Anatomo-histologic characterization of myometrium at the end of pregnancy and in labor

It is important to note that studies aimed at studying the architecture of the myometrium and anatomical and histological studies of the structure of the uterus have shown that by the end of pregnancy the uterus increases in length to 36 cm, its width reaches 25 cm, and the thickness (anterior-posterior diameter) of the body is up to 24 cm.

The powerful layer of smooth muscles, which makes up the middle layer of the uterus along the course and direction of the fibers, is represented by three layers: the outer and inner - longitudinal and the middle - annular. The same layers continue into the cervix, gradually thinning, with the annular layer being especially thin.

It has been established that, starting from the 2nd month of pregnancy, the lumen of the isthmus begins to gradually expand, participating in the formation of the ovarian cavity, and this expansion is normally completed by the beginning of the 5th month, and from this moment until the end of pregnancy (in the absence of contractions), the internal os forms the boundary between the fetal receptacle, in which the lower uterine segment also participates, and the cervix, while the length of the isthmus by the end of pregnancy reaches 7 cm. The muscles of the uterus, starting from the 4th month of pregnancy, both in the area of the body and in the area of the lower segment of the uterus are located in parallel plates, and until the end of pregnancy, the muscles of the lower segment are little different from the muscles of the body, although in the latter it is still thicker. With the onset of contractions, the lower uterine segment progressively thins and stretches. The stretching zone of the uterus reaches the place of tight attachment of the peritoneum to the anterior wall of the organ. At the height of this place is located what is called the "contraction ring". Despite the stretching, the muscles of the lower segment of the uterus actively contract during and after childbirth. The tissues of the cervix change significantly during pregnancy, and the cervix itself turns into a cavernous body. The isthmus, which turns into the lower uterine segment during pregnancy, is an independent section of the uterus with certain boundaries, both macro- and microscopic, and certain anatomical and functional features. The upper border of the lower segment of the uterus corresponds to the site of tight attachment of the peritoneum to its walls. There is a significant functional difference between the properties of the muscle cells of the body of the pregnant uterus and its lower segment. It is believed that the muscle cells of these two sections belong to two different differentiated types, and this is seen as a certain functional parallelism with the data of anatomical studies. The muscle cells of the lower segment, or at least some of them, have the contractility characteristic of the muscle cells of the body of the uterus.

A number of interesting regularities have been revealed, which are confirmed in the clinical observations of modern authors. It has been shown that the round ligament has the form of a triangular band and is a muscular layer 5-7 mm thick in a non-pregnant state, while the ligament, unfolding when approaching the uterus, covers the anterior surface of its body almost completely, i.e., starting immediately below the attachment of the tubes and ending at the place where the peritoneum departs from the anterior surface of the uterus and which serves as the lower border of the body of the uterus. The muscular bundles of the ligament have a longitudinal direction in relation to the ligament.

If we follow further the distribution of the most superficial bundles that have passed from the ligament to the anterior surface of the uterus, we see that these bundles pass to the anterior surface of the uterus, being located in the transverse direction to its long axis. On the midline of the uterus, the muscle bundles of the ligaments of both sides, when meeting each other, bend mostly downwards and lie side by side. As a result, a large median bundle is formed along the midline of the anterior surface of the uterus, protruding above the level of the transverse bundles that have passed from the ligament.

In the peripheral parts of the right and left sides of the body of the uterus, the main direction of the bundles going from the outer layer of the anterior wall from front to back, perpendicular to the axis of the uterus. At the same time, the muscle bundles of the body of the uterus, located close to the border of the cervix, maintain this direction most consistently; it is here that these transverse bundles are the thickest, strongest and longest, so that they even go into the back wall of the cervix.

The same transverse muscle bundles are visible on a considerable thickness of the sides of the uterus, and they are especially abundant above the border between the body and the cervix.

The peculiarity of the arrangement of muscle bundles in the cervix is that the main mass of muscle bundles in the cervix is a direct continuation of the muscle bundles of the outer and vascular layers of the body of the uterus, and the entire complex of muscle bundles, occupying almost the entire thickness of the cervix, goes straight down. From this complex, muscle bundles depart individually inward, towards the mucous membrane, and they make bends, changing their direction to a more horizontal one, and such departure of individual muscle bundles inward is observed along the entire length of the cervix from top to bottom. The bent muscle bundles approach the mucous membrane in a direction perpendicular to it, wherever its surface is facing.

Due to such arrangement of muscle bundles during labor, the opening of the cervical canal is observed first and muscle bundles serve as real dilators of the cervix. At the same time, the submucosal layer of muscles, in the author's opinion, is so weak that it cannot, when contracting, counteract the stretching action of the described muscular system. In this case, it becomes clear why the opening of the cervix occurs gradually from above, starting from the internal os - the upper bundles are short and less curved, the first effect of the action of contraction of this group of muscles begins with them, as the cervix opens, the curved lower bundles gradually straighten, and only after their straightening does their stretching action begin. Such straightening of the muscles occurs sequentially, starting from the upper and ending with the lowest ones, opening the external os. The author makes a very important conclusion that there is no division into the contracting active (upper) and stretching passive parts.

It is believed that all parts of the uterus are active during labor: both during the opening of the cervix and during the forward movement of the fetus, the muscles of the lower segment and the cervix must take the most active part; the effect of the action of the muscles depends on the direction of the muscle bundles. In the peripheral layers of the sides of the uterus, a thick layer of transversely running strong muscle bundles, when contracting, narrows the cavity of the uterus in a direction transverse to its axis, and since the thickest and longest muscle bundles are at the level of the transition of the body into the cervix, and therefore the strongest action of this group of muscles should be immediately above the cervix.

The works of modern authors have shown that the uterine muscle is a complexly organized system of functionally ambiguous smooth muscle cells and it is necessary to take into account the possibility of its functional heterogeneity. From this position, the doctrine of the dependence of the functional state of the smooth muscle bundle on its spatial orientation in a separate layer is of particular interest, given that each layer of the female myometrium is represented by a three-dimensional network of muscle bundles. It is believed that obstetrician clinicians still do not sufficiently assess the state of the lower segment and cervix during pregnancy and childbirth, and the three layers have different spontaneous activity. At the same time, the inner and middle layers have similar activity, but in most cases the spontaneous activity of the inner layer is higher than that of the outer one. It was found that oxytocin has the effect of increasing the spontaneous uterine activity of all three layers. At the same time, the inner and middle layers (experiments on rats) contract with a high frequency and less intensity than the outer one. The author attributes this difference between the inner and middle layers to the fact that the outer layer has a different embryological origin. Based on these data, the author emphasizes that spontaneous uterine activity, including prenatal Braxton Hicks-type contractions of the uterus, initially begins in one or more areas of the inner layer and then transforms to other layers.

The latest studies on the physiology of the lower uterine segment during pregnancy, childbirth, the study of the histological picture of the neurovegetative receptors of the vaginal part of the cervix, the relationship between the structure of the myometrium and the effect of stimulation and suppression of contractile activity of the uterus have shown that the isthmus undergoes progressive hypertrophy and lengthening and its sphincter remains a very dense contraction ring, at least until the end of the 24th week of pregnancy. In this case, the lower segment of the uterus is formed entirely from the prolonged and hypertrophied isthmus. The upper isthmic sphincter begins to relax long before the lower sphincter and this is the result of a gradual unfolding of the isthmus from above downwards. In most primigravidas, the upper sphincter becomes completely relaxed approximately 3-4 weeks before childbirth. In women pregnant again, this is not observed until the first stage of labor and the head descends deep into the pelvic inlet as soon as the upper sphincter is completely relaxed. Changes are also noted during labor: smoothing of the cervix depends on the relaxation of the lower sphincter and with abnormal contractions of the isthmus, a slow advancement of the head and a slow opening of the cervix are observed. In this case, the resulting ring of constriction - cervical dystocia is the cause of local abnormal contractions of the upper or lower sphincter.

Currently, it is generally accepted that the myometrium is divided into 3 layers: subserous from longitudinal bundles, middle from circular bundles, and submucous from longitudinal bundles. The ideas about the direction of the muscle bundles in individual layers of the myometrium have changed somewhat in recent years. Thus, some authors indicate that the submucous (inner) muscle layer consists of circular (not longitudinal) bundles, and the middle (vascular) layer consists of muscle bundles running in various directions. Other authors have not found any pattern in the direction of muscle fibers in the uterine wall.

The study of the electromyographic activity of the cervix showed that its greatest activity was recorded at the time of contractions, basal activity - immediately after amniotomy and in the active phase of labor. With the least mature cervix, maximum electromyographic activity was noted after amniotomy, while no electromyographic discharges were recorded in the body of the uterus. When oxytocin is prescribed, these discharges are grouped, their intensity increases, they are synchronized with the onset of contractions. The ratio of discharges of the cervix and the body of the uterus is greater than one with an immature cervix and less than one with a mature one. As labor progresses, the electromyographic activity of the body of the uterus begins to predominate. At the beginning of labor after amniotomy, the greatest activity is noted in the cervix.

There are also two possible mechanisms for cervical dilation during labor:

• longitudinal contraction of the uterine walls, causing an increase in intrauterine pressure;
• radial tension as the head moves along the cervix.

Before this study, there was no method for separately measuring intrauterine pressure and radial tension. The authors designed a voltage transducer that responded minimally to an increase in intrauterine pressure. A probe with 4 such transducers was placed between the fetal head and the mother's cervix along the long axis of the fetus. The intrauterine pressure transducer at the end of the probe allowed for the simultaneous measurement of amniotic pressure. The possibility of radial tension in the opening of the cervix during labor was established.

Biochemical, biophysical, electron microscopic and X-ray structural characteristics of the contractile apparatus of the uterus at the end of pregnancy and during labor

A study of the main structural and functional substrate - uterine myocytes - showed that, compared to the end of pregnancy (38-40 weeks), during normal labor, myocytes are significantly increased in size, and “light” and “dark” cells are present in equal quantities.

An increase in the activity of respiratory enzymes - succinate dehydrogenase, cytochrome C oxidase and the total content of nucleic acids in the mitochondria was revealed, indicating a high level of oxidation-reduction processes in the cells of the myometrium during normal labor, as well as the possible participation of these organelles in enhancing the biosynthesis of cellular proteins.

The increase in creatine phosphokinase activity in the uterine muscle homogenate during normal labor activity, found in our studies, indicates the presence of this enzyme in the myometrium and its role during labor activity. Increased creatine phosphokinase activity in the uterine muscle mitochondria may indicate the location of this enzyme's action in the complex system of regulation of myometrium contraction processes during labor.

We monitored the changes occurring in the contractile apparatus of the uterine muscle on glycerinated muscle models and established that bundles of glycerinated cells, under the influence of ATP, develop the highest tension.

When studying the regulation of smooth muscle myosin, it is indicated that phosphorylation of smooth muscle myosin light chains is a key reaction necessary for tension development. Native thin filaments participate in the regulation of actomyosin interaction. Myosin phosphorylation begins with an increase in intracellular Ca ²⁺ concentration, which is mediated through a system of second messengers.

To determine the structural features of the contractile apparatus of the uterine muscle in its various functional states (late pregnancy, normal labor, weak labor, labor-stimulating therapy), we used the X-ray structural analysis method, which is highly informative and allows us to judge the interatomic and intermolecular distances in the substance. Our study of X-ray patterns of bundles of glycerinated cells prepared from the uterine muscle during normal labor showed the presence of weak (due to the significantly low content of myosin in the smooth muscle), but pronounced traces of the meridional arc of the corresponding periodicity of 5.1 A and compactions or spots on the equator of the arc with a periodicity of 9.8 A, which indicates the existence of an orientation of fibrillar proteins in the contractile apparatus of the myometrium cells, which should be associated with the development of high tension by bundles of these cells under the action of ATP, and the uterus as a whole - pronounced contractile activity. At the end of full-term pregnancy, the data from bundles of glycerinated cells indicate disorientation of fibrillar proteins in the contractile apparatus of myometrial cells, which is apparently one of the factors that determines the absence of high tension developed by bundles of these cells under the influence of ATP, and pronounced contractile activity of the uterus at these stages of pregnancy.

From the standpoint of perinatal protection of the fetus in the treatment of weak labor activity, a special place belongs to the study of the structure and function of the placenta. The development of the problem of placental insufficiency deserves a separate direction.

Our electron microscopic study of the placenta during normal labor showed that its ultrastructure differs little from that at the end of a full-term pregnancy. In the homogenate and mitochondria of the placenta tissue during normal labor, compared to a full-term pregnancy, the activity of succinate dehydrogenase, cytochrome C oxidase, creatine phosphokinase and the total content of nucleic acids are increased. Consequently, the direction of the indicated changes in the placenta corresponds to those in the uterine muscle.

The increase in the total content of nucleic acids in the blood can be explained by their increased formation in the myometrium and placenta during severe labor. For the same reason, the activity of creatine phosphokinase increases, the latter is apparently also due to the increased production of this enzyme in the tissue of the myometrium and placenta and its entry into the bloodstream.

With weak labor activity, a whole series of changes occur in the fine structure of uterine myocytes and in the mitochondrial fraction of these cells, the dominant ones being signs of disorganization mainly in the myofilaments and especially in the mitochondria, against the background of which changes in the activity of the studied enzymes and the content of nucleic acids are detected.

Thus, the uterine myocytes swell and have a "light" appearance. The intercellular spaces are expanded and filled with bundles of collagen fiber fibrils and heterogeneous amorphous material of varying electron-optical density. The slightly cleared main membrane is swollen and fragmented in some areas. Edema of the sarcoplasm is expressed in the perisarcolemmal region, accompanied by the appearance of empty spaces filled with edematous fluid in the zone of myocyte contraction. In this same zone, edema, swelling and disorientation of myofilaments with a higher electron-optical density are most pronounced.

In most myocytes, the number of organelles is reduced, and in those that remain, disorganization phenomena predominate. The membrane of the sarcoplasmic reticulum is sharply fragmented. Ergastoplasm is degranulated, unfixed ribosomes are rare. The Golgi complex is not detected in most cells. Most mitochondria have only remnants of cristae with blurred or granular contours.

Thus, the changes we have identified in the ultrastructure of uterine myocytes and in the mitochondrial fraction of these cells indicate the existence (with weak labor activity) of disorientation of myofilaments and disruption of the structure of mitochondria - the substrate of oxidative phosphorylation occurring in the cristae and elementary particles of these organelles.

In case of weak labor activity, bundles of glycerinated cells of the uterine muscle develop significantly less tension under the influence of ATP than during normal labor activity. Their X-ray images resemble those of uterine muscle cells at the end of a full-term pregnancy. Such a change in the X-ray diffraction pattern may indicate either a disruption in the structure of the molecules themselves or a disorder in the mutual orientation of the molecules with each other.

Consequently, disorientation of molecules or cells relative to each other can lead to a change in muscle contractility and a decrease in the tension developed by the muscle model prepared by glycerinization. We have found this at the end of full-term pregnancy and in cases of impaired uterine contractility with weak labor activity.

Electron microscopic examination of the placenta during weak labor activity reveals changes similar to those in uterine myocytes, which are as follows: flattening of the plasmodial trophoblast, basement membrane, and capillaries develops. The number of microvilli with their characteristic thickening and club-shaped expansion decreases. The number of mitochondria in the cytoplasm of the plasmodiotrophoblast decreases significantly, and the metric becomes darker. The amount of amorphous substance increases in the syncytiotrophoblast. Langerhans cells increase in size, but the number of mitochondria decreases in them, and the metric becomes clearer. The basement membrane thickens significantly. In all cellular elements, the endoplasmic reticulum is presented in the form of small vesicles covered with granules, RPN. The detection of young villi and capillary hyperplasia can be considered a characteristic sign of the development of compensatory reactions.

In the mitochondrial fraction of the placenta, organelles have different sizes, from small to large. As in the mitochondria of myocytes, only in some of them are the remains of cristae preserved and homogeneous inclusions are rarely found.

Consequently, a whole series of stereotypical changes occur in the placenta, among which, characteristic, along with disorganizational-functional shifts, is the presence of compensatory-adaptive reactions to a greater or lesser degree.

Determination of enzyme activity and nucleic acid content showed that in the homogenate and in the mitochondrial fraction of the uterine muscle and placental tissue with weak labor activity, compared to the norm, there is a decrease in the activity of creatine phosphokinase, succinate dehydrogenase, cytochrome C oxidase and the total content of nucleic acids, indicating a decrease in the level of oxidative processes, inhibition of tissue respiration and protein biosynthesis in the myometrium and placenta.

In the blood of women in labor, pronounced metabolic acidosis is determined, some decrease in the content of calcium and sodium in the blood plasma is noted, as well as a reliable increase in the activity of oxytocinase, a decrease in the activity of creatine phosphokinase and a decrease in the total content of nucleic acids.

When carrying out treatment (use of uterotropic agents for weak labor activity) according to accepted methods and intravenous administration of oxytocin, the ultrastructure of uterine myocytes does not undergo significant changes.

Most of the myocytes have a "light" appearance and are of unevenly large sizes. The intercellular spaces remain expanded due to the proliferation of fibrils of collaten fibers and amorphous matter. In most of the cells, unclear contours of the sarcolemma are preserved in the edematous, loosened, adjacent basal membrane. The randomly located myofilaments are narrowed or edematous. Ergastoplasm is degranulated over a significant area. The Golgi complex is absent in most of the myocytes. Unlike the uterine myocytes of women with untreated weakness of labor, the mitochondria in rare cases show a preserved composition of somewhat hypertrophied cristae and isolated osmiophilic inclusions. The size of the myocyte nucleus is somewhat increased, and their scalloped membranes have fairly clear contours.

The mitochondrial fraction is dominated by organelles with a clear structure, unclear, granularly degenerated contours of the cristae. Vacuole-shaped mitochondria with no internal structure are encountered somewhat less frequently.

Consequently, when using uterotropic agents to stimulate labor activity, the picture characteristic of untreated weakness of labor activity is generally preserved in the ultrastructure of uterine myocytes and their mitochondrial fraction. However, against the background of disorganization, after the noted treatment, compartments with sufficient preservation of myofilaments, membranes of the sarcoplasmic reticulum and mitochondria are somewhat more often detected, which is apparently associated with the action of estrogens, and may indicate some improvement in the course of oxidative processes in them.

A study of the X-ray structure of bundles of glycerinated myometrial cells (contractile models) also showed some improvement in the degree of orientation of fibrillar protein molecules compared to those in untreated labor weakness.

The ultrastructure of the placenta shows a flattening of the plasmodiotrophoblast with a cytoplasm filled with a large amount of structureless substance. In some of them, individual mitochondria are found with the absence of cristae and a clear matrix. The number and size of mitochondria in Langerhans cells is somewhat increased, and the amount of structureless substance in the basal membrane is reduced.

The mitochondrial fraction also contains organelles with a complete absence of cristae, and in some mitochondria the cristae are located near the inner membrane, and their matrix contains dense, osmiophilic inclusions.

Thus, during drug stimulation of labor, the disorganizational and functional changes that we found in untreated weakness of labor persist in the placenta. However, the differences that were found, although not particularly significant, may indicate some improvement in compensatory and adaptive reactions and the course of oxidative processes in the placenta, apparently also associated with the action of estrogens included in the treatment regimens for weakness of labor.

It can be assumed that the same factor (the influence of estrogenic hormones) is associated with a tendency to improve oxidation-reduction processes in the body of women in labor, which is primarily revealed by a slight increase in the content of the total amount of nucleic acids in the mitochondria of the uterine muscle and an increase in the activity of creatine phosphokinase in the mitochondria and placental homogenate, while maintaining a picture of pronounced metabolic acidosis in the blood of women in labor.

A comparative analysis of the results of electron microscopic studies of uterine myocytes and placental tissue showed that labor stimulation by intravenous drip administration of oxytocin in a buffer solution leads to the greatest increase in the number and size of myocytes, in which organelles are found, especially mitochondria and sarcoplasmic reticulum with clear membrane contours. In addition, myofilaments are located in them more parallel, and in some cases an increase in the number of unfixed ribosome grains and even a "rosette" of polyrbosomes is noted.

The mitochondrial fraction is dominated by organelles of increased size with preserved, but somewhat randomly located cristae. As for the ultrastructure of the placental tissue, flattening of the basal membrane and capillaries is not found in it. The cytoplasm contains osmiphilic granules, ribosomes, and the plasmodiotrophoblast has an anuclear and flattened-nuclear zone. Langerhans cells contain the Golgi apparatus with an increased number of mitochondria in them, etc. Ribosomes, the Golgi complex, and mitochondria appear in the cytoplasm of the capillary endothelial cells.

In the mitochondrial fraction of the placenta, organelles of particularly large sizes are less common, and in most of them the cristae structure is preserved.

In the homogenate and mitochondria of the uterine muscle and placental tissue, an increase in the activity of creatine phosphokinase, succinate dehydrogenase, cytochrome-C-oxinase and the total content of nucleic acids is detected, which in turn indicates the functional completeness of the uterine myocytes, placental cells and their mitochondria, against the background of the picture of compensated metabolic acidosis and an increase in the activity of creatine phosphokinase and the total content of nucleic acids existing at this time in the blood of women in labor.

The results of the conducted experimental studies also showed that intravenous administration of a buffer solution to animals, even with uncompensated metabolic acidosis, leads to normalization of the acid-base and electrolyte balance in the blood, the activity of mitochondrial respiratory enzymes and the total content of nucleic acids in the myometrium, and together with oxytocin significantly increases the severity of contractions of the uterine horns, whereas intramuscular administration of estrogenic hormones, as well as intravenous oxytocin in a 5% glucose solution, does not lead to the restoration of the studied parameters and a significant increase in the contractility of the myometrium. In addition, it was found that sodium succinate, which is part of the buffer solution, increases the activity of the mitochondrial enzymes succinate dehydrogenase and cytochrome C oxidase, which is apparently associated with the high efficiency and productivity of this acid in maintaining the energy potential of the cell. The inclusion of succinic acid in the metabolic cycle activates not only energy but also plastic processes, since the four-carbon skeleton of this acid is also used for the synthesis of all types of oxidative cell systems on a porphyrin basis (cytochromes, catalase, peroxidase, etc.). This mechanism also explains adaptation to hypoxia - increased regeneration of mitochondria when taking succinic acid.

A comparative analysis of the results of a study of the X-ray structure of the contractile apparatus of the uterine muscle of women showed that after intravenous administration of oxytocin in a buffer solution, the most distinct increase in the degree of ordering of fibrillar proteins, an improvement in the degree of orientation of contractile protein molecules, and an approximation of the diffraction pattern of these muscle models to those prepared from the uterine muscle of women with normal labor activity are noted.

Thus, during the labor-stimulating therapy with intravenous drip administration of oxytocin in a buffer solution, we have revealed a sharp increase in the reparative regeneration of mitochondrial membranes (cristae) and other membrane structures in uterine myocytes and placental cells, which may indicate an increase in the intensity of oxidative phosphorylation processes along with an increase in the intensity of oxidation-reduction processes. This is confirmed by our discovery of an increase in the activity of creatine phosphokinase, succinate dehydrogenase, cytochrome-C oxidase and an increase in the total content of nucleic acids in the homogenate and mitochondria of the myometrium and placenta, and in the contractile apparatus of the uterine muscle - the existence of a pronounced orientation of fibrillar protein molecules, leading to an increase in the tension developed under the action of ATP by bundles of glycerinated myometrium cells.

The new data we obtained on the structural and functional characteristics of the contractile apparatus of the uterine muscle and the subcellular formations of the myometrium and placental tissue allowed us to establish new, previously unknown aspects of the pathogenesis of weakness of labor and to substantiate a new complex method of labor-stimulating therapy with intravenous administration of oxytocin in a buffer solution, correcting the metabolic processes in the body of mothers and fetuses that are impaired in this pathology of labor.

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