Application of cellular technologies to improve the appearance of scars

Modern science is characterized by the rapid development of a number of related disciplines, united under the general name of "biotechnology". This section of science, based on the latest achievements in the field of biology, cytology, molecular genetics, genetic engineering, transplantology, aims to use the enormous potential inherent in plant and animal cells - the basic structural units of all living things. "A living cell is a ready-made biotechnological reactor in which not only the processes leading to the formation of the final product are realized, but also a number of others that help maintain the catalytic activity of the system at a high level," - John Woodward, 1992. The beginning of cell science was laid in 1665, when the English physicist R. Hooke created the first microscope and discovered cells - cellulae ("cells") in a cork. In 1829, M. Schleiden and T. Schwann substantiated the "cell theory", which proved that all living things consist of cells. In 1858, R. Virchow proved that all diseases are based on a violation of the structural organization and metabolism of cells. He became the founder of "cellular pathology". A fundamental contribution to the science of the cell was made in 1907-1911 by R. Harrison and A.A. Maximov, who proved the possibility of culturing cells outside the body. Their work showed that for cell cultivation, animal tissues and plant parts must be mechanically separated into small pieces. To isolate cells, tissues are cut with a sharp knife or microtome into thin sections, approximately 0.5-1.0 mm. Physical separation of cells is called immobilization. Isolated cells are obtained by enzymatic dispersion of pieces of plants or tissues. After grinding with sharp scissors, the pieces are treated with trypsin or collagenase to obtain a suspension - a suspension of individual cells or their microaggregates in a special medium. Alginate gels (calcium alginate) are widely used to immobilize plant cells. It has been proven that immobilized plant and animal cells retain the ability to biosynthesize. Cellular biosynthesis products accumulate in cells, their expression occurs either spontaneously or with the help of special substances that promote increased permeability of cell membranes.

Cultivation of animal cells is a much more complex process than culturing plant cells, requiring special modern equipment, high technology, the presence of various media, growth factors designed to preserve the viability of cells and maintain them in a state of high functional activity. It was found that most cells of solid tissues, such as kidney, liver, and skin tissues, are surface dependent, so they can be cultivated in vitro only in the form of thin sheets or monolayers directly associated with the surface of the substrate. The life span, proliferation, and functional stability of cells obtained by enzymatic dispersion of tissues largely depend on the substrate on which they are grown. It is known that all cells obtained from vertebrate tissues have a negative surface charge, so positively charged substrates are suitable for their immobilization. Isolated cells obtained directly from whole tissues can be maintained in a primary culture in an immobilized state while maintaining high specificity and sensitivity for 10-14 days. Immobilized, surface-dependent cells play a major role in biology today, especially in clinical research. They are used to study cell development cycles, regulation of their growth and differentiation, functional and morphological differences between normal and tumor cells. Immobilized cell monolayers are used in biotests, for quantitative determination of biologically active substances, as well as for studying the effect of various drugs and toxins on them. Doctors of all specialties have shown great interest in the cell as a therapeutic agent for decades. Cell technologies are currently rapidly developing in this direction.

The beginning of tissue and cell therapy is associated with the name of the famous Russian scientist V.P. Filatov, who in 1913 laid the foundations of the doctrine of tissue therapy, studying the results of corneal transplants from healthy donors to patients with cataracts. In the process of working with corneal transplants, he discovered that the cornea preserved in the cold for 1-3 days at a temperature of -2-4 degrees C takes root better than fresh. Thus, the property of cells to secrete some substances in unfavorable conditions that excite vital processes in transplanted tissues and regenerative ones in the recipient's tissues was discovered. Tissue and cells separated from the body are in a state of stress, that is, slow vital activity. Blood circulation in them stops, and therefore nutrition. Tissue respiration is extremely difficult, innervation and trophism are upset. Being in a new qualitative state, adapting to new conditions of existence, cells produce special substances with medicinal properties. These substances of non-protein nature were called biogenic stimulants by V.P. Filatov. He established together with V.V. Skorodinskaya that material from animals and plants can be freely autoclaved at t 120 degrees C for an hour after being kept in unfavorable conditions, and they not only did not lose activity, but on the contrary, increased it, which was explained by the release of biological stimulants from preserved tissues. In addition, they lost antigenic properties, which significantly reduced the possibility of rejection. Preserved sterile material was introduced into the body by implantation (plantation) under the skin or in the form of injections of extracts, with adequate results. It was also discovered that fetal tissues contain a significantly larger number of biologically active substances than the tissues of adult individuals, and some factors are found only in embryos. Inoculated fetal tissues are not perceived by the recipient's organism as foreign due to the absence of proteins responsible for species, tissue and individual specificity (proteins of the major histocompatibility complex) in the cytoplasmic membranes. As a result, inoculation of animal fetal tissues into the human organism does not trigger mechanisms of immune protection and reactions of incompatibility and rejection. V.P. Filatov widely used human placenta and skin in his medical practice. Treatment courses consisted of 30-45 injections of tissue extracts and 1-2 implantations of autoclaved tissues.

Having started his research with human and animal tissues and cells, he transferred his generalizations to the plant world. Conducting experiments with living parts of plants (aloe, plantain, agave, beet tops, St. John's wort, etc.), he created unfavorable conditions for them, placing cut leaves in a dark place, since the plant needs light for its vital functions. He also isolated biogenic stimulants from estuary mud and peat, due to the fact that mud and peat are formed with the participation of microflora and microfauna.

Tissue therapy received a new round of its development in the late 70s, when the knowledge and experience accumulated over decades allowed using animal and plant tissues and cells at a qualitatively new level to treat humans and prolong their active longevity. Thus, in some domestic clinics and a number of foreign ones, women in physiological menopause with climacteric syndrome or against the background of ovariectomy began to undergo tissue therapy with fetal tissues of the placenta, hypothalamus, liver, ovaries, thymus and thyroid glands to slow down the aging process, the development of atherosclerosis, osteoporosis, dysfunctions of the immune, endocrine and nervous systems. In one of the most prestigious gerontocosmetology clinics in Western Europe, injections of extracts obtained from fetal tissues of the gonads of rams have been used for the same purposes for several decades.

In our country, biostimulating treatment has also found wide application. Until recently, patients with various diseases were actively prescribed injections of placenta extracts, aloe, kalanchoe, sedum major (biosed), FiBS, peloid distillate, peloidin, peat, humisol prepared according to the method of V.P. Filatov. At present, it is almost impossible to buy these highly effective and inexpensive domestic tissue preparations of animal, plant and mineral origin in pharmacies.

The basis for obtaining various biogenic preparations from human tissues and organs of imported production, such as rumalon (from cartilage tissue and bone marrow), actovegin (from calf blood), solcoseryl (cattle blood extract), as well as domestic preparations - vitreous body (from the vitreous body of the eye of cattle), kerakol (from the cornea of cattle), splenin (from the spleen of cattle), epithalamin (from the epithalamic-epiphyseal region) are also the research of V.P. Filatov. The unifying property for all tissue preparations is the general effect on the entire body as a whole. Thus, “Tissue Therapy” by Academician V.P. Filatov formed the basis for most modern developments and directions in surgery, immunology, obstetrics and gynecology, gerontology, combustiology, dermatology and cosmetology related to the cell and the products of its biosynthesis.

The problem of tissue transplantation has concerned mankind since ancient times. Thus, in the Ebers papyrus, dated 8,000 BC, there is already a mention of the use of tissue transplantation to compensate for defects in individual areas of the body. In the "Book of Life" of the Indian scientist Sushruta, who lived 1,000 years BC, there is a detailed description of the restoration of the nose from the skin of the cheeks and forehead.

The need for donor skin grew proportionally to the increase in the number of plastic and reconstructive surgeries. In this regard, cadaveric and fetal skin began to be used. There was a need to preserve donor resources and find ways to replace human skin with animal tissues, and various skin modeling options. And it was in this direction that scientists worked when in 1941 P. Medovar first demonstrated the fundamental possibility of keratinocyte growth in vitro. The next important stage in the development of cellular technologies was the work of Karasek M. and Charlton M., who in 1971 carried out the first successful transplantation of autologous keratinocytes from a primary culture onto rabbit wounds, using collagen gel as a substrate for culturing CC, which improved cell proliferation in culture. J. Rheinvvald. H Green. developed a technology for serial cultivation of large quantities of human keratinocytes. In 1979, Green and his co-authors discovered the prospects for the therapeutic use of keratinocyte cell culture in the restoration of skin in cases of extensive burns, after which this technique, constantly being improved, began to be used by surgeons in burn centers abroad and in our country.

In the process of studying living cells, it was found that cells produce not only biogenic stimulators of non-protein origin, but also a number of cytokines, mediators, growth factors, polypeptides, which play an important role in regulating the homeostasis of the entire organism. It was found that various cells and tissues contain peptide bioregulators. which have a wide range of biological action and coordinate the processes of development and functioning of multicellular systems. The era of using cell culture as a therapeutic agent began. In our country, transplantation of fibroblast suspension and multilayered keratinocyte cell layers has been adopted in combustiology in recent decades. Such an active interest in transplanting skin cells to burnt patients is explained by the need for rapid closure of large burn surfaces and a shortage of donor skin. The possibility of isolating cells from a small piece of skin capable of covering a wound surface 1000 or even 10,000 times larger than the area of the donor skin has proven to be very attractive and important for combustiology and burn patients. The percentage of keratinocyte layer engraftment varies depending on the burn area, age and health of the patient from 71.5 to 93.6%. Interest in keratinocyte and fibroblast transplantation is associated not only with the possibility of quickly closing a skin defect, but also with the fact that these transplants have a powerful biologically active potential for improving the appearance of tissues obtained as a result of transplantation. New vessel formation, hypoxia relief, improved trophism, accelerated maturation of immature tissue - this is the morpho-functional basis for these positive changes occurring due to the release of growth factors and cytokines by the transplanted cells. Thus, due to the introduction of progressive cellular technologies for transplantation of multicellular layers of autologous and allogenic keratinocytes and fibroblasts onto large wound surfaces into medical practice, combustiologists were able not only to reduce the mortality rate of burn victims with a high percentage of skin lesions, but also to qualitatively improve the scar tissue that inevitably occurs at the site of IIb and IIIa and b degree burns. The experience of combustiologists obtained in the treatment of wound surfaces in burn patients suggested the idea of using the already modified Green method in dermato-surgical practice for various skin and cosmetic pathologies (trophic ulcers, vitiligo, nevi, bullous epidermolysis, tattoo removal, age-related skin changes, and to improve the appearance of scars).

The use of allogeneic keratinocytes in surgery, combustiology and dermatocosmetology has a number of advantages over the use of autologous keratinocytes, since the cellular material can be prepared in advance in unlimited quantities, preserved and used if necessary. It is also known that allogeneic CCs have reduced antigenic activity, since when cultivated in vitro they lose Langerhans cells, which are carriers of the HLA complex antigens. The use of allogeneic CCs is also supported by the fact that they are replaced by autologous ones after transplantation, according to various authors, within 10 days to 3 months. In this regard, cell banks have been created in many countries today, thanks to which it is possible to obtain cell transplants in the required quantity and at the right time. Such banks exist in Germany, the USA and Japan.

Interest in the use of cellular technologies in dermatocosmetology is due to the fact that "cellular compositions" carry a powerful bioenergetic and information potential, thanks to which it is possible to obtain qualitatively new treatment results. Autokines secreted by transplanted cells (growth factors, cytokines, nitric oxide, etc.) primarily act on the body's own fibroblasts, increasing their synthetic and proliferative activity. This fact is especially attractive to researchers, since the fibroblast is a key cell of the dermis, the functional activity of which determines the condition of all skin layers. It is also known that after skin injury with a cautery, laser, needle and other instruments, the skin is replenished with fresh stem precursors of fibroblasts from bone marrow, adipose tissue and capillary pericytes, which contributes to the "rejuvenation" of the pool of body cells. They actively begin to synthesize collagen, elastin, enzymes, glycosaminoglycans, growth factors and other biologically active molecules, which leads to increased hydration and vascularization of the dermis, improving its strength,