Nanostructures based on derived inverse opal films promote the osteogenesis of mesenchymal stem cells

Abstract

Event Abstract Back to Event Nanostructures based on derived inverse opal films promote the osteogenesis of mesenchymal stem cells Qian-Ru Xiao1, Ning Zhang1 and Ning-Ping Huang1 1 State Key Laboratory of Bioelectronics, School of Biological Science and Medical Engineering, China Introduction: Targeted osteogenesis stimulated by nanotopography alone has been demonstrated in mesenchymal stem cells (MSCs)[1]. MSCs are multipotent cells that can differentiate into various lineages such as those of adipocytes, chondrocytes, myocytes, and osteoblats under chemical or physical stimuli[2],[3]. A variety of nanotopographical surfaces such as nanotubes, nanopits, and nanofibres have been demonstrated to be osteoinductive[4],[5]. Nevertheless, it seems difficult to find the surface commonality and there is a need to keep looking for optimal topographical surface for the targeted osteogenesis. In addition, the underlying mechanism of topographical guidance of stem cells is still poorly understood. In this study, we produced nanostructures based on derived inverse opal films in a much simpler and more cost-effective way to induce MSCs to differentiate into osteoblasts in the absence of inducing media. We also studied Smad-dependent signaling and MAPK pathways, key pathways involved in nanotopography-driven osteogenesis. Materials and Methods: Fabrication of Derived Inverse Opal Films: First, SiO2 opal templates were made through vertical deposition method, then the polystyrene (PS) solution was poured onto the templates. After desiccation and removal of the templates, PS inverse opal films were obtained. To get the derived inverse opal films, the inverse opal films were stretched in hot water. Scanning electron microscope (SEM) was used to characterize the topography of the above films. MSCs Isolation and Culture: Primary MSCs were isolated from the bone marrow of tibiae and the femurs of young adult female Sprague-Dawley rats. Cells of passage 2-4 were used for the experiments. MSCs were seeded with density of 2 x 104 cells/cm3 onto the above films and cultured for several days. Examination of osteogenic differentiation: Cell were collected as different time points for immunofluorescence staining and qPCR analysis. Results and Discussion: SEM images showed that the derived films had oriented mesh-like surfaces, as shown in Figure 1. It is notable that during the early stage of MSC differentiation on both the 3- and 6-times stretched films, higher levels of RUNX2, ALP, COL, and OCN were observed compared to the other substrata, while at the late stage, only the 3-times stretched films showed very high transcript levels of these osteogenic genes. Figure 2 shows that the phosphorylated Smad1/5 could hardly be observed at day 3 whereas it was more easily found at day 7, and the phosphorylation level seemed higher on both the inverse opal films and especially so on the 3-times stretched films. Furthermore, higher levels of phosphorylated ERK and phosphorylated p38 MAPK were also observed on the 3-times stretched films at day 10 (Figure 3). These results indicate that stretched film-initiated osteogenesis is reliant on both the Smad-dependent BMP signaling and MAPK pathways. Conclusion: Here, for the first time, we show that the derived inverse opal films can guide MSCs to differentiate into osteoblasts without the use of exogenous inducing factors, meaning they could be of great importance to the design of scaffold biomaterials which can stimulate stem cells to differentiate into osteoblasts in the absence of inducing media without compromising material properties. National Science Foundation of China; National High Technology Research & Development Program of China