Conclusions
iRGD MTX ICG ELIP has a suitable particle size and can effectively target HUVECs and the joints with rheumatiod arthritis. With a good drug entrapment efficiency and acoustic responsiveness, the drug loaded liposome shows enhanced inhibitory effect on MH7A cells combined with ultrasound in vitro, suggesting its potential in the treatment of rheumatoid arthritis. 目的: 制备集治疗与成像为一体的载甲氨喋呤（MTX）和吲哚菁绿（ICG）iRGD靶向载药声学脂质体（iRGD-MTX-ICGELIP），观察其靶向性及联合低频超声体外抑制滑膜细胞（MH7A）增殖的效果。 方法: 采用薄膜水化法和冷冻冻干法制备iRGD-MTX-ICG-ELIP，检测其一般特性及声学响应性，通过细胞摄取实验验证iRGD-MTX-ICG-ELIP的体外靶向结合性能，构建类风湿关节炎小鼠模型，通过靶组织的药物荧光强度验证iRGD-MTX-ICG-ELIP的体内靶向性；CCK8法检测iRGD-MTXICG-ELIP联合超声体外抑制MH7A增殖的效果。 结果: 制备的iRGD-MTX-ICG-ELIP粒径为134.4±17.6 nm，电位为-10.07±4.28 mv，iRGD-MTX-ICG-ELIP中MTX和ICG的包封率分别为（62.56±0.77）%和（95.13±0.82）%；其联合低频超声控释药物发现，随超声作用强度增加和作用时间的延长，MTX与ICG释放均增多。细胞摄取实验表明血管内皮细胞HUVECs对iRGDMTX-ICG-ELIP的摄取效率比对MTX-ICG-ELIP摄取效率高1.89倍，差异有统计学意义（P < 0.05），活体成像实验显示iRGDMTX-ICG-ELIP在RA发病关节的荧光强度明显强于非靶向组；CCK8检测结果显示，iRGD-MTX-ICG-ELIP联合超声组的MH7A存活率为（32.49±3.04）%，与未联合超声组比较差异有统计学意义（P < 0.05）。 结论: 本研究制备的iRGD-MTX-ICGELIP粒径小、均一性好，对HUVECs和RA病变关节组织具有较好的靶向性。较佳的药物包封率和声学响应性增强了iRGDMTX-ICG-ELIP联合超声抑制MH7A的增殖作用，为后续更精准有效的治疗类风湿关节炎提供前期基础。
Methods
iRGD MTX ICG ELIP was prepared by the thin film rehydration and freeze-lyophilization method and its general characteristics and acoustic responsiveness were assessed. The targeting effect of the prepared liposome was observed by assessing its cell uptake in vitro. In a mouse model of rheumatiod arthritis, the targeting effect of the prepared liposome was determined by detecting the fluorescence intensity of the drug in arthrosis. The inhibitory effect of iRGD MTX ICG ELIP combined with ultrasound on synovial MH7A cells in vitro were investigated using CCK8 test.
Objective
To prepare internalized RGD (iRGD) modified echogenic liposomes containing methotrexate (MTX) and indocyanine green (ICG) (iRGD MTX ICG ELIP) and evaluate its targeting efficiency and inhibitory effect combined with ultrasound on synovial cells.
Results
The average diameter and zeta potential of iRGD MTX ICG ELIP was 134.4∓17.61 nm and 10.07∓4.28 mV, and the entrapment efficiency of MTX and ICG was (62.56∓0.77)% and (95.13∓0.82)%, respectively. With ultrasound exposure, the release of MTX and ICG from iRGD MTX ICG ELIP increased with the ultrasound intensity and with the exposure time. In HUVECs, the uptake efficiency of iRGD MTX ICG ELIP was 1.89 times higher than that of non targeted MTX ICG ELIP (P<0.05). In vivo imaging of mouse joint with rheumatiod arthritis showed that the fluorescence intensity of iRGD MTX ICG ELIP was significantly stronger than that of the non targeted liposome. CCK8 assay showed that iRGD MTX ICG ELIP combined with ultrasound resulted in a survival rate of MH7A cells of (32.49∓3.04)%, significantly lower than the rate of cells treated with iRGD MTX ICG ELIP but without ultrasound (P<0.05). Conclusions: iRGD MTX ICG ELIP has a suitable particle size and can effectively target HUVECs and the joints with rheumatiod arthritis. With a good drug entrapment efficiency and acoustic responsiveness, the drug loaded liposome shows enhanced inhibitory effect on MH7A cells combined with ultrasound in vitro, suggesting its potential in the treatment of rheumatoid arthritis. 目的: 制备集治疗与成像为一体的载甲氨喋呤（MTX）和吲哚菁绿（ICG）iRGD靶向载药声学脂质体（iRGD-MTX-ICGELIP），观察其靶向性及联合低频超声体外抑制滑膜细胞（MH7A）增殖的效果。 方法: 采用薄膜水化法和冷冻冻干法制备iRGD-MTX-ICG-ELIP，检测其一般特性及声学响应性，通过细胞摄取实验验证iRGD-MTX-ICG-ELIP的体外靶向结合性能，构建类风湿关节炎小鼠模型，通过靶组织的药物荧光强度验证iRGD-MTX-ICG-ELIP的体内靶向性；CCK8法检测iRGD-MTXICG-ELIP联合超声体外抑制MH7A增殖的效果。 结果: 制备的iRGD-MTX-ICG-ELIP粒径为134.4±17.6 nm，电位为-10.07±4.28 mv，iRGD-MTX-ICG-ELIP中MTX和ICG的包封率分别为（62.56±0.77）%和（95.13±0.82）%；其联合低频超声控释药物发现，随超声作用强度增加和作用时间的延长，MTX与ICG释放均增多。细胞摄取实验表明血管内皮细胞HUVECs对iRGDMTX-ICG-ELIP的摄取效率比对MTX-ICG-ELIP摄取效率高1.89倍，差异有统计学意义（P < 0.05），活体成像实验显示iRGDMTX-ICG-ELIP在RA发病关节的荧光强度明显强于非靶向组；CCK8检测结果显示，iRGD-MTX-ICG-ELIP联合超声组的MH7A存活率为（32.49±3.04）%，与未联合超声组比较差异有统计学意义（P < 0.05）。 结论: 本研究制备的iRGD-MTX-ICGELIP粒径小、均一性好，对HUVECs和RA病变关节组织具有较好的靶向性。较佳的药物包封率和声学响应性增强了iRGDMTX-ICG-ELIP联合超声抑制MH7A的增殖作用，为后续更精准有效的治疗类风湿关节炎提供前期基础。