Evaluation effects of ascorbic acid leads to activate and induce osteogenic protein marker expression: in silico and in-vitro study

Abstract

Background: Mesenchymal stem cells (MSCs) are multipotent cells that can differentiate into several cell types including chondrocytes and osteocytes. The process of differentiation into such tissues is aided by transcription factors and collagen gene markers. Collagen biosynthesis is influenced by the presence of vascular endothelial growth factor (VEGF) and ascorbic acid cofactors of the prolyl hydroxylase 2 (PHD2) enzyme found in human cell receptors. The focus of this study is evaluation of how in silico methods can elucidate the effect of ascorbic acid on PHD2 and its expression. This study aimed to evaluate the role of ascorbic acid in its interaction with PHD2 (in silico) and the expression of type 1 collagen (COL1), type 2 collagen (COL2), and VEGF. Methods: The technique of in silico molecular docking consisted of three steps: preparation of PHD2 receptors using AutoDock tools, preparation of ligand ascorbic acid using AutoDock tools, and molecular docking using AutoDock Vina. The results of in silico molecular docking were visualized using py-MOL and BIOVIA discovery studio. Furthermore, in vitro MSCs from adipose tissue were isolated and characterized based on protein marker expression. The effect of ascorbic acid supplementation (50 and 100 µg/mL) on stem cell differentiation was evaluated based on the mRNA ratio of VEGF/COL1, VEGF/COL2, and COL2/COL1. Results: Molecular docking of ligand ascorbic acid with PHD2 resulted in nine poses. The third pose was chosen because it had an affinity of -5.7 kcal/mol and a root-mean-square deviation (RMSD) of 1.96, indicating a valid docking result. Meanwhile, the 2D conformer showed van der Waals bonds on Tyr303 and Try329 residues. MSCs were differentiated into adipocytes, chondrocytes, and osteocytes. Regarding the marker ratio of gene expression, at 50 µg/ml ascorbic acid there was an increase in the VEGF/COL1 and VEGF/COL2 ratios and a decrease in the COL2/COL1 ratio compared with the control; at 100 µg/ml ascorbic acid there was a decrease in both ratios (VEGF/COL1 and COL2/COL1), and the VEGF/COL2 ratio continued to increase. Conclusions: In silico molecular docking between PHD2 and ascorbic acid, which was studied via bioinformatics, resulted in a stable interaction with good ligand position. Further in vitro studies would support the influence of ascorbic acid on transcription factors, collagen gene markers, and the differentiation of MSCs.

The Effectivity of Honey Bee Towards Mesenchymal Stem Cells Proliferation: A Systematic Review

“…This potential effect is similar to platelet-rich plasma supplementation in adipose-derived stem cells (ADSCs) media that increased the rate of cell proliferation as well as their doubling time (15,16). In addition, ascorbic acid that has high anti-oxidant activity showed increased proliferation (15), induced chondrogenic and osteogenic differentiation (17,18), and delayed aging process (19).…”

Role of vitamins beyond vitamin D3 in bone health and osteoporosis (Review)

“…Osteoblastogenesis has been shown to be mediated by VC-induced OSX expression through the activation of PHD and subsequent proteasomal degradation of OSX gene transcriptional repressors ( 251 ). The activation of osteogenesis by VC has been shown to involve its direct interaction with PHD2 ( 252 ).…”