Li Kanglu, Lin Hui, Yu Yihan, Liu Yiran, Yang Wenbo, Chen Songfeng, Xu Li, Huang Wei, Wang Hong, Meng Chunqing, Shao Zengwu, Wei Yulong, Zhao Lei, Peng Yizhong
Department of Orthopaedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China.
Department of Orthopedics, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, China.
Acta Biomater. 2025 May 1;197:357-373. doi: 10.1016/j.actbio.2025.02.060. Epub 2025 Mar 13.
Traditional methods of plasmid delivery, including viral vectors, lipofection, and electroporation, are widely used for gene editing but have limitations, such as cellular toxicity, limited transfection efficiency in primary cells, and nonspecific side effects. Here, we report the development of nucleus pulposus cell (NPC)-mimicking nanoparticles (HIF1A@NNP) with an NPC membrane as the shell and pcDNA3.1-rHIF1A encapsulated in the core via extrusion. HIF1A@NNP exhibited a protein expression pattern similar to that of the NPC membrane and displayed a typical vesicle profile. Compared to liposomes and lentiviruses, HIF1A@NNP overexpressed HIF1A in NPCs while improving cell viability. HIF1A@NNP was more readily internalized by NPCs than by other cell types, with fewer effects on vascularization, nerve growth, and macrophage polarization than HIF1A overexpression using lipo3000. HIF1A@NNP reduced the apoptotic rate and inhibited the senescent phenotype, as evidenced by reduced DNA damage, lower levels of senescence-related proteins, and fewer SA-β-Gal-positive cells. HIF1A@NNP induced a senescence-associated secretory phenotype (SASP), which enhanced macrophage migration and M1 polarization. Additionally, HIF1A@NNP activated autophagy in NPCs. In summary, HIF1A@NNP demonstrated satisfactory biocompatibility, alleviated the SASP, and inhibited SASP-mediated macrophage recruitment and inflammatory polarization, leading to reduced disc degeneration and providing a promising strategy for combating intervertebral disc degeneration. STATEMENT OF SIGNIFICANCE: Conventional plasmid delivery methods like viral vectors, lipofection, and electroporation struggle with cellular toxicity and inefficiency in primary cells. Non-cell-specific HIF1A activation via these methods may exacerbate inflammation and pain, as HIF1A drives angiogenesis and dendritic ingrowth into the disc. Thus, a cell-specific delivery strategy could circumvent such adverse effects. Our study introduces HIF1A@NNP, a nanoparticle mimicking nucleus pulposus cells (NPCs), with an NPC membrane shell encapsulating pcDNA3.1+-rHIF1A. It preferentially targets NPCs, achieving superior HIF1A overexpression and cell viability compared to liposomes and lentiviruses. This represents a highly promising and potentially transformative approach against intervertebral disc degeneration.
传统的质粒递送方法，包括病毒载体、脂质体转染和电穿孔，广泛用于基因编辑，但存在局限性，如细胞毒性、原代细胞中转染效率有限以及非特异性副作用。在此，我们报告了一种模仿髓核细胞（NPC）的纳米颗粒（HIF1A@NNP）的研发，其以NPC膜为外壳，通过挤压将pcDNA3.1-rHIF1A包裹在核心。HIF1A@NNP表现出与NPC膜相似的蛋白质表达模式，并呈现典型的囊泡形态。与脂质体和慢病毒相比，HIF1A@NNP在NPC中过表达HIF1A，同时提高细胞活力。HIF1A@NNP比其他细胞类型更容易被NPC内化，与使用Lipofectamine 3000过表达HIF1A相比，对血管生成、神经生长和巨噬细胞极化的影响更小。HIF1A@NNP降低了凋亡率并抑制了衰老表型，DNA损伤减少、衰老相关蛋白水平降低以及SA-β-Gal阳性细胞减少证明了这一点。HIF1A@NNP诱导了衰老相关分泌表型（SASP），增强了巨噬细胞迁移和M1极化。此外，HIF1A@NNP激活了NPC中的自噬。总之，HIF1A@NNP表现出令人满意的生物相容性，减轻了SASP，并抑制了SASP介导的巨噬细胞募集和炎症极化，从而减少椎间盘退变，为对抗椎间盘退变提供了一种有前景的策略。重要性声明：病毒载体、脂质体转染和电穿孔等传统质粒递送方法存在细胞毒性以及在原代细胞中效率低下的问题。通过这些方法进行的非细胞特异性HIF1A激活可能会加剧炎症和疼痛，因为HIF1A会驱动血管生成和树突长入椎间盘。因此，一种细胞特异性递送策略可以规避此类不良反应。我们的研究引入了HIF1A@NNP，一种模仿髓核细胞（NPC）的纳米颗粒，其具有包裹pcDNA3.1+-rHIF1A的NPC膜外壳。与脂质体和慢病毒相比，它优先靶向NPC，实现了更高的HIF1A过表达和细胞活力。这代表了一种对抗椎间盘退变的极具前景且可能具有变革性的方法。
