By Keletso Mahlangu
Abstract
Modern technology simplifies solutions to a vast array of complex problems. Some of this
technology has been adopted into the medical world to introduce ambitious solutions to formally
insurmountable problems. This article focuses on exploring biotechnological advances in 3D
tissue bioprinting as well as its prospective effects on healthcare provision.
Figure 1: Example of a bioprinter(1).
History of bioprinting
The technology of bioprinting is still in its infancy. In fact, the first 3D printer termed the Stereolithography Apparatus was invented by Charles Hull in 19841(1). From this, Hull patented the stereolithographic method of printing(2). Bioprinting was first introduced in 1988 which used an inkjet printer to show 2-dimensional micro-positioning of cells(1). In 2001 a synthetic human bladder scaffold was 3D printed(1). The first Extrusion-based bioprinter was used and commercialized in 2002 and the first inkjet bioprinter in 2003(1). Then in 2004, 3D printed tissue was printed without the need of a scaffold(1). Between the early 2000s and recent times, ample advances have been presented to develop 3D tissue printing which has led to various tissue types being printed including cartilage, blood vessels, heart tissue, and more.
Mechanisms of bioprinting
Levels of bioprinting
Bioprinting can be divided into four levels categorized by the type of structures being produced.
Level 1 refers to bioprinting to produce structures without bio-compatibility requirements, an example being tools used in surgical path planning. Level 2 refers to the production of non-degradable products that have bio-compatible requirements, an example being titanium alloy joints and silicone prostheses. Level 3 refers to the production of degradable bio-compatible, an example of which would be a biodegradable vascular stent and bioactive ceramic bone. Level 4 would then refer to the typically thought of bioprinting involving the manipulation of living cells and production of biomimetic 3D tissues such as blood vessels and other tissues(1).
Strategies of bioprinting
Bioprinting strategies have been curated to fit the structural requirements of the different tissue types being produced. Some of the commonly used strategies are listed below.
Stereolithography
The above mentioned stereolithography method discovered by Hull involves altering the state of matter of a substance by transforming it from liquid to solid and manipulating the shape using a resevoir(3). The integration of stereolithography in bioprinting involves the use of a bioink component which would introduce living cells as well as cellular material in printing. These bioprinters selectively solidify bioink using photo-polymerization on a per-layer based process along a z-axis(3). This method has been used in cranial reconstruction during reconstructive head surgeries.
Pros(3)
• The z-axis printing allows for the printing of 3D structures without the need for x and y-axis traveling nozzles. Hence it is faster than other methods.
• Selective cross-linking of bioink by the polymerizing light reduces shear stress to cells immensely improving cell health.
Cons(3)
• The liquid is required to be transparent to reduce the scattering of the polymerizing light. Hence bioink cell density is limited.
Inkjet Bioprinting
A method based on commercial 2D inkjet printers. In this method, the bioink is stored in an extruder where it is temporarily deformed by piezoelectric actuation or digitized thermal actuation to disperse droplets of the bioink. This is responsible for printing in axis x, y, and z on the printing platform(3). This method has been used in drug formulations, tissue repair cancer research, and other procedures(4).
Pros(3)
• The system is non-complex and inexpensive
• Print heads can be paired parallel to increase the resolution of the tissue
• Inkjet printing results in high cell viability
Cons(3)
• Inkjet printing results in low cell densities and limited viscosities of bioinks.
Laser-assisted bioprinting
This method utilizes a laser and harnesses the energy to deposit bioink. The system involves a donor layer, an energy absorbing layer, and a bioink layer. When the donor layer is vaporized by the laser, a high-pressure bubble leads to droplets falling onto the collection platform. The Z stage can then be manipulated to produce a 3D printout.
Pros(3)
• Cells do not experience high shear stress, hence a high cell viability.
Cons(3)
• Expensive.
Extrusion-based bioprinting
Extrusion-based printing, also known as micro-extrusion printing, is derived from inkjet printing with the alteration of the droplets being exchanged for ongoing filaments of bioink through continuous extrusion force. This is one of the most used methods(3). It is divided into three methods depending on the force driving the extrusion process: pneumatic, piston driven and screw driven extrusion-based bioprinting.
Pros(3)
• Offers continuous bioink extrusion.
• Non-complex use and affordability.
Cons(3)
• Slow printing and nozzle size-dependent resolution.
• Reduced cell viability due to shear stress.
Figure 2: Different strategies of bioprinting(9).
Application of bioprinting in medicine
The role of bioprinting in organ transplantation
The increasing demand for organ transplantation remains a burden on the healthcare system. With the difficulty of acquiring compatible donors for transplantation and the high expenses of each transplantation, along with intensive follow-up procedures(4), the need for a more sustainable solution is dire. Bioprinting presents a promising approach to the world of tissue and organ transplantation. Bioprinting has the potential to allow for organ production on prompt. Recent studies have been conducted to research organoid [3D cell cultures characteristic of their derived organs which can recreate certain biological processes of their “parent organs” from their development(5)].Organoids are developed from pluripotent stem cells or adult stem cells which allows some of these cells to form mimicking cell proliferation in vivo(5). This allows for recipient curated organ printing, producing biocompatible, histocompatible and size compatible organ printing. The bioprinting of organoids is however currently limited by certain criteria. Hence although research is being conducted in organ bioprinting, not much progress has been made in printing full-scale organs due to the complexities of the organ structure attributed to the combination of systems (nerves, musculature, etc) within organs. Over the years, a vast array of organoids has been produced including models derived from the heart, kidney, liver, and other organs(1).
Tissue bioprinting
Much of the progress in bioprinting in recent years is in tissue printing and studies have been conducted to produce specific tissues. Cartilage has been produced using bioinks containing mesenchymal stem cells which proliferated into mature chondrocytes(1). More studies have been exploring the use of bioprinting in the process of nerve regeneration. In 2015, a study was conducted to produce Nerve Guidance Conduits(1) (which guide axonal nerve growth) to test their effect on peripheral nerve and spinal cord regeneration. These studies revealed complete regeneration of the spinal cord and sciatic nerve in response to the conduits. Vasculrisation has also been a point of interest, and refers to the use of prevascularised tissues where different cell types were compiled into hydrogels to control cell layering. Endothelial cells mimicked a lumen in vivo implantation in vitro and there was survival of the endothelial network in the prevascularized tissue during in vivo transplantation(1). The challenges posed in vascularisation pose a major threat to large tissue printing and complete organ generation.
Figure 3: Vascularization in bioprinting(1).
Pharmacological research
3D printed tissues are also used in in-vitro drug testing(1). The biomimicry of the cells allows for comprehensive drug screening and drug delivery is possible in these living tissues. These provide a safe space to test the effect of drugs on specific tissues without the risk of harming a living host. Current uses in clinical medicine The recent use of 3D printing is mostly to produce tools and devices that are required to be modified before surgery(6). Hence the most applied levels of bioprinting are levels 1-3. Level 4 has seldom been used in the mainstream clinical setting due to the newnessy of the technology. One of the few documented cases of its use was research and creation of a liver-inspired 3D detoxification device bioprinted in 2014(7).
Hinderances in bioprinting
The main hinderance of large scale bioprinting is considered to be the networking of the vasculiture(8). Large scale tissue bioprinting does not allow for printing of complex branching blood vessels. Hence bioprinted tissue would not be able to undergo metabolic and respiratory processes facilitated by blood vessels and would therefore die. Stem cell technology is part of the foundation of bioprinting. This technology is however not fully developed or understood by researchers(8). Hence the behaviour of stem cells can be unpredictable whilst printing leading to the risk of tissue rejection. Stem cell technology limits the progression of bioprinting. The shear stress experienced by cells leads to DNA and structural modifications which damages cells during the bioprinting process(8). Complete organs are least likely to be composed with low cell viability (number of healthy cells). Another challenge comes with the maturation of the progenitor cells which have been printed(8). These cells are placed in a bioreactor for them to mature. This process is delicate and violent vibrations or movements have the potential to ruin the development of the tissue.
Conclusion
It is thus evident that 3D bioprinting has the potential to revolutionize medicine in surgery, clinical practices and drug research. The potential is however limited by the technology required to apply advanced bioprinting to some medical requirements, especially clinical medicine. Bioprinting currently has several hinderances including the limited knowledge of stem cells and technological deficits which contribute to challenges in clinical introduction. As more research is done to discover different approaches in methods of bioprinting, the biotechnological community gets nearer to overcoming these hinderances.
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