Department of Urology, The Affiliated Hospital of Qingdao University, Qingdao, 266003, People's Republic of China.
Department of Pharmaceutics, School of Pharmacy, Qingdao University, Qingdao, 266021, People's Republic of China.
Int J Nanomedicine. 2022 May 2;17:1987-2000. doi: 10.2147/IJN.S363168. eCollection 2022.
This study aimed to construct a delivery system based on L-arginine-modified calcium phosphate (CaP) to load eNOS plasmids (peNOS), which could amply nitric oxide (NO) to repair endothelial damage, promote angiogenic activities and alleviate inflammation.
pDNA-loaded CaP nanocomplex (CaP/pDNA) were prepared by co-precipitation method, subsequently modified by L-arginine. The gene transfection efficiency, pro-angiogenic and anti-inflammatory ability were investigated in vivo and in vitro. The therapeutic effect on ischemic hindlimb in vivo was assessed.
L-arginine modification augmented the transfection efficiency of CaP/peNOS to elevate the eNOS expression, and then served as NO substrate catalyzed by eNOS. At the same time, calcium ions produced by degradation of CaP carriers enhanced the activity of eNOS. In vitro experiments, the loading capability and transfection performance of R(L)-CaP were confirmed to be superior to that of CaP. Additionally, HUVECs treated with R(L)-CaP/peNOS showed the strongest NO release, cell migration, tube formation and the lowest inflammatory levels compared to the CaP/peNOS and R(D)-CaP/peNOS groups. We also demonstrated the advantages of R(L)-CaP/peNOS in increasing blood reperfusion in hindlimb ischemia mice by accelerating angiogenesis and reducing inflammation, which can be attributed to the highest eNOS-derived NO production.
The combination strategy of peNOS transfection, L-arginine supplement and calcium ions addition is a promising therapeutic approach for certain vascular diseases, based on the synergistic NO production.
本研究旨在构建基于精氨酸修饰的磷酸钙（CaP）的递送系统来负载 eNOS 质粒（peNOS），以充分提供一氧化氮（NO）来修复内皮损伤、促进血管生成活性和减轻炎症。
采用共沉淀法制备载 pDNA 的 CaP 纳米复合物（CaP/pDNA），随后用 L-精氨酸进行修饰。在体内和体外研究了基因转染效率、促血管生成和抗炎能力。评估了体内缺血后肢的治疗效果。
精氨酸修饰增强了 CaP/peNOS 的转染效率，从而提高了 eNOS 的表达，然后作为 eNOS 催化的 NO 底物。同时，CaP 载体降解产生的钙离子增强了 eNOS 的活性。体外实验证实，R(L)-CaP 的载药能力和转染性能优于 CaP。此外，与 CaP/peNOS 和 R(D)-CaP/peNOS 组相比，用 R(L)-CaP/peNOS 处理的 HUVECs 表现出最强的 NO 释放、细胞迁移、管形成和最低的炎症水平。我们还证明了 R(L)-CaP/peNOS 通过加速血管生成和减少炎症来增加后肢缺血小鼠的血液再灌注的优势，这归因于最高的 eNOS 衍生的 NO 产生。
基于协同产生的 NO，peNOS 转染、精氨酸补充和钙离子添加的联合策略是治疗某些血管疾病的有前途的方法。
