Micro-Nanostructured Polymeric Scaffolds for Bone Tissue Engineering

Abstract

While bone tissue allograft and autograft are commonly used in bone healing, their application is limited by factors such as availability, donor site morbidity, and immune response to the grafted tissue. Tissue-engineered implants, such as acellular or cellular polymeric structures, offer a promising alternative, and are a current trend in tissue engineering. Leveraging recent advancements in bone tissue engineering (BTE), we utilize 3D printing to develop biodegradable scaffolds that combine mechanical strength and bioactivity to facilitate bone repair and regeneration. This study focuses on the design and fabrication of mechanically competent 3D printed poly (L-lactic acid) (PLLA) micro-structured scaffolds. These scaffolds are enhanced with collagen type I nanofibrils to create bioactive scaffolds that promote tissue regeneration. The performance of these mechanically competent, micro-nanostructured polymeric matrices, in combination with bone marrow stromal cells (BMSCs), is evaluated in PLLA and PLLA-collagen scaffolds. The resulting micro-nanostructured PLLA-Collagen scaffolds mimic trabecular bone architecture, mechanical strength, and the extracellular matrix environment found in native bone tissue. The composite PLLA-collagen scaffolds exhibit mechanical properties in the mid-range of human trabecular bone. Both PLLA and PLLA-Collagen scaffolds support human BMSCs adhesion, proliferation, and osteogenic differentiation. A significantly higher number of implanted host cells are distributed in the PLLA-Collagen scaffolds with greater bone density, more uniform cell distribution, and attachment compared to the PLLA microstructure. Additionally, the biomimetic collagen nanostructure potently induces osteogenic transcription evidenced by increased alkaline phosphatase activity and upregulation of bone markers such as sialoprotein and collagen type I, ultimately guiding stem cell-mediated formation of a mature, mineralized bone matrix throughout the interconnected scaffold pores. This study underscores the benefits of micro-nanostructured scaffolds in successfully generating the inductive microenvironment of native bone extracellular matrix, triggering the cascade of cellular events required for functional bone regeneration, repairing critical-sized bone defects, and ultimately serving as an alternative material platform for bone regeneration, thereby instilling confidence in the potential of our research.

Keywords: Bone tissue engineering; collagen type I nanofibrils; human mesenchymal stem cells; osteogenic differentiation; poly (L-lactic acid).

Fig. 1.

(a) and (b) Optical images of 3D printed scaffolds PLA, (c) and (d) SEM images of 3D printed scaffolds PLA (scale bar = 2 mm), (e) collagen coated 3D printed scaffolds (top view, scale bar = 200 μm) and (f) collagen coated 3D printed scaffolds (cross-sectional view, scale bar = 50 μm).

Fig. 2.

Torsional analysis of 3 printed scaffolds break angle (◦), peak torque (NM).

Fig. 3.

Live/dead images of 3 printed scaffolds (a) PLA-collagen, (b) PLA (day 1, 3, 7 and 14) scale bar = 100 μm.

Fig. 4.

In vitro phenotype development by hMSCs seeded onto scaffolds under osteoinduction: (a) DNA content and (b) alkaline phosphatase activity over 3–28 days. Multiple t test statistical significance (*P < 0.05).

Fig. 5.

Alizarin red images for the PlA-collagen coated and PLA 3D printed scaffold for day 14, 21 and 28.

Fig. 6.

Immunostaining images for the PlA-collagen coated and PLA 3D printed scaffold for days 14 and 21.