川芎嗪PEG-PE纳米胶束的体外评价、细胞摄取及抗心肌细胞凋亡研究

In Vitro Assessment, Cellular Uptake and Anti-cardiomyocyte Apoptosis of Tetramethylpyrazine PEG-PE Micelles

目的制备川芎嗪PEG-PE纳米胶束,并评价该纳米胶束的细胞摄取和抗心肌细胞凋亡效果。方法采用薄膜水化法制备川芎嗪PEG-PE纳米胶束,并进行表征。采用体外释药、细胞摄取和细胞凋亡试验对该载药系统进行评价。结果川芎嗪PEG-PE纳米胶束粒径为(15.8±0.9)nm,Zeta电势为-(20.5±0.4)mV,载药量为(5.7±0.3)%,包封率为(87.2±5.4)%。电镜结果表明川芎嗪PEG-PE纳米胶束呈形态规则的圆球型结构;采用芘测定法测定PEG-PE纳米胶束的临界胶束浓度约为5.3μg·mL-1;细胞摄取试验结果表明,PEG-PE纳米胶束可以增强药物的细胞摄取量,细胞外残留量减少;川芎嗪PEG-PE纳米胶束在10%胎牛血清DMEM培养基稳定性良好,采用异丙肾上腺素诱导心肌细胞凋亡,Hoechst染色提示凋亡心肌细胞出现了大量形态学改变,而川芎嗪PEG-PE纳米胶束可以明显减少凋亡细胞和促凋亡Caspase-3活性、抑制促凋亡蛋白Bax表达,提高抗凋亡蛋白Bcl-2表达,均显著优于川芎嗪(P<0.01)。结论川芎嗪PEG-PE纳米胶束具有粒径小,载药量高,释药缓慢等优势,可很大程度上提高川芎嗪的心肌细胞摄取量,增强药物的抗心肌细胞凋亡作用。

OBJECTIVE To prepare tetramethylpyrazine PEG-PE(TMP-PEG-PE)micelles,and to evaluate the cellular uptake and anti-cardiomyocyte apoptosis of the nanomicelle.METHODS TMP-PEG-PE micelles were prepared by thin film hydration method and characterized.The drug delivery system was evaluated by in vitro release,cell uptake and apoptosis assays.RESULTS The particle size of TMP-PEG-PE micelles was(15.8±0.9)nm,the Zeta potential was-(20.5±0.4)mV,the drug loading was(5.7±0.3)%and the encapsulation efficiency was(87.2±5.4)%.The results of electron microscopy showed that the TMP-PEG-PE micelles were in the shape of a regular spherical structure.The critical micelle concentration of PEG-PE micelles was determined by Pyrene assay to be about 5.3μg·mL-1.The results of cell uptake assay showed that PEG-PE nanomicelles could enhance the cellular uptake of the drug and reduce the extracellular residual amount.TMP-PEG-PE micelles had good stability in 10%fetal bovine serum DMEM medium.Isoproterenol was used to induce myocardial apoptosis,Hoechst staining indicated that there were a lot of morphological changes in apoptotic cardiomyocytes.TMP-PEG-PE micelles could significantly reduce the pro-apoptotic caspase-3 activity,inhibit the expression of the pro-apoptotic protein Bax,and increase the expression of anti-apoptotic protein Bcl-2,which were significantly better than the test results of TMP(P<0.01).CONCLUSION TMP-PEG-PE micelles have the advantages of small particle size,high drug loading and slow drug release,which can greatly improve the myocardial cell uptake of TMP and enhance its anti-cardiomyocyte apoptosis effect.