Oil body bound oleosin-rhFGF9 fusion protein expressed in safflower (Carthamus tinctorius L.) stimulates hair growth and wound healing in mice

Jingbo Cai,Haiyan Li,Xiuran Wang,Shanyong Yi,Xiaokun Li,Haishan Tian,Chao Jiang,Wenqing Li,Ruicheng Wen,Linbo Zhang

doi:10.1186/s12896-018-0433-2

Abstract

BackgroundFibroblast growth factor 9 (FGF9) is a heparin-binding growth factor, secreted by both mesothelial and epithelial cells, which participates in hair follicle regeneration, wound healing, and bone development. A suitable source of recombinant human FGF9 (rhFGF9) is needed for research into potential clinical applications. We present that expression of oleosin-rhFGF9 fusion protein in safflower (Carthamus tinctorius L.) seeds stimulates hair growth and wound healing.ResultsThe oleosin-rhFGF9 expressed in safflower seeds, in which it localizes to the surface of oil bodies. The expression of oleosin-rhFGF9 was confirmed by polyacrylamide gel electrophoresis and western blotting. According to BCA and Enzyme-linked immunosorbent assay (ELISA) assay, the results show that the expression level of oleosin-rhFGF9 was 0.14% of oil body protein. The oil body bound oleosin-rhFGF9 showed mitogenic activity towards NIH3T3 cells in a methylthiazolyldiphenyl-tetrazolium bromide (MTT) assay. The efficacy of oil body bound oleosin-rhFGF9 in promoting hair growth and wound healing was investigated in C57BL/6 mice. In a hair regeneration experiment, 50 μg/μl oil body bound oleosin-rhFGF9 was applied to the dorsal skin of mice in the resting phase of the hair growth cycle. After 15 days, thicker hair and increased number of new hairs were seen compared with controls. Furthermore, the number of new hairs was greater compared with rhFGF9-treated mice. The hair follicles of mice treated with oil body bound oleosin-rhFGF9 expressed β-catenin more abundantly. In a wound healing experiment, dorsal skin wounds were topically treated with 50 μg/μl oil body bound oleosin-rhFGF9. Wound healing was quicker compared with mice treated with rhFGF9 and controls, especially in the earlier stages of healing.ConclusionsThe oil body bound oleosin-rhFGF9 promotes both hair growth and wound healing. It appears to promote hair growth, at least in part, by up-regulating β-catenin expression. The potential of oil body bound oleosin-rhFGF9 as an external drug can treat the alopecia and wounds or use in further clinical application.

Highlights

Fibroblast growth factor 9 (FGF9) is a heparin-binding growth factor, secreted by both mesothelial and epithelial cells, which participates in hair follicle regeneration, wound healing, and bone development
Safflower transformation and transgenic plant regeneration via grafting The pOTBar-oleosin-recombinant human FGF9 (rhFGF9) plasmid was transformed into the safflower genome using the Agrobacterium tumefaciens-mediated infection method, and grafted transgenic safflower plants were obtained (Fig. 1)
Grafted T0 safflower plants were identified by polymerase chain reaction (PCR) analysis of genomic DNA to determine whether oleosin-rhFGF9 had integrated into the genome

Summary

Introduction

Fibroblast growth factor 9 (FGF9) is a heparin-binding growth factor, secreted by both mesothelial and epithelial cells, which participates in hair follicle regeneration, wound healing, and bone development. We present that expression of oleosin-rhFGF9 fusion protein in safflower (Carthamus tinctorius L.) seeds stimulates hair growth and wound healing. Fibroblast growth factor 9 (FGF9), known as Gliaactivating factor (GAF), was originally isolated from human glioma cells [1, 2]. There are four tyrosine kinase fibroblast growth factor receptors (FGFR1–4), which undergo alternative splicing to produce isoforms with high affinity, ligand-dependent responses to different FGFs. The specificity of FGF ligand receptor interaction is augmented by heparin, heparin sulfate, or other glycosaminoglycan chains to ensure stable binding and to activate diverse physiological functions. FGF9 can bind and activate FGFR3IIIc, FGFR3IIIb and FGFR2IIIc, but can activate other receptors with lower affinity, such as FGFR1IIIc and FGFR4; FGF9 has highest affinity to bind and activate FGFR3 but shows no activity toward FGFR1b or -2b [6, 7]

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Conclusion