Inducing macrophage M2-Type polarization via anodized anisotropic TiO₂ nanoporous topography on titanium surfaces

Dental implant success critically depends on soft tissue integration (STI) to prevent peri-implantitis. Macrophages, key immune regulators, exhibit distinct pro-inflammatory (M1) or anti-inflammatory/pro-healing (M2) phenotypes, significantly impacting peri-implant tissue responses. While implant surface nanostructures modulate macrophage polarization, the effects of specific topographies and underlying mechanisms require further elucidation. This study investigated the ability of an anodized anisotropic titanium dioxide nanoporous (TNP) surface to direct human macrophage polarization toward the M2 phenotype and explored the associated mechanisms.

An anisotropic nanoporous titanium topography was fabricated via electrochemical anodization followed by annealing. Surface characterization included morphological analysis (FE-SEM, AFM), wettability assessment (contact angle measurement), and evaluation of mechanical properties (nanoindentation, nanoscratch testing). THP-1-derived macrophages were utilized for in vitro evaluation. Cell viability and adhesion were measured using the CCK-8 assay. Macrophage morphology was observed via scanning electron microscopy (SEM). Polarization status was assessed by flow cytometry (CD68, CD86, CD206) and RT-qPCR (CD86, CD206, TNF-α, IL-1β, iNOS, TGF-β). Subsequently, transcriptomic profiling through RNA sequencing (RNA-Seq) was conducted to analyze underlying molecular pathways.

The TNP surface featured uniform 30 nm diameter nanopores, significantly increased hydrophilicity and surface energy, and adequate mechanical properties. Macrophage early adhesion and spreading (enhanced pseudopodia formation) were significantly increased on TNP compared to control. Flow cytometry and RT-qPCR revealed that TNP induced M2 polarization, as evidenced by significantly upregulated CD206 and TGF-β expression, while downregulating pro-inflammatory markers (CD86, TNF-α, IL-1β, and iNOS). RNA-Seq analysis further identified significant alterations in genes associated with cell projection membranes, collagen fiber organization, and actin binding.

Anodization followed by annealing successfully created a mechanically stable, hydrophilic titanium nanoporous surface. The in vitro study demonstrated that the specific nano-topographical structure enhanced macrophage adhesion and spreading, inducing a shift towards the anti-inflammatory M2 phenotype characterized. The observed polarization is potentially associated with cytoskeletal reorganization and the activation of mechanotransduction pathways triggered by the nanostructure. This surface modification strategy offers significant potential for enhancing dental implant abutment soft tissue integration by favorably modulating the local immune response.