Chronic endometritis. Taking API‑NORM® suppositories during clinical therapy.

Institute of Pediatrics, Obstetrics and Gynecology of the National Academy of Sciences of Ukraine

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Report on the conduct of a clinical study.

An open controlled randomized comparative study of the clinical effectiveness and safety of complex therapy in combination with the drug "Api-Norm" and complex therapy in patients undergoing treatment for chronic endometritis.

1. General information

2. The purpose and tasks of the research

The purpose of this study is a comparative assessment of the effectiveness and tolerability of the drug "Api-Norm" as part of complex therapy in comparison with standard therapy for chronic endometritis.

Objectives of the study:

to study the effectiveness of the drug "Api-Norm" in patients with chronic endometritis;

to compare the effectiveness of complex therapy in combination with the drug "Api-Norm" and standard complex therapy in the treatment of chronic endometritis;

to study the tolerability and identify possible side reactions of the drug "Api-Norm".

3. Information about the study drug

The researched drug "Api-Norm", vaginal suppositories - a balanced composition based on natural epiphyte-preparations. Providing a combined prophylactic effect (anti-inflammatory, anti-infectious, antiviral, antispasmodic), "Api-Norm" prevents the development of complications and possible relapses in diseases of the pelvic organs.

The composition of the drug includes: activated propolis extract, beeswax, St. John's wort extract, calendula extract, sea buckthorn oil

Currently, "Api-Norm" vaginal suppositories are registered in Ukraine according to the following indicators:

atrophy of the lower parts of the genitourinary tract;

phenomena of discomfort in the genital area, accompanied by itching, burning, and dryness;

cystitis, cervicitis, endometritis, fungal and bacterial etiology as part of complex therapy;

as a preventive measure, after operative surgical and diagnostic manipulations on the organs of the small pelvis.

Method of application and dosage. In case of acute inflammatory processes, suppositories are inserted deep into the vagina after hygienic procedures, one suppository once a day at night for 5-10 days. In the case of chronic diseases, the drug is used one candle per night in three courses of 5 days, with a break between them of 5 days. For preventive purposes, the drug is used for 3-5 days, depending on the state of health.

Contraindication. Children's age, the period of pregnancy, and lactation increased sensitivity to the components of the drug.

Side effects. Allergic reactions are possible: hyperemia, rash, redness, and itching of the skin.

Drug interactions: Compatible with other prophylactic and therapeutic drugs.

4. General research plan

4.1 Research design.

This clinical study was conducted as an open, randomized, controlled, parallel.

4.2 General description of the study and principles of group allocation.

The study included 90 patients who were receiving outpatient treatment in the clinic of the Department of endocrine gynecology of the State University "Institute of Pediatrics, Obstetrics and Gynecology of the National Academy of Medical Sciences of Ukraine" and who met the inclusion/exclusion criteria described in this protocol. Patients who gave written informed consent to participate in the study were included in the trial.

All patients who participated in the study were randomly divided into the main and control groups.

During the study, each patient underwent a clinical and laboratory examination according to the scheme.

Scheme of the study. The study included the following stages: screening from 0 to 15 days, treatment - 8 weeks. Periodicity of observations and registration of patient examination data was carried out according to the following scheme.

Notes:

1 - includes determination of hemoglobin, erythrocytes, leukocytes, leukocyte formula, ESR.

2 - includes determining the levels of glucose, protein, creatinine, aspartate aminotransferase (AST), alanine aminotransferase (ALT), total bilirubin (if there are deviations - bilirubin fractions).

Terms of termination of the study. The study was not interrupted.

5. Selection of patients

5.1. Criteria for including patients in the study:

age 18-45 years;

the presence of artificial abortions and reproductive losses in the anamnesis;

histological and immunohistochemical confirmation of chronic endometritis;

the patient's ability to adequately cooperate in the research process;

informed written consent of the patient to participate in the study.

5.2. Exclusion criteria:

atypical hyperplasia, endometrial cancer;

submucosal myoma of the uterus for more than 10 weeks;

II degree endometriosis;

ovarian tumors that require surgical treatment;

malignant neoplasms;

pregnancy, lactation;

any diseases and clinical conditions associated with a risk to life;

increased sensitivity to the components of the studied drugs;

impaired liver and/or kidney function;

participation in any other clinical trial;

any concomitant diseases or acute conditions, the presence of which can significantly affect the results of the study;

the need to take non-recommended medicines.

8. Conducting the test and the methods used

Before inclusion in the study and in its process, each patient underwent an examination in accordance with the scheme:

I. General clinical research methods:

objective examination (measurement of heart rate, blood pressure, palpation and percussion of the abdomen, examination of the skin and visible mucous membranes);

gynecological examination, palpation of mammary glands;

assessment of the volume of blood loss with the implementation of the RVAS technique;

general blood analysis: erythrocytes, hemoglobin, leukocytes, ESR;

general analysis of urine (specific gravity, pH, protein, glucose, leukocytes, epithelial cells, erythrocytes, cylinders);

biochemical blood analysis (protein, bilirubin, ALT, AST, creatinine, glucose).

II. Special research methods:

ultrasound of the pelvic organs;

hysteroscopy;

separate diagnostic endometrial scraping or biopsy using Pipelle de Cornie;

morphological examination of a scraping from the cervix and endometrium;

morphological study of the endometrium;

immunohistochemical study of the endometrium.

General clinical research methods

When collecting anamnesis, attention was paid to heredity, transferred infectious and somatic diseases. The menstrual cycle was evaluated in detail (age of menarche, regularity, duration of the cycle, nature of menstruation, dysmenorrhea, and presence of prolonged scanty bleeding before and after menstruation).

The PBAC (Pictorial Blood Assessment Chart) technique proposed by Higham (1990) was used to objectively assess the amount of menstrual blood loss. During menstruation, each patient used the table (Fig. 1), where he indicated the number of tampons or pads and the degree of their blood-wetting. A tampon or pad that is slightly stained is rated at 1 point, if it is half wet, it is rated at 5 points, if it is completely soaked, it is rated at 20 points, and the presence of clots after removing the tampon or pad is rated at an additional 5 points.

The obtained number of points was multiplied by the amount of sanitary material that the patient changed per day and summed over the entire period of menstrual bleeding. An indicator of more than 100 points during the period of menstruation was considered excessive uterine bleeding.

Pic. 1

The table for estimating the amount of blood loss according to the RBAS method

The number, outcome, and complications of previous pregnancies were also evaluated; nature of sexual life, number of sexual partners, and use of contraceptive methods. Particular attention was paid to the presence of the anamnesis of surgical interventions, including on the organs of the small pelvis, inflammatory diseases of the genital organs, and infections transmitted mainly sexually.

During a special gynecological examination, the degree of development and features of the external genitalia, the nature of the pubic hair, the condition of the vagina, the cervix, the properties of the cervical mucus, the size and shape of the uterus, its mobility, tenderness, displacement, the condition of the appendages, the enlargement of the ovaries, the presence of tuboovarian formations, adhesions were evaluated, the state of the sacro-uterine ligaments, the nature and amount of discharge from the vagina.

Special research methods

Special research methods were carried out before the start of treatment and 2 months after its end.

Ultrasound examination of the pelvic organs.

Pelvic ultrasound was performed using a series of longitudinal and transverse sections with "Nemio XG" devices from Toshiba (Japan) using a multi-frequency transvaginal sensor with a frequency of 4.0 - 7.5 MHz. The study was conducted on days 7-9 and 17-21 of the menstrual cycle before treatment and on the same days of the menstrual cycle 2 months after the end of treatment. The condition and dimensions of the uterus (length, anterior-posterior size, width) were determined, the structure of the myometrium, the presence and nature of myomatous nodes were evaluated. The M-echo structure was specifically studied: endometrial thickness, echomorphology, and echogenicity. When examining the ovaries, their size was measured, the diameter and number of follicles were estimated.

Endoscopic research methods.

Diagnostic hysteroscopy was performed on the 7-10th day of the menstrual cycle using the equipment of the company "K.Storz" (Germany) according to the generally accepted method. The sterile Turusol irrigation solution was used as the optical medium. During hysteroscopy, the size and shape of the uterine cavity, the presence of deformation, and endometrioid processes were assessed. The color, uniformity of color, folding, and non-uniformity of the thickness of the endometrium, the presence of hemorrhages, polypoid formations, intrauterine synechiae, and foreign bodies in the uterine cavity were evaluated. Under hysteroscopic control, separate diagnostic scraping of the endocervix and endometrium was performed, as well as removal of various pathological formations or endometrial biopsy. A control biopsy of the endometrium was performed in all patients using a Pipelle de Cornie on day 7-10 of the menstrual cycle, 2 months after the end of treatment.

Morphological study.

In order to verify the diagnosis of chronic endometritis, all patients underwent an endometrial biopsy:<

46 (51%) patients underwent a separate diagnostic curettage of the uterus after hysteroscopy in hospital conditions;

44 (49%) patients underwent endometrial biopsy during hysteroscopy using biopsy forceps.

In view of the possible influence of the type of surgical intervention on the expression of the Ki67 marker and, as a result, the result of treatment, during the randomization of patients, the groups were evenly distributed according to this feature.

An endometrial biopsy was performed on the 7th to 10th day of the menstrual cycle, that is, in the middle phase of proliferation according to the classification of R.W. Noyes (1950). Endometrial biopsy in patients of the control group was performed on day 7-10 of the menstrual cycle using a Pipelle de Cornie aspiration curette.

In the main groups, 2 months after the end of the treatment course, on the 7-10th day of the menstrual cycle, the patients underwent a control biopsy of the endometrium using a Pipelle de Cornier aspiration curette. Serial paraffin sections stained with hematoxylin and eosin were used for morphological studies. The obtained endometrial tissue samples were fixed with a 10% formalin solution. After appropriate histological processing, endometrial samples were embedded in paraffin, paraffin sections were prepared with a thickness of 6 microns, and stained with hematoxylin and eosin.

Morphological signs of chronic endometritis were considered to be:

the presence of plasma cells;

the presence of inflammatory infiltrates in the endometrium, which consists mainly of lymphoid elements with an admixture of macrophages and is located more often around glands and blood vessels, less often diffusely;

localized stroma fibrosis;

sclerotic changes in the walls of the spiral arteries of the endometrium.

Immunohistochemical study of the endometrium.

Immunohistochemical examination of the endometrium was performed in dynamics in women with CE without combined gynecological pathology before treatment and 2 months after the end of treatment.

Serial paraffin sections were used for immunohistochemical examination. The research was carried out on the unmasking of antigens in a microwave oven at 600 W in a citrate buffer at pH = 6.0.

The study was conducted by the method of double antibodies using the Ultra Vision Quant detection system, peroxidase polymer, and DAB plus chromogen. The following monoclonal antibodies produced by Thermo Fisher Scientific Anatomical Pathology (UK) were used as primary antibodies:

CD138 Ab-2 (Clone Mi 15) is a plasma cell marker;

CD31/PECAM-1 (Endothelial Cell Marker) Ab-1 (Clone JC/70A) is a platelet-endothelial cell adhesion molecule;

Ig A (α-Heavy Chain) Ab-2;

Ig G (γ-Heavy Chain) Ab-1;

Ig M (μ-Heavy Chain) Ab-2;

MMP-1 (Collagenase-1) Ab-6;

MMP-2 (Collagenase IV) Ab-4 (Clone A-Gel VC2);

Epidermal Growth Factor Receptor (EGFR, Clone EP 384);

Ki-67 (Clone SP6) is a marker of cell proliferation;

Estrogen Receptor (Clone SP1) — estrogen receptors;

Progesterone Receptor (Clone YR85) — progesterone receptors.

Universal secondary antibodies conjugated with a polymer, labeled with an enzyme that detects mouse and rabbit immunoglobulin were used as secondary antibodies. The polymer complex was then visualized using the appropriate system, the substrate-chromogen.

Control tests for immunohistochemical reactions were performed:

as a negative test, reactions were performed according to the protocol without the use of primary antibodies;

as positive tests, histological sections of tonsils, sections of a lymph node, and placenta with known expression of the studied markers were used.

The results of the reactions (CD31, EGFR, MMP-1, MMP-2) were evaluated semiquantitatively in points according to the generally accepted method:

0 - no staining;

1 - less than 10% of positively stained cells;

2 - more than 10% and less than 50% of positively stained cells;

3 - homogeneous staining of more than 50% of cells.

The results of the Ki-67, CD138, ER, and PR reactions were evaluated by the percentage of stained cells (nuclei) of a certain type (epithelial, plasma cells, vascular endothelium) when counting 100 cells of this type.

9. Evaluation of efficiency

9.1 Main performance criteria

The effectiveness was evaluated based on the results of the morphological examination of the endometrium:

9.2. Secondary performance criteria

• relief of clinical symptoms of the disease;

• restoration of the echographic picture of the endometrium;

• normalization of cell proliferation and apoptosis processes in the endometrium (Ki67, EGFR), stabilization of the extracellular matrix (MMP);

• restoration of reproductive function.
  

10. Tolerability assessment

The tolerability of the drug was assessed on the basis of objective data obtained during the study, subjective sensations reported by the patient, and laboratory examination data performed before and after the course of treatment with the studied drug.

The tolerability of the drug was assessed by the researcher and the patient according to the following scale:

11. Registration of side effects/reactions

11.1 Terms

Adverse reaction.

As part of a pre-registration clinical trial of a new drug or when studying it for new indications, especially if the therapeutic doses of the drug have not been precisely established, adverse reactions should include all negative or unexpected reactions associated with the administration of any dose of the drug. The term "related to the administration of the drug" means that there is at least a minimal probability of a cause-and-effect relationship between the drug and the adverse reaction, that is, a relationship cannot be excluded. In relation to registered medicinal products, this term means all negative or unpredictable reactions associated with the use of a medicinal product in normal doses for the purpose of prevention, diagnosis, or treatment of diseases, restoration, correction, or influence on physiological functions.

An unexpected adverse reaction is an adverse reaction whose nature or severity is not consistent with the available information about the drug (eg, the investigator's brochure for an unregistered drug or the package insert and summary for a registered drug).

Adverse event (AE) - any adverse medical manifestation in a test subject that does not necessarily have a causal relationship with the use of the study drug (changes in laboratory data, symptoms or diseases that coincide in time with the use of the study drug, etc.).

Serious adverse reaction or serious adverse event - any adverse medical manifestation when using a medicinal product (regardless of dosage), that leads to death, poses a threat to life, requires hospitalization or an increase in the duration of hospitalization; leads to long-term or significant loss of working capacity or disability; or is manifested by congenital anomalies and malformations.

11.2 Adverse Event Reporting

No side effects were detected during the study.

12. Static processing of received results

The obtained indicators of the examined patients were processed by the method of variational mathematical statistics. For the creation of a database and mathematical processing of statistical material, the package of modules for statistical data processing STATISTICA ® for Windows, Release 7.0 of the company Statsoft®Inc, USA, was chosen as the main software.

Quantitative signs were entered into the database without changes. The difference in means was taken as reliable at p < 0.05 (T > 2). To assess the reliability of differences in the distribution of quality indicators, after creating conjugation tables, the χ2 agreement criterion (chi-square) and confidence interval values (p) were calculated. Arithmetic mean (M), arithmetic mean error (m), and root mean square deviation were calculated for quantitative traits.

To compare the parametric data (after checking the quantitative data for normal distribution), we used the Student's t-test for 2 independent samples. For non-parametric data, the U-Mann-Whitney method (for 2 groups) was used for unrelated populations. For dynamic data analysis, the paired Student's t-test was used for dependent samples and the Wilcoxon test for indicators that do not conform to the law of normal distribution.

13. Analysis and presentation of data

13.1 Clinical characteristics of women included in the study

The study included 90 women who applied to the Center with various disorders of reproductive function: infertility and miscarriage, in whom chronic endometritis was diagnosed during the examination. All patients gave written consent to participate in the study.

To evaluate the effectiveness of complex therapy, all women were randomly divided into 2 groups.

In the 1st group (n = 45) complex treatment of XE was carried out. At the first stage of the basic treatment, taking into account the results of the microbiological examination, etiotropic drugs were used: ornidazole 500 mg 2 times a day for 10 days in combination with levofloxacin 500 mg 1 time a day for 14 days. At the same time, anti-inflammatory drugs were prescribed in the form of rectal suppositories diclofenac sodium 100 mg once a day for 10 days. In order to restore and prevent vaginal microbiocenosis disorders, all women received a multi-strain probiotic containing 2.0 x 109 CFU of Lactobacillus acidophilus, Lactobacillus rhamnosus, Lactobacillus reuteri, and 1.0 x 109 CFU of Lactobacillus casei, Lactobacillus plantarum, Lactobacillus fermentum , Bifidobacterium bifidum.

At the second (restorative) stage, as an additional treatment, the drug "Api-Norm" was prescribed 1 suppository vaginally at night once a day for 10 days, and then 2 times a week for a total course of 15 suppositories.

In the II group (n = 45), a similar full course of basic treatment was carried out, consisting of etiotropic, anti-inflammatory, and antibacterial drugs. There is no additional restorative treatment, the women of this group did not use the drug "Api-Norm".

In the main groups of patients, 2 months after the end of the entire course of treatment, on the 7-10th day of the menstrual cycle, a complete examination and a control biopsy of the endometrium were performed using a Pipelle de Cornier aspiration curette.

The control group for the immunohistochemical and morphological study of the endometrium consisted of 15 healthy fertile women of reproductive age who did not have gynecological diseases and did not use intrauterine or hormonal methods of contraception. Endometrial biopsy in patients of the control group was performed on days 7-10 of the menstrual cycle using a Pipelle de Cornier aspiration curette, after obtaining appropriate informed consent.

The groups were comparable in terms of age, somatic and gynecological anamnesis

Tab 13.1.1

In the course of the study, a thorough analysis of the complaints of patients with HE was carried out (Table 13.1.2).

In the structure of complaints, the clear dominance of various disorders of reproductive function (infertility and miscarriage), which made up a total of 89%, attracts attention. At the same time, complaints about other clinical manifestations of the disease (pain syndrome, acyclic bleeding from the genital tract, etc.) were presented by no more than 1/2 of women with CE. 3 (6.7%) women of the I group and 1 (2.2%) of the II group had no complaints. There was no statistically significant difference in the complaints of patients of the examined groups.

Tab 13.1.2

13.2 Morphological and immunohistochemical characteristics of the endometrium in women with chronic endometritis.

In order to verify the diagnosis of XE, all patients underwent endometrial biopsy. In all cases, the endometrial biopsy was performed on the 7-10th day of the menstrual cycle, that is, in the middle phase of proliferation according to the Noyes classification.

Light microscopy of sections stained with hematoxylin and eosin revealed lymphohistiocytic inflammatory infiltrates of various degrees in the endometrium in all cases. Infiltrates were located in the form of foci more often periglandular and perivascular or diffusely in the basal and functional layers of the endometrium. Inflammatory infiltrates were represented by mononuclear cells and consisted of lymphocytes, plasma cells, and macrophages surrounded by fibroblasts.

13.2.1 Immunohistochemical features of the endometrium in women with chronic endometritis.

For the purpose of a detailed study of the endometrium in all women with CE and without accompanying gynecological pathology, an immunohistochemical study was performed using markers of inflammation, cell proliferation, apoptosis, angiogenesis, as well as the extracellular matrix and steroid receptors.

Expression of inflammatory markers in the endometrium.

During light microscopy of sections stained with hematoxylin and eosin, plasma cells were not detected in all focal lymphohistiocytic infiltrates, which did not allow to immediately clearly verify the diagnosis of chronic endometritis.

In order to clarify the diagnosis, immunohistochemical reactions were carried out with antibodies to the plasma cell marker, surface glycoprotein - CD138 (syndecan-1).

In the case of an immunohistochemical reaction, CD138 was detected in the form of stained membranes or cytoplasm of cells in brown color. This marker was detected both when staining the plasma cells of the infiltrate, and in the form of individual cells with scattered lymphohistiocytic infiltration. In all cases, the reaction was positive, which made it possible to confirm the presence of classic chronic endometritis in all patients with a previous diagnosis of HE (table 13.2.1, photo 1). In the control group, the CD138 marker was not detected in any case.

Tab 13.2.1

Processes of proliferation, cell apoptosis, and angiogenesis in the endometrium.

The level of cell proliferation in the endometrium was assessed by immunohistochemical reactions based on the expression of the Ki-67 marker.

Ki-67-positive cells were located mainly in the basal layer of the endometrium. The reaction product was localized in the cell nuclei and had a brown color. With CHE, the expression of Ki-67 in the cells of the glandular and covering epithelium was significantly higher than the control values in both groups (p <0.05), and there was also a tendency to increase the expression of Ki-67 in the cells of the endometrial stroma (table 13.2.2; photo 8 , 9.).

The study assessed the expression of the epidermal growth factor receptor EGFR. EGF itself induces the proliferation of epithelium and stromal cells, as well as vascular endothelium. The brown reaction product was localized in the cytoplasm of mainly epithelial cells of the glands throughout the thickness of the endometrium. In representatives of groups I and II, the expression level of the EGF growth factor receptor was 1.5 times higher than the similar indicator in the control group (p <0.05) (Table 13.2.2).

Against the background of chronic inflammation, endometrial angiogenesis also acquires pathological features. In this regard, in our research, we also provided for the study of its parameters.

CD31 (PECAM-1) - platelet endothelial cell adhesion molecule - an adhesion molecule belonging to the family of immunoglobulins. It is believed that under physiological conditions, the endothelial cell does not express adhesion molecules. An increase in their concentration on the cell surface is the result of the influence of various factors. However, in normal endometrium, CD31 is expressed in the endothelium of stromal vessels and plays the role of a trigger in intravascular cytotrophoblastic invasion (G. Yurdakan et al., 2008; Smirnova T.L., 2009). Decreased CD31 in the endometrium reflects incomplete decidualization of the endometrial stroma, failure of the first wave of cytotrophoblast invasion, and incomplete transformation of endometrial segments of spiral arteries. In the presented study, the expression of the cytokine CD31 in the endometrium of women with XE, both in the I and II groups, was significantly reduced in comparison with the endometrium of healthy women (p <0.05) (Table 13.2.2).

Tab 13.2.2

Estrogen and progesterone receptors

To assess the functional potential of the endometrium, immunohistochemical reactions were performed with antibodies to steroid receptors: estrogen receptors (type a) and progesterone receptors. The brown reaction product was localized in the nuclei of epithelial cells in large quantities in the functional layer. At the same time, a significant spread in the expression of steroid receptors in different patients drew attention.

Despite the numerous scientific data available in the literature about dysfunctional disturbances of tissue reception in chronic endometritis, which are manifested in the increased expression of steroid receptors in the glands and stroma, no such pattern was found in our study. On the contrary, the study of the spectrum of the distribution of receptors showed that the expression of steroid receptors to estrogens and progesterone in the nuclei of cells of the glandular epithelium in XE was not significantly different from the control group (p ~ 0.05) (Table 13.2.3).

Tab 13.2.3

13.3 Dynamics of clinical and laboratory indicators after treatment of chronic endometritis.

All women with XE initially represented a clinically complex contingent of patients with gynecologically complicated and long-term unsuccessful treatment of reproductive function disorders. At the same time, the majority of women had not previously been examined hysteroscopically and immuno-morphologically, which made it difficult to verify the diagnosis and was an important factor in failures in the implementation of reproductive function. In this regard, assessment of the adequacy of therapeutic measures in this group of patients previously represented certain difficulties.

In general, the assessment of the effectiveness of the therapy included the analysis of several factors: the dynamics of clinical symptoms, and changes in the morphofunctional and immunological characteristics of the endometrium.

13.3.1 Clinical indicators after XE treatment.

When analyzing the dynamics of clinical manifestations of the disease against the background of therapy, it was shown that a full course of XE therapy leads to significantly more pronounced changes than an incomplete course of therapy without a recovery phase. Data on the dynamics of clinical symptoms of XE and the nature of menstrual function in different groups after treatment are presented in tables 13.3.1.1 and 13.3.1.2.

Tab 13.3.1.1

Table 13.3.1.2

The average duration of menstruation after treatment was 5.1 ± 0.4 days in patients of group I, varying from 2 to 7 days, in patients of group II 4.8 ± 0.3 days, varying from 3 to 8 days. There were no significant differences in this indicator between the groups and in comparison with the initial data.

When analyzing the intensity of bleeding during menstruation, it was noted that in patients of group I, blood loss during menstruation significantly decreased, while heavy menstruation remained in 8.8% of women and in all cases was associated with the presence of combined pathology (myomas of the uterine body and adenomyosis).

In the II group, this indicator also decreased from 20.8% to 4.4%, but this difference was statistically insignificant (p ~ 0.05).

Premenstrual bleeding after treatment was detected in 4.4% of women in group I and in 6.6% of women in group II (p <0.05 compared to baseline data). Complaints about intermenstrual bleeding for several days persisted only in 2.2% of women of both groups (p <0.05).

All women who previously complained of pain noted partial or complete relief of the pain syndrome. Constant pains in the lower part of the abdomen, not related to the cycle, after treatment were noted by 4.4% of women in group I and 6.7% in group II (p <0.05 compared to data before treatment). At the same time, the pains, as a rule, were periodic, were of moderate intensity, without irradiation, and did not require the use of analgesic drugs.

8.9% of women in group I and 13.3% of group II complained of dysmenorrhea after treatment (p <0.05 compared to baseline data), while patients noted pain of moderate or medium intensity. After treatment, 2.2% of II group patients complained of dyspareunia, but the data were considered statistically unreliable (p> 0.05).

During a bimanual gynecological examination, it was established that there was no tenderness during cervical traction and during palpation of the uterine body in any woman in all groups, tenderness during palpation of the uterine appendages remained in 5.7% of women in the II group (p <0.05 in compared with the original data). Pain during palpation of the walls of the small pelvis was not noted in any case.

13.3.2 Dynamics of morphological and immunohistochemical characteristics of the endometrium after treatment of chronic endometritis.

In the course of the work, the morphological and immunohistochemical features of the endometrium after treatment were analyzed in comparison with the endometrium before treatment and normal endometrium in the middle phase of proliferation.

With light microscopy of sections stained with hematoxylin and eosin, it was noted that after treatment 86% of patients in group I and 77% in group II showed complete restoration of the normal structure of the endometrium. In most cases, focal periglandular lymphohistiocytic inflammatory infiltrates disappeared in the functional layer of the endometrium, but in the stroma of the endometrium, weak scattered infiltration by lymphocytes and single macrophages remained. Plasma cells under light microscopy and with the help of immunohistochemical reactions (marker CD138) after treatment in the I group were not detected in any case. In the II group, the number of plasma cells decreased by 2.7 times (from 12.4 ± 2.13 to 4.6 ± 0.72; p ~ 0.05), CD138 was also detected in the membranes in a significantly lower amount than before treatment (table 13.3.2.1, graphs 13.1 and 13.2, diagrams 1-4).

Graph. 13.1

Dynamics of CD138 (plasma cells) against the background of XE treatment, M ± m

Graph. 13.2

Dynamics of CD138 (membranes) against the background of XE treatment, M ± m

Evaluation of the dynamics of indicators of humoral protective factors in the endometrium after treatment did not reveal any statistically significant improvement. However, the tendency to decrease the levels of IgA, IgG, IgM in both I and II groups is clearly visible (tab. 13.3.2.1, diagram igA, igG, igM)

Diagram igA

Dynamics of IgA after completed courses of XE treatment, M ± m

Diagram igG

Dynamics of IgA after completed courses of XE treatment, M ± m

Diagram igG

Dynamics of IgM after completed courses of XE treatment, M ± m

Tab 13.3.2.1

After treatment, normalization of cell proliferation processes was noted in the endometrium, which was confirmed by an immunohistochemical study based on the level of expression of Ki-67 and EGF markers. In group I, the expression of Ki-67 decreased (p <0.05), but it approached the control values as much as possible only in the stroma. Ki-67 also decreased in the endometrial epithelium of women of the I group, but it was higher than in the I group (p <0.05). In the stroma, the dynamics of this marker were similar to those in group I (tab. 13.3.2.3, diagrams ki-67-gland and ki-67-stroma, photos 5-8).

When evaluating the expression level of epidermal growth factor receptor (EGFR), which induces the proliferation of epithelium, stromal cells, and vascular endothelium, no significant changes were detected. A slight tendency towards its decrease was observed in women of group I, in group II the expression of EGFR in the glands remained at a high level and was significantly higher than in the control group (p <0.05).

Tab 13.3.2.3

Diagram ki-67-gland

Dynamics of Ki-67 in glands after XE treatment courses, M ± m

Diagram ki-67-stroma

Dynamics of Ki-67 in the stroma after XE treatment courses, M ± m

Monitoring the content of CD31 in the endometrium of women who received treatment did not reveal statistically significant changes in the indicator in both groups. The increase of this marker in the examined women turned out to be insignificant. Perhaps a longer period is required to reach physiological levels of CD31.

After the course of treatment in both studied groups, there were no significant differences in the expression of estrogen and progesterone receptors both in the epithelium and in the stroma (tab. 13.3.2.5).

Tab13.3.2.5

The changes that occurred during treatment with MMP1 and MMP2 are presented below (graph 13.3, table 13.3.2.6). In both groups, the level of MMP1 decreased statistically significantly and reached the physiological level (p <0.05). Of particular interest are the data obtained on MMP2. In a number of scientific studies of the last decade, it was proved that chronic endometritis, in combination with infertility and miscarriage, is associated with a decrease in the content of MMP2 in the endometrium. A similar correlation was found by us. Against the background of treatment in group II, the level of MMP2 did not change, and in the endometrium of women of group I, a slight increase in MMP2 activity was registered, but the indicators were considered statistically insignificant. Despite this, the emerging trend towards an increase in MMP2 in the group with the study drug is, in our opinion, of scientific interest and requires further study.

Graph. 13.3

Dynamics of MMP1 in epithelium after XE treatment courses

Tab 13.3.2.6

13.3.3 Analysis of information obtained during the research

Analyzing the mechanism of action of the drug "Api-Norm" on the endometrium, we can conclude that it potentiates the anti-inflammatory effect of basic therapy. Complex treatment, including a restorative stage (in our study in the form of the drug "Api-Norm"), blocks the formation of inflammatory infiltrates by reducing the levels of immunocompetent cells (CD138). At the same time, influencing more gently, bringing them as close as possible to the values in a healthy endometrium.

As a result of the research, normalization of tissue regeneration processes, and reduction of cell apoptosis (Ki-67, EGFR) was noted. It is interesting that the potential for pathological tissue regeneration, which is hidden in chronic inflammation, is not completely eliminated by the appointment of standard antibiotic therapy (II group in the study). It was much more effective to reduce active proliferation in the endometrium by prescribing complex therapy, which included the drug under study.

In general, against the background of complex therapy with the inclusion of the drug "Api-Norm", the disturbed tissue homeostasis of the endometrium was restored, which led to the removal of clinical symptoms and laid the foundation for the successful implementation of the reproductive function.

According to the results of the positive dynamics of the clinical symptoms of the disease, the restoration of the morphological structure of the endometrium, the normalization of the processes of proliferation, apoptosis, and immune balance against the background of treatment with the drug "Api-Norm", the clinical effectiveness of its use has been proven.

The tolerability of the drug was evaluated according to the proposed scoring system, which is shown in Table 13.3.3.1. As can be seen from the table, in 6 patients of the II group, the researcher assessed the tolerability of the drug as satisfactory, since the patients noted a feeling of irritation due to an increase in the number of secretions, which was interpreted as a minor side reaction that did not cause serious problems for the patient and did not require the withdrawal of the drug.

Tab 13.3.3.1

Based on the data in the table, the majority of the examinees rated the tolerability of the drug as good, a satisfactory rating in 13.3% of patients, possibly due to a greater concentration of attention on the drug.

Conclusion. The conducted clinical trial of the drug "Api-Norm" testifies to its effectiveness as part of the complex therapy of chronic endometritis and the good tolerability of this drug, which gives reason to recommend allowing it for clinical use to treat this pathology.

The responsible executor of the study is Doctor of Medical Sciences, Professor T. F. Tatarchuk.