Characterization of decellularized chicken skin as a tissue engineering scaffold.

Decellularization has been applied to many tissues and organs to obtain biomaterials for applications in tissue engineering. In this study, decellularization and characterization of chicken skin was performed to provide comprehensive information and in-depth details on this material as a potential tissue scaffold. Application of Triton X-100 and sodium dodecyl sulfate (SDS) on tissues at different time intervals as two decellularization protocols were compared according to various aspects, such as removal of cellular components, DNA quantification, protection of extracellular matrix (ECM), mechanical properties, and cytocompatibility, to find the optimum technique during preparation of decellularized scaffolds. The results showed that treatment with SDS revealed better results when compared with Triton X-100 regarding the preservation of tissue structure and morphology, although there was no difference in the efficiency of decellularization. In general, the tissues decellularized with SDS demonstrated higher levels of cytocompatibility and better mechanical properties in comparison with samples treated with Triton X-100. In conclusion, this study revealed that decellularized chicken skin is a cheap, abundant, and biocompatible material that supports cell attachment, growth, and proliferation. Therefore, it could be used as a proper candidate to prepare scaffolds for further studies on tissue engineering, especially for skin tissue engineering.

Fluorescence labelled XT5 modified nano-capsules enable highly sensitive myeloma cells detection.

Accurate diagnosis of cancer cells in early stages plays an important role in reliable therapeutic strategies. In this study, we aimed to develop fluorescence-conjugated polymer carrying nanocapsules (NCs) which is highly selective for myeloma cancer cells. To gain specific targeting properties, NCs, XT5 molecules (a benzamide derivative) which shows high affinity properties against protease-activated receptor-1 (PAR1), that overexpressed in myeloma cancer cells, was used. For this purpose, 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[carboxy(polyethylene glycol)-2000]-carboxylic acid (DSPE-PEG2000-COOH) molecules, as a main encapsulation material, was conjugated to XT5 molecules due to esterification reaction using N,N'-dicyclohexylcarbodiimide as a coupling agent. The synthesized DSPE-PEG2000-COO-XT5 was characterized by using FT-IR and1H NMR spectroscopies and results indicated that XT5 molecules were successfully conjugated to DSPE-PEG2000-COOH. Poly(fluorene-alt-benzothiadiazole) (PFBT) conjugated polymer (CP) was encapsulated with DSPE-PEG2000-COO-XT5 due to dissolving in tetrahydrofuran and ultra-sonication in an aqueous solution, respectively. The morphological properties, UV-vis absorbance, and emission properties of obtainedCPencapsulatedDSPE-PEG2000-COO-XT5(CPDP-XT5) NCs was determined by utilizing scanning electron microscopy, UV-vis spectroscopy, and fluorescent spectroscopy, respectively. Cytotoxicity properties of CPDP-XT5 was evaluated by performing MTT assay on RPMI 8226 myeloma cell lines. Cell viability results confirmed that XT5 molecules were successfully conjugated to DSPE-PEG2000-COOH. Specific targeting properties of CPDP-XT5 NCs and XT5-free NCs (CPDP NCs) were investigated on RPMI 8226 myeloma cell lines by utilizing fluorescent microscopy and results indicated that CPDP-XT5 NCs shows significantly high affinity in comparison to CPDP NCs against the cells. Homology modeling and molecular docking properties of XT5 molecules were evaluated and simulation results confirmed our results.

Spatiotemporally controlled, aptamers-mediated growth factor release locally manipulates microvasculature formation within engineered tissues.

Spatiotemporally controlled growth factor (GF) delivery is crucial for achieving functional vasculature within engineered tissues. However, conventional GF delivery systems show inability to recapitulate the dynamic and heterogeneous nature of developing tissue's biochemical microenvironment. Herein, an aptamer-based programmable GF delivery platform is described that harnesses dynamic affinity interactions for facilitating spatiotemporal control over vascular endothelial GF (VEGF165) bioavailability within gelatin methacryloyl matrices. The platform showcases localized VEGF165 sequestration from the culture medium (offering spatial-control) and leverages aptamer-complementary sequence (CS) hybridization for triggering VEGF165 release (offering temporal-control), without non-specific leakage. Furthermore, extensive 3D co-culture studies (human umbilical vein-derived endothelial cells & mesenchymal stromal cells), in bi-phasic hydrogel systems revealed its fundamentally novel capability to selectively guide cell responses and manipulate lumen-like microvascular networks via spatiotemporally controlling VEGF165 bioavailability within 3D microenvironment. This platform utilizes CS as an external biochemical trigger for guiding vascular morphogenesis which is suitable for creating dynamically controlled engineered tissues.

Preparation and characterization of decellularized rooster comb as a scaffold for tissue engineering applications.

Decellularization is a method that has been widely used in tissue engineering especially in the last 20 years. In this study decellularized rooster comb was prepared and characterized for using it as a tissue scaffold. Treatment of tissues with sodium dodecyl sulfate (SDS) and Triton X-100 as two decellularization procedures in different time points were compared according to different parameters such as cytocompatibility, cell removal, preservation of extracellular matrix (ECM), and mechanical properties to find the optimum technique. Even though there was no difference regarding to efficiency on cell removal, SDS demonstrated better results on protection of tissue morphology in comparison with Triton X-100. Therefore, in general the samples treated with SDS showed higher levels of mechanical properties and cytocompatibility in comparison with Triton X-100 applied tissues. In the cuisines of many countries, rooster comb is discarded as a waste material however, in this study it was demonstrated that decellularized rooster comb could be utilized as a cheap, easily obtainable, and biocompatible scaffold. In conclusion, it was revealed that decellularized rooster comb is a promising biomaterial for using as scaffold and it is expected to be utilized for the further studies in particular on skin tissue engineering.

Decellularized inner body membranes for tissue engineering: A review.

Body membranes are thin sheets/layers of cells or tissues which cover the surface of internal organs, the outside of the body and lines various body cavities. These membranes are separated into two main groups which are epithelial membranes and connective tissue membranes. Decellularized forms of inner body membranes in the groups of epithelial membranes (amniotic membrane, mesentery, omentum, pericardium, peritoneum, pleura) and connective tissue membranes (fascia, periosteum, synovial membrane) have been used in tissue engineering studies for preparation and regeneration of various tissues such as bone, tendon, cartilage, skin, cornea, ocular surface, uterine, periodontium, vascular and cardiovascular structures. Decellularized inner body membranes have high biocompatibility and support cell attachment, cell growth and angiogenesis which are desired properties for using as versatile tools in tissue engineering applications. Even though, decellularized forms of these membranes have been used in many studies, it is necessary to develop new decellularization methods for more effective cell removal and less destructive properties on tissue structures. Moreover, development of decellularization agents which target removal of antigens of donor tissues is also essential because these antigens are one of the main reasons for tissue-organ rejections in allogeneic and xenogeneic tissue-organ implantations. This review provides comprehensive information and analysis about the current state of the art in the literature on decellularized inner body membranes and applications of these membranes in tissue engineering.

Critical-size alveolar defect treatment via TGF-ß3 and BMP-2 releasing hybrid constructs.

In the present study a combination of Transforming Growth Factor Beta 3 (TGF-ß3) and Bone Morphogenetic Protein-2 (BMP-2) loaded gelatin films sandwiched between poly (L-lactide) (PLLA)/poly (ε-caprolactone) (PCL) matrices were produced to enhance bone formation in alveolar bone defects. Osteogenic properties of tissue constructs were tested in alveolar bone defect model in rats. Bone healing was assessed by osteogenic gene expression levels of bone sialoprotein (BSP), alkaline phosphatase (ALP), osteonectin (ON, SPARC), osteocalcin (OC), runt-related transcription factor 2 (RUNX2), bone specific alkaline phosphatase (BALP) activity, histomorphometry and microtomography. Increase in osteogenic gene expression levels and BALP activity results showed that new bone formation was significantly accelerated in TGF-ß3 + BMP-2 loaded scaffold group compared to growth factor free and only BMP-2 loaded groups. The micro-computed tomography (µ-CT) data from the 4th months revealed that (TGF-ß3+ BMP-2) loaded scaffolds displayed increased bone formation and was able to fulfill 84% of the defect area (p < 0.05). Accelerated bone formation in the S-GF-B-T group compared to that of the S-GF group at the end of the 4th month was further verified via histomorphometric analysis (p = 0.008). Gene expression, BALP activity, microtomography and histomorphometry analysis indicated that (TGF-ß3 + BMP-2) loaded PLLA/PCL scaffolds increased the new bone formation. BMP-2 loaded scaffolds were less effective than combination of TGF-ß3 and BMP-2 loaded scaffolds. These findings demonstrated that focusing on the PLLA/PCL hybrid scaffolds combined with (TGF-ß3 + BMP-2) may lay the groundwork for future therapy-oriented efforts to enhance bone formation in alveolar defects.

Bioinks for 3D bioprinting: an overview.

Bioprinting is an emerging technology with various applications in making functional tissue constructs to replace injured or diseased tissues. It is a relatively new approach that provides high reproducibility and precise control over the fabricated constructs in an automated manner, potentially enabling high-throughput production. During the bioprinting process, a solution of a biomaterial or a mixture of several biomaterials in the hydrogel form, usually encapsulating the desired cell types, termed the bioink, is used for creating tissue constructs. This bioink can be cross-linked or stabilized during or immediately after bioprinting to generate the final shape, structure, and architecture of the designed construct. Bioinks may be made from natural or synthetic biomaterials alone, or a combination of the two as hybrid materials. In certain cases, cell aggregates without any additional biomaterials can also be adopted for use as a bioink for bioprinting processes. An ideal bioink should possess proper mechanical, rheological, and biological properties of the target tissues, which are essential to ensure correct functionality of the bioprinted tissues and organs. In this review, we provide an in-depth discussion of the different bioinks currently employed for bioprinting, and outline some future perspectives in their further development.

3D bioprinting of biomimetic aortic vascular constructs with self-supporting cells.

Cardiovascular diseases are the leading cause of deaths throughout the world. Vascular diseases are mostly treated with autografts and blood vessel transplantations. However, traditional grafting methods have several problems including lack of suitable harvest sites, additional surgical costs for harvesting procedure, pain, infection, lack of donors, and even no substitutes at all. Recently, tissue engineering and regenerative medicine approaches are used to regenerate damaged or diseased tissues. Most of the tissue engineering investigations have been based on the cell seeding into scaffolds by providing a suitable environment for cell attachment, proliferation, and differentiation. Because of the challenges such as difficulties in seeding cells spatially, rejection, and inflammation of biomaterials used, the recent tissue engineering studies focus on scaffold-free techniques. In this paper, the development of novel computer aided algorithms and methods are developed for 3D bioprinting of scaffold-free biomimetic macrovascular structures. Computer model mimicking a real human aorta is generated using imaging techniques and the proposed computational algorithms. An optimized three-dimensional bioprinting path planning are developed with the proposed self-supported model. Mouse embryonic fibroblast (MEF) cell aggregates and support structures (hydrogels) are 3D bioprinted layer-by-layer according to the proposed self-supported method to form an aortic tissue construct.

The treatment of segmental bone defects in rabbit tibiae with vascular endothelial growth factor (VEGF)-loaded gelatin/hydroxyapatite "cryogel" scaffold.

The aim of this study was to investigate the effectiveness of a novel hydroxyapatite containing gelatin scaffold--with and without local vascular endothelial growth factor (VEGF) administration--as the synthetic graft material in treatment of critical-sized bone defects. An experimental nonunion model was established by creating critical-sized (10 mm. in length) bone defects in the proximal tibiae of 30 skeletally mature New Zealand white rabbits. Following tibial intramedullary fixation, the rabbits were grouped into three: The defects were left empty in the first (control) group, the defects were grafted with synthetic scaffolds in the second group, and synthetic scaffolds loaded with VEGF were administered at bone defects in the third group. Five rabbits in each group were killed on 6th and 12th weeks, and new bone growth was assessed radiologically, histologically and with dual-energy X-ray absorptiometry (DEXA). At 6 weeks, VEGF-administered group had significantly better scores than the other two groups. The second group also had significantly better scores than the control group. At 12 weeks, while no significant difference was noted between the second and third groups, these two groups both had significantly better scores in all criteria compared with the control group. There were no signs of complete fracture healing in the control group. The administration of hydroxyapatite containing gelatin scaffold yielded favorable results in grafting the critical-sized bone defects in this experimental model. The local administration of VEGF on the graft had a positive effect in the early phase of fracture healing.

Gelatin cryogels crosslinked with oxidized dextran and containing freshly formed hydroxyapatite as potential bone tissue-engineering scaffolds.

Gelatin-based cryogels were prepared by using a novel crosslinker, oxidized dextran, which was synthesized and used in the study. The cryogels were also loaded with freshly formed hydroxyapatite (HA) particles. These cryogels are opaque, spongy and highly elastic and have a pore structure with large interconnected pores. They swell about 500% in aqueous media and within a few minutes reach their final swollen forms. The elastic moduli of HA-containing cryogels were 18.5 ± 3.0 kPa, which is suitable for non-load-bearing bone tissue-engineering (TE) applications, especially for the craniofacial area.

Enzyme-catalyzed crosslinking in a partly frozen state: a new way to produce supermacroporous protein structures.

In this study a new way to produce supermacroporous protein structures was investigated. Enzyme-mediated crosslinking of gelatin or casein was performed in a partly frozen state, which yielded stable, protein-based cryogels. The reaction kinetics for the formation of cryogels were found to be fairly slow, most likely due to the low temperature (-12 °C) used or due to an increased viscosity owing to the cryo-concentration taking place. The produced cryogels were characterized with regards to their physical properties and in vitro degradation. Furthermore, cryogels produced from gelatin and casein were evaluated as potential scaffolds by fibroblast cultivation to confirm their in vitro biocompatibility. Gelatin- and casein-based scaffolds both supported cell proliferation and migration through the scaffold.