Organs from the laboratory

Yulia Kondratenko, "Biomolecule"Special project on the mechanisms of cellular aging, longevity and transhumanism

Artificial organs are needed not only for transplants. Drugs can still be tested on them and intercellular interactions can be studied. Depending on the purposes for which an artificial organ is obtained, it may resemble a natural organ to varying degrees. Therefore, different strategies for reproducing the work of organs and their systems are suitable for different tasks. Our review is devoted to the basic principles of these strategies.

What is more like a human mouse or human cells living separately from the body is a controversial issue. But we can definitely say that it is easier, cheaper and more ethical to conduct experiments on cells, and not on whole animals. In addition, cells can be used to check whether a certain treatment method is suitable for a particular patient – and this is the way to individual medicine. Therefore, many scientists are working on systems in which human cells could feel the same as inside a native organism. The better we can simulate the living conditions of cells inside the human body in such systems, the safer it will be possible to allow drugs tested in these systems to be tested in human clinical trials. 

Modeling conditions is a basic requirement for drug test systems. It is not necessary for them to accurately reflect the shape and natural structure of real organs. Thanks to this, test systems can be made, firstly, compact, and secondly, modular - combined with each other in different combinations. These capabilities are used by the developers of "organs on chips" – matrices the size of a credit card, the cells of which are populated with human cells. The cells are connected by channels simulating the vascular network. The chip must be contained in a reactor that drives nutrient solutions through the "vessels" at the correct pressure. Some reactors even simulate the beating of the heart – for greater reliability in modeling blood flow. In addition, the reactor maintains the correct temperature of the chip, and also pumps gas into the solution to simulate breathing.

The chips have already reproduced the work of many organs and tissues – liver, kidneys, lung, adipose tissue, muscles [1]. It was also possible to reproduce the structure of the blood-brain barrier, which prevents many drugs and toxic substances from entering the brain from the blood. Russian scientists have also achieved success in designing organs on chips. Researchers from the company "Bioclinicum" have set themselves an ambitious task – to reflect on one chip the work and interaction of all organs that can be affected by the drug [2]. They have already received a system that simulates the six most important organs, the condition of which is primarily of concern to drug developers: intestines, liver, lungs, heart, brain and skin. In such a system, you can study where the drug will get if it is administered intravenously, orally or applied to the skin, as well as monitor the lifetime of its molecules in the body. For example, after passing through the liver cells, the drug must be metabolized so that its concentration in the blood decreases over time.

Organs on chips are not only useful, but also stylish: the model developed at Harvard (Fig. 1) even received the London Design Museum Award for the best design solution of this year [3].

Unlike chips, this technology allows you to obtain organs of the desired shape, including those ideally suited to a particular patient. And so that the organ does not cause rejection, it needs to be printed from the patient's own cells. To do this, stem cells are carefully selected from his adipose tissue, which can be reprogrammed into cells of various necessary specializations. In the "cartridges" of the bioprinter, the cells are contained in a special gel that does not allow them to stick together. And when the cells come out of it on a special substrate, which serves as a "paper", they stick together under the action of surface tension forces.

The bioprinter prints the organ in accordance with the computer model loaded into it, which must be worked out in detail. The most advanced bioprinters are already able to print several different types of cells, which is absolutely necessary to recreate complex organs. In addition to the main working cells of the organ, it is necessary to print at least vessels that will supply it with nutrients and remove waste products. 3D printing of vessels is described in detail in the article [4].

The printed organ is placed in a special reactor that supports its vital activity. While the organ is in the reactor, its vascular network continues to develop.

3D Bioprinting Solutions is engaged in bioprinting in our country, which developed the first Russian bioprinter (Fig. 2) [7, 8]. 

To date, the thyroid gland is one of the most complex organs created using 3D printing. Basically, this method now produces simpler structures – cartilage and skin fragments. According to scientists, it will take 10-15 years to print such complex organs as the liver and kidneys, for the functioning of which microstructure is very important. So far, scientists can print only small fragments of such organs or their mini-variants – organoids (not to be confused with intracellular structures!). You can watch the beating of heart organoids printed on a 3D printer right now:

Ideally, if the conditions are chosen correctly, the embryo will develop independently in the same way as in the embryo [9]. It is unlikely that it will be possible to grow ready–made organs in this way, because the conditions will have to be constantly changed as it grows. Nevertheless, a sufficiently developed rudiment can be transplanted to the recipient, in whose body it will form completely, at the same time adapting to the new environment. The plan looks complicated, but successful examples of its application are already known. For example, in 2013, Japanese researchers managed to simulate the conditions of embryonic development of the rudiment of the liver [10]. The secret of success was to select the right ratio of different cell types (liver endoderm cells, mesenchymal stem cells and vascular cells), as well as a suitable substrate. In a few days, on a regular Petri dish, researchers managed to grow liver rudiments resembling embryonic ones. Such rudiments were transplanted into laboratory mice. A few days later, the germ vessels were combined with the vessels of the mouse, and proteins produced by the new "liver" began to flow into the bloodstream of the animal. When hepatitis started in the mouse's "native" liver, the "new" organ helped them cope better with the disease.

The most interesting thing is that by imitating embryonic development, even the rudiments of the human brain can be obtained [11]. Scientists from the Austrian Academy of Sciences started with stem cells and selected the conditions in which to start their differentiation. Under the right conditions, it is possible to obtain the rudiments of the brain with cells of various specializations – neurons and glia. On the grown rudiments the size of a pea, you can see the emerging forebrain and even the developing retina (Fig. 3).

For example, when researchers tried to grow organoids from the cells of people with microcephaly, they found that the cells of such patients divided less and prematurely began to differentiate into neurons. By introducing CDK5RAP2 protein into organoids, the gene of which carries a mutation in microcephals, scientists managed to obtain normal organoids from defective cells. Thus, when trying to reproduce the embryonic development of a diseased organ, you can learn more about the causes of violations of its structure and functions.

And right now, and not when bioprinters develop enough to learn how to print all the necessary organs, including the most complex ones. These scientists are going their own way, not easy, but also promising. To get a full-fledged artificial organ, first prepare an anatomically shaped frame, and then populate it with the cells of the recipient for whom the organ is intended (read about growing organs on special frames in articles [5] and [6]). The first work on such a scheme was carried out back in the early 90s - then scientists grew an artificial ear [12]. A live mouse was used as an incubator for growing cells, which had a polyester ear frame seeded with calf chondrocytes implanted under the skin. In the process of cell growth, the polyester base degraded, so that as a result, pure cartilage in the shape of a human ear turned out.

Now such cruel methods of growing artificial organs are no longer used, and anatomically shaped incubators are being designed for each of them. The inner chamber of the incubator should fit the shape of the cell-populated base. The reactor must subtly regulate the flow of nutrient solutions through different parts of the organ, because local cell densities depend on it. The delicate and complex process of forming a full-fledged organ in an incubator goes on for several weeks, and it seems impossible to simplify it fundamentally. Anatomically shaped human bones obtained using human fibroblasts were grown in such an incubator [13]. Fibroblasts are located on the surface of the skin, and it is much easier to get them than stem cells.

Therefore, recently developed methods of reprogramming specialized cells into stem cells have greatly facilitated the procedures for growing artificial organs from patients' cells. The skeletons for artificial human bones were obtained from calf bone completely stripped of cells. Its extracellular material is quite suitable as a replacement for human. Then the bases were populated with mesenchymal cells obtained from the patient's fibroblasts by reprogramming, and placed in a special incubator. After implantation, the bones took root well – cartilage capsules formed around them, vessels sprouted there and host cells-osteoclasts, engaged in bone tissue reconstruction, crawled in (Fig. 4). Similarly, by populating the bases of animal organs, cleaned of their cells, with human cells, you can get more complex organs: for example, American scientists have grown a heart from human cells.

In 2012, for example, in Krasnodar Regional Clinical Hospital No. 1, a trachea and part of the larynx were transplanted from the cells of a patient who damaged the trachea in a car accident [14]. The work was carried out with the participation of the staff of the Karolinska Institute, in which the specialists of the clinic were trained. The base of the trachea, made of nanocomposite material, was seeded with cells obtained from the patient's bone marrow and placed in an incubator for 48 hours so that the cells would be fixed. Further development of the organ took place already in the patient's body after the operation.

Accidental injuries of the most important parts of the body will cease to be tragedies, giant queues for donor organs will disappear. What is most remarkable is that an individual approach is developing – it is already possible to grow anatomically shaped bones, and sometime, probably, it will be possible to reproduce the necessary organs down to the details of the cellular structure. Then all parts of a person will become potentially replaceable. Let us leave to the philosophers the questions of whether a man will be the same, all the aged parts of which have been replaced with new ones (see "Theseus' ship"). And we will be glad that this person will definitely not die because of a ridiculous accident, without waiting for his turn for a donor organ.