Recombinant human osteopontin expressed in Nicotiana benthamiana stimulates osteogenesis related genes in human periodontal ligament cells

Tissue engineering aims to utilise biologic mediators to facilitate tissue regeneration. Several recombinant proteins have potential to mediate induction of bone production, however, the high production cost of mammalian cell expression impedes patient access to such treatments. The aim of this stud...

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Bibliographic Details
Published in:	Scientific reports Vol. 7; no. 1; pp. 17358 - 9
Main Authors:	Rattanapisit, Kaewta, Abdulheem, Supaniga, Chaikeawkaew, Daneeya, Kubera, Anchanee, Mason, Hugh S., Ma, Julian K-C, Pavasant, Prasit, Phoolcharoen, Waranyoo
Format:	Journal Article
Language:	English
Published:	London Nature Publishing Group UK 11-12-2017 Nature Publishing Group
Subjects:	45/23 45/29 45/77 631/61/2035 631/61/447/2311 82/16 82/29 82/80 82/83 96/106 Affinity chromatography Bone growth Cell Differentiation Cell Proliferation Cells, Cultured Enzyme-linked immunosorbent assay Host plants Humanities and Social Sciences Humans Ligaments multidisciplinary Nicotiana - genetics Nicotiana - metabolism Nicotiana benthamiana Osteogenesis Osteopontin Osteopontin - genetics Osteopontin - metabolism Periodontal ligament Periodontal Ligament - cytology Periodontal Ligament - metabolism Recombinant Proteins - genetics Recombinant Proteins - metabolism Regeneration Science Science (multidisciplinary) Stem cells Structural analysis Tissue Engineering
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Summary:	Tissue engineering aims to utilise biologic mediators to facilitate tissue regeneration. Several recombinant proteins have potential to mediate induction of bone production, however, the high production cost of mammalian cell expression impedes patient access to such treatments. The aim of this study is to produce recombinant human osteopontin (hOPN) in plants for inducing dental bone regeneration. The expression host was Nicotiana benthamiana using a geminiviral vector for transient expression. OPN expression was confirmed by Western blot and ELISA, and OPN was purified using Ni affinity chromatography. Structural analysis indicated that plant-produced hOPN had a structure similar to commercial HEK cell-produced hOPN. Biological function of the plant-produced hOPN was also examined. Human periodontal ligament stem cells were seeded on an OPN-coated surface. The results indicated that cells could grow normally on plant-produced hOPN as compared to commercial HEK cell-produced hOPN determined by MTT assay. Interestingly, increased expression of osteogenic differentiation-related genes, including OSX , DMP1 , and Wnt3a , was observed by realtime PCR. These results show the potential of plant-produced OPN to induce osteogenic differentiation of stem cells from periodontal ligament in vitro , and suggest a therapeutic strategy for bone regeneration in the future.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23
ISSN:	2045-2322 2045-2322
DOI:	10.1038/s41598-017-17666-7