Hu Haiyang, Sheng Qianglong, Yang Fan, Wu Xinyi, Zhang Youlai, Wu Shuling, Liu Yihu, Hu Ningyan, Fu Chenhong, Leong Jialin, Deng Rufei, Jiang Zhenyu, Chen Jiaxin, Wang Zhenxing, Chen Chunyuan, Chen Fei, Luo Yixuan, Zeng Yuanlin, Yu Yin, Xie Hui, Wang Gang, Zou Lijin
Jiangxi Provincial Key Laboratory of Trauma, Burn and Pain Medicine, Medical Center of Burn Plastic and Wound Repair, The First Affiliated Hospital, Jiangxi Medical College, Nanchang University, Nanchang, Jiangxi, China.
Department of Pharmacology, Innovative Institute of Basic Medical Sciences of Zhejiang University, Zhejiang, Hangzhou, China.
Int Wound J. 2025 May;22(5):e70143. doi: 10.1111/iwj.70143.
Efficient wound healing remains a formidable medical challenge in clinical practice, due to the prevalence of skin defects arising from diverse etiological factors. It is indisputable that epidermal growth factor (EGF) plays a pivotal role in wound repair. However, its clinical application through recombinant proteins encounters challenges, including a short half-life in vivo and high production costs. Addressing these limitations, recent advancements in chemically modified mRNA (cmRNA) technologies offer a promising alternative. This study explores the utilisation of cmRNA in a biocompatible citrate-saline formulation to encode EGF for therapeutic purposes, capitalising on the advantages of cmRNA's inherent stability and the formulation's compatibility with biological systems. CmRNA demonstrated high transfection efficiency in human immortalised keratinocyte (HaCaT) and normal human dermal fibroblasts (NHDF) cells (93.97% ± 1.25% and 90.37% ± 0.97%, respectively), resulting in efficient production of biologically active EGF protein. In vitro, EGF cmRNA significantly promoted HaCaT and NHDF cell cycle, proliferation and migration. In vivo, in vivo imaging system (IVIS) imaging of murine skin confirmed localised and sustained expression of Luciferase cmRNA, with signals detectable up to 11 days post-injection. Immunohistochemistry revealed protein expression in both epidermal and dermal layers as early as 1 h post-injection, peaking at 48 h, further corroborated by enzyme-linked immunosorbent assay (ELISA). In a full-thickness skin defect mouse model, EGF cmRNA significantly accelerated wound healing, with superior re-epithelialisation observed compared to controls by Day 6. Mitogen-activated protein kinase (MEK)/Extracellular signal-regulated kinase (ERK) and Ki67 mRNA expression levels were markedly increased, both in vitro and in vivo. By Day 14, histological and immunohistochemical analyses revealed that EGF cmRNA outperformed recombinant human EGF (rhEGF), as indicated by enhanced formation of hair follicles and cutaneous glands, better-organised collagen fibres, and a reduced collagen Type I/III ratio. No adverse effects were observed in major organs, confirming cmRNA's biosafety. These results highlight the therapeutic potential of EGF-encoding cmRNA as an effective and safe alternative for enhancing wound healing.
由于多种病因导致的皮肤缺损普遍存在，高效的伤口愈合在临床实践中仍然是一项艰巨的医学挑战。表皮生长因子（EGF）在伤口修复中发挥关键作用，这是无可争议的。然而，通过重组蛋白进行临床应用面临挑战，包括在体内半衰期短和生产成本高。为了解决这些限制，化学修饰mRNA（cmRNA）技术的最新进展提供了一种有前景的替代方案。本研究探索了在生物相容性柠檬酸盐-盐溶液制剂中利用cmRNA编码EGF用于治疗目的，利用cmRNA固有的稳定性优势以及该制剂与生物系统的兼容性。cmRNA在人永生化角质形成细胞（HaCaT）和正常人真皮成纤维细胞（NHDF）中表现出高转染效率（分别为93.97%±1.25%和90.37%±0.97%），从而高效产生具有生物活性的EGF蛋白。在体外，EGF cmRNA显著促进HaCaT和NHDF细胞周期、增殖和迁移。在体内，小鼠皮肤的体内成像系统（IVIS）成像证实了荧光素酶cmRNA的局部和持续表达，注射后长达11天可检测到信号。免疫组织化学显示，注射后1小时表皮和真皮层均有蛋白表达，48小时达到峰值，酶联免疫吸附测定（ELISA）进一步证实了这一点。在全层皮肤缺损小鼠模型中，EGF cmRNA显著加速伤口愈合，与对照组相比，在第6天时观察到更好的重新上皮化。丝裂原活化蛋白激酶（MEK）/细胞外信号调节激酶（ERK）和Ki67 mRNA表达水平在体外和体内均显著增加。到第14天，组织学和免疫组织化学分析表明，EGF cmRNA优于重组人EGF（rhEGF），表现为毛囊和皮肤腺体形成增强、胶原纤维排列更有序以及I型/III型胶原比例降低。主要器官未观察到不良反应，证实了cmRNA的生物安全性。这些结果突出了编码EGF的cmRNA作为增强伤口愈合的有效且安全的替代方案的治疗潜力。
