Engineering High-Resolution Micropatterns Directly onto Titanium with Optimized Contact Guidance to Promote Osteogenic Differentiation and Bone Regeneration

Abstract

Topographical cues play an important role in directing cell behavior, and thus, extensive research efforts have been devoted to fabrication of surface patterns and exploring the contact guidance effect. However, engineering high-resolution micropatterns directly onto metallic implants remains a grand challenge. Moreover, there still lacks evidence that allows translation of in vitro screening to in vivo tissue response. Herein, we demonstrate a fast, cost-effective, and feasible approach to the precise fabrication of shape- and size-controlled micropatterns on titanium substrates using a combination of photolithography and inductively coupled plasma-based dry etching. A titanium TopoChip containing 34 microgrooved patterns with varying geometry parameters and a flat surface as the control was designed for a high-throughput in vitro study of the contact guidance of osteoblasts. The correlation between the surface pattern dimensions, cell morphological characteristics, proliferation, and osteogenic marker expression was systematically investigated in vitro. Furthermore, the surface with the highest osteogenic potential in vitro along with representative controls was evaluated in rat cranial defect models. The results show that microgrooved pattern parameters have almost no effect on osteoblast proliferation but significantly regulate the cell morphology, orientation, focal adhesion (FA) formation, and osteogenic differentiation in vitro. In particular, a specific groove pattern with a ridge width of 3 μm, groove width of 7 μm, and depth of 2 μm can most effectively align the cells through regulating the distribution of FAs, resulting in an anisotropic actin cytoskeleton, and thereby promoting osteogenic differentiation. In vivo, microcomputed tomography and histological analyses show that the optimized pattern can apparently stimulate new bone formation. This study not only offers a microfabrication method that can be extended to fabricate various shape- and size-controlled micropatterns on titanium alloys but also provides insight into the surface structure design of orthopedic and dental implants for enhanced bone regeneration.

The Synergy of Topographical Micropatterning and Ta|TaCu Bilayered Thin Film on Titanium Implants Enables Dual‐Functions of Enhanced Osteogenesis and Anti‐Infection

“…After further lithographic exposure (Karl Suss MA6) for 7 s, and developing for 30 s, the discs were rinsed in deionized (DI) water and dried with nitrogen gas. [ 24 ] The photoresist pattern with the designed dimension was transferred into the underlying Ti substrates using ICP‐based dry etching in Cl 2 /BCl 3 /Ar (volume ratio = 3:1:6) plasma for 3 min.…”

“…The surgical procedure is illustrated in Figure S11 (Supporting Information) and detailed in the previous study. [ 24 ] In brief, the skin of the anesthetized SD rats beyond the cranium were first shaved and sterilized by 10% povidone‐iodine solution, a 3 cm midline skin incision was then created over the cranial bone, followed by separation of the periosteum from the bone surface. A circular defect with Φ = 3 mm was drilled under a constant saline rinse.…”

“…[ 23 ] Recently, we have developed a feasible approach to the precise fabrication of shape‐ and size‐controlled micropatterns on Ti substrate and found the optimized pattern which can stimulate new bone formation using high‐throughput screening. [ 24 ] However, the surface topography for enhanced osteogenic differentiation fails in prevention of bacteria attachment and colonization, partially due to the length scale difference between osteoblasts and bacteria. [ 25 ] Therefore, developing smart implant surfaces with simultaneously enhanced osteogenesis and antibacterial activity is in urgent need.…”

“…[52] Herein, we describe a surface design strategy to achieve dualfunctions of enhanced osteogenesis and antimicrobial activity on Ti implant through the synergistic effect of topographical micropatterning and a bilayered nanostructured metallic thin film. Specific microgrooved patterns showing optimized contact guidance [24] were fabricated directly on Ti implant (PTi), followed by sequential deposition of nanostructured Ta-5 wt% Cu (TaCu) and pure Ta thin films (Ta|TaCu|PTi), respectively. The underlying layer provides the source of the antibacterial agent Cu, while the top layer is used to prevent burst release of Cu ions and serves as an osteoinductive layer.…”

Properties improvement of titanium alloys scaffolds in bone tissue engineering: a literature review

“…and surface microstructure accurately. Zhu et al (28) reported a combination method of photolithography. They inductively coupled plasma-based dry etching for surface design, which can precisely control the microstructure pattern of the titanium surface and screen appropriate patterns and parameters to achieve the best osteogenesis effect.…”