Stimulants and cell growth factors. Kinins - plant growth hormones Stimulation of cell division

The optimal stage for studying chromosomes is the metaphase stage, when the chromosomes reach maximum condensation and are located in one plane, which allows them to be identified with high accuracy. To study the karyotype, several conditions must be met:

Stimulation of cell divisions to obtain the maximum amount dividing cells,

- blocking cell division in metaphase;

- cell hypotension and preparation of a chromosome preparation for further research under a microscope.

Chromosomes can be studied cells from actively proliferating tissues(bone marrow cells, testicular walls, tumors) or cell cultures, which are obtained by cultivation under controlled conditions on special nutrient media of cells isolated from the body (peripheral blood cells *, T lymphocytes, red bone marrow cells, fibroblasts of various origins, chorion cells, tumor cells)

* The technique of obtaining chromosome preparations from peripheral blood lymphocytes cultivated in isolated conditions is the most simple method and consists of the following steps:

Sampling of venous blood under aseptic conditions;

Adding heparin to prevent blood clotting;

Transfer of material to vials with a special nutrient medium;

Stimulation of cell divisions by adding phytohemagglutinin;

Culture incubation for 72 hours at a temperature of 37 0 C.

Blocking cell division at the metaphase stage achieved by introducing into the environment colchicine or colcemid – substances - cytostatics that destroy the spindle of division. Receipt preparations for microscopic analysis includes the following steps:

- hypotonization of cells, which is achieved by adding a hypotonic solution of potassium chloride; this leads to swelling of the cell, rupture of the nuclear envelope and dispersion of chromosomes;

- cell fixation to stop the life of the cell while maintaining the structure of chromosomes; for this, special fixatives are used, for example, a mixture of ethyl alcohol and acetic acid;

- staining of the drug according to Giemsa or the use of other staining methods;

- analysis under a microscope in order to identify numerical disturbances (homogeneous or in mosaic) and structural aberrations;

- photographing and excision of chromosomes;

- Identification of chromosomes and drawing up a karyogram (idiogram).

Stages of karyotyping Differential staining of chromosomes

At present, along with the routine methods of studying the karyotype, differential staining methods are used, which make it possible to reveal the alternation of stained and unstained bands in chromatids. They're called bands and havespecific andexact distribution due to the peculiarities of the internal organization of the chromosome

Differential staining methods were developed in the early 1970s and became an important milestone in the development of human cytogenetics. They have a wide practical application, because:

The alternation of bands is not random, but reflects internal structure of chromosomes for example, the distribution of euchromatic and heterochromatic regions rich in AT or GC DNA sequences, chromatin regions with different concentrations of histones and non-histones;

The distribution of bands is identical for all cells of one organism and all organisms of a given species, which is used for accurate species identification;

The method allows exactly identify homologous chromosomes, that are genetically the same and have a similar distribution of bands;

The method provides accurate identification of each chromosome, because different chromosomes have a different distribution of bands;

Differential staining reveals many structural abnormalities of chromosomes(deletions, inversions), which are difficult to detect by simple staining methods.

Depending on the method of chromosome pretreatment and staining technique, there are several methods of differential staining (G,Q,R,T,C). Using them, one can obtain an alternation of colored and uncolored bands - bands that are stable and specific for each chromosome.

Characterization of different methods of differential staining of chromosomes

Method name	Dye used	The nature of bands	Practical role
		Painted - heterochromatin; unpainted - euchromatin	Detection of numerical and structural anomalies of chromosomes
	Quinacrine (fluorescent dye)	Painted - heterochromatin; unpainted - euchromatin
Method R (reverse)		Stained - euchromatin; unpainted - heterochromatin	Detection of numerical and structural anomalies of chromosomes
	Giemsa or fluorescent dye	Stained centromeric heterochromatin	Analysis of chromosome polymorphism
	Giemsa or fluorescent dye	stained - telomeric heterochromatin	Analysis of chromosome polymorphism

Cell metabolism stimulators and regeneration stimulants: placenta extract, extract amniotic fluid, panthenol, extract medicinal leeches, salmon milk, marine plankton, pollen, bone marrow, embryonic cells, royal jelly of bees (apilak), DNA, RNA, growth factors, thymus, umbilical cord, bone marrow organ preparations, sea buckthorn oil, phytesestrogens, etc.

Growth factors are proteins and glycoproteins that have a mitogenic effect (stimulate division) on various cells. Growth factors are named after the cell type that was first shown to be mitogenic, but they are more a wide range actions and are not limited to one group of cells. Keratinocyte growth factor stimulates the division of keratinocytes. Appears with skin wounds. Epidermal growth factor - stimulates regeneration. Suppresses differentiation and apoptosis, provides re-epithelialization of wounds. May induce tumor growth. Heparin-binding growth factor has an antiproliferative effect on keratinocytes. growth factor nerve cells stimulates the division of keratinocytes. At present, growth factors capable of activating human cell division have been isolated from milk whey, animal amniotic fluid, placenta, tissues of human embryos, invertebrate gonads, and mammalian sperm. Growth factors are used to activate mitoses in aging skin, accelerate the renewal of the epidermis and regenerate the skin.

Which substances stimulate cell renewal?

• vitamins,
• trace elements,
• amino acids,
• enzymes,

It can be: vit. A, E, C, F, zinc, magnesium, selenium, sulfur, silicon, vit. group B, biotin, glutathione, protease, papain, etc.

Substances that increase skin turgor and elasticity, elastostimulants (sulfur, vitamin C, chondroitin sulfate, hyaluronic acid, collagen, silicon, glucosamines, retinoids and retinoic acid, fibronectin, phytoestrogens, cellular cosmetics, etc.).

Retinoids

Retinoids are natural or synthetic compounds that exhibit a similar effect to retinol (Vit. A). The effect of retinoids on the skin: exfoliating, brightening, increasing firmness and elasticity, smoothing wrinkles, reducing inflammation, wound healing, side effect- annoying. Retinoids cause simultaneous thickening of the epidermis and exfoliation of the stratum corneum, accelerating the renewal of keratinocytes. Retinoid groups:

• Non-aromatic retinoids - retinaldehyde, tretinoin, isotretinoin, trans-retinol in - glucuronide, fentretinide, esters of retinoic acid (retinyl acetate, retinyl palmitate).
• Monoaromatic retinoids - etretinate, trans-acitretin, motretinide.
• Polyaromatic retinoids - adapalene, tazarotene, tamibarotene, arotinoid methylsulfone.

In external medicinal and cosmetics ah, retinol, retinol palmitate, retinaldehyde, tretinoin, retinoic acid esters, isotretinoin are used to correct aging; tretinoin, isotretinoin, arotinoid methylsulfonate, fenretinide are used to correct photoaging; tretinoin, isotretinoin, motretinide, adapalene are used to correct acne.

In single-celled organisms such as yeast, bacteria, or protozoa, selection favors each individual cell to grow and divide as quickly as possible. Therefore, the rate of cell division is usually limited only by the rate of absorption of nutrients from environment and processing them into the substance of the cell itself. In contrast, in a multicellular animal, the cells are specialized and form a complex community, so that the main task here is the survival of the organism, and not the survival or reproduction of its individual cells. In order for a multicellular organism to survive, some of its cells must refrain from dividing, even if there is no shortage of nutrients. But when the need arises for new cells, for example, when repairing damage, previously undivided cells must quickly switch to the division cycle; and in cases of continuous "wear and tear" of the tissue, the rates of new formation and cell death must always be balanced. Therefore, there must be complex regulatory mechanisms more high level than the one that operates in such simple organisms as yeast. This section is devoted to such "social control" at the level of a single cell. In ch. 17 and 21 we will get acquainted with how it functions in a multicellular system to maintain and renew body tissues and what its violations occur in cancer, and in ch. 16 see how even more a complex system controls cell division in the processes of individual development.

13.3.1. Differences in the frequency of cell division are due to the different duration of the pause after mitosis

Cells human body, whose number reaches 1013, are divisible with very different speeds. Neurons or skeletal muscle cells do not divide at all; others, such as liver cells, usually divide only once every one or two years, and some epithelial cells intestines,

Rice. 13-22. Cell division and migration in the epithelial lining small intestine mice. All cell divisions occur only in the lower part of the tubular invaginations of the epithelium, called crypts. The newly formed cells move up and form the epithelium of the intestinal villi, where they digest and absorb nutrients from the intestinal lumen. Most of the epithelial cells have a short life span and are shed from the tip of the villus no later than five days after leaving the crypt. However, a ring of approximately 20 slowly dividing "immortal" cells (their nuclei are isolated over dark color) remain associated with the base of the crypt.

These so-called stem cells give rise to two daughter cells during division: on average, one of them remains in place and then functions again as undifferentiated stem cell, while the other migrates upward, where it differentiates and becomes part of the epithelium of the villus. (Amended from C. S. Pptten, R. Schofield, L G. Lajtha, Biochim. Biophys. Acta 560: 281-299, 1979.)

to ensure constant renewal of the internal lining of the intestine, they divide more than twice a day (Fig. 13-22). Most vertebrate cells are located somewhere in this time range: they can divide, but usually do not do it as often. Almost all differences in the frequency of cell division are due to the difference in the length of the interval between mitosis and S-phase; slowly dividing cells stop after mitosis for weeks and even years. On the contrary, the time it takes a cell to go through the series of stages from the beginning of the S-phase to the end of mitosis is very short (usually 12 to 24 hours in mammals) and surprisingly constant, whatever the interval between successive divisions.

The time spent by cells in a non-proliferating state (the so-called G0 phase) varies depending not only on their type, but also on the circumstances. Sex hormones induce cells in the uterine wall to divide rapidly over several days each. menstrual cycle to replace tissue lost during menstruation; blood loss stimulates progenitor proliferation blood cells;

damage to the liver causes the surviving cells in that organ to divide once or twice a day until the loss is replaced. Similarly, the epithelial cells surrounding the wound begin to rapidly divide to repair the damaged epithelium (Fig. 13-23).

To regulate the proliferation of cells of each type in accordance with the need, there are carefully debugged and highly specific mechanisms. However, although the importance of such regulation

Alberts B., Bray D., Lewis J., Raff M., Roberts K. Watson J.D. Molecular biology of the cell: In 3 vols. 2nd ed. revised and additional T. 2.: Per. from English. – M.: Mir, 1993. – 539 p.

Rice. 13-23. Proliferation of epithelial cells in response to injury. The lens epithelium was damaged with a needle and after certain time 3H-thymidine was added to label cells in S phase (highlighted in color); then again fixed and prepared preparations for r.dioautography. In the schemes on the left, the areas with cells in the S phase are highlighted in color, and those with cells in the M phase are marked with crosses; the black spot in the center is the site of the wound. The stimulation of cell division gradually propagates from the wound, involving resting cells in the G0 phase, leading to an unusually strong response to relatively little injury. On a 40-hour preparation, cells far from the wound enter the S phase of the first division cycle, while cells near the wound itself enter the S phase of the second division cycle. The figure on the right corresponds to the area enclosed in the diagram on the left in a rectangle; it was taken from a photograph of a 36-hour preparation stained to reveal cell nuclei. (After C. Harding, J. R. Reddan, N. J. Unakar, M. Bagchi, Int. Rev. Cytol. 31: 215-300, 1971.)

obvious, its mechanisms are difficult to analyze in the complex context of the whole organism. Therefore, a detailed study of the regulation of cell division is usually carried out in cell culture, where it is easy to change the external conditions and long time watch the cells.

13.3.2. When conditions for growth become unfavorable, animal cells, like yeast cells, stop at a critical point in G1 - at the restriction point.

When studying cell cycle In vitro, in most cases, stable cell lines (Section 4.3.4) are used, which can multiply indefinitely. These are lines specially selected for maintenance in culture; many of them are so-called untransformed cell lines are widely used as models for the proliferation of normal somatic cells.

Fibroblasts (such as various types of murine 3T3 cells) usually divide faster if they are not too densely packed in the culture dish and culture media is used that is rich in nutrients and contains serum - fluid obtained during blood clotting and purified from insoluble clots and blood cells. When there is a shortage of some important nutrients, such as amino acids, or when an inhibitor of protein synthesis is added to the medium, the cells begin to behave in much the same way as the yeast cells described above with a lack of nutrition: average duration phases GT increases, but all this has almost no effect on the rest of the cell cycle. Once a cell has passed through G1, it inevitably and without delay passes through the S, G2, and M phases, regardless of environmental conditions. This transition point in the late G1 phase is often referred to as restriction point(R) because this is where the cell cycle can still be suspended if external conditions prevent it from continuing. The restriction point corresponds to the starting point in the yeast cell cycle; as in yeast, it may partly serve as a mechanism for regulating cell size. However, in higher eukaryotes, its function is more complex than in yeast, and in the phase G 1, there may be several slightly different restriction points associated with different mechanisms for controlling cell proliferation.

Alberts B., Bray D., Lewis J., Raff M., Roberts K. Watson J.D. Molecular biology of the cell: In 3 vols. 2nd ed. revised and additional T. 2.: Per. from English. – M.: Mir, 1993. – 539 p.

Rice. 13-24. The scatter in cell cycle durations commonly observed in homogeneous population of cells in vitro. Such data is obtained by observing individual cells under a microscope and directly marking the time between successive divisions.

13.3.3. The duration of the cycle of proliferating cells, apparently, has a probabilistic character.

Individual cells dividing in culture can be continuously observed using time-lapse filming. Such observations show that even in genetically identical cells, the duration of the cycle is highly variable (Fig. 13-24). Quantitative analysis shows that the time from one division to the next contains a randomly changing component, and it changes mainly due to the G1 phase. Apparently, as cells approach the restriction point in GJ (Fig. 13-25), they must “wait” for some time before moving on to the rest of the cycle, and for all cells, the probability per unit time to pass the point R about the same. Thus, cells behave like atoms when radioactive decay; if in the first three hours half of the cells passed through point R, in the next three hours half of the remaining cells will pass through it, after another three hours - half of those that remain, and so on. A possible mechanism explaining this behavior was proposed earlier, when it was about the formation of the S-phase activator (Sec. 13.1.5). However, random changes in the duration of the cell cycle mean that the initially synchronous cell population will lose its synchrony after several cycles. This is inconvenient for researchers, but may be beneficial for a multicellular organism: otherwise, large clones of cells could undergo mitosis at the same time, and since cells during mitosis usually round and lose their strong connection with each other, this would seriously damage the integrity of the tissue, which consists of such cells.

It is known that some cells are continuously dividing, for example bone marrow stem cells, cells of the granular layer of the epidermis, epithelial cells of the intestinal mucosa; others, including smooth muscle, may not divide for several years, and some cells, such as neurons and striated muscle fibers, are not able to divide at all (except in utero).

In some tissue deficiency of cell mass eliminated by rapid division of the remaining cells. So, in some animals, after surgical removal of 7/8 of the liver, its mass is restored almost to the initial level due to cell division of the remaining 1/8 part. Many glandular cells and most cells of the bone marrow, subcutaneous tissue, intestinal epithelium and other tissues have this property, with the exception of highly differentiated muscle and nerve cells.

So far, little is known how the body maintains the necessary number of cells different types . Nevertheless, experimental data indicate the existence of three mechanisms of cell growth regulation.

Firstly, division of many types of cells is under the control of growth factors produced by other cells. Some of these factors come to the cells from the blood, others - from nearby tissues. Thus, the epithelial cells of some glands, such as the pancreas, cannot divide without a growth factor produced by the underlying connective tissue.

Secondly, most normal cells stop dividing when there is not enough room for new cells. This can be observed in cell cultures, in which cells divide until they begin to contact each other, then they stop dividing.

Thirdly, many tissue crops stop growing if even a small amount of substances produced by them gets into the culture liquid. All these mechanisms of cell growth control can be considered as variants of the negative feedback mechanism.

Cell size regulation. Cell size depends mainly on the amount of functioning DNA. So, in the absence of DNA replication, the cell grows until it reaches a certain volume, after which its growth stops. If colchicine is used to block the formation of the fission spindle, then mitosis can be stopped, although DNA replication will continue. This will lead to the fact that the amount of DNA in the nucleus will significantly exceed the norm, and the volume of the cell will increase. It is assumed that overgrowth cells in this case is due to increased production of RNA and protein.

Cell differentiation in tissues

One of growth characteristics and cell division is their differentiation, which is understood as a change in their physical and functional properties during embryogenesis in order to form specialized organs and tissues of the body. Consider interesting experiment to help explain this process.

If from eggs frogs using a special technique to remove the nucleus and place the nucleus of a cell of the intestinal mucosa instead, then a normal frog can grow from such an egg. This experiment shows that even highly differentiated cells, such as cells of the intestinal mucosa, contain all the necessary genetic information for the development normal organism frogs.

It is clear from the experiment that differentiation is not due to loss of genes, but due to selective repression of operons. Indeed, on electron micrographs one can see that some DNA segments “packed” around histones are condensed so strongly that they can no longer be untwisted and used as a template for RNA transcription. This phenomenon can be explained as follows: at a certain stage of differentiation, the cellular genome begins to synthesize regulatory proteins that irreversibly repress certain groups of genes, so these genes remain inactivated forever. Howbeit, mature cells The human body is capable of synthesizing only 8,000-10,000 different proteins, although if all the genes were functioning, this figure would be about 30,000.

Embryo experiments show that some cells are able to exercise control over the differentiation of neighboring cells. Thus, the chordomesoderm is called the primary organizer of the embryo, since all other tissues of the embryo begin to differentiate around it. Transforming during differentiation into a segmented dorsal mesoderm consisting of somites, the chordomesoderm becomes an inductor for the surrounding tissues, triggering the formation of almost all organs from them.

As another example of induction can lead to the development of the lens. When the eye vesicle comes into contact with the head ectoderm, it begins to thicken, gradually turning into a lens placode, and this, in turn, forms an invagination, from which the lens is formed as a result. Thus, the development of the embryo is largely due to induction, the essence of which is that one part of the embryo causes the differentiation of the other, and that one causes the differentiation of the remaining parts.
So, although cell differentiation in general still remains a mystery to us, many of the regulatory mechanisms that underlie it are already known to us.

The rejuvenation procedures performed by a cosmetologist to improve the appearance of the face and eliminate wrinkles are based on the regeneration of skin cells, which must be stimulated. To do this, there are many cosmetic products and procedures, the action of which is aimed at activating cellular processes in such layers of the skin as the epidermis and dermis, as well as accelerating the production of collagen and elastin. Methods and means of rejuvenation are selected taking into account the ability of the skin to respond to stimulating effects.

Some Causes of Slow Skin Cell Regeneration

Slow cell renewal in aging skin occurs due to a decrease in the rate of their division in the basal layer, as well as due to a violation of the process of desquamation of the stratum corneum scales. As a result, the barrier function of the skin is disrupted, the number of defective cells in the epidermis increases, and the overall appearance skin.
The dermis suffers from external damaging environmental factors not much less than the epidermis, and therefore also needs to be updated. Fibroblasts of this skin layer constantly synthesize elastin and collagen fibers, hyaluronic acid, other glycosaminoglycans and also constantly destroy them, supporting skin regeneration processes. Over time, fibroblasts lose the ability to synthesize intercellular substance as quickly as before, and the rate of renewal of the dermis slows down.

Possible ways to stimulate skin cell regeneration

Today, studies of the possibilities of stem cells, capable of almost endless division, have become promising. It is generally accepted that epidermal stem cells are located in the bulge hair follicle, which is confirmed by some experiments, during which scientists managed to grow a fragment of a full-fledged skin from the cells of a hair follicle.

In addition, the cells of the basal layer of the skin are capable of intensive division, and it is precisely those cells that are located in areas of the epidermis deep into the dermis. The rate of renewal of the epidermis depends on the rate of cell division of the basal layer, but not directly, since they divide much faster than necessary. This feature of the basal layer is explained by the need to create some reserves in case of skin damage and the need for immediate regeneration of skin cells. Under normal conditions, the epidermis inhibits this process by producing chalons that inhibit cell division and maintains the optimal thickness of the stratum corneum.

With any damaging effect on the skin, the rate of division of basal cells increases. If the damage occurs in a small area, thickening of the skin occurs locally (a vivid example of this is the formation of calluses on the worn area of ​​the foot). Damage to the skin over a large area causes acanthosis - a general thickening of the epidermis (for example, after excessive insolation, the skin on the body becomes rougher and denser).

Methods and means of stimulating the regeneration of skin cells

In cosmetology, one of the ways to stimulate the skin to regenerate - peeling - is based precisely on this property of the skin to respond to its damage by active cell division of the basal layer. Another way to signal them to reproduce intensively is to use cytokines and retinoids.

Cytokines are mediators of a protein nature that are involved in intercellular signaling, regulate cell proliferation and differentiation. Retinoids are able to directly stimulate epidermal cells to divide and differentiate, as well as weaken the bonds between the cells of the stratum corneum, which contributes to their exfoliation.

Phytoestrogens- Another remedy that stimulates the regeneration of skin cells. Phytoestrogens can speed up cell renewal if cells have become slower to divide due to insufficient hormonal stimulation.

Stimulation of epidermal cells for renewal promotes the activation of dermal fibroblasts, which leads to an increase in the synthesis of collagen and elastin. The following substances can act as such stimulants in the composition of cosmetics that accelerate the regeneration of skin cells and help smooth out certain types of wrinkles:

• N-acetyl-L-cysteine ​​(sulfur-containing amino acid);
• gamma-aminobutyric acid;
• unsaponifiable fractions of avocado oil, soybean oil;
• yeast wall polysaccharides;
• purified aloe gel polysaccharides;
• L-ascorbic acid.

The choice of means and method for the regeneration of skin cells and its overall rejuvenation, as well as for its treatment in case of damage or UV radiation, depends on the severity of the signs of aging or the nature of the damage, as well as the ability of the skin to respond to stimulating actions. If cell degradation due to age or influence external factors has gone too far and the skin does not respond to cosmetic treatment, more intensive anti-aging treatments or the help of plastic surgery will be needed.