**Abstract**

Tissue engineering (TE) is an interdisciplinary field that was introduced from the necessity of finding alternative approaches to transplantation for the treatment of damaged and diseased organs or tissues. Unlike the conventional procedures, TE aims at inducing the regeneration of injured tissues through the implantation of customized and functional engineered tissues, built on the so-called 'scaffolds'. These provide structural support to cells and regulate the process of new tissue formation. The properties of the scaffold are essentials, and they can be controlled by varying the biomaterial formulation and the fabrication technology used to its production. Pectin is emerging as an alternative biomaterial to non-degradable and high-cost petroleum-based biopolymers commonly used in this field. It shows several promising properties including biocompatibility, biodegradability, non-toxicity and gelling capability. Pectin-based formulations can be processed through different fabrication approaches into bidimensional and three-dimensional scaffolds. This chapter aims at highlighting the potentiality in using pectin as biomaterial in the field of tissue engineering. The most representative applications of pectin in preparing scaffolds for wound healing and tissue regeneration are discussed.

**Keywords:** pectin, tissue engineering, scaffolds, bioprinting, biofabrication, tissue regeneration

## **1. Introduction**

Tissue engineering (TE) is an interdisciplinary field whose first definition dates back to 1987. It combines the knowledge from different research areas including medicine, material science and engineering to develop engineered biological substitutes able to restore, maintain or improve tissue functions [1]. TE was introduced from the necessity of finding alternative methodologies to organ transplantations due to their increasing demand in clinical medicine. Furthermore, TE emerged as a promising approach to overcome the limitations of the conventional surgical approaches for the treatment of tissue damages caused by injuries, diseases and congenital disorders [2, 3]. These surgical procedures are based on replacing the injured tissues or organs with a healthy one harvested from the same patient (autograft), or a compatible donor (allograft). Although these approaches have been revolutionary and lifesaving, there are still some drawbacks that need to be addressed. The surgical procedures used to harvest both autografts and allografts are often invasive and painful. The risk of post-surgical limitations in

the donor's body due, for example, to infections and hematomas is, in fact, quite high. Moreover, when allografts are transplanted, the chance of inflammatory and immune responses in the patient body together with the transmission of diseases from the donor to the patient is significant [4].

TE aims at overcoming the complications associated with the conventional techniques used during organ transplantation by inducing the complete regeneration of the damaged tissues instead of replacing them [2, 3]. Several approaches to promote *de novo* tissue formation have been implemented in TE so far. These are mainly based on the use of biodegradable and biocompatible engineered tissues, based on the so-called '*scaffolds*'. A scaffold is a structure that provides temporary mechanical support and a guiding template to cells during the synthesis of new tissue. With the desired shape, architecture and functions. Concurrently, the scaffold biodegrades leaving space for new tissue in-growth. Notably, the biodegradability of the scaffold is what differentiates it from permanent implants. The complete biodegradation of the scaffolds prevents, the need for additional surgical interventions to remove it or, eventually, substitute it. The scaffold can be directly implanted into the injured site to induce the regeneration of the tissues *in vivo.* Otherwise*,* prior to implantation, the scaffold can be initially cellularized with cells isolated from the patient, subsequently cultured *in vitro* to synthesize tissues that will finally be transplanted into the defect to restore its functions (**Figure 1**). In this case, scaffolds can be further

**Figure 1.** *Illustration of TE paradigm (figure created with BioRender.com).*

cultivated in bioreactors, namely, devices able to apply biophysical stimuli to cells (e.g., mechanical or chemical) to better mimic the dynamic physiological conditions. In both approaches, the scaffold can be loaded with drugs, growth factors, micro- and/or nano-particles to further facilitate the recovering capabilities of tissues [5, 6].

The scaffold plays an essential role in regulating the process of new tissue formation. An ideal scaffold should be biocompatible and should degrade with kinetics compatible with the rate of tissue regeneration. It should be highly porous (< 75% [7]) with adequate pore size to promote cell migration/scaffold colonization and nutrient transfer throughout the scaffold. A scaffold should mimic the features of biological tissues in terms of topological properties (e.g., shape, size), mechanical properties (e.g., stiffness), and the biochemical processes that control and regulate the functionalities of the tissues. Moreover, it should not alter the normal functions of cells, which should adhere, migrate and proliferate within the scaffold before producing new tissue [5, 6, 8, 9]. Depending on their applications, scaffolds with different shapes, compositions and properties have been developed so far.

The biomaterial formulations used to produce the scaffold strongly affect its properties [10, 11]. Thus, the selection of the proper biomaterial formulation is pivotal for inducing the regeneration of the tissue in a controlled manner avoiding any undesired side-effects (e.g., cytotoxicity, apoptosis, carcinogenicity). The most used biomaterial formulations in TE are mainly based on synthetic biopolymers, natural biopolymers and composites [12, 13]. Synthetic biopolymers, like polycaprolactone, can be produced on a large scale under controlled conditions with predictable and reproducible physicochemical properties (e.g., mechanical properties, biodegradability) [6, 14, 15]. However, many synthetic biopolymers that have been developed so far are mainly derived from petroleum and coal, which make them not compatible with the environment [16]. Natural biopolymers include animal-derived proteins (e.g., gelatin, hyaluronic acid, collagen, silk) and animal- and vegetalderived polysaccharides (e.g., cellulose alginate, chitosan). One of the advantages of this class of biopolymers is their biological similarity to native tissues which is beneficial for supporting cell functionalities (e.g., cell adhesion). Nonetheless, the use of animal-derived biopolymers may be associated with a high risk of transmission of diseases from animal to patient [10, 17, 18]. Therefore, the use of naturally occurring biopolymers from vegetal sources represents an attractive alternative to overcome these limitations. Moreover, they represent an ecological alternative to synthetic biopolymers in the preparation of sustainable and green scaffolds.

In recent years particular attention has been paid to the adoption of methodologies to derive biopolymers from renewable sources, such as industrial by-products, such as pectin from fruit pomace produced from the fruit processing industry [19] and cellulose nanofibers obtained from paper waste [20]. The application of more ecologically viable biomaterials in TE may, in fact, strongly contribute to reduce the polluting impact of producing and using un-recyclable synthetic biopolymers. Among the renewable and natural biopolymers, pectin is gaining particular attention in TE for its advantageous properties including biocompatibility, biodegradability and non-toxicity [21, 22]. In addition, the versatility in processing pectin-based formulations allows to produce scaffolds with diverse properties and for different applications (Section 2).

This chapter aims at highlighting the applications of pectin as the building block of bidimensional (2D) and three-dimensional (3D) scaffolds for TE applications. With this aim, in Section 2 the properties of pectin as biomaterial are provided. Section 3 reports the most representative applications of pectin-based formulations for producing scaffolds for tissue regeneration in the shape of 2D films for wound healing and 3D scaffolds for tissue regeneration.
