Evaluation of histological and biomechanical properties on engineered meniscus tissues using sonication decellularization.

Sonication decellularization treatment requires proper evaluations on its ability to decellularize meniscus tissue efficiently. This study was done to evaluate the histological and biomechanical properties within meniscus scaffolds. Van Gieson staining was done to evaluate the efficiency of cell removal in meniscus tissues. The consequences of treatment on viscoelastic properties are vital for scaffolds quality and were properly investigated. Picrosirius red and Safranin-O/Fast green staining was carried out to detect extracellular matrix materials (ECM). Sonication decellularization treatment has the ability to demonstrated complete nuclei removal compare to control samples as well as maintaining viscoelastic properties, namely stiffness, compression and residual force. Thus, sonication decellularization treatment had successfully produced and prepared a meniscus bioscaffold candidate in which its biomechanical strength is sustained through protection of ECM properties.

Evaluation of recellularization on decellularized aorta scaffolds engineered by ultrasonication treatment.

Aortic scaffolds prepared using sonication decellularization treatment has provided a successful medium for repopulation with vascular smooth muscle cells (VSMCs). The objective of this study is to explore the potential of tissue decellularization using ultrasonication treatment and its recellularization before implantation of the cell-seeded scaffolds into host. Aorta tissue samples are decellularized in 2% SDS with sonication for 10 hours and compared with the native tissues. The 4',6-diamidino-2-phenylindole (DAPI) staining was used to evaluate the decellularization and Hematoxylin-Eosin (H-E) staining was used to compare the VSMCs infiltrations onto the decellularized tissues at day-0 and day-6 after cell-seeding. The results histologically showed complete DNA removal from scaffolds after decellularization and subsequent recellularization resulted in successful VSMCs infiltration. Accordingly, the decellularized tissues treated with 2% SDS in sonication demonstrated successful VSMCs repopulation afterward and is speculated to have less toxicity and able to be effectively implanted into host.

Photodynamic therapy mediates innate immune responses via fibroblast-macrophage interactions.

Antibacterial photodynamic therapy (PDT) has come to attract attention as an alternative therapy for drug-resistant bacteria. Recent reports revealed that antibacterial PDT induces innate immune response and stimulates abundant cytokine secretion as a part of inflammatory responses. However, the underlying mechanism how antibacterial PDT interacts with immune cells responsible for cytokine secretion has not been well outlined. In this study, we aimed to clarify the difference in gene expression and cytokine secretion between combined culture of fibroblasts and macrophages and their independent cultures. SCRC-1008, mouse fibroblast cell line and J774, mouse macrophage-like cell line were co-cultured and PDT treatments with different parameters were carried out. After various incubation periods (1-24 h), cells and culture medium were collected, and mRNA and protein levels for cytokines were measured using real-time PCR and ELISA, respectively. Our results showed that fibroblasts and macrophages interact with each other to mediate the immune response. We propose that fibroblasts initially respond to PDT by expressing Hspa1b, which regulates the NF-κB pathway via Tlr2 and Tlr4. Activation of the NF-κB pathway then results in an enhanced secretion of pro-inflammatory cytokines (TNF-α, IL-6 and IL-1ß) and neutrophil chemoattractant MIP-2 and KC from macrophages.

The use of sonication treatment to decellularize aortic tissues for preparation of bioscaffolds.

A novel decellularization method using sonication treatment is described. Sonication treatment is the combination of physical and chemical agents. These methods will disrupt cell membrane and release cell contents to external environments. The cell removal was facilitated by subsequent rinsing of sodium dodecyl sulfate detergents. Sonication treatment is used in the preparation of complete decellularized bioscaffolds. The aim of this study is to confirm the usefulness of sonication treatment for preparation of biological scaffolds. In this study, samples of aortic tissues are decellularized by sonication treatment at frequency of 170 kHz in 0.1% and 2% sodium dodecyl sulfate detergents for 10-h treatment time. The relation between decellularization and sonication parameters such as dissolved oxygen concentration, conductivity, and pH is investigated. Histological analysis and biomechanical testing is performed to evaluate cell removal efficiency as well as changes in biomechanical properties. Minimal inflammation response elicit by bioscaffolds is confirmed by xenogeneic implantation and immunohistochemistry. Sonication treatment is able to produce complete decellularized tissue suggesting that these treatments could be applied widely as one of the decellularization method.

Preparation of decellularized meniscal scaffolds using sonication treatment for tissue engineering.

Scaffolds play a key role in the process of regeneration and morphogenesis of tissue or organ. We have developed a novel sonication decellularization system to prepare decellularized bio-scaffolds in a short treatment time. The aim of the study is to investigate sonication decellularization condition that completely decellularize meniscus can be changed as well as to maintain the biomechanical parameters of scaffolds. The meniscus samples were decellularized using sonication treatment. The treated samples were evaluated histologically by EVG for cell removal, picrosirius red for content of collagen type I and III, and safranin-O/fast green staining for content of glycosaminoglycan, and SEM for observation of scaffold surface. Indentation apparatus was used to analyze the unconfined deformation under load of native and decellularized menisci. The load parameters which are stiffness, compression and residual force were not significantly different compare with native and sonicated scaffolds. However, the content of extracellular matrix and its fiber alignment changed significantly due to sonication treatment as observed by SEM and safranin-O/fast green staining, respectively. The removal of immunogenic cell components by sonication decellularization as well as maintain its biomechanical strength of decellularized scaffolds, so that it has potential to use as an implant material for tissue engineering of menisci.

The use of sonication treatment to completely decellularize blood arteries: a pilot study.

We have developed a novel sonication decellularization system to prepare completely decellularized bioscaffolds in a short treatment time. The aim of the study is to investigate the sonication decellularization efficiency and its relation with ultrasonic power output and dissolved oxygen (DO) concentration in different detergent solution. In the study, we used aorta samples to evaluate sonication decellularization efficiency, which assessed treatment duration, sonication power and SDS detergent with/without saline. The treated samples were evaluated histologically by Hematoxylin Eosin (HE) staining and scanning electron microscopic (SEM) photographs. The concentration of DO was monitored to identify the effect of sonication on cavitation-related DO concentration in the solution. From histological results, the sonication decellularization efficiency was better than the other preparation methods. Decellularization efficiency was tended to increase significantly when DO value decreasing after 6 hours of treatment. In conclusion, we conclude that sonication treatment can be used to prepare the complete decellularized scaffolds in short treatment time.