Document Type : Original Article
.School of Chemical Engineering, College of Engineering, University of Tehran, Tehran, Iran;.Department of Biomedical Engineering, Research Center for New Technologies in Life Science Engineering, University of Tehran, T
4.Department of Surgery and Radiology, Faculty of Veterinary Medicine, University of Tehran, Tehran, Iran;5.Institute of Biomedical Research, University of Tehran, Tehran, Iran
.Amsterdam UMC-location VUMC and Academic Centre for Dentistry Amsterdam (ACTA), Vrije Universiteit Amsterdam, Department of Oral and Maxillofacial Surgery/Oral Pathology, Amsterdam Movement Sciences, Amsterdam, The Neth
6.Department of Oral Cell Biology, Academic Centre for Dentistry Amsterdam (ACTA), University of Amsterdam and Vrije Universiteit Amsterdam, Amsterdam Movement Sciences, Amsterdam, The Netherlands
This study investigated whether short stimulation (30 minutes) of human adipose stem cells (hASCs) with 1,25-dihydroxyvitamin D3(calcitriol or 1,25-(OH)2VitD3), fitting within the surgical procedure time frame, suffices to induce osteogenic differentiation, and compared this with continuous treatment with 1,25-(OH)2VitD3.
Materials and Methods
In this experimental study, hASCs were pretreated with/without 10 nM calcitriol for 30 minutes, seeded on biphasic calcium phosphate (BCP), and cultured for 3 weeks with/without 1,25-(OH)2VitD3. Cell attachment was determined 30 minutes after cell seeding. AlamarBlue assay, alkaline phosphatase (ALP) assay, ALP staining, real-time polymerase chain reaction (PCR), and protein assay were used to evaluate the effect of short calcitriol pretreatment on proliferation and osteogenic differentiation of hASCs up to 3 weeks.
Pretreatment with 1,25-(OH)2VitD3enhanced the attachment of hASCs to BCP by 1.5-fold compared to non- treated cells and increased the proliferation by 3.5-fold at day 14, and 2.6-fold at day 21. In contrast, continuous treatment increased the proliferation by 1.7-fold only at day 14. After 2 weeks, ALP activity was increased by 18.5-fold when hASCs were pretreated with 1,25-(OH)2VitD3for 30 minutes but increased only 2.6-fold when compared with its continuous counterpart. Moreover, after 14 days, pretreatment resulted in significant upregulation of the osteogenic markers RUNX2 and SPARC by 3.6-fold and 2.2-fold, respectively, while this was not observed upon continuous treatment. Finally, 30 minutes pretreatment of hASCs with 1,25-(OH)2VitD3increased VEGF189 expression, which may contribute to the process of angiogenesis.
This study is the first research showing that 30 minutes pretreatment of hASCs with 1,25-(OH)2VitD3, not only enhanced cell attachment to the scaffold at seeding time, but also promoted the proliferation and osteogenic differentiation of hASCs more strongly than continuous treatment, suggesting that short pre-treatment with 1,25-(OH)2VitD3is a promising approach for the regeneration of bones in a one-step surgical procedure.