Meetings

2005

2006

2007

Talks and seminars



How to print Organ?   [slideshow (~3MB PDF)]


by Vladimir Mironov  MD, PhD
Director of MUSC Bioprinting Center
Medical University of South Carolina,
Charleston, South Carolina
Organ printing technology includes three main technological steps:
  1. preprocessing or computer aided design or "blueprints" of organs;
  2. processing or an actual printing and fast solidification of tissue or/and organ constructs;
  3. postprocessing or accelerated tissue ad organ maturation.
Preprocessing
In order to build anything; a car, airplane, bridge or house, one must at first develop a blueprint. This is also true in the case of organ printing. Before building an organ, we must have a blueprint in the form of a computer-aided design of the designed organ. We might define this as computer compatible precise spatial information about the localization of cells in the 3D organ or, in other words, the "address" of each cellular or extracellular component of the tissue or organ that we want to built. There are several ways in which we can get the information about the anatomy, histological structure, composition and topology of human organs necessary for computer-aided design of printed organs. Recent progress in clinical bioimaging and ultrasound make it possible for us to discern the gross anatomical characteristics of organs even while they are still inside their owner. The advantage of this approach lies in its the capacity to demonstrate the patient's specific anatomical information as well that of his organs, not to mention the fact that we do not need to remove the individual's organ in order to examine it (a fact that many patients might well appreciate). However, resolution of this technique has not yet reached the histological and cellular level. More importantly, tissue composition and cell redistribution cannot be precisely identified as yet. In short, this method is not yet refined enough to be utilized in the process of organ printing. A second approach is based on computer-aided reconstruction of a histological section. This method provides a high level of resolution and information about the size and shape of the organ, as well as details about its composition. The problem inherent in this method lies in the fact that human organs are available for this sort of inspection only after death, and are hence subject to change and distortion. Other limitations of histological approach are that it is enormously labor-intensive, time consuming and is not patient specific. However, considering that organs have a polymeric structure and consist of repeating structural functional units, one can reconstruct only one two typical organ unit and then assemble the whole organ in silico by adding a reconstructed unit based on the gross anatomical structure or by filling the available space. A third approach is based on a mathematical computational anatomical model. For example, by knowing the mathematics of vascular branching it is possible to reconstructed in synthetic materials a very realistic model of the vascular tree found inside the organ. In fact, several commercially available pieces of software permit the creation of a realistic anatomical model from bioimages, and several laboratories around the world have developed virtual cadavers with gross anatomical and microanatomical level resolution, in the public domain and available through the internet. These successes suggest that the computer aided design of printed organs is feasible, although prior to finalizing the task of printing a viable organ, the existing software will need to be upgraded to embody more capacities and greater flexibility. We also need to decide what the basic building block for printing cells should be. Some believe that organs could be printed using inkjet printers from single cells piece by piece because of the technique†¢s high speed, printing 6000 or more drops per second with one cell per drop. However, there is some doubt as to whether fragile cell aggregates can resist the harsh environment and stress forces inherent in the printing process without falling apart. Others believe that printing would be achieved more easily by using a cell aggregate as the basic building block. In this case, cell aggregates also need to be manufactured prior to printing, and several methods are available, including shaking, centrifugation, hanging drop and monolayer wadding. Arguments in favour of using cell aggregates include faster printing due to the dramatic reduction of printing cycles, better cell survival due to the social effect and maximally high cell density and the opportunity to use predesigned cell aggregates. It is possible to create cell aggregates of different sizes and shapes, including aggregates of different types of cell, of different types, of cells combined with non-cellular material and even in the form of hollow spheres. The variability of cell aggregates will reduce the time necessary for post-printing cell adhesion, tissue self-assembly and polarization. Finally, the resistance of cell aggregates to stress can be dramatically enhanced by using aggregates with internal polymeric microscaffolds or highly porous sponge-like microspheres. As to which will provide the best outcome, we must await the efforts of future research. We have already shown that closely placed cell aggregate in hydrogel can fuse in the histological structure of the desired geometrical form. We refer to this combination of cells or cell aggregates together with hydrogel as "bioink". While there are various types of hydrogel, they all share a capacity for rapid solidification during and after printing. Making these hydrogels more cell friendly with more biomimetic properties while remaining suitable for fast solidification and postprinting tissue remodeling is a goal, but certainly an achievable one.
Processing
Processing or actual printing of tissue and organ constructs will be physically carried out by one of a range of material transfer dispensing and deposition devices. One of most promising technologies currently on the cards is represented by inkjet printers, because they are inexpensive and operate at a very fast speed of 6000 drops per second or more. Several research groups have already been able to demonstrate both that cells survive the process of printing and that one drop can contain a single cell. Utilizing cell aggregates in inkjet printers is a trickier proposition, but may be possible if the former are enhanced by the inclusion of a highly porous scaffold or spheroids. As well as inkjet, rapid prototyping technologies offer another possibility, although the use of high temperatures, toxic resin or resin and plastic with toxic catalyst is a counter-indication. At the time of writing, rapid prototyping presents as a tool that can be used for designing scaffolds for tissue engineering, before the scaffold is seeded with cells. The biggest problem of this technology is the limited extent to which we can control the position of cells in the 3D scaffold. What makes organ printing different from scaffold-based techniques is the method employed: simultaneous (one step procedure) layer by layer deposition of cells and stimuli sensitive hydrogel. The rapid prototyping technology that matches this description is stereolitography, using cells in photosensitive hydrogel. Positioning the cells in the gel is carried out by using a special mask and dielectrophoresis, or by directing the cells by using a special laser with a different light frequency to that used for the polymerization of photo-sensitive hydrogel. A popular method in the works for printing organs is based on using an automatic robotic deposition device; basically a syringe used in combination with a robotic hand. This allows the deposition of biological material in a very precise matter. A problem with this technique is that the manufacturing system is extremely expensive, but the laser ablation capacities of these devices dramatically enhance the carving or sculpting of tissue or organ engineered constructs The future will show us which type of bioprinting or dispensing system is the most practical for clinical applications. When a conclusion has been reached, we will be well on our way towards changing how surgeons of the XXI century carry out surgery. Recent developments indicate that organ printing technology is feasible and that bioprinting technology is coming of age, although we must continue towards mastering the challenges of achieving the effective vascularization of printed constructs, as well the maintenance of the printed shape of 3D tissue and organ construct, which otherwise tend to melt and distort. Possibilities to solve these problems include, respectively, the introduction of temporary, removable needles with pores that can both provide temporary mechanical support and perfusion for the printed scaffold, and the automatic printing of large vascular-like tissue tubes from self-assembling cell aggregates and of lumenized vascular spheroids capable of fusing into intermediate diameter vascular like tube. In order for organs to survive, it is crucial that the problem of vascular systems be solved.
Postprocessing
After printing, we have on our hands nothing more than printed tissue and organ constructs. They are not yet mature, functional tissues and organs. In order to be functional organs, they must undergo a rapid process of self-assembly, maturation and differentiation. Biophysically, they must have the physical properties of visco-elastic fluid, whereas mature organs usually have physical properties of elastic solid. The process of becoming solid organs may be defined as "accelerated tissue maturation". If printed constructs are to become viable organs, they require a wet environment that can only be achieved by using a special perfusion device, a bioreactor that allows the cells to survive. As yet, we have not solved the problem as to whether or not the bioreactor should be an essential, integrated component of the bioprinter. From both the perspective of cost and engineering, it is preferable that the organ be removed from the printer and placed in separate environment for further postprocessing in order to use the bioprinter more productively. Another factor essential to accelerated tissue maturation is the chemical and mechanical conditioning of the printed tissue and organ construct. In this case, each specific organ will require a specially designed perfusion media and regime. Finally, it must be possible, post-printing, to provide for the non-invasive biomonitoring of maturation of printed tissue and organ construct. Eventually, postprocessing or accelerated printed tissue ad organ maturation could be performed in the human body in case of in vivo printing.