The Key to Spinal Cord Recovery: Harnessing p21 Inhibition to Boost Neural Stem/Progenitor Cell Proliferation

Spinal cord injury (SCI) is a devastating traumatic condition
of
the central nervous system (CNS), usually resulting in irreversible
motor and sensory deficits that severely compromise patients’
quality of life. Harnessing the untapped potential of endogenous neural
stem/progenitor cells (NSPCs) may yield revolutionary therapeutic
techniques for overcoming the limited NSPC proliferative capacity
following SCI in adults. Single-cell sequencing results demonstrated
that the limited proliferative capacity of NSPCs after SCI is associated
with the upregulation of the p21. Herein, we developed a cationic
liposome-based delivery system encapsulating p21 small interfering
RNA (P21siRNA@LP) to
enhance NSPC proliferation following SCI. P21siRNA@LP significantly increased the primary
NSPC proliferation rate (145.4% on day 1 and 144.7% on day 3, respectively)
while maintaining differentiation capacity in vitro. Transcriptomic and functional characterization showed that P21siRNA@LP modulated the
expression of cyclin-dependent kinases in NSPCs, enhancing cell cycle
pathways (enrichment score: 0.6691) and proliferation, with extracellular
matrix reorganization (col1a1, col5a1) and gliogenesis (olig1/2) identified
as key regulation pathways. Gelatin hydrogels incorporating P21siRNA@LP promoted dense tissue
cable formation in T9 SCI rats, facilitating NSPC migration and proliferation
at lesion sites, which accelerated locomotor function recovery. These
findings emphasize cell cycle manipulation as a promising method to
spinal cord regeneration, providing a basis for future therapeutic
advances in CNS disorders.

脊髓损伤（Spinal cord injury, SCI）是一类极具破坏性的中枢神经系统（central nervous system, CNS）创伤性疾病，常引发不可逆的运动与感觉功能障碍，严重损害患者的生活质量。激活内源性神经干细胞/祖细胞（endogenous neural stem/progenitor cells, NSPCs）的未发掘潜能，有望催生革命性治疗手段，以克服成年个体脊髓损伤后NSPC增殖能力受限的瓶颈。单细胞测序结果显示，脊髓损伤后NSPC增殖能力不足与p21基因的上调表达密切相关。本研究构建了一种包载p21小干扰RNA（small interfering RNA, siRNA）的阳离子脂质体递送系统（P21siRNA@LP），用于增强脊髓损伤后NSPC的增殖活性。体外实验证实，P21siRNA@LP可显著提升NSPC的初始增殖率（第1天达145.4%，第3天达144.7%），同时维持其体外分化潜能。转录组学与功能表征分析表明，P21siRNA@LP可调控NSPC内细胞周期蛋白依赖性激酶的表达，增强细胞周期通路（富集得分：0.6691）与细胞增殖能力，并确定细胞外基质重塑（col1a1、col5a1）与神经胶质生成（olig1/2）为关键调控通路。将P21siRNA@LP载入明胶水凝胶后，可促进T9节段脊髓损伤大鼠形成致密的组织索带，助力损伤部位的NSPC迁移与增殖，进而加速运动功能恢复。本研究结果证实，细胞周期调控是一种极具前景的脊髓再生治疗策略，为中枢神经系统疾病的后续治疗进展提供了理论依据。