Nanovibrational Stimulation of Mesenchymal Stem Cells Induces Therapeutic Reactive Oxygen Species and Inflammation for Three-Dimensional Bone Tissue Engineering

Abstract

There is a pressing clinical need to develop cell-based bone therapies due to a lack of viable, autologous bone grafts and a growing demand for bone grafts in musculoskeletal surgery. Such therapies can be tissue engineered and cellular, such as osteoblasts, combined with a material scaffold. Because mesenchymal stem cells (MSCs) are both available and fast growing compared to mature osteoblasts, therapies that utilize these progenitor cells are particularly promising. We have developed a nanovibrational bioreactor that can convert MSCs into bone-forming osteoblasts in two- and three-dimensional, but the mechanisms involved in this osteoinduction process remain unclear. Here, to elucidate this mechanism, we use increasing vibrational amplitude, from 30 nm (N30) to 90 nm (N90) amplitudes at 1000 Hz and assess MSC metabolite, gene, and protein changes. These approaches reveal that dose-dependent changes occur in MSCs’ responses to increased vibrational amplitude, particularly in adhesion and mechanosensitive ion channel expression and that energetic metabolic pathways are activated, leading to low-level reactive oxygen species (ROS) production and to low-level inflammation as well as to ROS- and inflammation-balancing pathways. These events are analogous to those that occur in the natural bone-healing processes. We have also developed a tissue engineered MSC-laden scaffold designed using cells’ mechanical memory, driven by the stronger N90 stimulation. These mechanistic insights and cell-scaffold design are underpinned by a process that is free of inductive chemicals.

A Hydrogel Platform that Incorporates Laminin Isoforms for Efficient Presentation of Growth Factors – Neural Growth and Osteogenesis

“…Results showed upregulated osteopontin expression when compared to the gels cultured in osteogenic media at 14 days (Figure 4D), as previously observed. [ 69 ] OPN and OCN expression were further evaluated after 21 days, indicating that OCN expression level did not show any statistically significant difference between the two tested conditions (Figure S8B, Supporting Information). The lack of statistical significance in the mRNA expression values of OPN and OCN at 21 days might be due to the temporal expression profile of both genes, [ 69 ] where OPN reaches a maximum around 14 days and then the expression is attenuated, whereas OCN shows a monotonic increase in its expression values, reaching a maximum after 21 days.…”

“…[ 69 ] OPN and OCN expression were further evaluated after 21 days, indicating that OCN expression level did not show any statistically significant difference between the two tested conditions (Figure S8B, Supporting Information). The lack of statistical significance in the mRNA expression values of OPN and OCN at 21 days might be due to the temporal expression profile of both genes, [ 69 ] where OPN reaches a maximum around 14 days and then the expression is attenuated, whereas OCN shows a monotonic increase in its expression values, reaching a maximum after 21 days. Even if the amount of BMP‐2 retained in the gel after 5 days is similar (220 and 200 ng corresponding to LM/PEG and PEG), the biological effects of the BMP‐2 bound to LM332 on the hMSC differentiation are significantly different.…”

The use of nanovibration to discover specific and potent bioactive metabolites that stimulate osteogenic differentiation in mesenchymal stem cells

“…Note that our activity metabolite testing data have been performed on cells in 2D, and there will be differences when scaled to 3D. For nanovibrational MSC stimulation, there are similarities in known mechanism in 2D and 3D in that intracellular tension from adhesion and mechanoresponsive channels (e.g., transient receptor potential cation and piezo channels) have been identified under both conditions ( 7 , 15 , 52 ). However, in 2D, the bias appears to be toward adhesion and tension regulation of MSC osteogenesis, and in 3D, the bias is toward mechanoresponsive channel regulation ( 7 , 15 , 52 ).…”

Electrical Stimulation of Adipose-Derived Stem Cells in 3D Nanofibrillar Cellulose Increases Their Osteogenic Potential

“…Osteogenic differentiation of ASCs and MSCs can be induced by a combination of soluble factors such as dexamethasone, ascorbic acid, and β-glycerophosphate and further increased by a wide range of biophysical factors including nano-, micro-, and macrotopography [ 15 , 16 ], substrate stiffness [ 17 ], fluid shear stress [ 18 ], nanovibration-induced displacement [ 19 , 20 ], and ES [ 21 , 22 ]. The effects of ES on cells have been recently reviewed and they appear to include a profound impact on cellular proliferation and osteogenic differentiation [ 23 ].…”