Assessing the biocompatibility of bovine tendon scaffold, a step forward in tendon tissue engineering.

Tendon is a collagen-enriched, tough, and intricately arranged connective tissue that connects muscle to the bone and transmits forces, resulting in joint movement. High mechanical demands can affect normal tissues and may lead to severe disorders, which usually require replacement of the damaged tendon. In recent decades, various decellularization methods have been studied for tissue engineering applications. One of the major challenges in tendon decellularization is preservation of the tendon extracellular matrix (ECM) architecture to maintain natural tissue characteristics. The aim of the present study was to create a decellularized bovine Achilles tendon scaffold to investigate its cytocompatibility with seeded hAd-MSCs (human adipose derived-mesenchymal stem cells) and blastema tissue in vitro. Here, we describe a reliable procedure to decellularize bovine Achilles tendon using a combination of physical and chemical treatments including repetitive freeze-thaw cycles and the ionic detergent SDS, respectively. The decellularization effectiveness and cytocompatibility of the tendon scaffolds were verified by histological studies and scanning electron microscopy for up to 30 days after culture. Histological studies revealed hAd-MSC attachment and penetration into the scaffolds at 5, 10, 15 and 20 days of culture. However, a decrease in cell number was observed on days 25 and 30 after culture in vitro. Moreover, migration of the blastema tissue cells into the scaffold were shown at 10 to 25 days post culture, however, destruction of the scaffolds and reduction in cell number were observed on 30th day after culture. Our results suggest that this decellularization protocol is an effective and biocompatible procedure which supports the maintenance and growth of both hAd-MSCs and blastema cells, and thus might be promising for tendon tissue engineering.

Biocompatible Films of Collagen-Procyanidin for Wound Healing Applications.

The study investigated the effect of polyphenols present in Cassia auriculata (CA) leaves in enhancing the stability of the collagen protein and the wound healing potential of collagen films. The crude ethanol extract of CA was analyzed for the presence of phytochemicals and purified by column chromatography using solvents with increasing polarity. The ethanol eluted active fractions (EEAF) that precipitated gelatin was characterized using HP-TLC, FTIR spectroscopy, ESI-FT-MS/MS, and 1H NMR spectroscopy. The active compound was identified to be procyanidin B belonging to the proanthocyanidins group. The wound healing property of EEAF and collagen type I extracted from Clarias batrachus fish skin and the bovine tendon was assessed by in vitro scratch assay on L929 mice fibroblast cell lines. The EEAF-treated collagen coating enhanced in vitro wound closure in comparison with the uncoated dish. It was observed that EEAF treatment improved the physical strength of collagen films. The in vivo wound healing of the EEAF-treated collagen film was examined in male Wister rats and the wound site tissues were assessed. In vivo wound examination showed enhanced healing with EEAF incorporated collagen films. Comparatively, the EEAF-treated bovine tendon collagen films showed improved physical properties and better wound healing property than fish collagen films.

Fabrication and characterization of a decellularized bovine tendon sheet for tendon reconstruction.

Obtaining a performing decellularized tendon scaffold with proper dimensions and adequate availability is highly desirable. However, the combined study of complete decellularization and detailed characterization of native tendon extracellular matrix (ECM) from large animals is still lacking. In the present study, we developed a new decellularization protocol, including physical methods and enzymatic solutions for processing bovine Achilles tendons, and produced a decellularized bovine tendon sheet (DBTS) scaffold for tendon reconstruction. The decellularization effectiveness was demonstrated by DNA quantification and histological qualification. The removal of the alpha-gal epitopes was confirmed by ELISA analysis and immunohistochemical staining. After decellularization, there were no significant alterations of the native tendon extracellular matrix (ECM) properties, including the internal ultrastructure, biochemical compositions such as collagen, glycosaminoglycans (GAGs), basic fibroblast growth factor (bFGF) and transforming growth factor-ß1 (TGF-ß1), fibronectin and decorin, as well as substantial mechanical strength. Furthermore, the DBTS scaffold showed no cytotoxic and promoted the proliferation of NIH-3T3 fibroblasts in vitro. When implanted into rat subcutaneous tissue, the DBTS scaffold displayed excellent histocompatibility in vivo. Our results, while offering a new decellularization protocol for large tendons, can provide a promising biologic scaffold with a combination of mechanical strength and tendon ECM bioactive factors that may have many potential applications in tendon reconstruction. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 2299-2311, 2017.