Differential diagnostic technique of leiomyoma growth patterns

FIELD: medicine.

SUBSTANCE: differential pre-operative diagnosis of the fast growth patterns of leiomyoma is ensured by measuring the relative content of CD3+CD56+CD158i lymphocytes in the peripheral venous blood of a woman with the fast-growing leiomyoma. If the derived value is equal to 0.8% or less, the true growth of leiomyoma is diagnosed, while the value of more than 0.8% shows the false growth of myoma.

EFFECT: using the given technique enables the preoperative diagnosis of the fast growth patterns of leiomyoma in the females of reproductive age that enables developing the optimum management of the patient in good time and selecting the conservative and surgical methods of treating.

1 tbl, 3 ex

 

The invention relates to medicine, namely to gynecology, and can be used as a preoperative differential diagnosis types of rapid growth of leiomyoma of the uterus in women of reproductive age.

The relevance of the proposed method is determined by the fact that fibroids is one of the most common gynecological diseases. Developing in women of reproductive age, uterine fibroids violates their reproductive function, leads to infertility, miscarriage, perinatal loss, which negatively affects the demographic situation in the country (Immune mechanisms of rapid growth of fibroids / A. I. Malyshkina, N. Y. Sotnikova, Y. S. Antsiferov, A. K. Krasilnikov. Ivanovo, 2010. - 272 S.).

There is the concept of rapid growth of fibroids, which refers to the increasing size of the uterus during the last year of observation on the value corresponding to 4-5 weeks of pregnancy. When studying morphological criteria of rapid growth of fibroids distinguish "true" type tumor growth, caused by the proliferation of myocytes, and "false" when the node increases due to secondary changes (edema, hyalinosis, sclerosis, petrification) associated with circulatory disorders (Immune mechanisms of rapid growth of fibroids / A. I. Malyshkina, N. Y. Sotnikova, Y. S. Antsiferov, A. K. Krasilnikov. - vinovo, 2010. - 272 S.). It is known that proliferating uterine fibroids has a less favorable course. Patients with this type of tumor growth is often a malignancy of the tumor and its recurrence. Women with proliferative type of growth of fibroids belong to the risk group for the development of hyperplastic endometrium and mammary glands, as well as on the development of cystic benign and malignant tumors in the ovaries (Fibroids / Ed. by I. S. Sidorova. - M.: MIA, 2003. - 256 S.).

When choosing tactics and treatment of patients is important for defining the types of tumor growth. In this regard, we need to search for new criteria that allow to accurately differentiate between "true" and "false" type growth of leiomyoma of the uterus.

A known method of differential diagnosis of a rapidly growing leiomyoma of the uterus on the content of epidermal growth factor in endometrial. When the value of 6 ng/ml or less, diagnose "false" growth of fibroids, while values greater than 6 ng/ml is the "true" growth of fibroids (patent No. 2258222. Of The Russian Federation. The method of differential diagnosis of rapidly growing leiomyoma of uterus / L. C. Moiseeva, A. I. Malyshkina, A. K. Krasilnikova, etc. // Invention. The utility model. - 2005. No. 22). However, this method has several disadvantages.

1. The method is invasive.

2. Taking endometrial investigated the e is carried out during the hysteroscopy in stationary conditions, which is associated with hospitalisation and use of expensive equipment.

3. The method is extremely time-consuming, because its implementation requires the allocation of endometrial tissue enriched population of mononuclear cells with their subsequent cultivation for 24 hours. For mycobacterial culture required expensive nutrient and CO2-incubator.

A method of differential diagnosis of rapidly growing leiomyoma of the uterus in women of reproductive age, by definition, in the blood content of TNF-α, the absolute content of CD4+ lymphocytes, the relative content of T-cytotoxic lymphocytes CD8+, results of spontaneous NBT-test, and serum total complement activity and calculation of index Y by the formula. When the Y value is less than the forecast of 4.6 nephrolithiasis option growth of uterine fibroids; when Y is greater 6,8 predict proliferating option growth of uterine fibroids; for Y from 4.6 to 6.8 - uncertain prognosis, requiring the use of the method of descriptive statistics, using quality anamnestic and clinical criteria (patent No. 2400757. Of The Russian Federation. The way to diagnose morphological types of uterine leiomyoma / Ie; but, Y. O. Sevastyanova, I. A. Gazieva, etc. // Invention. The utility model. - 2010. No. 27). However, this method it is no several disadvantages.

1. The method is extremely time-consuming, as it involves a large number of techniques and, consequently, has a higher cost.

2. There is a range of values from 4.6 to 6.8, which is regarded as an uncertain prognosis, requiring the use of additional methods of research that has an impact on the accuracy of the results. According to the mathematical formula of this study, the reliability of the results is 70%. The closest technical solution to the claimed method is a method of differential diagnosis of the type of growth of leiomyoma of the uterus in women of reproductive age. In the peripheral blood of women with growing leiomyomas uterine determine the relative number of lymphocytes with the phenotype CD4+IL-10+ and if the value of this rate of 5.8%, or more, diagnose the "true" growth of uterine leiomyoma, and a value of less than 5.8% - "false" growth (patent No. 2361216. Of The Russian Federation. The method of differential diagnosis of the type of growth of uterine leiomyoma / N. Y. Sotnikova, L. C. Moiseeva, O. M. Boyko, etc. // Invention. Utility models. - 2009. - №19).

However, it was shown that the level of this pool T-helper cells increased in patients with several diseases. It is established that the CD4+IL-10+ increased in patients with connective tissue dysplasia (Regulatory T cells in eripheral blood of patients with mixed connective tissue disease / S. Barath, S. SipKa, M. AleKsza et al. // Scand. J. Rheumatol. - 2006. - Vol.35, No. 4. - P. 300-304), with autoimmune diseases such as systemic lupus erythematosus (Monocytes and T lymphocytes contribute to a predominance of interieukin 6 and interleukin 10 in systemic lupus lupus / S. Mellor-Pita, M. J. Citores, R. Castejon et al. // Cytometry In Clin. Cytom. - 2009. - Vol.76, No. 4. - P. 261-270), and with antiphospholipid syndrome (Immunological features of primary anti-phospholipid syndrome in connection with endothelial dysfunction / P. Soltesz, H. Der, K. Veres et al. // Rheumatology (Oxford). - 2008. - Vol.47, No. 11. - P. 1628-1634). In addition, there are a number of studies A. Bourn, confirming the change of this indicator during infection with herpes simplex virus (Dysregulated expression of IFN-gamma and IL-10 and impaired IFN-gamma-mediated responses at different disease stages in patients with genital herpes simplex virus-2 infection / R. Singh, A. Kumar, W. D. Creery et al. // Clin. Exp.Immunol. - 2003. - Vol.133, No. 1. - P. 97-107). In addition, there is an increase in the pool of T-helper cells in the blood of allergic patients after a course of allergen-specific immunotherapy (Induction of T regulatory cells by standardized house dust mite immunotherapie: an increase in CD4+CD25+ interieukin-10+ T cells expressing peripheral tissue trafficking / L. M. Gardner, F. C. Thien, J. A. Douglass et al. // Clin. Exp. Allergy. - 2004. - Vol.34, No. 8. - P. 1209-1219). Thus, this indicator varies in a wide range of diseases, and therefore the specificity of the method is reduced. The accuracy of the method is 79,1%. These drawbacks are proposed to be eliminated in the present method.

The technical solution of the proposed method is to improve the accuracy of cooperation the th differential diagnosis types growth of leiomyoma of the uterus in women of reproductive age 88,2%, sensitivity to 85.0%, specificity up of 92.9%. The claimed technical result is achieved by the fact that in the peripheral blood of women with growing leiomyomas uterine determine the relative number of lymphocytes with the phenotype CD3+CD56+CD158i and if the value of this indicator more than 0.8% diagnose "false" the growth of uterine leiomyoma, and a value equal to or less than 0.8%, the "true" growth.

The novelty of the proposed method lies in the fact that for the first time are encouraged to differentiate between "true" and "false" type rapid growth of uterine leiomyoma by definition in the peripheral blood expression of the receptor CD158i on natural killer T-lymphocytes (EC-T), markers which are CD3+CD56+. Previously it was shown that the level of this indicator decreased in endometrial tissue in women with rapidly growing fibroids (Character expression of KIR receptors of natural killer T-lymphocytes in the endometrium uterine fibroid different growth / A. I. Malyshkina, N. Y. Sotnikov, A. N. Voronin and other // problems of reproduction. - 2012. - Special. release. - S. 186). In other works studied receptors (CD1581) on natural killer cells (CD56+). There was an increased CD56+CD158i lymphocytes during HIV infection (Perturbation of the natural killer cell compartment during primary human immunodeficiency virus 1 infection primarily involving the CD56 bright subset / P. Mantegani, G. Tambussi, L. Galli et al. / Immunology. - 2010. - Vol.129, No. 2. - P. 220-233) and chronic hepatitis (High frequency of polyfunctional natural killer cells with high expression of NKG2D is linked to HBV-specific T-helper type 1 immune control / I. Carey, A. Mendes, S. Bansal et al. // Journal of Hepatology. - 2010. - Vol.52. - P. 176). In addition, the marked increase in CD56+CD158i lymphocytes in cancer of blood - leukemia T-cell large granular lymphocyte leukemias have multiple phenotypic abnormalities involving pan-T-cell antigens and receptors for MHC molecules / R. Lundell, L. Hartung, S. Hill et al. // Am J. Clin. Pathol. - 2005. - Vol.124, No. 6. - P. 937-946). For the differential diagnosis of types of rapid growth of uterine leiomyoma content of CD3+CD56+CD158i lymphocytes have not used.

The population of lymphocytes with the phenotype CD3+CD56+ referred to as natural killer T-lymphocytes (EC-T). It is established that the targets for these cells are transformed (infected) and rapidly proliferating cells (Yarylo A. A. Immunology. - M.: GEOTAR-Media, 2010. - 752 C.). Cytotoxic activity of EC-T is regulated by activating and inhibitory KIR receptors that interact with ligands on the target cells (Activating and inhibitory receptors of natural killer cells / J. Pegram Hollie, Daniel M. Andrews, Mark J. Smyth et al. // Immunology and Cell Biology. - 2011. - Vol.89. - P. 216-224). Receptors CD158i are activators of cytotoxic lymphocytes directed against target cells, because of their involvement in tumor immunity (Activating and inhibitory receptors of natural killer cells CD158 Receptor Controls Cytotoxic T-Lymphocyte Susceptibility to Tumor-Mediated Activation-Induced Cell Death by Interfering with Fas Signaling / A. Gati, N. Guerra, C. Gaudin et al. //Cancer Research. - 2003. - Vol.63, No. 1. - P. 7475). Changes in the expression of activating CD158i Rotz is tori, ligands are molecules HLA-Cw on the surface of transformed cells of the myometrium, reflect the state of the anti-tumor surveillance by the immune system. Thus, the EC-T, expressing the activating KIR receptors are involved in the mechanisms of rapid growth of fibroids.

The method is performed by a standard method as follows.

1) To 3 ml of heparinized venous blood obtained from the cubital vein, add 3 ml of medium 199 and produce an enriched population of mononuclear cells standard method of high-speed centrifugation (Bourn A. Isolation of mononuclear cells and granulocytes from human blood and bone marrow // Scand. J. Clin. Invest. - 1968. - Vol.21, No. 97. - P. 77).

2) Suspension of mononuclear cells washed twice with saline, adjusting the cell concentration to 1×106KL/mn. To 0.1 ml of the resulting suspension at the same time add 20 ál of monoclonal antibodies anti-D3 labeled PC 5, anti-CD56, labeled D, anti-SW labeled FICE (JSC "Sorbent", Moscow). Cells incubated for 30 min at room temperature in the dark, washed in 1 ml of saline.

3) Determine the relative (percentage) number of triple-positive CD3+CD56+CD158i lymphocytes in lymphocytic gate on a flow cytometer.

The use of the proposed method with high accuracy 88,2% to differentiate between "true" and "false" is the IPA rapid growth of leiomyoma of the uterus.

Distinctive features of the process are: the establishment of differential diagnostic parameter of the relative content of CD3+CD56+CD158i lymphocytes, when the value of which is 0.8% or less diagnosed "true" growth of uterine leiomyoma, and a value of more than 0.8% is "false" the growth of leiomyoma of the uterus.

The essence of the proposed method is illustrated by the following examples.

Example 1. Patient Z., 39 years old, was admitted for planned surgery. Was observed in the antenatal clinic with uterine myoma of the small sizes (6 weeks). During the next inspection one year after the first treatment discovered the rapid growth of fibroids. The patient experienced pain in the lower abdomen and lower back, heavy menstruation, accompanied by a decrease of hemoglobin. History 1 miscarriage, 3 medical abortion. Method of contraception - barrier. The patient planning to conceive.

During examination: cervix without mucosal defects, the body of the uterus is increased to 18 weeks of pregnancy due to myoma node, located on the rear wall of the uterus. The body of the uterus is thick, movable, painless. Appendages to palpation without features.

According to CCS: the body of the uterus heterogeneous patterns, sizes together with site - 142×130×122 mm In the rear wall of the uterus is intramurals-subserous site 132×123×107 mm, the uterus is not deformed. The thickness of the end the Tria 13 mm The right ovary 30×18 mm Left ovary 38×30 mm Both ovaries normal patterns.

Blood analysis: hemoglobin - 94 g/l, erythrocytes-4.0 t/l, colour index-0.6, leukocyte - 5,2 g/l, stab neutrophils - 2, segmented neutrophils - 59, monocytes - 8, lymphocytes - 28, ESR - 15 mm/h

Clinical diagnosis: intramurals-subserous leiomyoma of the uterus. Rapid growth. Hypermenorrhea, leading to anemia. Pain syndrome.

According to immunological examination by the present method it is established that the content in peripheral blood CD3+CD56+CD158i lymphocytes was 0.55%, which is lower than the proposed option (0,8%).

Conclusion: diagnosed "true" type rapid growth of leiomyoma of the uterus.

Given the age of the woman and unrealized reproductive function, scheduled organ-preserving volume of surgical intervention. In connection with the results of the growth of uterine fibroids, and since this type of tumor has a less favorable for the patient as a pre assigned agonist RH-RG - buserelin of 3.75 mg of Drug was administered once in 28 days intramuscularly for 2 months. In the second stage, 3 weeks after the last injection, the woman admitted for surgical treatment.

According to CCS after treatment: the body of the uterus heterogeneous patterns, sizes together with node - 125×122×102 mm On Adna of the uterus is intramurals-subserous site 120×108×87 mm The uterus is not deformed. Endometrial thickness of 8 mm Right ovary 30×18 mm Left ovary 38×30 mm Both ovaries normal patterns.

Blood test (after treatment): hemoglobin - 120 g/l, erythrocytes-4.2 t/l, leucocytes - 5,3 g/l, stab neutrophils - 1, segmented neutrophils - 63, monocytes - 8, lymphocytes - 26, ESR -18 mm/h

Transaction - laparotomy, myomectomy.

Histological examination of the surgical material from the proliferating leiomyoma uterus". Thus, the diagnosis of "true" type growth of leiomyoma of the uterus was confirmed.

Example 2. Patient B., 38 years. During the year, was observed in female consultations on uterine myoma of the small sizes (6-7 weeks). For a specified period of menstruation became more abundant, there is a weakness after menstruation. History 1 abortion, 1 timely delivery without complications. Method of contraception - barrier.

At vaginal examination: cervix cylindrical, without defects mucosa. The uterus is increased to 12-13 weeks of pregnancy, due to multiple hosts, much too soft to palpation, mobile, painless. The appendages are not palpable, vaults available.

According to the ultrasonic scanning (CCS): the body of the uterus dimensions 120×80×115 mm, heterogeneous structure. In the front wall of the uterus is intramurals-subserous site diameter is rum 48 mm in the right side wall with a diameter of 38 mm, in the left corner of the node with a diameter of 48 mm, the rear node diameter 57 mm, All nodes of heterogeneous structure. The uterus is not deformed. Endometrial thickness 10 mm Right ovary 38×26 mm, left ovary 37×20 mm Both ovaries normal patterns.

Blood analysis: hemoglobin - 63 g/l, erythrocytes - 2,78 t/l, colour index of 0.7, the leukocyte count was 3.9 g/l, stab neutrophils - 5, segmented neutrophils - 50, monocytes - 9, lymphocytes - 31, ESR - 20 mm/h

Clinical diagnosis: multiple intramural-subserous leiomyoma uterus large sizes. Rapid growth. Hypermenorrhea, leading to anemia.

According to immunological examination by the present method it is established that the content in peripheral blood CD3+CD56+CD158i lymphocytes amounted to 1.35%, which is higher than the proposed option (0,8%).

Conclusion: diagnosed "false" type rapid growth of leiomyoma of the uterus.

Transaction: Subtotal hysterectomy without appendages.

Histological examination of the surgical material was diagnosed: "leiomyoma uterus with severe edema and focal hyalinosis stroma". Thus, the diagnosis of "false" type growth of leiomyoma of the uterus was confirmed.

Example 3. Patient B., 41. Was observed in the antenatal clinic for 4 years with uterine myoma of the small sizes (5-6 weeks is). In the past, there were profuse menstruation, accompanied by a decrease of hemoglobin. History 1 timely delivery without complications, 3 medical abortion. Method of contraception - barrier.

At vaginal examination: cervix without defects mucosa, uterus increased to 15 weeks of pregnancy, due to a host that is located in the anterior wall of the uterus. The uterus is a dense, mobile, painless. The appendages are not palpable, vaults available.

According to CCS: the body of the uterus dimensions 120×75×100 mm, heterogeneous structure. The front wall is visualized intramurals-subserous site 95×65×80 mm with the phenomenon of swelling. The uterus is not deformed. Endometrial thickness 12 mm Right ovary size 38×25 mm, left ovary size 37×20 mm, regular patterns.

Blood analysis: hemoglobin - 121 g/l, erythrocytes - 4,55 m/l, color index is 0.8, leukocyte - 4.0 g/l, segmented neutrophils - 62, monocytes - 6, lymphocytes - 26, ESR - 19 mm/h

Clinical diagnosis: intramurals-subserous leiomyoma of the uterus. Rapid growth. Hypermenorrhea. Pain syndrome.

According to immunological examination by the present method it is established that the content in peripheral blood CD3+CD56+CD158i lymphocytes was 0.8%, which corresponds to the edge value of the requested parameter.

Conclusion: diagnosed "true" type b is strictly growth of leiomyoma of the uterus.

Transaction: Subtotal hysterectomy without appendages.

Histological examination of the surgical material obtained result is: "proliferating leiomyoma with a moderate number of vessels and stromal edema". Thus, the diagnosis of "true" type growth was confirmed.

In this way were examined 34 women of reproductive age with growing leiomyomas of the uterus. These studies are listed in table 1.

Table 1
IndexThe number of the examined
Total female34
true-positive result17
false-positive result1
a true-negative result13
a false-negative result3
Total: the accuracy of the proposed method - 88,2%, the sensitivity of the proposed method - 85,0%, specificity of the proposed method - of 92.9%

The advantages of the proposed method.

1. The way n which increases accuracy, sensitivity, specificity differential diagnosis types growth of leiomyoma of the uterus.

2. The method expands the Arsenal of tools for the differential diagnosis of types of growth of leiomyoma of the uterus.

The method of differential diagnosis types rapid growth of uterine leiomyoma by immunological studies of peripheral venous blood in women of reproductive age, characterized in that to determine the relative number of CD3+CD56+CD158i lymphocytes, and if the value of this indicator, equal to 0.8% or less, diagnose the "true" growth of uterine leiomyoma, and a value of more than 0.8% is "false" the growth of uterine leiomyoma.



 

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3 ex, 1 tbl

FIELD: medicine, pediatrics.

SUBSTANCE: in 5-10-d-aged neonatals one should detect relative content of CD8+HLA-DR+ lymphocytes in peripheral blood and at its value being below 2.4% it is possible to predict the healing at different types of neonatal infections.

EFFECT: higher accuracy of prediction.

3 ex, 1 tbl

FIELD: medicine, gynecology.

SUBSTANCE: invention relates to a method for diagnosis of internal endometriosis in peripheral venous blood of women wherein the relative content of lymphocytes CD25+ is determined. Internal endometriosis is diagnosed at values of this index 6% or above. Proposed method provides carrying out diagnosis of internal endometriosis in women with high precision, sensitivity and specificity that allows carrying out the correct and well-timed necessary complex of curative-prophylactic treatment.

EFFECT: improved method for diagnosis.

1 tbl, 3 ex

FIELD: medicine, immunological laboratory diagnostics.

SUBSTANCE: at terms from 6 to 12 wk of gestation one should study relative content of CD3+CD16+ lymphocytes in peripheral venous blood and at its values being either equal or above 5.4% one should predict the development of light-degree gestosis to carry out the complex of curative-prophylactic means.

EFFECT: higher efficiency of prediction.

3 ex, 1 tbl

FIELD: medicine, laboratory diagnostics.

SUBSTANCE: during the 1st trimester of pregnancy (6-13 wk) in peripheral venous blood in women at risk of failed pregnancy one should detect relative content of CD16+CD56- lymphocytes and at its value being either equal or below 11% it is possible to predict the development of infectious diseases in full-term neonatals during the first 7-10 d of their lives. The innovation enables to predict the development of local form of infectious-inflammatory diseases in full-term neonatals.

EFFECT: higher accuracy of prediction.

3 ex, 1 tbl

FIELD: medicine, clinical laboratory diagnostics.

SUBSTANCE: in the sample of peripheral venous blood one should determine relative content of CD45RO+ lymphocytes and at its value being equal to 31% or lower it is possible to diagnose external genital endometriosis. The method is atraumatic and enables to diagnose external genital endometriosis at high accuracy.

EFFECT: higher efficiency of diagnostics.

3 ex, 1 tbl

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