**1. Introduction**

34 Biomedicine

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xenotransplantation. *Biomaterials*, Vol.26, No.4, pp.403-412, ISSN 0142-9612 Yamamoto, T., Mita, A., Ricordi, C., Messinger, S., Miki, A., Sakuma, Y., Timoneri, F.,

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in vitro and in vivo function of three different shaped bioartificial pancreases made

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nanoencapsulation of insulin-producing beta-cells grown as pseudoislets for potential cellular delivery of insulin. *Biomacromolecules*, Vol.11, No.3, pp.610-616,

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Brunnenmeier, F., Zimmermann, H., Westphal, I., Fuhr, G., Nöth, U., Haase, A., Steinert, A., & Hendrich, C. (2001). A novel class of amitogenic alginate microcapsulesfor long-term immunoisolated transplantation. *Ann. N. Y. Acad. Sci*., Tissue loss or organ failure caused by injury or damage is one of the most serious and costly problems in human health care. Tissue engineering, proposed by Langer *et al.* in the early 1990's (Langer & Vacanti, 1993), is an emerging strategy of regenerative biomedicine that holds promise for the restoration of defect tissues and organs. The concept of tissue engineering is defined as "the application of the principles and methods of engineering and the life sciences towards the fundamental understanding of structure-function relationships in normal and pathological mammalian tissues and the development of biological substitutes that restore, maintain or improve tissue function" (Langer & Vacanti, 1993). In order to accomplish these goals by tissue engineering, three essential components are required, that is, cells for the generation of new tissues, scaffolds for supporting the cell growth and the regeneration of new tissues, and bioactive factors capable of stimulating biological signals *in vivo* for cell proliferation, dfferentiation and tissue growth. Among these, the scaffolds play an important role in the success of tissue regeneration since they serve as temporary temples to mimick the excellular matrix for cell growth and interim mechanical stability for tissue regneration and integration,.

Hydrogels are one of most used bio-scaffolds in the field of tissue enginereering. They are three-dimensional, water-swollen, crosslinked networks of hydrophilic polymers. Wichterle and Lim for the first time reported on hydrogels based on the hydroxyethyl methacrylate (HEMA) for biological use in 1960 (Wichterle & Lim, 1960). Due to their unique tissue-like properties, such as high water content and good permeability to oxygen and metabolites, hydrogels have been widely studied as biomimetic extracellular matrixes for tissue regeneration. Hydrogels may be used by implantation or injection, which corresponds to socalled preformed hydrogels or in-situ forming hydrogels. From the clinical point of view, insitu forming hydrogels are highly desirable since they gain advantages over preformed hydrogels: (1) Enabling minimally invasive surgeries for implantation; (2) Formation in any desired shape in good alignment with surrounding tissue defects; (3) Easy encapsulation of bioactive molecules and progenic cells. Therefore, in-situ forming hydrogels have received much attention in recent years.

In-Situ Forming Biomimetic Hydrogels for Tissue Regeneration 37

incorporated to yield biofunctional or bioreponsive hydrogels, enhancing cell adhesion and matrix production (Seliktar et al., 2004). This indicates that Michael-type addition is an ideal

Radical polymerization is one of the most frequently used crosslinking methods to prepare robust and stable in-situ forming hydrogels. Radicals are created from initiator molecules through thermal, redox or photointiated mechanisms. Then, the radicals propagate through unreacted double bonds during polymerization to form long kinetic chains, and the chains react further with each other to form crosslinked polymeric networks (Ifkovits & Burdick, 2007). In general, macromers bearing vinyl groups are relatively biocompatible and more favourable as compared to monomers. Since the reaction takes place in aqueous solutions, the conversion of double bonds is high due to the high mobility of reacting species during gel formation. This also decreases the potential toxicity of the materials. PEG and PEG-based copolymers are commonly used synthetic biomaterials (Nguyen & West, 2002) (Fig. 2). They can be functionalized with acryl chloride and further crosslinked in the presence of freeradical initiators under a physiological environment to form hydrogels. Multi-arm polymers such as 4-arm and 8-arm PEG were also employed to increase the crosslinking density because of their increased functionality as compare to linear analogues. Other types of polymers are natural polymers such as dextran, hyaluronic acid and collagen (Dong et al., 2005; S.H. Kim et al., 1999; Y.D. Park et al., 2003). As compared to synthetic polymers, they have different functional groups (hydroxyl, amine or carboxylic groups) on their polymer backbones and are amenable to various chemical modifications. The number of double bonds introduced can be precisely controlled on demand. Thus, the properties of freeradical polymerized hydrogel such as gelation time, mechanical properties and degradation

Fig. 2. Commonly used poly(ethylene glycol)-based polymers for in-situ forming hydrogels

In-situ hydrogels formation using enzymes have emerged recently. Enzymes are known to exhibit a high degree of substrate specificity, which potentially avoids side reactions during crosslinking. Another advantage of the enzymatic crosslinking is of mild gelation conditions

method for the preparation of in-situ crosslinked hybrid hydrogels.

profiles can be adjusted for use in different tissue engineering.

(e.g. physiological conditions), favourable for tissue regeneration.

**2.1.2 Radical polymerization** 

**2.1.3 Enzymatic crosslinking** 
