RU2540499C1 - Method for prediction of anembryonic pregnancy - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: method involves dropping a supernatant of uterine fluid aspirate or menstrual fluid in an amount of 0.2 ml on a slide surface, drying at room temperature for one day, and microscoping the prepared samples (phases). The presence of a complex of morphological elements, including toxic plaques, wedge and disk-shaped structures enables predicting a high risk of anembryonic pregnancy.

EFFECT: simplifying and providing higher sensitivity of the method for the prediction of anembryonic pregnancy and enables predicting the clinical course and outcome of a pregnancy within a short time and with a small amount of the biological material available.

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Description

The invention relates to medicine, namely to laboratory methods of research, and can be used to predict non-developing pregnancy.

Over the past decade, among the most important problems of practical obstetrics and gynecology, one of the first places has been the problem of miscarriage (V.M. Sidelnikova, G.T. Sukhikh. Miscarriage: A Guide for Practitioners. - M.: Medical Information Agency LLC , 2010 - 536 p.).

According to the classification of T.Yu. Pestrikova (1994), non-developing pregnancy (NL) is considered one of the pathogenetic options for miscarriage (T.Yu. Pestrikova. Miscarriage: spontaneous abortion, premature birth, non-developing pregnancy. Management of pregnancy and childbirth at high risk. M: Relax, 1994). In recent years, among cases of spontaneous miscarriages in the early stages, the frequency of this pathology has increased from 10-20% to 45-88.6% (V.E. Radzinsky, V.I. Dimitrova, I.Yu. Mayskova. Non-developing pregnancy - M. : GEOTAR - Media, 2009. - 200 p.), Which makes us consider the NB as an urgent medical and socially significant problem.

One approach to solving this problem when planning pregnancy is to assess the risk of developing NI already at the stage of pregravid preparation (V.E. Radzinsky, V.I. Dimitrova, I.Yu. Mayskova. Non-developing pregnancy - M .: GEOTAR - Media, 2009 g. - 200 p.).

Thus, modern medicine needs new methods for predicting NB in order to identify risk groups for its development at the pregravid preparation stage.

Sonohisterography and transvaginal sonography (BS Demidov. Clinical significance of dopplerometry in the diagnosis and prediction of placental insufficiency in the second and third trimesters of pregnancy are used for screening examination of patients with NB at the stage of pregravid preparation: Abstract, Ph.D. in medical sciences. M ., 2000). Using these methods, it is possible to predict NB. But these methods have significant disadvantages:

- require the availability of expensive equipment;

- statement of the presence of NB;

- a high frequency of false-positive results in the early stages;

- impossible early preclinical prediction of non-developing pregnancy.

The most common is a method for predicting NB, based on a histological examination of removed tissues (M. Burston. Histochemistry of enzymes, transl. From English. M., 1965; R Lilly. Pathological technique and practical histochemistry, transl. From English., M., 1969; E. Pearce. Histochemistry, trans. From English, M., 1962; Principles and methods of histocytochemical analysis in pathology, under the editorship of A.P. Avtsyn et al., L., 1971; O. I. Topchieva, V. .A. Pryanishnikova, ZP Zhemkova. Endometrial biopsy - M .: Medicine. - 1978).

The disadvantages of the histological method are:

- establishment of possible causes of NB only after diagnostic manipulation;

- the complexity of technical performance;

- the need for a specially equipped room;

- the need to use expensive equipment;

- the duration of the study until the issuance of the final conclusion (10-14 days).

A special place in the prognosis of NB is occupied by the method for determining the level of β - chorionic gonadotropin (CG) (instructions for using the enzyme-linked immunosorbent assay for the quantitative determination of human chorionic gonadotropin "HCG-ECO-test", manufactured by LLC "Diateh-EM").

The disadvantages of the method are:

- use of special test systems;

- a high frequency of false-positive results (an increase in the level of chronic hepatitis can be observed with a number of gynecological diseases - trophoblastic tumors, dysgerminoma);

- impossible early preclinical prediction of non-developing pregnancy.

A method for predicting NB by the level of placental growth factor in blood serum from 5 weeks of pregnancy (A.Yu. Ponamoreva, V.V., Avrutskaya, II Krukier and others. - “Method for predicting an undeveloped pregnancy” - Patent for the invention No. 2313793. - FGU Rostov Research Institute of Obstetrics and Pediatrics of the Federal Agency for Health and Social Development, 2006) has a number of disadvantages, namely:

- forecasting of NB in the period of 5 weeks or more;

- the need for expensive test systems;

- inefficiency of use at the pregravid stage of preparation.

After the implantation process, peripheral trophoblast cells penetrate the mother’s endometrium and provoke an immune response mechanism, as a result of which embryotropic autoantibodies arrive (N.T. Hakhva, 2003; S.V. Apresyan, 2003; V.V. Misnik, 2005). To assess their influence on the formation of chorion and prognosis of the development of NB, an ELI-P test (ELISA-detected Probability of pathology in Pregnancy, 1998 (A.B. Poletaev, N.K. Vabishchevich, S.G. Morozov) was proposed. examination of women of childbearing age using the ELI-P test system to predict the development of the embryo and fetus and the birth of a healthy or abnormal child. RF patent for the invention No. 2107913 dated March 27, 1998).

The disadvantages of this method are:

- the need for expensive test systems, a cytogenetic laboratory;

- with NB, all three types of immunoreactivity can occur, the method is not specific;

- the influence of somatic pathology on the test results - shifts in immunoreactivity predicted in extragenital diseases (N.T. Hakhva, 2003).

There is a method of assessing the level of glycodeline in menstrual blood for predicting NB in women with a history of hormonal imbalance (Y. S. Balkhanov. "Possibilities of restoring the morphofunctional ability of the endometrium in women with miscarriage." - Abstract of thesis ... Candidate of Medical Science. - Perm, 2009).

The disadvantages of this method are:

- the need for expensive reagents and the technical complexity of the method;

- strict specificity of the method (informative only for the prognosis of NB hormonal origin);

- a change in glycodelin level can be observed with various pathologies of the endometrium that are not associated with pregnancy;

Our invention is aimed at simplifying and increasing the sensitivity of the method for predicting NB.

The technical result of the invention is achieved by the fact that the supernatant of aspiration fluid from the uterine cavity or menstrual flow in the amount of 0.2 ml is applied to the surface of a glass slide in the form of a drop, dried at room temperature for 24 hours, after which the obtained samples (facies) are microscopic and, if any a complex of morphological elements, including: toxic plaques, linguistic and disc-shaped structures, predict a high risk of developing an undeveloped pregnancy.

The essence of the invention is illustrated by drawings, where:

figure 1 - a fragment of the facies of the supernatant of aspiration fluid from the uterine cavity in a woman with a progressing pregnancy;

figure 2 - a fragment of the facies supernatant of the menstrual flow of a healthy woman;

figure 3 - a fragment of the facies supernatant of aspiration fluid from the uterine cavity in a woman with a non-developing pregnancy, where:

1. lingual structures

2. disk-shaped structures

3. toxic plaques.

The proposed method was successfully tested in gynecological departments of OA GBUZ "Clinical Maternity Hospital", Astrakhan, JSC GBUZ AIOCB Astrakhan, women's consultation "Clinical Maternity Hospital", Astrakhan in 2012-2013.

We examined 80 women aged 16 to 43 years, 30 (40%) women with a diagnosis of NB, the control group consisted of 50 (60%) women with a normal pregnancy in the period from 5 to 12 weeks with no signs of threat that turned to terminate pregnancy wish to gynecological hospitals in Astrakhan. Along with the traditional clinical and laboratory examination, all of the subjects underwent pelvic ultrasound, transvaginal access, mammary glands, cytological examination of endo-exocervix and morphological analysis of facies of the studied biological fluids. The results were compared with clinical data, histological findings of pregnancy and the morphological picture of the obtained facies.

Testing results are given below.

Example No. 1

Pregnant M., born in 1989, history No. 3495, was admitted to the gynecological department on 23/12/12 with a diagnosis of pregnancy within 5 weeks of gestation (for an artifact abortion).

No complaints at admission.

Menstrual and reproductive functions: menstruation from 14 years to 4 days, regular, moderate, 30 days cycle. Last menstruation 2/11/12 g.

In the anamnesis: childbirth - 2, abortion - 2.

Concomitant pathology: chronic adnexitis, remission.

Somatic history is not burdened.

Objectively:

P.V. The external genitalia are developed correctly, the vaginal mucosa is not changed. The body of the uterus is increased to 5 weeks of pregnancy, soft, painless. Appendages on both sides without features. Parametries are free.

Allocations are white.

Ultrasound from 12.21.12 - the uterus is 54 × 46 × 48 mm in size, enlarged to 5 weeks of pregnancy, the contours are clear, even, the structure is heterogeneous. M-echo 0.9 mm. In the uterine cavity, the fetal egg KTR is 0.45 mm, SVD is 15 mm, the fetal heart rate is determined. The right ovary is 30 × 21 mm, the left ovary is 18 × 25 mm. Material sampling: after examining a woman on the mirrors under aseptic conditions, vacuum aspiration of the contents of the uterine cavity was performed. The resulting aspiration fluid from the uterine cavity was centrifuged for 15 minutes at a frequency of 3000 rpm. The suction fluid supernatant was used for further work.

With a semi-automatic pipette-dispenser, the supernatant of the aspiration fluid from the uterine cavity in the amount of 0.2 ml in the form of a drop was applied to the surface of previously prepared glass slides. After 24 hours, the obtained samples (facies) are microscopic in transmitted light and the morphological picture of the obtained facies is evaluated.

Morphological picture of the facies of aspiration fluid from the uterine cavity: symmetrical arrangement of cracks, clear rectangular parts, small round nodules, abnormal structures were not found. Conclusion: the morphological picture is characteristic of a progressing uterine pregnancy (see Fig. 1).

The result of histological examination No. 0967-67: decidual tissue, chorionic villi, characteristic of progressive uterine pregnancy.

Example No. 2

Patient F., born in 1980, history No. 8831, applied to the maternity ward of the Clinical Maternity Hospital on 18/12/12 with the aim of planning pregnancy.

No complaints at admission.

Menstrual and reproductive functions: menstruation from 13 years to 4 days, regular, moderate, 30 days cycle. The last menstruation on 17/12/12

In the anamnesis: childbirth - 2, abortion - 2.

Concomitant pathology: chronic adnexitis, remission

Somatic history is not burdened.

Objectively:

P.V. The external genitalia are developed correctly, the vaginal mucosa is not changed. The body of the uterus is not enlarged, soft, painless. Appendages on both sides without features. Parametries are free. The discharge is bloody (2nd day of menstruation).

Ultrasound from 12/18/12 - the uterus is 42 × 40 × 34 mm in size, the contours are clear, even, the structure is heterogeneous. M-echo 0.8 mm. The right ovary is 28 × 25 mm, the left ovary is 34 × 29 mm.

Material sampling was carried out similarly, see example No. 1.

Morphological study of menstrual flow from the uterine cavity: symmetrical type of cracks, clear rectangular sections, rounded nodules, lack of abnormal structures.

The morphological picture of the facies of menstrual flow: a symmetrical arrangement of cracks, clear rectangular parts, round small nodules, no abnormal structures were found.

Conclusion: the morphological picture corresponds to the norm (see Fig. 2).

The result of histological examination No. 0965-89: proliferative endometrium.

Example No. 3

Pregnant L., born in 1978, history No. 41935, was admitted to the gynecological department on 11/03/13 with a diagnosis of non-developing pregnancy at 6 weeks of gestation according to the type of death of the embryo.

Complaints upon receipt of a delay in menstruation, urogenital discharge from the genital tract during the day.

Menstrual and reproductive functions: menstruation from 15 years to 5 days, regular, moderate, cycle 25 days. Date of last menstruation 3 \ 02 \ 13 g. History: 1 abortion. In 2008 - laparotomy, cystectomy.

Somatic history: mild iron deficiency anemia.

Objectively:

P.V. The external genitalia are developed correctly, the vaginal mucosa is not changed. The body of the uterus is increased to 5 weeks, soft, painless. Appendages on both sides without features. Parametries are free. Allocations are vernacular.

Ultrasound of the pelvic organs from 03/10/13, the uterus is 62 × 52 × 52 mm in size, increased to 5 weeks of pregnancy, the contours are clear, even, the structure is heterogeneous. M-echo 12 mm. In the uterine cavity, the deformed fetal egg KTR is 4.7 mm, SVD is 3 mm, the fetal heart rate is not determined. The right ovary is 37 × 31 mm, the left ovary is 24 × 20 mm.

Material sampling was carried out similarly, see example No. 1.

Morphological picture of the facies of aspiration fluid from the uterine cavity: asymmetric arrangement of large cracks, the presence of a complex of morphological elements, including toxic plaques, linguistic and disc-shaped structures (see Fig. 3).

Conclusion: the morphological picture is characteristic of an undeveloped pregnancy.

The result of histological examination No. 0197709: decidual tissue, chorionic villi with lymphocytic infiltration, dead fetal egg

Example No. 4

Pregnant T., born in 1978, history No. 41435, applied to the antenatal clinic "Clinical Maternity Hospital" on 20/02/13 with the aim of planning pregnancy.

Complaints at admission does not show.

Menstrual and reproductive functions: menstruation from 13 years to 5 days, regular, moderate, cycle 28 days. The last menstruation on 02/18/13. History: 1 abortion, 1 birth, 1 non-developing pregnancy

Concomitant pathology: chronic adnexitis, remission.

Somatic history: chronic gastroduodenitis remission, impaired fat metabolism 2 degrees.

Objectively:

P.V. The external genitalia are developed correctly, the vaginal mucosa is not changed. The body of the uterus is not enlarged, soft, painless. Appendages on both sides without features. Parametries are free. Discharge - bloody, moderate (3rd day of menstruation).

Ultrasound from 02/18/13 - the uterus is 35 × 49 × 36 mm in size, the M-echo is 10 mm, the contours are fuzzy, uneven, the structure of the endometrium is heterogeneous with single hyperechoic inclusions. The structure of the myometrium is heterogeneous, echogenicity is reduced. Dilated vessels of the arcuate plexus. The right ovary is 23 × 34 mm, the left ovary is 27 × 31 mm. Conclusion: signs of chronic metroendometritis in remission.

Material sampling was carried out similarly, see example No. 1.

Morphological study of menstrual flow from the uterine cavity: asymmetric arrangement of radial fissures, the presence of toxic plaques, lingual and disc-shaped structures (see Fig. 3).

Conclusion: the morphological picture indicates a high risk of developing NB.

The result of histological examination No. 065564-77: proliferative endometrium with leukocyte infiltration.

Thus, the present invention is aimed at simplifying and increasing the sensitivity of the method for predicting an undeveloped pregnancy. The study of the morphological features of the aspiration fluid or menstrual flow makes it possible to predict the course and outcome of pregnancy in a short time, on a small amount of biological material.

The inventive method is characterized by low invasiveness, technical simplicity, accessibility to perform in conditions of antenatal clinics, inpatient facilities.

Claims (1)

A method for predicting an undeveloped pregnancy by examining biological fluids, namely, that the supernatant of aspiration fluid from the uterine cavity or menstrual flow in an amount of 0.2 ml is applied to the surface of a glass slide in the form of a drop, dried at room temperature for a day, after which the obtained samples (facies) are microscopic and in the presence of a complex of morphological elements, including: toxic plaques, linguistic and disc-shaped structures, they predict a high risk developed I'm developing pregnancy.

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