Short Pretreatment with Calcitriol Is Far Superior to Continuous Treatment in Stimulating Proliferation and Osteogenic Differentiation of Human Adipose Stem Cells

Mokhtari-Jafari, Fatemeh; Amoabediny, Ghassem; Dehghan, Mohammad Mehdi; Helder, Marco N.; Zandieh-Doulabi, Behrouz; Klein-Nulend, Jenneke

doi:10.22074/cellj.2020.6773

Short Pretreatment with Calcitriol Is Far Superior to Continuous Treatment in Stimulating Proliferation and Osteogenic Differentiation of Human Adipose Stem Cells

Document Type : Original Article

Authors

¹ .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

² 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

³ .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

⁴ 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

10.22074/cellj.2020.6773

Abstract

Objective
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.
Results
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.
Conclusion
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.

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