Accelerating effect of Shilajit on osteogenic property of adipose derived mesenchymal stem cells (ASCs)

Abstract: Background Shilajit, as a herbomineral natural substance, has been most widely used remedy for treating a numerous of illness such as bone defects in Iran traditional folk medicine since hundreds of years ago. The aim of the present study was to explore the effect of Shilajit on the osteogenic differentiation of human adipose-derived mesenchymal stem cells (ASCs) in two- and three-dimensional cultures. Materials and methods ASCs were isolated and seeded in three-dimensional (3D) 1% alginate (Alg) hydrogel with or without Shilajit at a density of 3 × 10⁵ cell/mL. For two-dimensional (2D) cultures, 3 × 10⁴ ASCs /mL were seeded into culture plates and treated with 500µg/mL Shilajit. Then, characterization was done using electron microscopy (SEM)/energy dispersive X-ray spectroscopy (EDX), alkaline phosphatase (ALP) activity, Alizarin red staining, and Raman confocal microscopy. Results Adding Shilajit had no impact on the Alg scaffold degradability. In the 3D Alg hydrogel and in the present of osteogenic medium (OM), Shilajit acted as enhancer to increase ALP activity, also showed osteoinductive property in the absence of OM compared to the 2D matched groups at all time points (days 7 and 21 both P < 0.001, for 14 days P < 0.001 and P < 0.05, respectively). In addition, calcium deposition was significantly increased in the cultures exposed to Shilajit compared to 2D matched groups on days 14 (P < 0.0001), and 21 (P < 0.001and P < 0.01, respectively). In both 3D and 2D conditions, Shilajit induced osteogenic differentiation, but Shilajit/Alg combination starts osteogenic differentiation in a short period of time. Conclusion As Shilajit accelerates the differentiation of ASCs into the osteoblasts, without changing the physical properties of the Alg hydrogel, this combination may pave the way for more promising remedies considering bone defects.