30 March 2015

Thyroid for the mouse

For the first time , Russian scientists succeeded
to print the thyroid gland on a bioprinter

Anna Govorova, Infox.ruRussian scientists from the laboratory of biotechnological research "3D Bioprinting Solutions" for the first time in the world managed to print a real organ on a bioprinter – the thyroid gland of a mouse, Infox reports.

In general, the creators themselves call it not an organ, but an organ structure. "There is a certain classification of the structural organization of the levels of the hierarchy of the living," says the scientific director of the laboratory, PhD Vladimir Mironov. – This is a molecule, cell, tissue, organs, organ systems and an organism. If we take this system for granted, then, based on it, what we have printed has no place in this classification. That's why we decided to call it an organ construct. And here's why. A tissue is a group of cells of the same type. An organ is a group of tissues. The organ structure created by us is much closer to the organ than to the tissue, since it consists of several types of tissues, has vascularization (vessels) and function at the body level. The organ structure of the mouse thyroid gland printed by us is also much more complex than the so-called tissue organ-like structures or organoids that do not have vascularization before implantation. This is a fundamentally important difference."

Scientists hope that they will soon be able to print a human thyroid gland. In the foreseeable future, this technology will help solve the acute problem of the shortage of donor organs for transplantation.

According to Vladimir Mironov, their opponents, skeptical about the technology of three-dimensional bioprinting of organs, argued that organs cannot be printed because they are very complex.

And indeed, until quite recently, this idea looked like a real fantasy. "The idea of bioprinting real functional organs came to me 12 years ago. Last October, we announced that we would print the thyroid gland in March. And we did it," says Vladimir Mironov.

This technology can be used in clinical practice in the foreseeable future, says Andrey Polyakov, Ph.D., Head of the Department of Head and Neck Tumors and Microsurgery at the P.A. Herzen FMIC. "For clinical practice, the main thing is that this created organ has vessels – arterial and venous, which can provide blood supply and nutrition, as well as transport of hormones produced into the body. Now the most important stage is to show the functionality and safety of this method," says Andrey Polyakov.

Why did scientists decide to create the thyroid gland? The choice was not accidental.

"We had to choose an organ that is relatively simple. The thyroid gland does not have a complex system of ducts for removing the products of its activity. Hormones that synthesize the follicles of the gland enter directly into the networks of blood capillaries that entwine each follicle. That is, to create such an organ, you need follicles entwined with capillaries, an internal vascular bed that connects to the bringing artery and the carrying vein," explains Vladimir Mironov. In addition, it turns out that the thyroid gland is the first organ that was transplanted to a person.

Like any new technology, the method of 3D bioprinting of organs did not arise from scratch, but absorbed the achievements of many sciences and directions – science and biomaterials, genetics, developmental biology, cell biology and information technology, scientists say. They also managed to successfully combine these methods and show that it really works.

Printing with live cells Initially, with the help of a special program, a three-dimensional computer model of the mouse thyroid gland was created, which was incorporated into the bioprinter.

The role of "bio–paper" was performed by a hydrogel, on which "biochernila" was applied – a conglomerate of living cells, which are called tissue spheroids. Such spheroids can consist of a variety of cells.

As Vladimir Mironov explains, they used two types of spheroids to print the thyroid gland. Some spheroids were filled with follicles in which the synthesis of the hormone thyroxine occurs. And another type was used to create a network of blood vessels.

Mouse embryonic explants were used to create the follicles themselves. "We can say that these explants represent one of the stages of development of embryonic stem cells, when they turn into thyroid follicles. At the next stage of development, they are already able to synthesize the hormone thyroxine. What is especially important, at this stage these follicles are already vascularized – that is, they are enveloped in a network of blood capillaries that secrete thyroid hormones directly into the blood," says Vladimir Mironov.

The most difficult thing is to create a network of vesselsThe most difficult thing in creating any organ is to provide it with blood vessels – to vascularize.

This task is especially important when creating organs, scientists emphasize. Because a structure of tissues can be called an organ if there is not only a working part consisting of functional cells, but also a network of vessels. "For example, tissue structures of skin, cartilage, blood vessels and liver already created on bioprinters cannot be strictly considered printed organs, says Vladimir Mironov. "Since the functionality of such structures is not provided at the level of the whole organism, they do not have a network of blood vessels."

Providing an organ with a network of blood vessels is not only an important task, but also one of the most difficult in regenerative medicine. In general, vascularization hides the main difference between the method of organ bioprinting from other sections of regenerative medicine, when organs are grown from embryonic or induced pluripotent stem cells – cells that have unlimited possibilities to transform into almost any cells and tissues.

Bioengineers have already learned to obtain tissues or fragments of individual organs from stem cells – the so-called tissue organ-like structures or organoids. But, as Vladimir Mironov emphasizes, all organoids grown in this way do not have a vascular system before implantation. And it is necessary for clinical practice (if we talk about the fact that these organs can be transplanted to patients in the future).

"We are often asked, how did you manage to achieve vascularization? Currently, two approaches have been developed in tissue engineering to create vascular-like channels. This is a sacrificial hydrogel method and a method based on the principle of endothelial cells to assemble themselves into capillary networks and even large-diameter vessels. The possibility of formation of a vascular bed inside the organ structure in this way has been confirmed experimentally. We used two methods," says Vladimir Mironov.

How the bioprinter worksAs Vladimir Mironov explains, a bioprinter is a robotic device that allows you to accurately distribute biomaterial – living spheroid cells in three–dimensional space, in layers, according to a digital model.

And to put it more simply, it is a syringe that moves in three directions.

Currently, 16 companies from 12 countries are engaged in the development of bioprinters. But, according to Vladimir Mironov, the bioprinter they created in their laboratory is unique. "The Russian bioprinter has five syringes. The American company Organova, the market leader in the production of bioprinters, has a printer equipped with two nozzles. Our printer is multi–functional. We can print three different types of spheroids. Another proof that our printer is a pioneer development – we have recently applied for a patent," adds Mironov.

How to print the thyroid glandBioprinting begins with the fact that the printer sprays hydrogel, and then spheroids are planted into it "like onions in a garden" – first vascular, and then functional (follicles with a network of blood vessels).

About three thousand spheroids were used to print the mouse thyroid gland. Printing takes place according to a pre-created digital model, the program sets the movement of the injectors. Signals are transmitted from the computer to the printer via the control unit.

"After printing, we check the integration of the entire system with ink. It turns out that we have printed the thyroid gland, and it is vascularized," says Vladimir Mironov.

Now the most important task is to prove the functionality of such an organ construct – that is, to show that it can perform its main function – to synthesize the hormone thyroxine, scientists say.

An animated film created by the forces of the Laboratory of Biotechnological Research "3D Bioprinting Solutions" and "DA Media" with the financial support of INVITRO, tells in detail how bioprinting takes place and why this body – VM was chosen for printing.

And now an organ transplant is ahead "The next stage of our work is to transplant a construct printed on a bioprinter to a mouse whose thyroid gland does not work.

The experiment will look like this: we plant the construct under the kidney capsule and after four weeks we measure the level of the hormone thyroxine in the blood. If the thyroxine level returns to normal, or it may be slightly below normal, then the construct works, it is functional," says Elena Bulanova, head of the Laboratory of Biotechnological Research "3D Bioprinting Solutions".

...and the seal of the human thyroid glandScientists are also making quite ambitious plans.

As Vladimir Mironov adds, in the near future they plan to print a human thyroid gland.

"Last year Sabine Costagliola from the Free University of Brussels received thyroid follicles from mouse embryonic stem cells. Then these follicles were implanted into the animal's body, where they earned and began to secrete the hormone thyroxine. Terry Davis recently did the same thing from human embryonic stem cells. Now we are waiting for the results of research when thyroid tissue will be grown from induced pluripotent human stem cells," says Vladimir Mironov.

Then, scientists hope, they will be able to start developing methods for printing the human thyroid gland.

Portal "Eternal youth" http://vechnayamolodost.ru30.03.2015

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