RU2324184C1 - Method of evaluation of factor of erythrocytes stability in hypoxia - Google Patents

Abstract

FIELD: medicine; laboratory techniques.

SUBSTANCE: to evaluate the stability factor of erythrocytes in hypoxia, patient's blood sample is taken; heparin 0.1 ml is used as a coagulant. The sample is centrifuged at 4000 rpm for 10 min, and the erythrocyte suspension is received. The real part of dielectric constant

of erythrocyte suspension is measured immediately after it is obtained and in 60 min, at fixed frequencies -

1 and

The polarisation factors (PF₁ and PF₂) are calculated by formula:

, then the stability factor of erythrocytes in hypoxia is calculated (SF): SF=SF₂: SF₁, and if it is 1.0 and less, the affected erythrocytes stability to hypoxia is diagnosed.

EFFECT: method allows the dynamic evaluation of treatment efficiency in diseases, where the free-radical processes play the key role in pathogenesis.

4 tbl, 3 ex

Description

The invention relates to medicine and can be used for a spectrometric determination of the resistance of red blood cells to hypoxia in patients in whom free radical reactions are leading in the pathogenesis of the disease, for example, coronary heart disease (CHD). [Kogan A.Kh., Ershov V.I., Sokolova I.Ya. On the mechanism for enhancing free radical processes in patients with coronary heart disease - angina pectoris, depending on its severity. // Ter. archive - 1994. - No. 4. - S.32-36.]

Activation of free radical processes in organs and blood under the influence of hypoxia is accompanied by an increase in the concentration of free radicals of the seven-quinone type and manifests itself as an increase in the polarization coefficient (CP). [Abankin V.P., Pidemsky E.L. Polarization processes in isolated organs and blood are normal and when exposed to various chemical and physical factors. // Study of the biological effects of new products of organic synthesis of natural compounds. Perm, 1977. - S.15-22.] Based on the change in KP, it is possible to determine the coefficient of resistance (KU) of red blood cells to hypoxia.

In clinical medicine, the level of antioxidant enzymes, antioxidants themselves and lipid peroxidation products are used to detect disorders of oxidative metabolism in red blood cells as a result of activation of free radical reactions.

The disadvantage of the methods is that they require special equipment and expensive reagents. [Kagan V.E., Orlov O.V., Prilipko L.L. Problems of analysis of endogenous lipid peroxidation products. // M., Itogi Nauki i Tekhniki, 1986, VINITI, Ser. Biophysics. eighteen.]

Effect: simplification of the study with high accuracy of the method and the ability to evaluate the effectiveness of treatment.

эритроцитарной взвеси на фиксированных частотах -

и на

Вычисляют коэффициент поляризации (КП₁) по формуле:The task is achieved by the fact that blood is taken from the examined person from a cubital vein in a volume of 10.0 ml, heparin 0.1 ml is used as an anticoagulant. Centrifuged for 10 min at a speed of 4000 rpm and get an erythrocyte suspension. Measure the real part of the dielectric constant

erythrocyte suspension at fixed frequencies -

and on

The polarization coefficient (KP ₁ ) is calculated by the formula:

эритроцитарной взвеси повторяют через 60 мин на частотах

и

и вычисляют КП₂ по формулеMeasurement of the real part of the dielectric constant

erythrocyte suspension is repeated after 60 minutes at frequencies

and

and calculate KP ₂ according to the formula

Then determine the coefficient of resistance of red blood cells to hypoxia:

KU = KP ₂ : KP ₁ and when its value is 1.0 or less, a violation of the resistance of red blood cells to hypoxia is diagnosed.

эритроцитарной взвеси на двух частотах -

и на

Вычисляют КП 1 по формуле:The task is achieved as follows. The subject takes blood from a cubital vein in a volume of 10, 0 ml. As an anticoagulant, 0.1 ml heparin is used. Blood is centrifuged for 10 min at a centrifuge speed of 4000 rpm and an erythrocyte suspension is obtained. Part of the erythrocyte suspension is placed in a Teflon cell mounted on a micrometer with oxidized tantalum electrodes without through conductivity. Tantalum oxide dispersion is absent up to 1 MHz. The cell electrodes are connected to a two-channel dielspektrometer created at Perm State University (copyright certificate No. 1337827 of 05.15.87). Turn on the device and on its scale determine the real part of the dielectric constant

erythrocyte suspension at two frequencies -

and on

Calculate KP 1 by the formula:

1 и на

After 60 minutes, the remaining erythrocyte suspension is placed in the Teflon cell, the device is turned on, and the real part of the dielectric constant at two frequencies is determined on its scale -

1 and on

KP _{2 is} calculated by the formula:

Then determine the coefficient of resistance of red blood cells to hypoxia of the examined KU = KP ₂ : KP ₁ and with its value of 1.0 or less, a violation of the resistance of red blood cells to hypoxia is diagnosed.

Examples of specific performance of the method.

Example 1. Surveyed K., 18 years old, the diagnosis is healthy.

и на

Blood was taken from a cubital vein in a volume of 10.0 ml with the addition of 0.1 ml of heparin. It was centrifuged for 10 min at a centrifuge speed of 4000 rpm; an erythrocyte suspension was obtained. Part of the erythrocyte suspension was placed in a Teflon cell, the electrodes were connected to a two-channel dielspektrometer, the device was turned on and the real part of the dielectric constant was determined on a scale

and on

The polarization coefficient (KP ₁ ) was 6.17.

и на

After 60 minutes, the rest of the erythrocyte suspension was placed in a Teflon cell, the electrodes were connected to a two-channel dielspektrometer, the device was turned on and the real part of the dielectric constant was determined on a scale

and on

The polarization coefficient (KP ₂ ) amounted to - 7.22.

The coefficient of resistance of red blood cells to hypoxia was calculated by the formula:

KU = KP ₂ : KP ₁ = 7.22: 6.17 = 1.17

The value of KU, equal to 1.17, is greater than the threshold value of 1.0, so the conclusion is made: the resistance of red blood cells to hypoxia is normal.

Example 2. Surveyed E., 52 years old, the diagnosis is healthy.

и на

Коэффициент поляризации (КП₁) составил - 4,07.Blood was taken from a cubital vein in a volume of 10.0 ml with the addition of 0.1 ml of heparin and after centrifugation for 10 min at a centrifuge speed of 4000 rpm, an erythrocyte suspension was obtained. Part of the erythrocyte suspension was placed in a Teflon cell, the electrodes were connected to a two-channel dielspektrometer, the device was turned on and the real part of the dielectric constant was determined on a scale

and on

The polarization coefficient (KP ₁ ) was - 4.07.

и на

Коэффициент поляризации (КП₂) составил - 5,60.After 60 minutes, the rest of the erythrocyte suspension was placed in a Teflon cell, the electrodes were connected to a two-channel dielspektrometer, the device was turned on and the real part of the dielectric constant was determined on a scale

and on

The polarization coefficient (KP ₂ ) amounted to 5.60.

KU = KP ₂ : KP ₁ = 5.60: 4.07 = 1.38

The KU value of 1.38 is greater than the threshold value of 1.0, so the conclusion is made: the resistance of red blood cells to hypoxia is normal.

Example 3. Patient C., 58 years old. Diagnosis: Coronary heart disease, unstable angina (progressive). Hypertension stage III. A persistent form of atrial fibrillation. Chronic heart failure III functional class.

и на

Коэффициент поляризации (КП₁) составил - 8,23.Blood was taken from a cubital vein in a volume of 10.0 ml with the addition of 0.1 ml of heparin and after centrifugation for 10 min at a centrifuge speed of 4000 rpm, an erythrocyte suspension was obtained. Part of the erythrocyte suspension was placed in a Teflon cell, the electrodes were connected to a two-channel dielspektrometer, the device was turned on and the real part of the dielectric constant was determined on a scale

and on

The polarization coefficient (KP ₁ ) was 8.23.

и на

Коэффициент поляризации (КП₂) составил - 7,28.After 60 minutes, the rest of the erythrocyte suspension was placed in a Teflon cell, the electrodes were connected to a two-channel dielspektrometer, the device was turned on and the real part of the dielectric constant was determined on a scale

and on

The polarization coefficient (KP ₂ ) was 7.28.

The coefficient of resistance of red blood cells to hypoxia was calculated by the formula:

KU = KP ₂ : KP ₁ = 7.28: 8.23 = 0.88

The KU value of 0.88 is less than the threshold value of 1.0, therefore, the conclusion is made: the resistance of red blood cells to hypoxia is impaired.

16 days after standard drug therapy, a red blood cell study of red blood cells for hypoxia was again conducted.

и на

Коэффициент поляризации (КП₁) составил - 7,36.Blood was taken from a cubital vein in a volume of 10.0 ml with the addition of 0.1 ml of heparin. It was centrifuged for 10 min at a centrifuge speed of 4000 rpm; an erythrocyte suspension was obtained. Part of it was placed in a Teflon cell, the electrodes were connected to a two-channel dielspektrometer, the device was turned on and the real part of the dielectric constant was determined on a scale

and on

The polarization coefficient (KP ₁ ) was 7.36.

и на

Коэффициент поляризации (КП₂) составил - 7,50.After 60 minutes, the rest of the erythrocyte suspension was placed in a Teflon cell, the electrodes were connected to a two-channel dielspektrometer, the device was turned on and the real part of the dielectric constant was determined on a scale

and on

The polarization coefficient (KP ₂ ) was 7.50.

The coefficient of resistance of red blood cells to hypoxia was calculated by the formula:

KU = KP ₂ : KP ₁ = 7.50: 7.36 = 1.02

The value of KU, equal to 1.02, is greater than the threshold value of 1.0, so the conclusion is made: the resistance of red blood cells to hypoxia is not violated.

The KU indices were compared in 14 young subjects, the average age was 18.64 ± 0.32 (first group), in 10 subjects, the average age was 42.00 ± 0.97 (second group), in 10 subjects, the average age - 54.50 ± 1.85 years (third group), and in 17 patients with coronary artery disease with unstable angina, the average age is 59.53 ± 1.68. The results are presented in tables 1, 2, 3 and 4.

Table 1
Indicators of the coefficient of resistance of red blood cells to hypoxia in healthy individuals of the first group No. Healthy KP ₁ KP ₂ KU one Surveyed K., 18 years old 6.17 7.22 1.17 2 Examined S., 18 years old 6.92 6.48 0.94 3 Surveyed D., 18 years old 6.62 6.41 0.97 four Surveyed S., 20 years old 5.14 8.40 1,63 5 Surveyed M., 19 years old 6.00 7.30 1.22 6 Surveyed A., 21 years old 6.36 7.81 1.23 7 Surveyed S., 20 years old 4.26 7.13 1,67 8 Surveyed K., 19 years old 6.28 7.90 1.26 9 Surveyed B., 19 years old 5.53 7.17 1.30 10 Surveyed D., 17 years old 5.53 7.08 1.28 eleven Surveyed G., 17 years old 6.59 7.81 1.19 12 Surveyed S., 17 years old 6.03 7.04 1.17 13 Surveyed Ch., 19 years old 4.31 7.77 1.80 fourteen Surveyed X., 19 years old 4.80 6.10 1.27 M ± m 18.64 ± 0.32 5.75 ± 0.23 7.24 ± 0.17 1.29 ± 0.07 table 2
Indicators of the coefficient of resistance of red blood cells to hypoxia in healthy individuals of the second group No. Healthy KP ₁ KP ₂ KU one Surveyed G., 45 years old 6.09 5.96 0.98 2 Surveyed G., 35 years old 2.54 3.92 1,54 3 Surveyed Sh., 45 years old 5.73 8.89 1.55 four Surveyed Ch., 41 years old 6.30 7.26 1.15 5 Surveyed B., 39 years old 7.00 8.65 1.24 6 Surveyed U., 44 years old 6.00 7.04 1.17 7 Surveyed S., 43 years old 5.70 4.67 0.82 8 Surveyed K., 42 years old 5.43 10.0 1.84 9 Surveyed K., 44 years old 3.65 7.64 2.09 10 Surveyed A., 42 years old 3.06 4.38 1.43 M ± m 42.00 ± 0.97 5.15 ± 0.48 6.84 ± 0.65 1.38 ± 0.12 Table 3
Indicators of the coefficient of resistance of red blood cells to hypoxia in healthy individuals of the third group No. Healthy KP ₁ KP ₂ KU one Surveyed M., 63 years old 6.76 6.35 0.94 2 Surveyed E., 52 years old 4.07 5.60 1.38 3 Surveyed M., 49 years old 3.16 5.16 1,63 four Surveyed M., 59 years old 4.44 5.29 1.19 5 Surveyed N., 50 years old 3,50 5.30 1.51 6 Surveyed S., 50 years old 6.13 7.29 1.19 7 Surveyed K., 48 years old 7.90 8.32 1.05 8 Surveyed M., 63 years old 5.52 6.35 1.15 9 Surveyed P., 59 years old 6.64 7.24 1.09 10 Surveyed E., 52 years old 4.09 5.62 1.37 M ± m 54.50 ± 1.85 5.22 ± 0.50 6.20 ± 0.35 1.25 ± 0.07 Table 4
Indicators of the coefficient of resistance of red blood cells to hypoxia in patients with coronary artery disease with unstable angina No. Sick KP ₁ KP ₂ KU one Patient D., 50 years old 7.54 6.68 0.89 2 Patient N., 59 years old 8.40 8.19 0.98 3 Patient 0., 50 years old 8.35 8.14 0.97 four Patient P., 49 years old 8.40 7.41 0.88 5 Patient V., 58 years old 8.23 7.28 0.88 6 Patient V., 60 years old 7.12 7.69 1,08 7 Patient N., 59 years old 7.68 7.86 1,02 8 Patient D., 64 years old 7.45 7.50 1.01 9 Patient B., 59 years old 9.05 6.96 0.77 10 Patient K., 62 years old 8.60 6.40 0.74 eleven Patient M., 63 years old 6.37 7.56 1.19 12 Patient P., 60 years old 6.34 6.31 1.00 13 Patient S., 49 years old 5.21 6.51 1.25 fourteen Patient E., 71 years old 7.86 7.24 0.92 fifteen Patient A., 62 years old 8.94 8.30 0.93 16 Patient P., 65 years old 9.00 8.47 0.94 17 Patient Ch., 72 years old 8.66 8.66 1.00 M ± m 59.53 ± 1.68 7.84 ± 0.26 7.48 ± 0.18 0.97 ± 0.03

In the first group, the average value of the index of red blood cell count for hypoxia was 1.29 ± 0.07, in the second - 1.38 ± 0.12, in the third - 1.25 ± 0.07, however, these indicators did not differ significantly ( p> 0.05), i.e. erythrocyte resistance to hypoxia is the same in all age groups of healthy individuals.

In patients with coronary heart disease, the red blood cell KU for hypoxia was 0.97 ± 0.03 and significantly differed (p <0.05) from healthy individuals.

By the method of statistical analysis, the optimal diagnostic threshold of KU of red blood cells for hypoxia was determined, equal to 1.0. The sensitivity of the method was 70.6%, specificity was 85.3%, the prognostic value of a positive result was 70.6%, and the prognostic value of a negative result was 85.3%.

The advantages of the proposed method are simplicity and high speed, low cost, as it eliminates the need for expensive equipment and reagents, treatment control.

Claims (1)

A method for determining the resistance of red blood cells to hypoxia using a blood test, characterized in that the patient is taken blood from a cubital vein in a volume of 10.0 ml, heparin 0.1 ml is used as an anticoagulant, centrifuged for 10 min at a speed of 4000 rpm , the obtained erythrocyte suspension is placed in a teflon cell mounted on a micrometer with oxidized tantalum electrodes, the cell electrodes are connected to a two-channel dielspektrometer, and on the dielspektrometer scale determine the real part of the diel ktricheskoy permeability

erythrocyte suspension at two frequencies -

and on

calculate the polarization coefficient (KP ₁ ) by the formula

after 60 minutes, the remaining erythrocyte suspension is placed in a teflon cell, the real part of the dielectric constant at two frequencies is determined -

and on

calculate KP ₂ according to the formula

then determine the coefficient of resistance (KU) of red blood cells to hypoxia of the subject according to the formula KU = KP ₂ : KP ₁ and with a KU value of 1.0 or less, a violation of the resistance of red blood cells to hypoxia is determined.