Perichondrium-inspired permeable nanofibrous tube well promoting differentiation of hiPSC-derived pellet toward hyaline-like cartilage pellet

Abstract

The pellet formation has been regarded as a golden standard for in vitro chondrogenic differentiation. However, a spatially inhomogeneous chondrogenic microenvironment around a pellet resulted from the use of a traditional impermeable narrow tube, such as the conical tube, undermines the differentiation performance and therapeutic potential of differentiated cartilage pellet in defective articular cartilage treatment. To address this drawback, a perichondrium-inspired permeable nanofibrous tube (PINaT) well with a nanofibrous wall permeable to gas and soluble molecules is proposed. The PINaT well was fabricated with a micro deep drawing process where a flat thin nanofibrous membrane was transformed to a 3.5 mm deep tube well with a ∼50 µm thick nanofibrous wall. Similar to in vivo perichondrium, the PINaT well was found to allow oxygen and growth factor diffusion required for chondrogenic differentiation across the entire nanofibrous wall. Analyses of gene expressions (COL2A1, COL10A1, ACAN, and SOX9), proteins (type II and X collagen), and glycosaminoglycans contents were conducted to assess the differentiation performance and clinical efficacy of differentiated cartilage pellet. The regulated spatially homogeneous chondrogenic microenvironment around the human induced pluripotent stem cell-derived pellet (3 × 105 cells per pellet) in the PINaT well remarkably improved the quality of the differentiated pellet toward a more hyaline-like cartilage pellet. Furthermore, an accelerated chondrogenic differentiation process of the pellet produced by the PINaT well was achieved for 14 days, demonstrating a hyaline cartilage-specific marker similar to the control pellet differentiated for 20 days. Finally, the enhanced clinical efficacy of the hyaline-like cartilage pellet was confirmed using an osteochondral defect rat model, with the repaired tissue resembling hyaline cartilage rather than fibrous cartilage after 8 weeks of regeneration.

Advances in Regenerative Sports Medicine Research

“…To achieve a breakthrough in osteochondral regeneration, we must focus on preparing integrated bionic scaffolds that accurately simulate the microenvironment of osteochondral regeneration, and solve the following key scientific problems: 1) accurate simulation of 3D shapes, multi gradient structures, regional specific matrix components, and the microenvironment factors of joints to prepare a bionic osteochondral scaffold (Choe, et al, 2021;Lee, et al, 2021;Liu, et al, 2021); 2) achievement of osteochondral tissue regeneration and biological joint construction in vitro; and 3) realization of the industrialization of integrated bionic stents and clinical transformations of the biological joints. The accumulation of separate tissue regeneration research for the cartilage and bone and the application of emerging cutting-edge technologies in recent years has facilitated breakthroughs regarding these technical problems.…”

Preferential stimulation of melanocytes by M2 macrophages to produce melanin through vascular endothelial growth factor

“…The results were calculated using the LightCycler (Roche Diagnostics). Relative gene expression was calculated using the delta cycle-threshold (ΔCt) method, and the ΔCt values were converted to ratios 51 , 52 . The primer sequences are presented in Supplementary Table S1 .…”

Recent Progress in Fabrication of Electrospun Nanofiber Membranes for Developing Physiological In Vitro Organ/Tissue Models

“…Similar to in vivo perichondrium which contains a collagenous fibrous layer around the elastic and hyaline cartilages, the fabricated PCL NF wells promoted the diffusions of oxygen, nutrients, and growth factor across the permeable nanofibrous wall, which accelerated and enhanced chondrogenic differentiation of a human-induced pluripotent stem cell (hiPSC)-derived cartilage spheroid (Figure 12c). [142] Shim et al generated hiPSCderived insulin-producing spheroids in a thermoformed PCL NF microwell array, which showed enhanced differentiation capacity compared with the traditional impermeable PDMS microwell array. [101] Interestingly, in the studies of Kim et al, Leet et al, and Shim et al, they integrated the 2.5D NFM composed of NF microwells with a cell culture insert, which not only provided a user-friendly cell culture platform but also supplied enough oxygen and nutrients/growth factors to spheroids from both the apical and basolateral sides of the well plate.…”

“…[143] Furthermore, cells in a bio- License. [142] Copyright 2022, The Authors, published by SAGE Publications. b) HepG2 spheroid constructed on flat NFM-bottomed microwell array and concave NFM-bottomed microwell array.…”

“…Graph of proliferation and amylase activity level of salivary gland spheroids showing enhanced functionality of salivary gland spheroids cultured on the PCL NFM-bottomed microwell array compared to monolayer culture. Reproduced under the terms of the Creative Commons Attribution 4.0 InternationalLicense [142]. Copyright 2022, The Authors, published by SAGE Publications.…”