The loss or failure of an organ or tissue resulting from injury or other type of damage is one of the most recurrent, devastating and costly problems in human health care. Tissue or organ transplantation is a golden standard therapy to treat these patients (Brasile et al., 2002). However, it is severely limited by a critical donor shortage as evident from the data recorded by organ transplantation registries, such as Indian Transplant Registry (India), United Network for Organ Sharing (USA), Eurotransplant (Austria, Belgium, Croatia, Germany, Luxembourg, the Netherlands and Slovenia), Global Observatory on Donation and Transplantation (Spanish National Transplant Organization in association with World Health Organization), etc. (Figure 1.1). The organ shortage, virtually a universal problem, is worsening every year, and a large number of patients are dying while waiting for needed organs. Other approaches followed by physicians to treat organ or tissue loss are surgical reconstruction and use of mechanical / artificial intra/extracorporeal devices. However, the long-term problems limit the use of surgical reconstruction therapy. While, the devices are unsuccessful in performing all the functions of an organ, thus, they fail to prevent progressive patient deterioration (Langer and Vacanti, 1993). Overall, although these conventional therapies have saved and improved countless lives, they still remain imperfect solutions.
The persistent shortage of donor transplants coupled with high morbidity and mortality without transplantation, and limitations associated with other conventional therapies has spurred the development of tissue engineering (TE) and regenerative medicine (RM) as a promising alternative therapy for tissue / organ failure (Fuchs et al., 2001). TERM is an interdisciplinary field that applies the principles of engineering and the life sciences toward the development of biological substitutes that restore, maintain or improve tissue function (Langer and Vacanti, 1993; Langer, 2000). TE and RM are considered as two sides of the same coin, however, TE often focuses on in vitro engineering of tissues using a combination of materials, cells, biochemical/biomechanical cues and bioreactors, while the RM stretches beyond the definition of TE and often aims to stimulate and support the body’s own self-healing capacity by several converging technological approaches including the use of soluble molecules, gene therapy, stem and progenitor cell therapy, tissue engineering and the reprogramming of cell and tissue types (Daar and Greenwood, 2007; Williams, 2006).
Figure 1.1. Screenshots of Indian Transplant Registry, United Network for Organ Sharing, Eurotranplant, and Global Observatory on Donation and Transplantation websites that maintain the statistics of organ transplantations (Accessed on Dec, 2011).
TERM adopts following three strategies for the creation of new tissues (Langer and Vacanti, 1993):
a) Cell based therapy using isolated cells or cell substitutes: This approach allows replacement of only those cells that supply the needed function, and permits manipulation of cells before infusion. Though it avoids surgery related complications, it is often associated with severe immunological rejection and failure to proper maintenance of functions in the recipient. Also, methods to deliver the cells to the target may be required.
b) In situ regeneration using tissue-inducing substances: This approach involves the use of appropriate signal molecules, such as growth factors for in situ regeneration of cells/tissues. It avoids the complications of surgery and immunological rejection.
However, it greatly depends on availability of sufficient amounts of pure signal molecules and methods to deliver these molecules to their targets. In situations where the regeneration process involves multi-step and multi-component pathways, this approach becomes complex and difficult to adopt.
c) In vitro engineering using matrices: This approach involves the in vitro culture of cells on or within three-dimensional (3D) artificial extracellular matrices (ECM), well known as scaffolds, to form a functional tissue. Such in vitro engineered tissues may be implanted directly into the patient body or be used in developing bioartificial extracorporeal devices.
Amongst these three approaches, however, the in vitro engineering using artificial ECM matrices has gained much significance as it offers control over every aspect of the process and promises to provide biomimetic functional artificial tissue. And, the use of term ‘TE’ in recent times often refers to the scaffold mediated in vitro engineering of tissues. The major components of the TE are (a) cells, (b) instructive signals (biochemical, biomechanical, etc.), (c) bioreactors and (d) scaffolds (Figure 1.2) (Langer, 2000). And the process involves (a) identification, isolation and production (in sufficient numbers) of appropriate cells, (b) isolation or synthesis of appropriate biocompatible material(s) and manufacture of scaffold with desired shape and dimensions, (c) culture of cells on or within scaffold in a bioreactor under controlled conditions along with appropriate instructive signals, and lastly, (d) the in vitro engineered cell-scaffold construct is placed into the appropriate in vivo site (Figure 1.3) (Langer, 2000).
Figure 1.2. A schematic of different components of tissue engineering.
Figure 1.3. A schematic of the process of tissue engineering.
1.1.1 Cells
TE strategies utilize living cells and thus availability of sufficient amount of cells is obviously a critically important issue. Autologous healthy somatic cells derived from the patient would be the ideal source; but, biopsies from patients with extensive end-stage organ failure may not yield enough normal cells, moreover, this is often restricted by the limitations associated with the primary cell culture strategies (Atala, 2006). However, with the continuous developments in human stem cell biology, the use of stem cells in TE has started and gained momentum in recent times (Koch et al., 2009). The high proliferative capacity and pluripotency or ability to differentiate into cells of multiple lineages makes stem cells attractive for deriving large cell quantities. Human embryonic stem cells (derived from discarded human embryos), adult stem cells and progenitor cells (derived from a variety of adult tissues) are envisioned as being potential alternative sources (Berthiaume et al., 2011). More recently, the induced pluripotent stem (iPS) cells that are derived by reprogramming of somatic cells represent potential components of TERM (Badylak and Nerem, 2010).
1.1.2 Biochemical and biomechanical cues
Significant breakthroughs in the field of developmental biology and mechanobiology have started strengthening the current understanding of how cells read and adopt native ECM. It is revealed that the cells are extremely sensitive to the native ECM composition and microenvironment, and can be switched between different functional states (i.e.
growth versus differentiation) by altering biochemical stimuli, or by changing the biomechanical balance and thereby altering cell shape and cytoskeletal structure (Badylak, 2005; Ghosh and Ingber, 2007; DuFort and Paszek, 2011). Thus, to engineer a tissue in vitro, the scaffold should ideally mimic the features of native ECM. Instead of using the scaffold as a bio-inert temporary template, the scaffold must incorporate such biochemical and micromechanical signals. In recent times, several studies have been reported on the development of bioactive scaffolds which are modified by coupling instructive biochemical signals (ex. adhesive ligands, growth factors, etc.) on the surface or the bulk of the matrix (Chung and Park, 2007). While, various scaffold design configuration and bioreactor systems are being used to micromechanical signaling between cells and scaffold (Freed et al., 2006; Ingber et al., 2006).
1.1.3 Scaffold
Besides cells and corresponding cues, the major and the most critical component of tissue engineering is the ‘scaffold’, made of biomaterials, that tries to mimic the natural ECM and acts as a template for in vitro culture of human cells (Hutmacher and Cool, 2007).
The surface and bulk properties of the materials used, and the architecture of the scaffold are central to its success (Liu et al., 2007; Sachlos and Czernuszka, 2003). A wide variety of scaffolds have been developed and evaluated for use in TE. The initial TE studies have used the scaffold as a mere bio-inert template; however, as researchers develop a greater understanding of the biology underlying fundamental structure-function relationships, a mere bio-inert scaffold may not yield a tissue with desired functions (Chan and Leong, 2008; Williams, 2008). Several signals from the in vivo microenvironment such as cell–
cell interactions, cell – ECM interactions, soluble signals and mechanical forces are found to influence the cell behavior in a 3D context (Tsang and Bhatia, 2004). Thus, in recent times, the factors that influence cell fate (proliferation, differentiation, apoptosis) and function (migration, gene expression, morphogenesis) are being incorporated with the TE scaffold to generate a functional tissue (Grayson et al., 2009; Biondi et al., 2008; Ghosh and Ingber, 2007).
1.1.4 Bioreactors
Although significant progress is made in the isolation and culture of cells and fabrication of scaffolds, the success of TE is hindered by the complexity of the process. Bioreactors are devices that allow operation of bio-processes under closely monitored and tightly controlled conditions and offers automation and scale-up of the process with a high degree of reproducibility (Plunkett and O'Brien, 2011). By providing a comprehensive level of monitoring and control over specific environmental factors (including biochemical and biomechanical) in 3D cultures, bioreactors can provide the technological means to understand the parameter-function relationships, and subsequently to engineer a functional tissue (Ratcliffe and Niklason, 2002; Martin et al., 2004; Pörtner et al., 2005).
However, besides the generation of grafts for implantation purposes, the bioreactors are playing an important role in the form of extracorporeal devices, such as bioartificial liver and kidney, to sustain in the life of a patient till a transplant is made possible (Iwahori et al., 2005).