**4. Biomaterials for scaffold production**

Biomaterials for tissue engineering have a considerable importance for the success of a tissue replacement or regeneration. In addition to interacting with the implant site, they have the ability to influence biological processes that are important for tissue regeneration.

Different kinds of biomaterials have been used for scaffold production such as ceramics and polymers, naturals and synthetics, metals, composites, and hydrogels.

#### **4.1 Ceramics**

For several decades, ceramic biomaterials have been used to reconstruct damaged body parts and for skeletal repair.

Ceramic biomaterials are inorganic compounds of natural or synthetic origin, which may contain metallic and nonmetallic elements. These biomaterials are generally made of polycrystalline solids, rarely of monocrystals and sometimes have an amorphous structure. Generally, their mechanical properties, including hard surface, high mechanical stiffness, low elasticity, low thermal expansion, chemicalphysic refractoriness, depend on the way they are produced or extracted, but their properties can also depend on the composition and particle size of the starting powders.

Ceramic scaffolds are commonly used for bone regeneration practices because they are highly biocompatible, rarely evoke an immune response, and hardly cause the formation of fibrous tissue around the scaffold; instead they are osteoinductive, considering their high ability to recruit cells from the biological environment and promote osteogenic differentiation. Although the ceramics present these advantages, their use in tissue engineering applications is limited due to their fragility and slow degradation [1, 13, 14].

On the basis of their main features, they can be distinguished into three categories: (a) bio-inert ceramics: completely inert to biological environment; (b) resorbable materials: subjects to *in vivo* degradation for phagocytosis or dissolution of the material in biological fluids; and (c) bioactive ceramics: able to form chemical bound with the cell surface [15].

The most common ceramic biomaterials used for tissue regeneration are: (1) CaP, including hydroxyapatite (HA) (Ca10[PO4]6[OH]2), beta-tricalcium phosphate (BTF) (Ca3[PO4]2), biphasic calcium phosphate (mixture of hydroxyapatite and beta-tricalcium phosphate), (2) bioglass, (3) alumina (Al2O3), and (4) zirconia oxide (ZrO2).

CaP biomaterials are often selected for bone graft since they mimic bone tissue composition. One of the first used ceramic biomaterials for skeletal repair was BTF in 1920 by Albee and Morrinson [16].

HA may be natural or synthetic. Natural HA derives from particular species of coral or bovine bone and can contain traces of other elements such as Mg, Na,

#### *Innovative Biomaterials for Tissue Engineering DOI: http://dx.doi.org/10.5772/intechopen.83839*

CO3, and F. Synthetic HA is prepared by sintering in dense or macroporous form as granules or blocks [17]. Ray and Ward, first, showed the high biocompatibility and biomimicry of synthetic HA in their study in which they used this material for bone tissue engineering application in the long bones and iliac wings of dogs [18]. Later, numerous other studies on HA have been carried out. Calabrese *et al.* in their studies tested a composite bi-layer type-1 collagen-HA/Mg scaffold for osteochondral regeneration, both *in vitro* and *in vivo*. They showed that the combination of this scaffold with mesenchymal stem cells (MSC) derived from adipose tissue (hAD-SCs) in the presence of specific differentiation conditions induce osteochondro differentiation both *in vitro* and *in vivo* [19–23].

Bioglass is composed by 45 wt% SiO2, 24.5 wt% CaO, 24.5 wt% Na2O, and 6.0 wt% P2O5. The first one (45S5 Bioglass) has been developed by Hench*,* which used it for biomedical applications *in vitro* and *in vivo* [24].

Bioglass materials can be synthesized through different methods such as polymer foam replication, thermal bonding of particles or fibers, and sol-gel processing. Similarly to HA, it is suitable for bone graft due to the high ratio of calcium to phosphorus promoting the formation of apatite crystals on its surface after grafting. Bioglass materials offer high osteoinductivity, control of rate of degradation, and excellent bioactivity even if they can present poor mechanical properties such as low strength and toughness [25, 26].

Alumina (Al2O3) is a ceramic biomaterial with a crystalline structure. Generally, a low porosity and reduced grain size increase its mechanical strength. Like other ceramic materials, alumina is fragile but it has good tribological properties such as resistance to wear.

Zirconia is characterized by a polymorphic structure and offer has a hard surface, a low thermal conductivity, and a high coefficient of thermal expansion. Its excellent biocompatibility and high breaking load make it a good candidate for prosthesis and bone grafting.

### **4.2 Polymers**

Various biological polymers such as collagen, alginate, proteoglycans, chitin, and chitosan have been used to produce scaffolds. They are biocompatible and bioactive promoting cellular adhesion and growth on their surface. However, they often show poor mechanical properties and fast biodegradability, which limit their use.

Collagen and its derivatives are good candidates for osteochondral regeneration but also tendon and ligament reconstruction since the extracellular matrix of these tissues is mostly made of type-1 collagen fibers [27–29].

Collagen scaffolds are highly bioactive ensuring excellent cellular adhesion to their surface. However, since they have low resistance to mechanical stress often are coupled with other materials, which improve their mechanical properties.

Several studies have been focused on the use of collagen scaffolds for tissue engineering strategies. Aravamudhan *et al.,* for example, reported the fabrication and characterization of cellulose and collagen-based micro-nanostructured scaffolds exhibiting mechanical features similar to those of trabecular bone that promoted good adhesion of human osteoblasts to their surface. Moreover, they underwent a progressive calcium deposition process compared to control polyester micro-nanostructured scaffolds [30].

In another study, Schneider *et al.* developed a collagen I/III hydrogel scaffold and used it to seed hMSC isolated from bone marrow of femoral head spongiosa and from umbilical cord. When stimulated with osteogenic induction medium, both cell types showed comparable osteogenic gene expression, migration, and

scaffold colonization [31]. Collagen scaffolds may also be used to deliver osteogenic differentiation factors as demonstrated by Lu H. *et al,* who immobilized BMP4 in a collagen-PLGA hybrid scaffold to promote osteogenesis [32].

Polysaccharides such as chitin, chitosan, and alginate are suitable for both hard and soft tissue regeneration. In particular, chitosan scaffolds can be manufactured by freeze-drying techniques, which allow obtaining a porous scaffold with high pore interconnectivity. Chitosan ensures good cellular adhesion and thank to its positive charges can interact with glycosaminoglycans and proteoglycans present in living tissues. Costa-Pinto *et al.* cultured human bone marrow MSC on melt-based porous chitosan scaffolds using an osteogenic differentiation medium. They found an increase of cell viability and ALP activity after 21 days. They also investigated the capacity of the cell seeded scaffold to repair a cranial defect in mouse, and 8 weeks after implantation bone formation in the scaffold was analyzed using Bone μCT [33]. Chitosan may also be used as an injectable biomaterial as demonstrated by Bi *et al.,* who produced a composite scaffold of tricalcium phosphate (TCP), chitosan, and platelet rich plasma (PRP). MSC seeded on injectable biomaterial was used in vivo to test its capacity to repair bone fracture in goat femora [34].

Synthetic polymers are high molecular weight compounds composed of a series of monomeric units. On the basis of their structure, they can be linear, branched, or cross-linked. Considering their thermo-mechanical properties, they are thermoplastic or thermosetting. Polymeric materials can be produced in the form of fibers, films, bars, and viscous liquids, and they offer the important advantage to modulate their mechanical properties and biodegradation by varying synthesis process and reactants used. However, they could have low biocompatibility and mechanical strength and show *in vivo* toxicity due to the release of ions and other residual particles of polymerization.

 Among the different synthetic polymers, the most suitable for scaffold production is the bio-erodible. These kinds of polymers undergo surface degradation with production of nontoxic low molecular weight compounds.

Numerous synthetic polymers have already been used such as: polystyrene, thermoplastic aromatic polymer with a linear structure; poly-l-lactic acid (PLA), hydrophobic polymer with slow degradation rate due to microorganisms; polyglycolic acid (PGA), hydrophilic polymer with good mechanical properties and fast degradation; poly-dl-lactic-co-glycolic acid (PLGA), biocompatible copolymers with fast degradation rate; and polycaprolactone (PCL), highly hydrophobic polymer with good permeability.

 In particular, PGA and PLA and their copolymers are natural polyesters normally present in the organism and therefore well tolerated. They have been used for suture threads, orthopedic screws, and prostheses manufacture since 1970, and more recently, they have been evaluated for scaffold production and tissue engineering strategies. About this, Eğri *et al.* combined PLA and PGA to obtain a PLA-PEG-PLA scaffolds able to release VEGF and BMP-2 in bone tissue lesion. In relation to its chemical composition, the scaffold allows fast release of VEGF in about 1 week and slower constant release of BMP-2 [35].

#### **4.3 Metals**

Metals are particularly suitable for tissue engineering strategies for their good mechanical properties such as high elastic module, yield strength, and high ductility allowing them to bear a load without being deformed. If mechanical resistance makes them excellent candidates for scaffold production, however, the reduced cell adhesion to their surface could be a considerable limit to their use. Moreover, metal implants can release toxic metallic ions and/or particles, and biological fluids

#### *Innovative Biomaterials for Tissue Engineering DOI: http://dx.doi.org/10.5772/intechopen.83839*

can show corrosive action on their surface that can alter their function. Among the different metals used for scaffold production, there are stainless steel, cobalt, and titanium alloys.

Stainless steels are iron-based alloys with a low content of carbon and a high content of chromium. The presence of carbon ensures good mechanical properties but determines carbides formation that makes the scaffold subject to corrosion in a biological environment.

Cobalt-based alloys are of two types: cobalt/chromium/molybdenum alloy obtained with casting/melting methods and cobalt/nickel/chromium/molybdenum alloy worked by forging. Generally, the high level of chromium and molybdenum typical of these alloys increase granule size and improve mechanical properties.

Titanium alloys can be *alpha, beta or alpha/beta* biphasic. *Alpha* alloys contain alpha stabilizers such as aluminum and gallium and are characterized by good strength, hardness, resistance sliding, and weld ability; *Beta* alloys contain beta stabilizers such as vanadium, niobium, and tantalus molybdenum and show good ductility. *Alpha/beta* biphasic alloys show a mix of *alpha/beta* stabilizers, and they are quite ductile even if little resistant to high temperatures, and the most suitable one for biomedical application is Ti 6Al 4 V.

Wohlfahrt *et al.* tested the osteoinductivity and osteointegration capabilities of Ti and TiO2 scaffolds in rabbit tibia peri-implant osseous defects. After 4 weeks, the implant was removed and the new bone formation was observed. Moreover, a gene expression analysis was performed considering different osteogenesis differentiation markers such as osteocalcin and collagen-I [36].

In another study, Zuchuat *et al.* developed Cr-Co-Mo membranes and placed them in rabbit tibiae to analyze the volume of new bone formation. After the explant, histological analysis showed a huge number of osteoblasts and osteocytes on the scaffold [37].

### **4.4 Composites**

Composite scaffolds are developed combining different biomaterials such as natural or synthetic polymers (PGA, PLA, gelatin, chitin, and chitosan), ceramics (hydroxyapatite and beta-tricalcium phosphate or bioglasses), and metals. They have technological, industrial, and applicative importance since they combine biocompatibility, biodegradation, and appreciable mechanical strength. Moreover, these kinds of scaffolds could be applied for both hard and soft tissue regeneration and greatly mimic tissue architecture being composed of cells and extracellular matrix.

Several studies displayed the efficacy of composite scaffolds (polymers/ceramics and synthetic/natural polymers) for tissue engineering strategies [38, 39].

Other researchers demonstrated that another interesting solution may be the combination of metallic implants with polymer coating or metal/ceramic scaffolds [40, 41].

#### **4.5 Hydrogels**

Hydrogels are hydrophilic polymers rich of polar moieties such as carboxyl, amide, amino, and hydroxyl groups, held together by chemical bounds or physical intra-molecular and inter-molecular attractions. Their main feature is the ability to absorb enormous amounts of water or biological fluids and swell without dissolving.

According to their origin, hydrogel can be classified into natural (made of polypeptides and polysaccharides), synthetic (obtained by traditional polymerization),


#### **Table 1.**

*Advantages, disadvantages, and main clinical uses of different kinds of biomaterials.* 

 and semi-synthetic. Moreover, they can present an amorphous or semi-crystalline structure that can be cationic, anionic, neutral, or ampholytic. Depending on their stability in a biological system, they can be considered durable if they do not undergo chemical-physical modification or biodegradable if they degrade into oligomers, which are subsequently eliminated from the body. In the last decades, smart hydrogels have been developed featured by the possibility to modify their structure and mechanical properties according to environmental stimuli such as pH or temperature. Already 50 years ago, these materials have been appreciated for their chemicalphysical characteristics by Wichterle and Lim*,* who developed a poly(2-hydroxyethyl methacrylate)-based hydrogel for contact lens [42]. Since they present a soft and rubbery consistency very similar to that of ECM of different tissues, they have been recently studied for tissue engineering strategies. In particular, hydrogels used for scaffold production may respond to important requirements such as biocompatibility and controlled *in vivo* biodegradation. It is very important to modulate parameters such as hydrogel cross-linking density, porosity, pore size, and interconnectivity to obtain a suitable structural for cellular colonization and proliferation. Hydrogels can be modified at the surface by peptides or growth factor, which can promote cell attachment and differentiation process. Generally, natural hydrogels are less toxic

#### *Innovative Biomaterials for Tissue Engineering DOI: http://dx.doi.org/10.5772/intechopen.83839*

 and more tolerated than synthetic ones, and Pasqui *et al*, for example, developed a natural cellulose-hydroxyapatite hybrid hydrogel for bone tissue engineering. For the chemical synthesis procedure, the freeze-dried hydrogel was immersed in a solution containing HA microcrystals, and then an *in vitro* study demonstrated that MG63 osteoblast-like human cell seeded into hydrogel samples adhered and proliferated rapidly. Moreover, an increase of ALP activity was identified at 3, 7, and 14 days [43]. Synthetic hydrogels could have limitations in the biocompatibility, but they offer the possibility to modulate their mechanical features and rate of degradation in biological environment. Kinard *et al.* developed a biodegradable oligo[poly(ethylene glycol)fumarate] hydrogel to deliver demineralized bone matrix (DBM) in a rat bone defect. They found that the *in vivo* degradation rate of the hydrogel depend on the DBM content, higher was the rate of DBM faster was the degradation. Moreover, high content of DBM could affect the mechanical properties of the hydrogel even if it increases its osteoinductivity in vitro and in vivo [44] (**Table 1**).
