Microspheres containing decellularized cartilage induce chondrogenesis in vitro and remain functional after incorporation within a poly(caprolactone) filament useful for fabricating a 3D scaffold

Abstract

In this study, articular cartilage was decellularized preserving a majority of the inherent proteins, cytokines, growth factors and sGAGs. The decellularized cartilage matrix (dCM) was then encapsulated in poly(lactic acid) microspheres (MS+dCM) via double emulsion. Blank microspheres without dCM, MS(-), were also produced. The microspheres were spherical in shape and protein encapsulation efficiency within MS+dCM was 63.4%. The sustained release of proteins from MS+dCM was observed over 4 weeks in vitro. Both MS+dCM and MS(-) were cytocompatible. The sustained delivery of retained growth factors and cytokines from MS+dCM promoted cell migration in contrast to MS(-). Subsequently, chondrogenesis of hMSCs was upregulated in presence of MS+dCM as evidenced from immunohistochemistry, biochemical quantification and qPCR studies. Specifically, collagen II, aggrecan and SOX 9 gene expression were increased in the presence of MS+dCM by an order or more in magnitude compared to MS(-) with concomitant downregulation of hypertrophic genes (COL X) despite being cultured in the absence of chondrogenic media, (p<0.05). Lastly, microspheres containing alkaline phosphatase (MS+ALP), a surrogate to assess the thermal stability of dCM proteins, incorporated within poly(caprolactone) filaments showed that the enzyme remained functional after filament production by melt extrusion. The establishment of a novel, thermally stable process for producing filaments containing chondroinductive microspheres provides evidence supporting subsequent development of a clinically-relevant, 3D scaffold fabricated from them for osteochondral regeneration and repair.

Desktop-stereolithography 3D printing of a radially oriented extracellular matrix/mesenchymal stem cell exosome bioink for osteochondral defect regeneration

“…In this study, we printed bioink based on photo-crosslinked cartilage ECM and GelMA hydrogels, which exhibited proper mechanical properties (33.24 ± 8.8 kPa) compared with that of un-crosslinked hydrogels. We also showed that cartilage ECM hydrogels could enhance cell migration which was consistent with a previous report 40. In the ECM proteomics analysis, the cell migration-related proteins such as prelamin-A/C, chemerin, vinculin, apolipoprotein E, and decorin were detected.…”

Bone defect reconstruction via endochondral ossification: A developmental engineering strategy

“…130 Therefore, the use of dCM for endochondral bone engineering remains controversial, and studies have demonstrated that dCM promotes the chondrogenic differentiation but inhibits the hypertrophic differentiation of MSCs in vitro and subsequent ECO in vivo. [131][132][133] It has long been realized that viable hypertrophic cartilage will form bone in vivo; decellularized hypertrophic cartilage matrix (dHCM), which is manufactured from native epiphyseal plates or fractured callus tissue, can also trigger the natural ECO process upon implantation. 134 A major drawback of these types of naturally derived dECMs is the limited donor source, which hinders their clinical application.…”

A Review of the Use of Microparticles for Cartilage Tissue Engineering

“…Cells grew on all groups in the experiment, but the number of viable cells was much higher on microspheres with the dCM compared to microspheres without the dCM. Cells in the dCM group portrayed more prominent sulfated glycosaminoglycan staining, and they were organized in a high-density, compact structure [ 83 ]. This method could be a potential treatment for cartilage regeneration, but more testing in vivo is needed to demonstrate safety and efficacy.…”