Research Institute of Orthopedics, The Affiliated Jiangnan Hospital of Zhejiang Chinese Medical University, Hangzhou, China.
Hangzhou Xiaoshan Hospital of Traditional Chinese Medicine, Hangzhou, China.
J Cell Mol Med. 2023 Aug;27(15):2136-2149. doi: 10.1111/jcmm.17800. Epub 2023 Jun 1.
The preparation of biodegradable scaffolds loaded with cells and cytokine is a feature of tissue-engineered skin. IPSCs-based tissue-engineered skin treatment for wound repair is worth exploring. Healthy human skin fibroblasts were collected and reprogrammed into iPSCs. After gene modification and induction, CK19 /Integrinβ1 /CD200 VEGF gene-modified iPS-HFSCs were obtained and identified by a combination of immunofluorescence and RT-qPCR. Astragalus polysaccharide-containing 3D printed degradable scaffolds were prepared and co-cultured with VEGF gene-modified iPS-HFSCs , and the biocompatibility and spatial structure of the tissue-engineered skin was analysed by cell counting kit-8 (CCK8) assay and scanning electron microscopy. Finally, the tissue-engineered skin was transplanted onto the dorsal trauma of nude mice, and the effect of tissue-engineered skin on the regenerative repair of total skin defects was evaluated by a combination of histology, immunohistochemistry, immunofluorescence, RT-qPCR, and in vivo three-dimensional reconstruction under two-photon microscopy. CK19 /Integrinβ1 /CD200 VEGF gene-modified iPS-HFSCs , close to the morphology and phenotype of human-derived hair follicle stem cells, were obtained. The surface of the prepared 3D printed degradable scaffold containing 200 μg/mL astragalus polysaccharide was enriched with honeycomb-like meshwork, which was more conducive to the proliferation of the resulting cells. After tissue-engineered skin transplantation, combined assays showed that it promoted early vascularization, collagen and hair follicle regeneration and accelerated wound repair. VEGF gene-modified iPS-HFSCs compounded with 3D printed degradable scaffolds containing 200 μg/mL astragalus polysaccharide can directly and indirectly participate in vascular, collagen, and hair follicle regeneration in the skin, achieving more complete structural and functional skin regenerative repair.
负载细胞和细胞因子的可生物降解支架的制备是组织工程皮肤的一个特征。基于 IPSC 的组织工程皮肤治疗用于伤口修复值得探索。收集健康人皮肤成纤维细胞并将其重编程为 iPSCs。经过基因修饰和诱导,获得 CK19/Integrinβ1/CD200 VEGF 基因修饰的 iPS-HFSCs,并通过免疫荧光和 RT-qPCR 联合鉴定。制备含黄芪多糖的 3D 打印可降解支架,并与 VEGF 基因修饰的 iPS-HFSCs 共培养,通过细胞计数试剂盒-8(CCK8)测定和扫描电子显微镜分析组织工程皮肤的生物相容性和空间结构。最后,将组织工程皮肤移植到裸鼠背部创伤处,通过组织学、免疫组织化学、免疫荧光、RT-qPCR 以及双光子显微镜下的体内三维重建相结合,评估组织工程皮肤对全层皮肤缺损再生修复的作用。获得了接近人源性毛囊干细胞形态和表型的 CK19/Integrinβ1/CD200 VEGF 基因修饰的 iPS-HFSCs。含有 200μg/mL 黄芪多糖的制备的 3D 打印可降解支架的表面富含蜂窝状网格,更有利于细胞的增殖。组织工程皮肤移植后,联合检测表明其促进了早期血管化、胶原和毛囊再生,加速了伤口修复。含 200μg/mL 黄芪多糖的 3D 打印可降解支架复合 VEGF 基因修饰的 iPS-HFSCs 可直接和间接参与皮肤中的血管、胶原和毛囊再生,实现更完整的结构和功能皮肤再生修复。
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