Therapeutic effects of G90'-ES2 protein complex from regenerated Eisenia fetida on radiation-induced skin injury.

Radiation-Induced Skin Injury (RISI) is the major cutaneous side effect met by cancer patients who received radiotherapy. Disposed earthworm (Dilong) has been historically employed for treating various ailments in ancient China. Amputated-regeneration and high radiation-tolerant are two distinctive characteristics for earthworms. These features might due to their abundant biologically active substances (65 % composed of proteins), which make them attractive candidates for radioprotector development. This study aimed to investigate therapeutic mechanism of regenerated E. fetida protein extract (G90'-ES2) in mitigating RISI, and reveal therapeutic potential macromolecules by proteomic analysis. Protein salt precipitation method was firstly adopted to obtain G90'-ES2/G90-ES2 protein complex. Their radioprotective activities were evaluated in vitro by cell viability, migration, ROS accumulation, EdU proliferation, cell cycle and apoptosis assays. A combined radiogenic injury mice model was constructed to analyze radiotherapeutic function of G90'-ES2 in vivo. Its mechanism against RISI was illustrated by RNA-Seq analysis, qPCR, and HIF Luciferase Reporter Assay. Finally, a nanoElute UPLC-MS/MS method was used to elucidate therapeutic potential macromolecules in G90'-ES2. We discovered that G90'-ES2 displayed higher efficacy than G90-ES2 in mitigating RISI, mainly by activating radio-resistant HIF-1 pathway, redistributing cell cycle phases (S-phase preservation), accelerating re-epithelialization, and enhancing tissue angiogenesis. The significantly up-regulated proteins (including lumbrokinase, aquaporin, RAD51C, TCTP, and SCBP2) in G90'-ES2 might hold particular importance for its therapeutic effect against RISI. This investigation will broaden our understanding of the radiation-tolerant and regenerative capacities for earthworm; also provide medical candidates for the development of skin-specific radioprotector.