In Silico and In Vitro Insights of Clay-Integrin Interactions—Evaluating Na-Montmorillonite's Role in Biomineralization and Osteogenic Differentiation of Mesenchymal Stem Cells in Nanoclay Scaffolds

Montmorillonite (MMT) clay, a prevalent natural mineral, significantly influences biomineralization processes within biopolymer scaffolds, a crucial mechanism for bone tissue regeneration through osteogenesis. Various extracellular and intracellular signaling pathways facilitate the osteogenic differentiation of human mesenchymal stem cells (hMSCs) into mature osteoblasts. Extracellular minerals, including MMT clay, can alter the conformation of membrane proteins such as integrins, initiating intracellular signal transduction. However, the precise molecular mechanisms governing these signaling events during biomineralization within nanoclay-based polymer scaffolds remain largely unexplored. To address this challenge, the clay-integrin interface was analyzed to explore the interaction and activation of integrin (αVβ3), which subsequently triggers intracellular events leading to osteogenesis. Our molecular modeling study revealed that the non-bonded interactions between Na-MMT clay and integrin induce a hinge opening between the βA and hybrid domains of integrin, confirming conformational changes necessary for its activation. Moreover, we elucidated the cell adhesion mechanism facilitated by clay-integrin binding, which promotes the formation of a stable adhesion complex between cells and clay minerals through non-bonded interactions. The impact of clay mineral proximity on the mechanical behavior of integrin was further examined using steered molecular dynamics simulations, demonstrating that such proximity influences the mechanical stimuli applied to integrins, which regulates biological responses. Additionally, experimental studies were conducted to examine the activation of integrin (αVβ3) expression in relation to biomarkers associated with various stages of osteogenic differentiation in the presence of nanoclay scaffolds. The inclusion of Na-MMT clay in biopolymer scaffolds enhanced the expression of αVβ3 integrin and osteoblast marker genes, as well as run-related transcription factor 2, indicating improved osteogenic differentiation of hMSCs. Our combined computational and experimental approach provides new insights into clay-integrin interactions that promote biomineralization and influence cell adhesion mechanisms, paving the way for more effective biomaterial designs.