In a recent time, the prevalence of degenerative disorders is increasing irrationally. The rising level of stress, unhealthy lifestyle, and lack of proper nutrition are identified as reasons for this sudden increase. Consequently, the current healthcare industry is facing challenges like lack of proper treatment, shortage of organs, and shortage of donors; this further raised a demand for artificial substitutes for failing organs. With technological advancements like 3D bioprinting, organ-on-chip models, etc. researchers are aggressive toward bringing these technologies to routine treatments. However, it is very imperative to note that these advanced technologies can meet the growing demands of tissues and organs, only when enough resources and quality raw material are invested in its further development. Thus, regardless of the rapid advancements in technology, the pursuit of fully functional 3D organs is impossible without the availability of viable seed cells. 

The ideal cell source should be qualified with the presence of several important characteristics

• Printability: The cells that are to be used for 3D bioprinting should be resilient enough to withstand sheer stress, pressure, and temperature fluctuations; this needs to be achieved while maintaining viability and functionality. 
• Proliferation: Tissue fabrication is one of the important steps of 3D bioprinting; which can be achieved only with the help of the high proliferation rate of cells used. At the same time, it is also very essential to control the proliferation rate of cells; without compromising viability as well as functionality. 
• Functionality: The primary cells used for 3D bioprinting, should either be functionally competitive or differentiate into mature functional cells. This property is essential to establish the desired functionality of 3D bio-printed material. 
• Safety: When testing the efficacy of bio-printed tissues for therapeutic properties, it is very important to use cells that have normal karyotypes. Apart from the same, cells should be non-tumorigenic as well as devoid of phycological toxicity. 
• Economy: The construction of large-scale organs necessitates a substantial number of seed cells. This necessitates the large-scale expansion of primary cells. 
• Ability to Self–assemble: The extent of full functionality of 3D printed material depends upon its microstructure; which is formed by the organization of appropriate cells. Thus, the vessel network of vascular tissues developed with the help of 3D bioprinting largely depends upon the primary cells. 

Accordingly, the present article intends to provide a comprehensive overview of currently available options for suitable primary cell sources in 3D technologies. But before that let’s have a brief overview of the technique. 

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The following briefly discusses prominent 3D bioprinting tissue engineering techniques:

• Microextrusion 3D Bioprinting: This pressure-assisted technique employs a glass or plastic cartridge to dispense bio-inks through a nozzle via pneumatic or mechanical methods controlled by a robotic arm. The bioink forms a thin filament based on a CAD design, allowing the creation of large-scale constructs with structural integrity. While suitable for scale-up and utilizing various bio-inks, it suffers from low resolution
• (~100 μm) and high extrusion pressure, potentially impacting cellular viability and tissue structure.
• Inkjet 3D Bioprinting: A non-contact technique, inkjet bioprinting expels bioink droplets onto a substrate using thermal, piezoelectric, or electromagnetic forces, mimicking the CAD-based model. Originating from conventional 2D paper-based printing, it offers cost-effectiveness, high-speed printing with multiple nozzles and relatively high cell viability. However, it requires low-viscosity bioink, posing challenges for highly viscous hydrogels and extracellular matrix (ECM) deposition.
• Laser-Assisted 3D Bioprinting (LAB): Another non-contact method, LAB employs a laser to induce hydrogel droplets on a coated ribbon, forming a jet for layer-by-layer construction. With high-speed printing and real-time cell monitoring, LAB enables high-throughput cell and biomaterial patterning. However, the generated heat from laser energy may compromise cell viability in the printed tissue.
• Stereolithography-Based Bioprinting (SLB): Utilizing photopolymerizable liquid polymer, SLB directs UV light or laser in a predesigned pattern to cross-link and harden polymers layer by layer. Known for its high resolution and minimal shear stress, SLB exposes cells to intense UV radiation during cross-linking, potentially causing cell damage.
• These 3D bioprinting techniques offer diverse approaches to fabricating biological constructs, each with its advantages and challenges in terms of resolution, speed, and cell viability.

Cell Types for 3D Bioprinting

Cells serve as the fundamental unit for 3D Bioprinting tissues; however, several routinely used cell sources face multiple challenges to adapt to this advanced technology. With the rapid development in the field of regenerative medicine, several stem cells can be good alternatives, holding promise for successful integration into 3D bioprinting applications.  

Cell lines for 3D Bioprinting

Cell lines are continuously generated populations of cells, maintaining stable phenotypes and functions over a longer duration. These cell lines are commonly employed in 3D bioprinting models, particularly because viably and functional primary cells are difficult to maintain and expand in vitro. The use of these cell lines is preferred because of their easy maintenance and cost-effective production. Researchers also prefer them over primary cells because their culture does not require any specific expertise and it remains robust against mechanical stress and environmental changes. However, there are many disadvantages to using cell lines, and the most important is that the high proliferative capacity of cell lines may not always be advantageous. Moreover, many cell lines display abnormal karyotypes including expression of mutagenic oncogenes and expression of tumor suppressor genes. Another common disadvantage of cell lines is their inability to fully mimic their in vivo counterparts in terms of maturity and functionalities. Consequently, it is very important to explore other safer alternatives to cell lines that are capable of generating functionally mature cells with a regulated rate of proliferation. 

Stem cells

Stem cells are the native cells with remarkable differentiation properties and self-renew properties. The cells can efficiently differentiate into cells of various other lineages and hence are the potential candidates for 3D-printed tissues and organs. During the past couple of decades, stem cells have proven their potential in therapeutic applications; further highlighting their promising future in the science of regenerative medicine. In particular, embryonic stem cells, mesenchymal stem cells, and pluripotent stem cells can potentially give rise to various cell types in the body, including liver cells, pancreatic beta cells, etc. Thus, these cells can be extensively used for further development and research applications (Table 1). 

 (Table 1)

Thus, advancing applications of embryonic as well as iPSC-derived cells display great potential as cell sources for 3D bioprinting.  Interestingly, many other primary cells are acknowledged as valuable cell sources in 3D technology. However, their practical applications are limited due to the unavailability of viable cell sources. Studies have implicated the use of primary cells that are isolated directly from tissue sources with improved adaptability, viability, and quality; such as hepatocytes, muscle satellite cells, as well as lung stem cells. It has also been observed that these cells maintain normal post-expansion karyocytes with better functional maturity and the ability to differentiate into mature functional cells. Notably, these cells possess a better capacity to form aggregates of cells, further enabling the formation of intricate microstructures. 

Thus, the key benefits of utilizing stem cells for 3D bioprinting are as follows:

• Better Physiological Relevance: These primary cells retain the in vivo phenotype and functions of native tissues. 
• Tissue-specific Functions:  Since these primary cells are the direct replica of tissue structures in terms of physiology. These cells can serve as good alternatives for fabricating organ-specific structures and functions, offering more accurate and specialized tissue models. 
• Patient-specific Cells:  Cells can directly be sourced from the patient’s tissues, further facilitating the construction of personalized organs that closely replicate the unique biological structural as well as functional properties of the donor. Today, these personalized applications in the field of regenerative medicine hold great potential; and the remarkable integration properties of primary cells further minimize the risk of immune rejections; which in turn enhances the potential for successful transplantation. 

Accordingly, several different kinds of primary cells are used in constructing 3D-printed organs, the details of which are shown in Table 2. 

 (Table 2)

Conclusion 

The progress in healthcare-focused printing technologies offers a significant prospect for the precise production of medications and the efficient manufacturing of diverse tissues and organs. Ongoing advancements in 3D bioprinting hold the potential to create implantable organs and tissues, addressing organ shortages and potentially saving numerous lives through increased transplant availability. Concurrently, the evolution of 3D bioprinting establishes an optimal foundation for advancing personalized medicine, contributing to an overall enhancement in healthcare quality.