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Tissue engineering can be broadly defined as the combination of biology and engineering to repair or replace lost tissue function. From an industry perspective, the field encompasses implanted biomaterials, cell and tissue transplants and therapies, and even extracorporeal cellular devices. To achieve its goals, tissue engineering must effectively utilize not only multiple aspects of engineering but also several aspects of biology that govern mechanisms of organ development, repair and regeneration. The field has always had a strong focus on application yet the challenge of integrating biological science, engineering and medicine has kept many past efforts from reaching their therapeutic and commercial potential. This chapter covers the evolution of tissue engineering, looking at the change in emphasis from bioengineering to stem cell biology and the potential impact of this shift in focus from an industrial perspective. In addition, we have analyzed four major commercial thrusts from past to present: vascular tissue engineering, cartilage repair, liver-assist devices and skin constructs, paying particular attention to how the biomedical disciplines must be integrated to achieve commercial feasibility and therapeutic success. Each example yields one or more important and practical lessons learnt that could be instructive for most future medical and commercial efforts in tissue engineering.

In vivo animal models are currently the gold standard for testing the capacity of stem/progenitor cells, smart biomaterials and novel growth factors for successful tissue engineering. In vitro models ultimately fail to provide the appropriate physiologically relevant microenvironment and hence animal models are an essential pre-requisite in the translation of any new therapy to the clinic. The aim of this chapter is to consider the available animal models commonly in use for tissue engineering, with a particular focus upon bone and cartilage research. Factors driving the choice of a given animal model are reviewed, according to the requirements of experimental design, hypothesis and the specific parameters to be tested. A number of animal models, together with their respective advantages and limitations are described, ranging from relatively simple experimental designs such as the subcutaneous implant and muscle pouch models, through to the diffusion chamber model and chorioallantoic membrane assay, to the more complex in vivo bioreactors and (arguably the most clinically relevant) bone and cartilage defect models. The need to consider the ethical issues of using animal models and the principles of reduction, replacement and refinement are emphasised in selecting the final experimental model of choice.

No abstract received.

The most exciting discoveries in the last two decades have been the mapping, sequencing, and understanding of genes in our body. In this chapter, we will discuss how to specifically map expression profiles of gene products — mRNA, protein, and reporter gene (X-gal) — using in situ hybridization, immunohistochemistry, and X-gal staining, respectively.

Cartilage damage is irreversible due to its properties of avascularity and lack of undifferentiated cells. Studying the biology of chondrocytes and cartilage is therefore critical to understand the underlying mechanism and to explore the potential repair approaches. It has been reported that three-dimensional (3D) chondrocyte culture behaves very differently from two-dimensional (2D) monolayer culture. Therefore, a native 3D culture of chondrocytes is valuable for such a research purpose, and may also be an option for repairing cartilage defects. This chapter describes a detailed protocol for high-yield chondrocyte isolation and the modified pellet culture technique (using a 3D chondrocyte culture), which can synthesize a bioengineered tissue up to 8 mm in diameter. The authors aim at providing a platform for researchers to study chondrocyte behavior in a 3D environment and to explore the application of scaffold-free tissue for transplantation into a large cartilage defect model.

Perhaps the most difficult task in bone histology work is the preparation of a good specimen. It is impossible to obtain a good image of bone or cartilage without careful preparation and processing of specimens. This chapter will describe very basic skills in the preparation of specimens. These techniques include the paraffin method, plastic method, and frozen method.

Bone or cartilage cells are essentially colorless, and it is difficult to distinguish their morphologies under a light microscope. To better visualize these structures, various staining techniques have been developed; due to page constraints, we will only describe the most common methods for readers. These staining assays include hematoxylin and eosin (H&E) stain, used for general histology of the cell; Safranin O stain, used for the cartilage; and von Kossa and van Gieson stains as well as modified Goldner's trichrome stain, used for nondecalcified tissues.

Attrition and eventual loss of articular cartilage are crucial elements in the pathophysiology of osteoarthritis. Preventing the breakdown of cartilage is believed to be critical in order to preserve the functional integrity of a joint. Magnetic resonance imaging (MRI) and advanced digital postprocessing techniques have opened novel possibilities for in vivo quantitative analysis of cartilage morphology, structure, and function in health and disease. Techniques of semiquantitative scoring of human knee cartilage pathology and quantitative assessment of human cartilage have recently been developed. Though cartilage represents a thin layer of material relative to the size of voxels typically used for MRI, cartilage thickness and volume have been quantified in human and in small animals. MRI-detected cartilage loss has been shown to be more sensitive than radiography-detected joint space narrowing. Progress made in MRI technology in the last few years allows longitudinal studies of knee cartilage with an accuracy good enough to follow disease-caused changes and to evaluate the therapeutic effects of chondroprotective drugs.

The etiology of most of the degenerative changes in the spine continues to remain obscure. However, several lines of evidence suggest that genetic factors may play an important role in the onset of degenerative changes, in addition to various environmental factors. We have generated transgenic mice expressing mutant αl(IX) collagen in the cartilage matrix. They developed progressive intervertebral disc degeneration with age as well as joint degeneration. Both radiologic and histologic studies indicated that cervical and lumbar disc degeneration was more advanced in the transgenic mice than in control littermates. The initial degenerative changes included shrinkage and replacement of the nucleus pulposus with consolidated fibrous tissue, that resulted in a loss of nuclear-anular demarcation. Partial disruption in the lamellar structure of the anulus fibrosus also occurred at this stage. With age, the disc degeneration progressively advanced and sometimes caused herniation of disc material and mild osteophyte formation. These findings imply that genetic abnormalities of cartilage matrix components, such as type IX collagen, may be responsible for certain degenerative diseases in the spine.

Articular cartilage is hyaline cartilage which has very limited self-repairingcapacity after its degeneration or injury. Recently, three-dimensional (3D) bioprinting provides a promising method for repair and regeneration of articular cartilage. Significant progress has been made in 3D bioprinting for cartilage regeneration, particularly in printing hydrogels in combination of cells and growth factors. In this chapter, we reviewed recent progress in cartilage 3D bioprinting, including the use of various cell sources and growth factors for cartilage formation. We also discussed the challenges and the future research directions of cartilage regeneration.

Hydroxyapatite is the well known bioactive ceramic used in medical and dental fields as blocks and particles, while chondroitin-sulfate is a dominant polysaccharide component of cartilage. Novel composite materials consisting of hydroxyapatite [Ca₁₀(PO₄)₆(OH)₂] and chondroitin-sulfate were synthesized by a coprecipitation method with a H₃PO₄ solution and a Ca(OH)₂ suspension, one of which contain chondroitin-sulfate, and were consolidated under a cold isostatic pressure. The composites were evaluated by X-ray diffractometry, Fourier transformed infrared spectroscopy and transmission electron microscopy. In the composites, hydroxyapatite nanocrystals contained calcium-deficiency and a small amount of carbonate ions like human bone minerals and their c-axes were aligned along chondroitin-sulfate molecules by a self-assembly mechanism.

Implants (hemitrochlea) were prepared from nacre (mother of pearl) and implanted in the knees of sheep. Cartilage formed at the endoarticular surfaces of the implant, and new endochondral bone was also formed at the interface between the host cancellous bone and nacre. These phenomena resulted in the total integration of the implant.