Diagnostic technique for foetus growth inhibition

FIELD: medicine.

SUBSTANCE: amniotic fluid of a pregnant woman on her 16-19 weeks of pregnancy is analysed for the concentration of arginine and praline by capillary electrophoresis. An aspect ratio is calculated, and if its value is equal to 3.2 and higher, foetus growth inhibition is diagnosed.

EFFECT: higher diagnostic accuracy of foetus growth inhibition.

2 ex

 

The invention relates to medicine, namely to obstetrics and Perinatology, and will find wide application in the development of midwifery practice.

The problem of delayed fetal development refers not only to the field of Perinatology, but also closely related to obstetric pathology and questions obstetric tactics (Ainstream et al. The fetal development. Kyiv. Health. 1988). The frequency of this complication of pregnancy according to some authors ranges from 4.5%to 39%, ranking in the structure of causes of perinatal morbidity and mortality in third place (Athboy, Mavodones // Obstetrics and gynecology. 1985. No. 12. S-30; Emilia, Tshapaieva // Obstetrics and gynecology. 1995. No. 6. P.18-24).

The effectiveness of remedial measures when diagnosed, usually at the end of II-III trimesters of pregnancy the fetal development is extremely reduced and only aimed at the prolongation of pregnancy, not its overcoming. In connection with this interest is the elucidation of early diagnostic markers of delayed fetal growth (rgts) in order to conduct timely remedial measures.

Analysis of patent and scientific and medical literature made it possible to analyze the existing methods of diagnosis of growth retardation of the fetus. A more modern technique of diagnosis rgts is the use of multiple fotometricheskikh until the of Atala, but they are informative only in the third trimester of pregnancy - 26-38 weeks (Ainstream, Annonin, Mavodones // Obstetrics and gynecology. 1988. No. 6. P.32-35). Ultrasound diagnosis rgts based on the comparison fotometricheskikh the results of the study, with the target indicators for the duration of pregnancy. One of the most informative parameters is biparentally the size of the head of the fetus. However, the measurement of this indicator cannot be used for the detection of growth retardation of the fetus.

Recently, to identify the syndrome rgts used diplomaticheskii determination of volumetric blood flow in the aorta of the fetus, the umbilical vein or the internal carotid artery with subsequent analysis of the curves of the flow velocity. Given that qualitative assessment of blood flow using systolicdiastolic ratio (C/D) and the resistance index reflects the same patterns in wide clinical practice for the diagnosis rgts also use the definition of pulsation index (PI), an increase which occurs when rgts in umbilical Vienna and a sharp decrease in the internal carotid artery. However, all these studies are the most reliable in the period from 17 to 20 weeks of pregnancy. Diagnostic value of disposable dynamic scintigraphy when is America fetal growth is 75-79% (Emilia, Tshapaieva // Obstetrics and gynecology. 1984. No. 6. P.18-24).

A known method for the diagnosis of threatening the status of the foetus in patent No. 2076624 (1997), which allows simultaneous registration of the electrical activity of the heart of the mother and fetus, and in the absence of a correlative relationship diagnose threatening the fetus or the degree of violation of the condition of the fetus. Regarding this analogue can be emphasized that the use of it in clinical practice allows to assess the presence of abnormalities in fetal development significantly in late pregnancy.

A known method for the diagnosis of intrauterine fetal development, described in the patent No. 2086981 (1997), which in the venous blood of the woman is determined by the absolute content of b-lymphocytes (VA) and the relative content of neutrophils have receptors for C3 component of complement (EAC-RON), and when the indicators VA greater than or equal to 0.3×109µg/l, and EAC-RON greater than or equal to 16%, the fetus is assessed as non-compliant physiological development and diagnosed the intrauterine development of the fetus. This method is not specific, since changes in the content of b-lymphocytes, and neutrophils may change at various diseases of the mother, not related to anti-aircraft regiment, as well as the accuracy of the method sostav the em only 80%.

In patent No. 2172958 (2001) proposed a method for the diagnosis of anti-aircraft regiment, which allows the diagnostics of early stages of growth retardation of the fetus by analyzing peripheral blood, including DNA isolation, conducting polymerase chain reaction and determination of allele PL-AII gene glycoprotein IIIA. It should be noted that this method has a major drawback, especially the limited application because of the complexity of its implementation, expressed in mnogoetapnoe and duration of its execution.

Another way to early prediction of delay syndrome fetal development is defined in the peripheral venous blood content HLA-DR + monocytes (patent 2265224, 2004), and its value is equal to or smaller 57,9%, predict the development of the syndrome of rgts. It should be noted that this method is also limited in application because of the complexity and the duration of its execution.

In the patent of Russian Federation №2331365 (2007) proposed a method for the diagnosis of syndrome of intrauterine development of the fetus by definition in the blood of a pregnant woman content of allantoin, and when the values of its concentration from 2.9 to 5.8 mmol/l diagnose the syndrome of intrauterine development of the fetus. This method is not specific, as expressed by the decrease of this indicator is observed not only in anti-aircraft regiment, and when karasuyama the pregnancy and early spontaneous abortion.

The prototype of the present invention is elected as a method for predicting the intrauterine development of the fetus, described in the patent No. 2246733 (2003). The essence of the prototype method is that in the serum of pregnant determine the content insulinopenia growth factor and vascular endothelial growth factor, calculate the ratio of their relationship and the value of its 28.5 and below diagnosed fetal development.

This method has a number of imperfections:

1. The method is not sufficiently specific - level change growth factors observed for the various complications of pregnancy - eclampsia, non-developing pregnancy, threatened miscarriage, premature birth (Avilov et al. // Ross. Bulletin akush. and gin. - 2005. No. 3. P.7-9).

2. The method is laborious and time-consuming it is running.

3. The implementation of the method requires expensive equipment and import sets of reagents for immunoassay analysis, which hinders its widespread use in clinical practice.

These drawbacks are eliminated in the invention.

The objective of the invention: improving the accuracy and specificity through the development of a specific method for the diagnosis of growth retardation of the fetus (rgts).

The problem is solved in that in the amniotic fluid of women in pregnancy (16 weeks the ü of gestation) by capillary electrophoresis determine the content of arginine and Proline, calculate the ratio of their relationship and the magnitude of the coefficient, equal to 3.2 and above, diagnosed with fetal growth retardation.

The technical result is obtained during use of the invention is to improve the accuracy of diagnosis of growth retardation of the fetus, which allows you to assign a timely and adequate therapy in the earlier period.

The novelty of the proposed method lies in the fact that for the first time proposed to diagnose rgts pregnant at an early age, exploring the amniotic fluid (once) and specifying in it the ratio of the amino acids arginine and Proline.

It is known that amino acids perform the vital role of "construction material" in the biosynthesis of a specific tissue proteins, enzymes, peptide hormones, etc. the Role of amino acids as essential substrates in metabolic processes is inextricably linked with another no less important is their function as regulators of many biochemical and physiological reactions (plastic, energy metabolism, and others). Among the nitrogenous components of the amniotic fluid free amino acids are important, so how to effectively use the fruit as a plastic material (Snaaake, .Asai // Acta Obstet. Ginec. Jap., 1979. - 31. No. 2. P.226).

Special attention is currently given to the role of arginine in beremend the STI, and also to diagnose the condition of the fetus (Iaustralia et al. // The Bulletin of PFUR. - 2005. No. 4. - Pp.92-98; Avishalom // Izvestiya vuzov North KAV. the region. - The natural. science. - 2006. No. 8. - S.112-116; Svilenova et al. // Ross. Bulletin akush. and gin. - 2007. No. 2. - P.4-7). Arginine is one of the most effective stimulant product somatotropic pituitary hormone (growth hormone) and allows it to maintain its concentration at the upper normal limits, and the lack of arginine leads to stunted growth of children (Mcherron et al. // Pharmacy. - 2004. - T. No. 5. - P.18-20).

An important role is also played by another amino acid - Proline. Particularly rich in them the main protein of connective tissue - collagen, which in high concentrations is contained in the bones and muscle tissue. Proline is a non-polar amino acid, is a building material cellular proteins. Recently published data on the practical use of Proline. It is widely used in medicine in the composition of amino acid mixtures for parenteral nutrition, in cryomedicine (Y. Morita et. al. 2005) and as the basis for the creation of antipsychotic drugs, as well as in the food industry as an antioxidant (Sagamino // ABT. Diss. Kida. Biol. Sciences. - 2005. - 240 S.).

This method is carried out by examination of samples of the amniotic fluid.

Under aseptic conditions, t is academically access produce the capture of amniotic fluid under ultrasound for the diagnosis of chromosomal pathology.

Before analysis the samples of amniotic fluid is treated with a 10% solution of trichloroacetic acid to deproteinization. Next, the samples centrifuged for 15 minutes at 5500 rpm C. the Obtained supernatant solution was used for analysis.

Identification and quantitative determination of free form arginine and Proline performed by capillary electrophoresis using unmodified silica capillary ("Capel-105", "Lumex", SPb., Russia).

Before you begin the capillary sequentially washed with 1 M hydrochloric acid, distilled water, 0.5 M sodium hydroxide solution and the buffer solution (20 mm borate buffer, pH 9,15). The time of sample introduction 15 C. a Pressure of 30 mbar, operating voltage 20 kV. Use direct spectrophotometric detection at 200 nm. All studies are carried out at a temperature of 30°C.

Data processing is done using an IBM PC with the software "Multichrome" (JSC "ampersand character"). Then calculate the ratio of arginine to Proline.

The efficiency of this method is confirmed by the following clinical examples.

Example 1. The patient Peninsula, 20 years. Outpatient map No. 193. Addressed in the consultative polyclinic NIIAP (22.12.2008, in connection with suspected chromosomal abnormality of the fetus, the time taken is nasty 16 weeks.

25.12.2008, in aseptic conditions transabdominal access made taking amniotic fluid under ultrasound for the diagnosis of chromosomal abnormalities and determination of arginine and Proline by capillary electrophoresis. By capillary electrophoresis identified in amniotic fluid content of arginine, which amounted to 216 mg/ml, and Proline - 70 mg/ml

The ratio of arginine to Proline = 3,1, indicating the absence of the occurrence of delayed fetal growth. During the whole period of gestation the patient was under observation in the clinic NIIAP.

11.05.2009, the woman came to give birth in the maternity unit of NIIAP.

D-C: pregnancy 39 weeks. Birth spontaneous, ended in a live birth, full-term baby with a mass 3200,0.

This clinical example confirms the accuracy of diagnosis of the absence of the occurrence of rgts.

Example 2. Patient G to 30 years. Outpatient map No. 18. Addressed in the consultative polyclinic NIIAP (19.01.2009, in connection with suspected chromosomal abnormality of the fetus, gestational age 22 weeks.

21.01.2009, in aseptic conditions transabdominal access made taking amniotic fluid under ultrasound for the diagnosis of chromosomal abnormalities and determination of arginine and n is oline by capillary electrophoresis. By capillary electrophoresis identified in amniotic fluid content of arginine, which amounted to 61 µg/ml, and Proline - 19 mcg/ml the Ratio of arginine to Proline = 3,2.

The diagnosis of the possible occurrence of rgts.

29.04.2009, the woman came to give birth in the maternity unit of NIIAP.

D-C: pregnancy 37-38 weeks. Birth spontaneous, ended in a live birth with a mass 2200,0.

This clinical example confirms the ability to accurately diagnose rgts on the proposed factor.

By the present method were examined 38 women, among whom 29 ratio of arginine/Proline in the amniotic fluid is equal to 3.1 and below, and in 9 patients it was 3.2 and above. All pregnant women with factor of 3.2 and above identified fetal growth retardation, confirmed the birth of underweight children (2370-2530,0). Women with a ratio of arginine/Proline was equal to 3.1 and below, gave birth to children without signs of rgts.

The above confirms the diagnostic value of the claimed method.

Thus, the inventive method is more effective in comparison with the known and has a number of advantages:

1. It has high accuracy. The accuracy is 99%.

2. Method-specific (definition rgts this factor is a specific method).

3. the manual high-sensitivity, allows definition rgts at the preclinical stage.

Approbation of the invention carried out on a sufficient volume of clinical material, which confirmed the efficiency of the method and the possibility of its application in clinical practice.

A method for the diagnosis of growth retardation of the fetus by a study of the biological fluid of a pregnant woman, characterized in that a pregnant woman in terms 16-19 weeks in amniotic fluid by capillary electrophoresis determine the content of arginine and Proline, calculate the ratio of their relationship and the magnitude of the coefficient, equal to 3.2 and above, diagnosed with fetal growth retardation.



 

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