CAJ | 학술논문

目的制备新型负载阿霉素(Adriamycin)的海藻酸钙(CA)纳米微球(NPs),探讨阿霉素-CA-NPs对人肿瘤HeLa细胞生长的影响及其分子机制.方法采用离子交联方法,制备阿霉素-CA-NPs,检测其粒径和包封率,行体外释放实验,检测其缓释,并设置空白对照和CA-NPs组.细胞计数试剂盒(CCK-8)法检测阿霉素和阿霉素-CA-NPs对HeLa细胞增殖的影响,流式细胞术检测HeLa细胞凋亡,Westren blot检测HeLa细胞中半胱氨酰天冬氨酸特异性蛋白酶(Caspase)-3蛋白的表达.结果 (1)制备的阿霉素-CA-NPs的粒径为(157.3±8.57) nm,其缓释12 h的累积缓释率近70％.(2) CCK-8法检测各组抑制HeLa细胞增殖作用的结果表显示:阿霉素和阿霉素-CA-NPs的半数抑制剂量(IC50)分别为192.75和89.63 μg/L,阿霉素-CA-NPs的IC50不到阿霉素IC50的1/2,阿霉素-CA-NPs的作用效果更显著(P＜0.05).(3)流式细胞术检测结果显示,阿霉素组和阿霉素-CA-NPs组的细胞凋亡强度为(45.40±0.79)％和(60.67±1.03)％,与CA-NPs组比较差异有统计学意义(P＜0.05),且阿霉素-CA-NPs组的凋亡强度较阿霉素更显著(P＜0.05).(4)划痕实验结果显示:不同浓度的阿霉素-CA-NPs处理12h后HeLa细胞迁移能力显著降低,与空白组和CA-NPs组比较,其差异有统计学意义(P＜0.05),且呈剂量依赖性.(5)Western blot检测结果显示:与空白对照组比较,阿霉素组与阿霉素-CA-NPs组Caspase-3蛋白表达量均有明显升高(0.67 ±0.13和0.88±0.16,P＜0.05)],而阿霉素-CA-NPs处理组Caspase-3的表达量又显著高于阿霉素组(P＜0.05).结论具有缓释功能的阿霉素-CA-NPs,对HeLa细胞具有更加显著的抑制作用,其可能是通过促进Caspase-3蛋白的表达进而诱导肿瘤细胞凋亡来实现的.
목적 제비신형부재아매소(Adriamycin)적해조산개(CA)납미미구(NPs),탐토아매소-CA-NPs대인종류HeLa세포생장적영향급기분자궤제.방법 채용리자교련방법,제비아매소-CA-NPs,검측기립경화포봉솔,행체외석방실험,검측기완석,병설치공백대조화CA-NPs조.세포계수시제합(CCK-8)법검측아매소화아매소-CA-NPs대HeLa세포증식적영향,류식세포술검측HeLa세포조망,Westren blot검측HeLa세포중반광안선천동안산특이성단백매(Caspase)-3단백적표체.결과 (1)제비적아매소-CA-NPs적립경위(157.3±8.57) nm,기완석12 h적루적완석솔근70％.(2) CCK-8법검측각조억제HeLa세포증식작용적결과표현시:아매소화아매소-CA-NPs적반수억제제량(IC50)분별위192.75화89.63 μg/L,아매소-CA-NPs적IC50불도아매소IC50적1/2,아매소-CA-NPs적작용효과경현저(P＜0.05).(3)류식세포술검측결과현시,아매소조화아매소-CA-NPs조적세포조망강도위(45.40±0.79)％화(60.67±1.03)％,여CA-NPs조비교차이유통계학의의(P＜0.05),차아매소-CA-NPs조적조망강도교아매소경현저(P＜0.05).(4)화흔실험결과현시:불동농도적아매소-CA-NPs처리12h후HeLa세포천이능력현저강저,여공백조화CA-NPs조비교,기차이유통계학의의(P＜0.05),차정제량의뢰성.(5)Western blot검측결과현시:여공백대조조비교,아매소조여아매소-CA-NPs조Caspase-3단백표체량균유명현승고(0.67 ±0.13화0.88±0.16,P＜0.05)],이아매소-CA-NPs처리조Caspase-3적표체량우현저고우아매소조(P＜0.05).결론 구유완석공능적아매소-CA-NPs,대HeLa세포구유경가현저적억제작용,기가능시통과촉진Caspase-3단백적표체진이유도종류세포조망래실현적.
Objective To prepare novel adriamycin-calcium alginate (CA)-nanoparticles (NPs) , and to study the inhibitory effects of adriamycin-CA-NPs on proliferation of human cervical carcinoma HeLa cells.Methods The adriamycin-CA-NPs were prepared by the method of ionic crosslinking.The particle size and entrapment efficiency were detected, and the release of the drug was tested in vitro.The control group was set up.The effect of adriamycin-CA-NPs on the proliferation of HeLa cells was detected by cell counting kit-8 (CCK-8) assay, and apoptosis was detected by flow cytometry.The expression of cysteinyl aspartate-specific protease (Caspase)-3 protein in HeLa cells was detected by Westren blotting.Results (1) The average particle size of adriamycin-CA-NPs was (157.3 ± 8.57) nm, and the cumulative release rate at 12 h was nearly 70％.(2) CCK-8 assay showed that the proliferation of HeLa cells in adriamycin-CA-NPs and adriamycin group was significantly higher than that of CA-NPs group [50％ inhibitory dose (IC50) =192.75 and 89.63 μg/L], and the effect of adriamycin-CA-NPs group was more significant (P ＜ 0.05).(3) Flow cytometry showed that the apoptotic cell death intensity in adriamycin group and adriamycin-CA-NPs group was (45.40 ± 0.79) ％ and (60.67 ± 1.03) ％, which was significantly higher than that in CA-NPs group, and the effect of adriamycin-CA-NPs group was more significant (P ＜0.05).(4) The scratch test showed that the migration ability of HeLa cells treated with different concentrations of adriamycin-CA-NPs was significantly reduced in a concentration-dependent manner as compared with the control group (P ＜ 0.05).(5) Western blotting showed that the expression of Caspase-3 protein in adriamycin group (0.67 ± 0.13) and adriamycin-CA-NPs group (0.88 ±0.16) was significantly higher than that in control group (P ＜0.05), and the expression of Caspase-3 in adriamycin-CA-NPs group was significantly higher than that in adriamycin group (P ＜ 0.05).Conclusion The adriamycin-CA-NPs with high efficiency and sustained release properties can significantly inhibit HeLa cells, probably by promoting the expression of Caspase-3 protein in tumor cells and further inducing apoptosis.