short pretreatment with calcitriol is far superior to continuous treatment in stimulating proliferation and osteogenic differentiation of human adipose stem cells

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)2vitd3 enhanced the attachment of hascs to bcp by 1.5-fold compared to nontreated 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)2vitd3 for 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)2vitd3 increased 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)2vitd3 is a promising approach for the regeneration of bones in a one-step surgical procedure.