Commenced in January 2007
Frequency: Monthly
Edition: International
Paper Count: 5

Search results for: decellularization

5 Prospects of Low Immune Response Transplants Based on Acellular Organ Scaffolds

Authors: Inna Kornienko, Svetlana Guryeva, Anatoly Shekhter, Elena Petersen

Abstract:

Transplantation is an effective treatment option for patients suffering from different end-stage diseases. However, it is plagued by a constant shortage of donor organs and the subsequent need of a lifelong immunosuppressive therapy for the patient. Currently some researchers look towards using of pig organs to replace human organs for transplantation since the matrix derived from porcine organs is a convenient substitute for the human matrix. As an initial step to create a new ex vivo tissue engineered model, optimized protocols have been created to obtain organ-specific acellular matrices and evaluated their potential as tissue engineered scaffolds for culture of normal cells and tumor cell lines. These protocols include decellularization by perfusion in a bioreactor system and immersion-agitation on an orbital shaker with use of various detergents (SDS, Triton X-100) and freezing. Complete decellularization – in terms of residual DNA amount – is an important predictor of probability of immune rejection of materials of natural origin. However, the signs of cellular material may still remain within the matrix even after harsh decellularization protocols. In this regard, the matrices obtained from tissues of low-immunogenic pigs with α3Galactosyl-tranferase gene knock out (GalT-KO) may be a promising alternative to native animal sources. The research included a study of induced effect of frozen and fresh fragments of GalT-KO skin on healing of full-thickness plane wounds in 80 rats. Commercially available wound dressings (Ksenoderm, Hyamatrix and Alloderm) as well as allogenic skin were used as a positive control and untreated wounds were analyzed as a negative control. The results were evaluated on the 4th day after grafting, which corresponds to the time of start of normal wound epithelization. It has been shown that a non-specific immune response in models treated with GalT-Ko pig skin was milder than in all the control groups. Research has been performed to measure technical skin characteristics: stiffness and elasticity properties, corneometry, tevametry, and cutometry. These metrics enabled the evaluation of hydratation level, corneous layer husking level, as well as skin elasticity and micro- and macro-landscape. These preliminary data may contribute to development of personalized transplantable organs from GalT-Ko pigs with significantly limited potential of immune rejection. By applying growth factors to a decellularized skin sample it is possible to achieve various regenerative effects based on the particular situation. In this particular research BMP2 and Heparin-binding EGF-like growth factor have been used. Ideally, a bioengineered organ must be biocompatible, non-immunogenic and support cell growth. Porcine organs are attractive for xenotransplantation if severe immunologic concerns can be bypassed. The results indicate that genetically modified pig tissues with knock-outed α3Galactosyl-tranferase gene may be used for production of low-immunogenic matrix suitable for transplantation.

Keywords: decellularization, low-immunogenic, matrix, scaffolds, transplants

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4 Prospects of Acellular Organ Scaffolds for Drug Discovery

Authors: Inna Kornienko, Svetlana Guryeva, Natalia Danilova, Elena Petersen

Abstract:

Drug toxicity often goes undetected until clinical trials, the most expensive and dangerous phase of drug development. Both human cell culture and animal studies have limitations that cannot be overcome by improvements in drug testing protocols. Tissue engineering is an emerging alternative approach to creating models of human malignant tumors for experimental oncology, personalized medicine, and drug discovery studies. This new generation of bioengineered tumors provides an opportunity to control and explore the role of every component of the model system including cell populations, supportive scaffolds, and signaling molecules. An area that could greatly benefit from these models is cancer research. Recent advances in tissue engineering demonstrated that decellularized tissue is an excellent scaffold for tissue engineering. Decellularization of donor organs such as heart, liver, and lung can provide an acellular, naturally occurring three-dimensional biologic scaffold material that can then be seeded with selected cell populations. Preliminary studies in animal models have provided encouraging results for the proof of concept. Decellularized Organs preserve organ microenvironment, which is critical for cancer metastasis. Utilizing 3D tumor models results greater proximity of cell culture morphological characteristics in a model to its in vivo counterpart, allows more accurate simulation of the processes within a functioning tumor and its pathogenesis. 3D models allow study of migration processes and cell proliferation with higher reliability as well. Moreover, cancer cells in a 3D model bear closer resemblance to living conditions in terms of gene expression, cell surface receptor expression, and signaling. 2D cell monolayers do not provide the geometrical and mechanical cues of tissues in vivo and are, therefore, not suitable to accurately predict the responses of living organisms. 3D models can provide several levels of complexity from simple monocultures of cancer cell lines in liquid environment comprised of oxygen and nutrient gradients and cell-cell interaction to more advanced models, which include co-culturing with other cell types, such as endothelial and immune cells. Following this reasoning, spheroids cultivated from one or multiple patient-derived cell lines can be utilized to seed the matrix rather than monolayer cells. This approach furthers the progress towards personalized medicine. As an initial step to create a new ex vivo tissue engineered model of a cancer tumor, optimized protocols have been designed to obtain organ-specific acellular matrices and evaluate their potential as tissue engineered scaffolds for cultures of normal and tumor cells. Decellularized biomatrix was prepared from animals’ kidneys, urethra, lungs, heart, and liver by two decellularization methods: perfusion in a bioreactor system and immersion-agitation on an orbital shaker with the use of various detergents (SDS, Triton X-100) in different concentrations and freezing. Acellular scaffolds and tissue engineered constructs have been characterized and compared using morphological methods. Models using decellularized matrix have certain advantages, such as maintaining native extracellular matrix properties and biomimetic microenvironment for cancer cells; compatibility with multiple cell types for cell culture and drug screening; utilization to culture patient-derived cells in vitro to evaluate different anticancer therapeutics for developing personalized medicines.

Keywords: 3D models, decellularization, drug discovery, drug toxicity, scaffolds, spheroids, tissue engineering

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3 Decellularized Brain-Chitosan Scaffold for Neural Tissue Engineering

Authors: Yun-An Chen, Hung-Jun Lin, Tai-Horng Young, Der-Zen Liu

Abstract:

Decellularized brain extracellular matrix had been shown that it has the ability to influence on cell proliferation, differentiation and associated cell phenotype. However, this scaffold is thought to have poor mechanical properties and rapid degradation, it is hard for cell recellularization. In this study, we used decellularized brain extracellular matrix combined with chitosan, which is naturally occurring polysaccharide and non-cytotoxic polymer, forming a 3-D scaffold for neural stem/precursor cells (NSPCs) regeneration. HE staining and DAPI fluorescence staining confirmed decellularized process could effectively vanish the cellular components from the brain. GAGs and collagen I, collagen IV were be showed a great preservation by Alcain staining and immunofluorescence staining respectively. Decellularized brain extracellular matrix was well mixed in chitosan to form a 3-D scaffold (DB-C scaffold). The pore size was approximately 50±10 μm examined by SEM images. Alamar blue results demonstrated NSPCs had great proliferation ability in DB-C scaffold. NSPCs that were cultured in this complex scaffold differentiated into neurons and astrocytes, as reveled by NSPCs expression of microtubule-associated protein 2 (MAP2) and glial fibrillary acidic protein (GFAP). In conclusion, DB-C scaffold may provide bioinformatics cues for NSPCs generation and aid for CNS injury functional recovery applications.

Keywords: brain, decellularization, chitosan, scaffold, neural stem/precursor cells

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2 Vascularized Adipose Tissue Engineering by Using Adipose ECM/Fibroin Hydrogel

Authors: Alisan Kayabolen, Dilek Keskin, Ferit Avcu, Andac Aykan, Fatih Zor, Aysen Tezcaner

Abstract:

Adipose tissue engineering is a promising field for regeneration of soft tissue defects. However, only very thin implants can be used in vivo since vascularization is still a problem for thick implants. Another problem is finding a biocompatible scaffold with good mechanical properties. In this study, the aim is to develop a thick vascularized adipose tissue that will integrate with the host, and perform its in vitro and in vivo characterizations. For this purpose, a hydrogel of decellularized adipose tissue (DAT) and fibroin was produced, and both endothelial cells and adipocytes that were differentiated from adipose derived stem cells were encapsulated in this hydrogel. Mixing DAT with fibroin allowed rapid gel formation by vortexing. It also provided to adjust mechanical strength by changing fibroin to DAT ratio. Based on compression tests, gels of DAT/fibroin ratio with similar mechanical properties to adipose tissue was selected for cell culture experiments. In vitro characterizations showed that DAT is not cytotoxic; on the contrary, it has many natural ECM components which provide biocompatibility and bioactivity. Subcutaneous implantation of hydrogels resulted with no immunogenic reaction or infection. Moreover, localized empty hydrogels gelled successfully around host vessel with required shape. Implantations of cell encapsulated hydrogels and histological analyses are under study. It is expected that endothelial cells inside the hydrogel will form a capillary network and they will bind to the host vessel passing through hydrogel.

Keywords: adipose tissue engineering, decellularization, encapsulation, hydrogel, vascularization

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1 Biodegradable and Bioactive Scaffold for Bone Tissue Engineering

Authors: A. M. Malagon Escandon, J. A. Arenas Alatorre, C. P. Chaires Rosas, N. A. Vazquez Torres, B. Hernandez Tellez, G. Pinon Zarate, M. Herrera Enriquez, A. E. Castell Rodriguez

Abstract:

The current approach to the treatment of bone defects involves the use of scaffolds that provide a biological and mechanically stable niche to favor tissue repair. Despite the significant progress in the field of bone tissue engineering, several main problems associated are attributed to giving a low biodegradation degree, does not promote osseointegration and regeneration, if the bone is not healing as well as expected or fails to heal, will not be given a proper ossification or new bone formation. The actual approaches of bone tissue regeneration are directed to the use of decellularized native extracellular matrices, which are able of retain their own architecture, mechanic properties, biodegradability and promote new bone formation because they are capable of conserving proteins and other factors that are founded in physiological concentrations. Therefore, we propose an extracellular matrix-based bioscaffolds derived from bovine cancellous bone, which is processed by decellularization, demineralization, and hydrolysis of the collagen protein, these protocols have been successfully carried out in other organs and tissues; the effectiveness of its biosafety has also been previously evaluated in vivo and Food and Drug Administration (FDA) approved. In the specific case of bone, a more complex treatment is needed in comparison with other organs and tissues because is necessary demineralization and collagen denaturalization. The present work was made in order to obtain a temporal scaffold that succeed in degradation in an inversely proportional way to the synthesis of extracellular matrix and the maturation of the bone by the cells of the host.

Keywords: bioactive, biodegradable, bone, extracellular matrix-based bioscaffolds, stem cells, tissue engineering

Procedia PDF Downloads 39