Key Laboratory for Liquid-Solid Structural Evolution and Processing of Materials, Ministry of Education, School of Materials Science and Engineering, Shandong University, Jinan, Shandong, China.
Key Laboratory for Liquid-Solid Structural Evolution and Processing of Materials, Ministry of Education, School of Materials Science and Engineering, Shandong University, Jinan, Shandong, China; Shenzhen Research Institute of Shandong University, Shenzhen, Guangdong, China.
Biomater Adv. 2022 Jun;137:212822. doi: 10.1016/j.bioadv.2022.212822. Epub 2022 Apr 25.
The Boston Keratoprosthesis type I (B-KPro) is widely used in the world, but the lack of donor corneas limits its application. This study aims to prepare the acellular porcine cornea (APC) crosslinked with ultraviolet A (UVA)/riboflavin instead of donor corneas as the scaffold for B-KPro. Decellularization of freeze-thaw combined with biological enzymes resulted in approximately 5 ng/mg DNA residue, the a-Gal removal rate of 99%, and glycosaminoglycans retention at a high level of 46.66 ± 2.59 mg/mg. UVA/ riboflavin cross-linking was adopted to induce the formation of new chemical bonds between adjacent collagen chains in the corneal stroma to improve the mechanical properties and resistance to enzymatic hydrolysis. Through comprehensive analysis of the biomechanics, enzyme degradation, immunogenicity and histological structure of the APC crosslinked at different times, CL3 (irradiation conditions, 365 nm, 3 mW/cm, 80 min, both sides) was selected and transplanted into the rabbit cornea model through interlamellar keratoplasty and penetrating keratoplasty as the scaffold of the B-KPro. Compared with the native porcine cornea (NPC) and APC, the experiment of interlamellar pocket indicated that the structure of CL3 was homogeneous without degradation and vascularization in vivo at 12 weeks after surgery. Simultaneously, the results of transplantation of B-KPro showed complete epithelialization of CL3 within 1 week, and neovascularization of the cornea indicated rejection but could be controlled with immunosuppressants. At 3 months postoperatively, the lens of B-KPro remained transparent, and the structure of CL3 was compact and uniform, accompanied by the migration and proliferation of a large number of stromal cells without degradation, suggesting the CL3 could be a promising corneal substitute.
波士顿角膜型人工角膜 I 型（B-KPro）在世界范围内广泛应用，但供体角膜的缺乏限制了其应用。本研究旨在制备去细胞猪角膜（APC），并用紫外线 A（UVA）/核黄素交联，代替供体角膜作为 B-KPro 的支架。通过冻融结合生物酶法处理，可使去细胞后角膜的 DNA 残留量约为 5ng/mg，α-Gal 去除率达到 99%，糖胺聚糖保留水平高达 46.66±2.59mg/mg。采用 UVA/核黄素交联诱导角膜基质中相邻胶原链之间形成新的化学键，以提高机械性能和抗酶解能力。通过对不同交联时间的 APC 的生物力学、酶降解、免疫原性和组织学结构进行综合分析，选择交联 3 次（辐照条件为 365nm、3mW/cm2、80min，双面）的 CL3 作为 B-KPro 的支架进行兔角膜模型的层间角膜移植和穿透性角膜移植。与天然猪角膜（NPC）和 APC 相比，层间袋实验表明，在手术后 12 周时，CL3 的结构均匀，无降解和血管化。同时，B-KPro 移植实验表明，CL3 在 1 周内完全上皮化，角膜新生血管化表明排斥反应，但可通过免疫抑制剂控制。术后 3 个月，B-KPro 的晶状体仍保持透明，CL3 的结构紧密均匀，伴有大量基质细胞的迁移和增殖，无降解，表明 CL3 可能是一种有前途的角膜替代物。
