Yi Xiao,1,2,* Shiyan Yang,2,3,* Yang Sun,2,* Robert L Sah,4,5 Jincheng Wang,2 Chunshan Han1 

1Thoracic Surgery Department, The China-Japan Union Hospital of Jilin University, Changchun, Jilin, 130000, People’s Republic of China; 2Orthopedic Medical Center, the Second Hospital of Jilin University, Changchun, Jilin, 130000, People’s Republic of China; 3Department of Head and Neck, Sun Yat-Sen University Cancer Center, Guangzhou, Guangdong, 510060, People’s Republic of China; 4Department of Bioengineering, University of California–San Diego, La Jolla, CA, 92037, USA; 5Center for Musculoskeletal Research, Institute of Engineering in Medicine, University of California–San Diego, La Jolla, CA, 92037, USA

*These authors contributed equally to this work

Correspondence: Chunshan Han, Thoracic Surgery Department, the China-Japan Union Hospital of Jilin University, Changchun, Jilin, 130000, People’s Republic of China, Email hancs@jlu.edu.cn

Abstract: Nanoscale morphologies on the surface of substrates/scaffolds have gained considerable attention in cartilage tissue engineering for their potential to improve chondrogenic differentiation and cartilage regeneration outcomes by mimicking the topographical and biophysical properties of the extracellular matrix (ECM). To evaluate the influence of nanoscale surface morphologies on chondrogenic differentiation of stem cells and discuss available strategies, we systematically searched evidence according to the PRISMA guidelines on PubMed, Embase, Web of Science, and Cochrane (until April 2024) and registered on the OSF (osf.io/3kvdb). The inclusion criteria were (in vitro) studies reporting the chondrogenic differentiation outcomes of nanoscale morphologies on the surface of substrates/scaffolds. The risk of bias (RoB) was assessed using the JBI-adapted quasi-experimental study assessment tool. Out of 1530 retrieved articles, 14 studies met the inclusion criteria. The evidence suggests that nanoholes, nanogrills, nanoparticles with a diameter of 10– 40nm, nanotubes with a diameter of 70– 100nm, nanopillars with a height of 127– 330nm, and hexagonal nanostructures with a periodicity of 302– 733nm on the surface of substrates/scaffolds result in better cell adhesion, growth, and chondrogenic differentiation of stem cells compared to the smooth/unpatterned ones through increasing integrin expression. Large nanoparticles with 300– 1200nm diameter promote pre-chondrogenic cellular aggregation. The synergistic effects of the surface nanoscale topography and other environmental physical characteristics, such as matrix stiffness, also play important in the chondrogenic differentiation of stem cells. The RoB was low in 86% (12/14) of studies and high in 14% (2/14). Our study demonstrates that nanomorphologies with specific controlled properties engineered on the surface of substrates/scaffolds enhance stem cells’ chondrogenic differentiation, which may benefit cartilage regeneration. However, given the variability in experimental designs and lack of reporting across studies, the results should be interpreted with caution.

Keywords: nanomorphology, surface engineering, chondrogenesis, stem cell, cartilage regeneration