What? Can a "human kidney" grow in a pig's body? Chinese scientists have done it!

In movies and TV dramas, we often see plots where the protagonist needs an organ transplant. These plots are actually not far away from us.

Every year, there are about 300,000 patients waiting for organ transplants due to end-stage organ failure, but the number of organ transplants is only about 20,000 per year. If we can produce enough suitable organs like producing auto parts, we may be able to save countless patients in need of treatment and even achieve "immortality".

In September 2023, Chinese scientists published a cover paper in the international academic journal Cell Stem Cell, stating that they had successfully cultivated a humanized mid-stage kidney in a pig. This is the first reported case of xenogeneic in vivo cultivation of a humanized functional organ in the world.

Image source: Cover of the current issue of Cell Stem Cell

How does one go from a cell to an organ?

A seed can grow into a towering tree through cultivation, and the "seed" that develops into a human is called a fertilized egg. The fertilized egg is an omnipotent cell formed by the fertilization of sperm and egg. The so-called omnipotence is because this cell can generate 40 to 60 trillion cells with the same genetic material but the same or different functions through cell proliferation and differentiation.

Among them, cells of the same type gather together to form tissues with certain structures and functions , such as our muscle tissue is formed by the aggregation of muscle cells. The organic combination and growth and development of different functional tissues will form organs with certain shapes and functions , such as the heart is composed of myocardial tissue, endocardial tissue and pericardial tissue.

Image source: pexels

Although organ formation is completed during the embryonic and fetal period, and most organs of adults no longer undergo large-scale regeneration and reconstruction, some stem cells are retained in the body for the repair of damaged organs . For example, our blood cells are derived from the division and differentiation of bone marrow hematopoietic stem cells, which is also the root of bone marrow transplantation for the treatment of leukemia. Skin injuries are supplemented by differentiation of stem cells in the basal layer of the skin, which is also the key to wound healing.

In principle, we can cultivate organs in vitro by obtaining stem cells , but organs are not a physical superposition of cells; they require precise physiological regulation.

How is organ culture in vitro achieved?

Understanding the developmental regulation mechanism of organs is the prerequisite for in vitro organ culture . Thanks to the efforts of countless scientists, we have a preliminary understanding of the developmental regulation mechanism of organs, and organs with relatively simple structures such as skin can also be cultured in vitro.

In 2022, the Burn Surgery Department of Shanghai Changhai Hospital successfully carried out the first "autologous epidermal cell transplantation technology" to repair large-area burn wounds in China. The surgical team first cut 5 square centimeters of normal skin from the patient and then cultured it in a cell culture matrix.

Under the action of growth regulatory factors, autologous epidermal cells expanded to more than 4,000 times their original number within 2 weeks, and eventually the formed autologous epidermal cell membrane was transplanted to the patient. The initial recovery time of 2 to 3 months for conventional patients was shortened to one month, and the skin damage repair rate was as high as 100%, and the patient was able to get out of bed and walk normally.

For organs that are more complex than the skin, scientists have used biomedical materials to complete in vitro or in vivo culture of organs such as muscles, bones and ovaries. Taking bone tissue regeneration as an example, scientists first use biomedical materials to build a cell scaffold that defines the three-dimensional space in which stem cells survive. Then, isolated autologous osteoblasts, bone marrow stromal stem cells, chondrocytes and other stem cells are attached to the cell scaffold for culture. Under the stimulation of regulatory factors and the action of nutrients, these stem cells grow on the prefabricated three-dimensional scaffold to form bone for transplantation.

Schematic diagram of tofu-based hydrogel scaffolds used for bone tissue regeneration and repair. Source: References [4][5]

For more complex organs, although scientists have successfully grown mini hearts and mini brains in the laboratory, due to the lack of thorough research on the development mechanism of organs, it is still impossible to fully grow a human organ of suitable size and complete functions for transplantation in the laboratory. Therefore, xenotransplantation and the use of other species to grow human organs have become one of the focuses of scientists' research.

However, there are serious safety issues in xenotransplantation or cultivation, the most important of which is immune rejection. The immune system is our body's defense department. Its function is to identify "self" and "non-self", protect "self" components such as organs in the body, and kill "non-self" components such as viruses, bacteria and xenotransplants. The main cause of death in patients who have received xenotransplantation is immune rejection.

With the development of biotechnology, gene editing technology provides a solution to reduce immune rejection and improve the survival rate of organ transplantation.

Pigs grew "human kidneys"?

Through continuous efforts, scientists have gained a deeper understanding of the functions of genes responsible for immune rejection and kidney formation.

In this study, scientists used gene editing technology to knock out the pig's gene responsible for immune rejection to obtain an immunodeficient pig model. But even without immune rejection, it is still a big problem to make human stem cells survive in pigs and differentiate into humanized kidneys in pigs.

In order to obtain chimeric stem cells with strong survival and differentiation abilities in pigs, the research team conducted in-depth research on the interspecies difference barrier mechanism and found that overexpression of the two genes MYCN and BCL2 can promote better survival of human stem cells in pigs. By adding differentiation-promoting compounds and regulatory factors to the stem cell culture medium, kidney stem cells (4CL/N/B cells) with both survival and differentiation abilities were finally obtained.

Image source: Reference [1]

After solving the key problem of cell survival, how to make human stem cells transplanted into pigs replace pig stem cells and develop into "humanized kidneys" is another difficult problem faced by scientists. The research team decided to knock out several key genes responsible for pig kidney development to obtain kidney-deficient pigs, thereby providing sufficient development space for human cells to "replace" them. This technology, called embryo compensation, has always been a key problem in xenotransplantation organ culture.

After close cooperation and repeated attempts by multiple research teams, the best solution for cultivating humanized kidneys was finally determined: injecting 3 to 5 human stem cells from the morula to the early blastocyst stage of embryonic development can maximize the possibility of obtaining chimeric embryos. At the same time, in order to solve the problem of difficulty in culturing chimeric embryos in vitro due to the different culture conditions of human and pig cells, the research team determined the best culture conditions for chimeric embryos through a lot of exploration. With the concerted efforts of multiple teams, the kidneys of 5 of the 1,820 transplanted embryos developed to the second stage, and the autopsy found that 70% of the kidney cells came from humans, indicating that the chimeric kidneys were successfully cultivated.

The process of organoid cultivation. Image source: Reference [1]

In fact, in addition to pigs, there are many animals used for organoid research, such as monkeys, mice, goats, etc. However, whether it is xenotransplantation or organ cultivation, pigs are the most common organ donors, because compared with other animals, pigs and humans have organs of similar size, similar structure and function, which is easy to match. At the same time, pigs, as mature livestock, have a shorter breeding cycle, lower breeding costs, and fewer ethical issues than primates.

What is the use of culturing organoids?

In addition to kidneys, other organoid studies include liver, heart, lungs, intestines, etc. Organoid research is still in its infancy, the technology is very immature, and there is still a long way to go before it can be applied to organ transplantation.

Heart organoids. Image source: Reference [2]

However, organoids have achieved initial success in the study of organ development and disease mechanisms, disease modeling, and drug screening . For example, in the field of drug screening, the development of conventional drugs requires multiple stages such as animal experiments and clinical research. Not only is the development time long and the cost high, but some drugs are highly toxic and may cause death to humans and animals.

Using organoids to study toxicity and efficacy is not only closer to the real effects in the human body than animal experiments, but also can shorten the R&D cycle and save R&D costs. For example, the dual-antibody drug MCLA-158, which is currently in clinical phase II, was screened through organoids, and the clinical results are excellent. The organoid model has also achieved initial results in the screening of drugs for colorectal cancer and pulmonary fibrosis.

Application of organoids, source: Reference [3]

Although organoids have broad application prospects, we also need to be aware of the challenges and limitations. For example, our understanding of organ development is still incomplete, organoids cannot simulate the physiological functions of real organs, and the technology for cultivating organoids is not yet mature.

But in general, organoids have broad application prospects in drug development, precision medicine, regenerative medicine, etc. due to their unique advantages and potential. They may play an important role in future medical research and contribute to human health.

References

[1] Cell Stem Cell. 2023 Sep 7;30(9):1235-1245.e6. doi: 10.1016/j.stem.2023.08.003.

[2] Lee, J., et al. In vitro generation of functional murine heart organoids via FGF4 and extracellular matrix. Nat Commun 11, 4283 (2020).

[3] Corrò, C., Novellasdemunt, L. & Li, VSW A brief history of organoids. Am J Physiol Cell Physiol 319, C151-C165 (2020).

[4] Huang Keqing; Liu Guiting; Gu Zhipeng; Wu Jun. Tofu as excellent scaffolds for potential bone regeneration. Chinese Chemical Letters (2020)

[5]Huang, Keqing; Gu, Zhipeng; Wu, Jun.Tofu-Incorporated Hydrogels for Potential Bone Regeneration.ACS BIOMATERIALS SCIENCE &ENGINEERING(2020)

Planning and production

Source: Shanghai Science and Technology Museum (ID: sstm01)

Author: Hu Zhiguo, PhD, Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences

Editor: Bai Li