Shanghai Jiao Tong University Affiliated Sixth People's Hospital, School of Biomedical Engineering, Shanghai Jiao Tong University, Shanghai 200030, China.
Department of Orthopaedics, Shanghai Key Laboratory for Prevention and Treatment of Bone and Joint Diseases, Shanghai Institute of Traumatology and Orthopaedics, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China.
J Control Release. 2023 Nov;363:585-596. doi: 10.1016/j.jconrel.2023.10.003. Epub 2023 Oct 11.
Delivering large therapeutic molecules via the blood-brain barrier to treat ischemic stroke remains challenging. NR2B9c is a potent neuroprotective peptide but it's safe and targeted delivery to the brain requires an efficient, natural, and non-immunogenic delivery technique. Small extracellular vesicles (sEVs) have shown great potential as a non-immunogenic, natural cargo delivery system; however, tailoring of its inefficient brain targeting is desired. Here, we coupled rabies virus glycoprotein 29 with sEVs surface via bio-orthogonal click chemistry reactions, followed by loading of NR2B9c, ultimately generating stroke-specific therapeutic COCKTAIL (sEVs-COCKTAIL). Primary neurons and Neuro-2a cells were cultured for in vitro and transient middle cerebral artery occlusion model was used for in vivo studies to evaluate neuron targeting and anti-ischemic stroke potential of the sEVs-COCKTAIL. Bio-clickable sEVs were selectively taken up by neurons but not glial cells. In the in vitro ischemic stroke model of oxygen-glucose deprivation, the sEVs-COCKTAIL exhibited remarkable potential against reactive oxygen species and cellular apoptosis. In vivo studies further demonstrated the brain targeting and increased half-life of bio-clickable sEVs, delivering NR2B9c to the ischemic brain and reducing stroke injury. Treatment with the sEVs-COCKTAIL significantly increased behavioral recovery and reduced neuronal apoptosis after transient middle cerebral artery occlusion. NR2B9c was delivered to neurons binding to post-synaptic density protein-95, inhibiting N-methyl-d-Aspartate receptor-mediated over production of oxidative stress and mitigating protein B-cell lymphoma 2 and P38 proteins expression. Our results provide an efficient and biocompatible approach to a targeted delivery system, which is a promising modality for stroke therapy.
通过血脑屏障递送大治疗分子以治疗缺血性中风仍然具有挑战性。NR2B9c 是一种有效的神经保护肽，但要将其安全且靶向递送至大脑，需要一种有效的、天然的、非免疫原性的递药技术。小细胞外囊泡（sEVs）已显示出作为非免疫原性、天然货物递药系统的巨大潜力；然而，需要对其向大脑的低效靶向作用进行修饰。在这里，我们通过生物正交点击化学反应将狂犬病毒糖蛋白 29 与 sEVs 表面偶联，随后装载 NR2B9c，最终生成针对中风的特异性治疗鸡尾酒（sEVs-COCKTAIL）。原代神经元和 Neuro-2a 细胞进行体外培养，短暂性大脑中动脉闭塞模型用于体内研究，以评估 sEVs-COCKTAIL 对神经元的靶向作用和抗缺血性中风的潜力。生物可点击的 sEVs 被神经元选择性摄取，但不被神经胶质细胞摄取。在体外氧葡萄糖剥夺缺血性中风模型中，sEVs-COCKTAIL 表现出对抗活性氧和细胞凋亡的显著潜力。体内研究进一步证明了生物可点击的 sEVs 的脑靶向性和半衰期延长，将 NR2B9c 递送至缺血性大脑，并减少中风损伤。sEVs-COCKTAIL 治疗可显著改善短暂性大脑中动脉闭塞后的行为恢复并减少神经元凋亡。NR2B9c 通过与突触后密度蛋白-95 结合递送至神经元，抑制 N-甲基-D-天冬氨酸受体介导的氧化应激过度产生，并减轻蛋白 B 细胞淋巴瘤 2 和 P38 蛋白的表达。我们的结果提供了一种高效且生物相容的靶向递药系统方法，为中风治疗提供了一种很有前途的治疗方式。
