Transplantation of donor tissue and organs entails a number of difficulties, the main of which is the deficit of donor material. The deficit of donor tissue and organs has stimulated the development of tissue engineering that designs bioartificial substitutes of donor material. Such substitutes are based on carcasses (scaffolds) that imitate the acellular component of connective tissue. Tissue-specific cells multiply and differentiate themsleves on the surface of the scaffolds, which results in the formation of tissue in properties similar to the lost tissue.
The first attempts to populate scaffolds with cells date back to the 1980s when enterocytes were used to grow intestinal tissue on biodegradable polymers. Simultaneously the issue came up to use stem cells of various origins in regenerative medicine. Further research revealed that stem cells circulating in the blood stream as well as the stem cells of bone marrow could differentiate into different types of cells: hepatocytes, epithelial skin cells and gastrointestinal tract cells. That led scientists to assume that stem cells could be used in tissue engineering structures.
Today regenerative medicine uses differentiated (mature) cells migrating from the surrounding tissue as well as progenitor cells, the type of stem cells that are aimed at differentiation into a specific cell type, as a source of cells that form the tissue of such structures.
The dangers of the “gold standard” of intestinal transplantology
The consequences of a partial enterectomy present one of the most serious medical problems. An extensive intestinal resection leads to a disruption in the absorption of nutrients and liquids, diarrhea, dehydration and progressive debilitation. Today, the most effective help to patients in need of an extensive intestinal resection and patients suffering from the consequences of such resection is transplanting engineered tissue intestines.
The “gold standard” of such surgeries implies the use of implants based on heterologous decellularized small intestine (all cells have been removed with preserved extracellular matrix). A donor organ is called heterologous (or alien) as it is taken from an animal (usually, a pig).
However, such method entails a risk of infecting the recipient’s organism with pathogens from the transplant’s biomaterial. Pathogen free animals are used as donors, but that only helps avoid bacterial infection, but not the risk of infecting the recipient with porcine endogenous retroviruses (PERV) or porcine cytomegalovirus (PCMV) capable of infecting human cells.
Viruses that are currently spread only among animals are also dangerous – for example, porcine circoviruses of type 1 and 2, virus that causes reproductive and respiratory syndrome, porcine encephalomyocarditis virus, swine flu virus, African fever virus, hepatitis E virus, herpes virus, parvoviruses and hamburger polyomaviruses. Viruses can acquire new characteristics during transplantation as a consequence of mutation and recombination with human viruses.
As a result there is a constant search for a mechanism for intestinal tissue engineering.
One of the promising materials for the development of bioartificial tissues is fibroin – carcass silk protein produced by silkworm Bombyx mori. Silk is widely used in medicine as sutures and has a number of unique properties – it is mechanically strong, is biocompatible with the body tissue (does not cause immune rejection, non-toxic) and most importantly, is capable of bio-resorption, i.e. with time is destroyed by the body cells and the products of its decomposition can be naturally metabolized by the body.
A number of studies revealed that fibroin is a good substratum for adhesion and proliferation of various types of cells. Scaffolds of various structures, of required form and size can be developed on the basis of fibroin.
The MSU School of Biology in cooperation with the surgeons from the Vladimirsky Scientific Research Clinical Institute of the Moscow Region have conducted a number of successful experiments using fibroin scaffolds preliminarily populated (vitalized) by bone marrow stem cells to regenerate the walls of jejunum in a model rat with damaged jejunum wall.
In the experiment the researchers took out a fragment of the experimental animals’ jejunum of the 0.8 x 0.6 cm size and sew in that place a fibroin implant with bone marrow stem cells. All animals survived the surgery. When a few months later a histological analysis of the transplantation area was performed the surgery area could be detected only by the remaining fragments of non-absorbable sutures.
The implantation area demonstrated complete restoration of all the layers of the intestines, the formation of the mucous with numerous crypts and villi, the muscular layer with transverse and longitudinal fibers necessary for intestinal motility, as well as with blood vessels. The most amazing thing the researchers observed was the restoration of the intestinal innervation, the formation of the nerve fibers and nerve plexus, which had been deemed impossible before.
Thus, silk fibroin implants represent an ideal matrix that provides intestinal impermeability and favorable mechanical and physiological conditions for reparative processes.
New transplants and new questions
One can assume that both the stem cells introduced into the engineered tissue structure and tissue-specific cells that migrated here from undamaged tissue took part in the formation of tissue in the place of tissue damage.
But the opportunity to form neurons de novo in regenerated tissue still remains debatable.
Until recently the possibility of neurogenesis in an adult organism was strongly denied until research proved the existence in certain part of the brain of stem nerve cells with mitotic activity, i.e. stem nerve cells with cell division and consequently proliferation.
Scientists easily came to agreement regarding the existence of nerve stem cells in nerve tissue, but the possibility of the formation of neuron of ectodermal origin from the bone marrow stem cells of mesenchyme origin is still being contested.
Ectoderm is the outer germ layer that serves to form epidermis and the nervous system, mesoderm is the middle germ layer that the skeleton, muscles and internal organs originate from, which the internal germ layer of endoderm turns into cells that coat the intestines from the inside. According to the dogma of embryology the borderline between them cannot be violated during the organism development.
However, there are already articles that describe not only the differentiation of mesenchyme stem cells into mesenchyme cells, but also into neuroectoderm and endoderm cells. In addition, populations of multipotent stem cells have been isolated from human and rodent intestines, which are capable of differentiating into neurons and glial cells. This means that there is at least a hypothetical possibility of neurogenesis de novo from bone marrow and intestinal stem cells, which explains the amazing success of using fibroin scaffolds populated by bone marrow stem cells.
Thus, the creation of the new engineered tissue structure and the demonstration of its capabilities in intestinal regeneration on the one hand opens the prospects for practical use in transplantology of fibroin based scaffolds and on the other hand opens a new area of research – the search for new mechanisms of regeneration of intestinal tissue and the role of stem cells in this process.
Anna Goncharenko, PhD in Biology, Moscow State University