**1. Introduction**

An organ is a combination of multiple tissues that provide specific physiological functions for human body. There are about 80 organs in the human body, dominating every physiological activity [1]. Clinically, organ failure is the leading cause of mortality all over the world, and it is often closely related to some chronic and acute diseases [2,3]. As a result, one failing organ can break down the whole physiological system of the human body.

At present, allogeneic organ transplantation is the only cure for the failing internal organs, such as the liver, kidney, and heart. However, it is facing many limitations in clinical applications. First, there is a desperate shortage of donor organs. In another word, the donor organ is seriously short. Take the year of 2013 as an example, there were 117,040 patients in the United States of America who needed organ transplantation, but only 28,053 of them are fortunate to ge<sup>t</sup> suitable donors [4]. Currently, there are over 34 million surgical procedures in America involved in the treatment of organ failures per year. Less than one out of ten patients can be saved by organ donations [5]. Second, there are serious side effects of immunosuppressive drugs. The donor organs are allogeneic sources and the patients need to take a life-long immunosuppressive treatment. Since the side effects of immunosuppressive complications are severe, the ability of resisting infectious diseases can be obviously weakened. Third, the surgical cost is very high. Organ transplantation can bring huge economic burdens to the ordinary patients and occupy enormous medical resources [6].

During the last several decades, the severity of donor organ shortage, the life-long treatment of allograft rejection, the side effect of immunosuppressive therapy, and the extremely high cost of allogeneic organ transplantation have activated numerous strategies for tissue repair and organ manufacturing [7–11]. One typical example is tissue engineering. It has undergone several circulations of ups and downs with the three elements, i.e., porous sca ffolds, cells, and growth factors. Nevertheless, organ manufacturing is a complex project that requires multi-disciplinary cooperation, involving a large scope of talents of technologies, such as biology, materials, chemistry, physics, mechanics, informatics, computers, and medicine. Designing and building the physical analogues of organs is only the first small step. It is more critical to make the multiple cell types/extracellular matrices (ECMs), hierarchical vascular, neural and/or lymphatic networks functional in a compacted construct [12–14].

Three-dimensional (3D) bioprinting has recently emerged as an extension of traditional rapid prototyping (RP), also named as solid freeform fabrication (SFF) and additive manufacturing (AM), technologies, by using bioactive or cellular components to build constructs in an additive or layer-by-layer methodology for encapsulation and culture of cells. These technologies allow for cell culture in controlled spatial environments. These environments can be tuned to simulate the complexity of in vivo cell growth environments with similar ECMs.

Polymers are large molecules or macromolecules, composed of many repeated subunits or small molecules, with molar masses ranging from thousands to millions. In another word, the units composing polymers derive from molecules of relatively low molecular mass [15]. There are two types of polymers: natural and synthetic. Natural polymers are naturally occurring polymers, such as cellulose, polysaccharide, protein, silk, and fibrinogen. Synthetic polymers are manmade polymers through chemically joining many small monomers together into one giant molecule. The list of synthetic polymers, roughly in order of worldwide demand, includes polyethylene, polypropylene, polystyrene, polyvinyl chloride, synthetic rubber, neoprene, nylon, polyacrylonitrile, phenol formaldehyde resin (or Bakelite), polyvinylbutyral (PVB), silicone, and many more. Because of their broad range of properties, both synthetic and natural polymers play essential and ubiquitous roles in everyday life. More than 330 million tons of these polymers are made every year (2015) [16]. Chitosan as a special natural polymer has attracted grea<sup>t</sup> attention both in tissue repair and organ 3D bioprinting areas.

### **2. Three-Dimensional (3D) Bioprinting**

### *2.1. The Concept of Organ 3D Bioprinting*

3D printing, traditionally termed as RP, SFF, and AM, is a series of material processing technologies based on the dispersion-accumulation principle of computer-aided manufacturing (CAM). Generally, an object can be divided into numerous two-dimensional (2D) layers before 3D printing with a defined thickness. The 2D layers are sequentially piled up by selectively adding the desired materials in a high reproductive additive manner under the instruction of computer-aided design (CAD) models [17–20].

Organ 3D bioprinting is the utilization of advanced 3D printing technologies to assemble multiple cell types, including stem cells/growth factors, along with other biomaterials in a layer-by-layer fashion to produce bioartificial organs that maximally imitate their natural counterparts with respect to anatomical structures, material components, and physiological functions (Figure 1) [12–14]. It mainly consists of four aspects, such as cell extraction (i.e., biological part), data collection (i.e., informatical part), starting material preparation (i.e., biomaterial part), and manufacturing (i.e., processing part). Patient-specific organ images, such as magnetic resonance imaging (MRI) and computerized tomography (CT) can be easily transferred into CAD models for customized organ manufacturing with predefined geometrical shapes, material (e.g., cells, growth factors, polymers, ECMs, drugs) components, and physiological functions [21–24]. Over the last decade, organ 3D bioprinting technologies have made a grea<sup>t</sup> contribution to various biomedical fields.

**Figure 1.** Graphical description of organ 3D bioprinting with four major aspects: cell extraction (i.e., biological), data collection (i.e., informatical), starting material preparation (i.e., biomaterial), and manufacturing (i.e., processing) parts.

### *2.2. Polymers as "Bioinks" for 3D Bioprinting*

As stated above, polymers are large molecules made up of many small and identical repeating units bonded by covalent bonds. Theoretically, any polymer solutions or hydrogels holding the sol-gel transition property can be printed in layers under the instruction of CAD models [25,26]. Living cells and growth factors can be encapsulated into the polymeric solutions or hydrogels for 3D tissue and organ construction. Actually, only few natural and synthetic polymers and their combinations can be used as "bioinks" for tissue and organ 3D bioprinting at mild temperatures or cell endurable conditions. The layer-by-layer construction processes depend largely on the liquid polymer solution transformation capabilities before, during, and after the 3D bioprinting. There are many different sol-gel transformation forms, such as physical (reversible), chemical (reversible or irreversible), and biochemical (i.e., enzymic) cross-linking. Compared with synthetic polymers, most of the natural polymeric hydrogels can provide cells with suitable environments to survive. Currently, natural polymeric hydrogels are the dominate components of "bioinks" for 3D bioprinting.

There are three major types of organ 3D bioprinting technologies (Figure 2): (A) multi-nozzle extrusion-based bioprinting (a: pneumatic; b: Piston); (B) multi-nozzle inkjet-based bioprinting (a: heater; b: piezoelectric actuator); (C) multi-channel laser-assisted bioprinting. Cellular behaviors are easily manipulated within the polymeric hydrogels, via adjusting the physical, chemical, biochemical, and physiological properties of the 3D printable polymers. It is surprising that all the bottleneck problems, such as large scale-up tissue/organ manufacturing, living tissue/organ preservation, hierarchical vascular/neural network construction, and partly/fully stem cell engagement, which have perplexed tissue engineers and other researchers for more than several decades, have been overcome by a single scientist, the corresponding author of this article herself, via several series of automatic and semiautomatic layer-by-layer material integration and step-by-step stem cell inducement strategies [12–14].

**Figure 2.** A schematic diagram of the three major types of organ 3D bioprinting technologies: (**A**) two-nozzle extrusion-based bioprinting (a: pneumatic; b: piston); (**B**) two-nozzle inkjet-based bioprinting (a: heater; b: piezoelectric actuator); (**C**) three-channel laser-assisted bioprinting. Image reproduced with permission from [26].

Thus, the purpose of organ 3D bioprinting is to design and manufacture bioartificial organs using polymeric materials, cells, bioactive agents, and advanced 3D printers. Defining and creating appropriate polymeric "bioinks" is a critical step in bioartificial organ 3D bioprinting [12–14]. Natural polymers, such as alginate, gelatin, hyaluronate, and chitosan, have been chosen as the preferable candidates for organ 3D bioprinting because of the specific properties, such as biocompatible, bioprintable, biodegradable, bioavailable, and biostable. Particularly, these natural polymers can be easily predesigned as the ECMs of each tissue in a natural organ.

### **3. Properties of Chitosan as a Natural Polymer**
