3D Bioprinting in Medicine

3D bioprinting of tissues and organs is game changer and promising technology in medicine. It has also great potential to be substitution of animal models as artificial tissue or organ platforms and can be used for transplantation to the patient directly. Natural and synthetic polymers can be used as bioinks in order to develop tissue or organ models and they can be applied from benches to clinical application. In this review, it is aimed to summarize 3D printing technology in medicine and bioinks.

Introduction

The idea of bioprinting tissues, organs is very promising and extraordinary since 1980’s. First commercial bioprinter and 3D printers were developed in late of 1980’s and development studies are still going on [1]. 3D printing technology is relied on Computer Aided Design (CAD) technology and digital images are returned to 3D structures with CAD tools. Then 3D structures at microscale and even nanoscale can be printed with 3D printers/ bioprinters with lower cost and higher flexibility and efficiency [2]. In last 20 years, this idea is evolved to printing of biological structures as bioprinting and several of bioprinting approaches have been carried out including stereolithography, extrusion, inkjet, laser and droplets based techniques for tissue and tissue substitutes and they were used from benches to clinical applications [3].

It is not possible to obtain fully functional and complex synthetic tissues or organs at all scales with a single bioprinting technique. It must be investigated and improved in terms of resolution, cell viability and bioinks. In literature, the most common used bioprinting techniques can be shown in Figure 1 [4].

Inkjet-based bioprinting is using the conventional inkjet process of inkjet desktop printers and droplets of dilute solutions are dispensed with a non-contacting printer [5]. Drop demand inkjet technology is commonly used for bioprinting applications with thermal, piezoelectric, electrostatic and electrohydrodynamic printing technologies [6].

Laser-assisted bioprinting technique utilizes a laser as the energy source to place biomaterials onto a substrate. This technique has mainly three part as; a pulsed laser source, a metal film layer, and a receiving substrate. The resolution of this techniques is differentiated in between pico- to micro-scale [7].

The extrusion based bioprinting method is combination of a fluid-dispensing piston and an automated robotic system. A deposition system is released the bioink material and all process is under control of a computer. The extrusion based bioprinting has advantages as rapid fabrication, better structural integrity and CAD software adaptability [8].

Briefly, the main strategy for bioprinting of a tissue, various cells are collected from the patient and cultured in a cell culture system. Then it is mixed with a suitable biomaterial and the developed bioink is fed to the bioprinting system. After the fedding, 3D bioprinting process has started due to 3D pattern of the desired tissues CAD/CAM image processing products. In Figure 2, 3D bioprinting steps of skin tissue were schematized [9].

Actually, 3D bioprinting process has several steps from starting to finalization. They can be classified as pre-printing, bioprinting and post-printing. In Table 1, this substeps are listed. Preprinting stage is extremely important in order to ensure that proper quality of the cells is obtained. Bioprinting stage is included bioink, bioprinter and bioprinting process which are effected by physicochemical, biological and other process based parameters. As a result of the bioprinting process, bioprinted materials are transferred to bioreactors in a part of post printing stage. In this stage, tissue maturation should be obtained by stimulation and developed tissue/organ can be monitored by biosensors with respect to functionality, rigidity and stability [10].

Recently, great efforts have been achieved in 3D cell culture development in order to fabricate of tissue constructs for application in research, tissue engineering, therapeutic, or drug screening. Bioprinting strategies will enable to construct a fully automated fabrication of matrix, cells, and bioactive layers instead of animal experiments [11]. 3D bio-printed tissues and organs are great alternatives to cell cultures and animal models as artificial platforms. This technology has already been used to fabrication and transplantation of important tissues such as bone, skin, heart tissue. Forming of 3D bio-printed model from 2D cell culture concludes complex steps such as cultivation in hydrogels, spheroids, membranes, 3D porous scaffolds, 3D fibrous scaffolds and so on [12].

3D bioprinting in medicial applications can include applications of dentistry, fabrication of tissue and organ models, fabrication of medical devices, fabrication of anatomical models and drug formulations [13]. 3D bioprinting is considered as a tool which is able to solve the problems for cancer patients by developing patient specific treatment via mimicking of in vitro models more closely real cancer conditions [14]. Medical 3D printing technology continues to develop rapidly in number of publications. Europe has the most publications followed by United State and China. Orthopedic applications are most published followed by otolaryngology, vascular and cardiac medicine. The leading applications in 3D printing of organs and tissues include vascular, skeletal, hepatic, and cardiac based, in the order of significance [15].

Nowadays, tissue specific functional 3D bioprinting is the new approach for transplantation applications in regenerative medicine. The overall purpose must be relied on fabrication of the tissues and organs with respect to desired shape and function and using them in vivo [16].

Bioinks

In 3D bioprinting technology, the material used to fabricate artificial tissue is called “bioink”. It can be stabilized or cross-linked in order to obtain the final morphological and chemical structure of the design. Bioinks are produced from natural or synthetic biomaterials as alone, or their composites [17]. Bioink’s requirements can be differentiate due to bioprinting techniques that will be used and substrate requirement can be taken shape according to process [18].

In bio-printing process, the major aim is defined as developing three dimensional complex and functional architectures. For this purpose, most suitable, stable and cell-responsive bioinks with chemical and physical properties are required. Researchers are commonly used naturally derived polymers or synthetic derived polymers such as gelatin methacryloyl (GelMA), Polyethylene Glycol Diacrylate (PEGDA) and Polyethylene Glycol Dimethacrylate (PEGDMA) [16,19]. The four types of bio-inks are listed as microcarriers, hydrogels, cell aggregates, and decellularized matrix components. Due to the chosen bioprinting technique, bio-inks should have certain properties such as viscoelasticity, flexibility, thermoresponsivity, shear-thinning and selfhealing [20].

Bio-inks which not include living cells are generally utilized for developing scaffold for cell culture. Typical scaffold materials are different forms of hydrogels [21]. Hydrogel based networks can be also performed as remodeled matrix and 3D environment for the normal development of functional tissues [22]. Hydrogels are generally biocompatible and they are commonly show non-Newtonian, shear thinning behavior, which is important for using in extrusion bioprinting [23]. In literature review, it can be seen that hydrogels were commonly used as bioinks for example bone [24], osteochondral grafts [25], cartilage [26] tissue engineering.

Bioinks may also contain cells in different forms and bioactive molecules or also biomaterials. It is understood that, bioinks is independent of biofabrication techniques and applications area [27]. There are several studies about bioink development and their cell/ tissue application in the literature. It is very popular and demanding topic in biomedical and tissue engineering field. It is hard to mention all researches about 3D bioprinting but in Table 2, most common bioink materials and some examples of their cell or tissue based applications were listed.

Conclusion

Three-dimensional (3D) printing of biological structures have great attention in last ten years and their application have started to utilize in regenerative medicine. This technology is in early progression step but it has huge possibility to implement in tissue engineering, drug delivery, cell/tissue and organ products, and other biomedical applications [42]. Despite of increasing articles, patent and researches of 3D bioprinting of living tissues or organs, it still faces significant challenges as unstable cellular behaviour and more complexities compared to non-biological printing process. In order to avoid these challanges, researchers need to work in multidisciplinary teams with engineers, biomedical scientists, basic scientist and medical doctors [43].

One of most important challenges of 3D bioprinting relies on in situ bioprinting as known as printing cells and biomaterials directly onto or in a patient [44]. However, the bioprinting technology is still being developed, it will be necessary to improve with ethical issues and related laws in order to use of 3D printers in industry, hospitals and academia. New regulations must be made before adopting developed prototypes into cilinical application [45,46].

In the future, it is foreseen that researches will focus on enhancing 3D printing materials with respect to biocompatibility, mechanical properties, in situ bioprintability and sustainability of printed cells, tissues and organs via vascularization process [47]. Maintaning of cell viability, vascularization and cell migrations are the main limiting factors in generating of large sized, fully functional, tissues and organs. Especially vascularization is related to the nutrient diffusion process so it must be developed for realization of in vivo applications of 3D bioprinting [48]. Also novel bioinks were still developing in order to use as new scaffolds or unique cell, tissue applications. With these improvements, 3D bioprinting technology will be a revolutionary approaches in medicine.

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