因表达，而在脾中未检测到。此外，当HVJ包膜载体与肝素混合直接注射到肌肉或脑中时，基因表达比没有肝素的增加约5倍。聚合物对载体缓释的影响正在研究中，但我们已经得到了初步的数据表明，阳离子聚合物可以增强HVJ包膜载体的缓释。少

We attempted to prolong release of NFkB oligodeoxynudeotid.es without degradation. First, we tested the efficiency of introduction of FITC-labeled oligonucleotides into cultured cells using atelocollagen. No fluorescence was detected at 48 hours after the transfer to HeLa, BHK-21 and human tongue cancer cell line SAS. Then, we developed HVJ (hemagglutinating virus of Japan ; Sendai virus) envelope vector which can introduce synthetic oligonucleotides, proteins, peptides and chemical drugs as well as genes. FITC-labeled oligonucleotides were incorporated into HVJ envelope vector by the treatment of mild detergent and centrifugation. Ten minutes after incubation of those cultured cells described above with the HVJ envelope vector, fluorescence was observed in almost all the nuclei of the cells. When NFkB decoy oligonucleotides were introduced into cancer cells such as SAS and HeLa cells using HVJ envelope vector, apoptosis after irradiation was enhanced by the suppression of NFkB-induced … More gene expression. When FITC-labeled NFkB decoy oligonucleotides were introduced into the joint space of Cynomolgus monkeys using HVJ envelope vector, fluorescence was detected at both synovium and articular cartilage. Next, HVJ envelope vector was embedded with various polymers. First, without polymer, HVJ envelope vector reaches spleen by intravenous injection and FITC-oligonucleotides were detected at the marginal zone of mouse spleen. Then, the vector containing luciferase gene was decorated with protamine suifate and the complex was injected into tail vein of mouse. Gene expression was detected exclusively in lung, not in spleen. Furthermore, when HVJ envelope vector mixed with heparin was injected into muscle or brain directly, gene expression was approximately 5 fold increased than that without heparin. The effect of polymers on slow release of the vector is being investigated, but we have got preliminary data suggesting that cationic polymers may enhance slow release of HVJ envelope vector. Less