Titanium nanotube arrays promote the activity of anastomotic healing-related cells by increasing fibronectin adsorption and activating the RGD–integrin pathway

Abstract

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The smooth titanium staples of stapling devices cannot reduce the incidence of gastrointestinal anastomotic leakage due to their bioinert nature and lack of active wound-healing promotion capability. This study aims to investigate whether titanium nanotube arrays (TNTs) can enhance the activity of cells involved in gastrointestinal anastomotic healing and further explore the potential mechanisms. TNTs were fabricated on pure titanium sheets via anodic oxidation, and characterized using scanning electron microscopy, roughness analysis, contact angle measurement, and x-ray photoelectron spectroscopy. Cell adhesion, proliferation, spreading, collagen secretion, and integrin expression were evaluated using methods such as CCK-8, immunofluorescence, qPCR, enzyme-linked immunosorbent assay (ELISA), and Western blot. Fibronectin (FN) adsorption and Arg-Gly-Asp tripeptide sequence (RGD domain) exposure were detected via bicinchoninic acid assay, fluorescent staining, and ELISA. The role of the RGD-integrin pathway was further investigated by supplementing serum-reduced medium with exogenous FN and using RGD-specific antagonists. The results showed that TNTs increased the roughness, hydrophilicity, and surface free energy of titanium surfaces. Compared with smooth pure titanium, TNTs promoted the adhesion, proliferation, spreading, and integrin expression of gastric mucosal epithelial cells and fibroblasts, while enhancing the collagen secretion capacity of fibroblasts. Moreover, TNTs adsorbed more FN and exposed more RGD domains, thereby upregulating integrinα5β1 expression. The RGD antagonist could reverse these enhanced cellular responses, confirming the pivotal role of the FN-RGD-integrin pathway. The conclusion indicates that TNTs enhance the adhesion, proliferation, and functional activity of gastrointestinal anastomosis-related cells by promoting FN adsorption and activating the RGD-integrin pathway, which demonstrates that TNT-modified titanium materials hold significant potential for developing bioactive anastomotic devices and promoting tissue healing.