RU2220423C1 - Method for predicting the flow of obstructive bronchitis in small children - Google Patents

Abstract

FIELD: medicine, pediatrics. SUBSTANCE: one should carry out the testing of biological liquid, moreover, in blood serum one should detect the values for dopamine, serotonin, cortisol and at higher dopamine value being 3,6±1,2 mcg/l (the norm being 1,2±0,7 mcg/l), decreased serotonin value up to 8,7±2,1 mcg/l (the norm being 19±2,3 mcg/l), increased cortisol values up to 1000±80,6 nM/l (the norm being 700±50,6 nM/l) it is possible to predict unfavorable result of obstructive bronchitis as transition into bronchial asthma phase. EFFECT: higher accuracy of prediction. 2 ex, 1 tbl

Description

 The invention relates to the field of health and will find application in pediatrics in diagnosing the nature of the course of obstructive bronchitis in young children. According to modern data, obstructive bronchitis occurs with a frequency of 75-200 per 1000 young children and in 50% of cases is the debut of bronchial asthma ("Modern problems of pulmonology in children." S. Yu. Kaganov, Pulmonology, 1992, 2, p. 6- 12).

 Bronchial asthma causes disability of children and adults, which leads to significant economic losses. The total cost of a disease in industrialized countries is 2% of all health care costs (WHO Materials "Bronchial asthma. Global strategy.", 1996). In this regard, the search for new methods for diagnosing and predicting the course of primary episodes of bronchial obstruction is of particular interest.

 Conducted research on the medical-scientific and patent literature revealed various methods for predicting the course of airway obstruction in young children.

 So, in the work of Sobkh Munisef Sobkhi "The clinical significance of the proteinase-inhibitory system in children of the first three years of life with recurrent obstructive bronchitis and bronchial asthma" (abstract of the dissertation of the candidate of medical sciences, Moscow, 1993), a method for assessing the nature of the course of obstructive bronchitis by the method of studying the activity of proteolytic enzymes (peptide hydrolases) of bronchial contents and their inhibitors (a 1-inhibitors).

 The disadvantage of this method is its high invasiveness, namely the use of bronchoscopic examination, accompanied by the use of anesthesia, mechanical trauma to the respiratory tract of the child.

 The well-known "Method for predicting the recurrence rate of infectious diseases of the respiratory tract and the persistence of recovery in preschool children" (RF patent 2169923, G 01 N 33/53, 33/48, BI 18, 2001) by determining the presence of conditionally pathogenic microflora in feces, leukocyte blood formulas, Ig A, M, G levels, the detection of a correlation with a correlation strength of less than 0.5 determine the strength of the immune system, and an unfavorable variant of the course of the disease is predicted. The disadvantage of this method is its low information content, a reflection of the reaction of only the immune system, which does not allow to predict the course of the disease.

 The well-known "Method for the differential diagnosis of bronchial asthma and obliterating bronchiolitis in children" (RF patent 2159935, G 01 N 33/52, 33/483, 33/68, BI 33, 2000). This method is taken as a prototype.

 The method consists in assessing the intensity of fluorescence of the fluid of bronchoalveolar lavage. Moreover, the intrinsic fluorescence intensity of intracellular pyridine nucleotides is measured at an excitation wavelength of 356 nm and a registration wavelength of 420-450 nm and flavproteids at an excitation wavelength of 440-480 nm and a registration length of 510-550 nm, the ratio of intrinsic fluorescence of intracellular pyridine nucleotides, and flavine protein is determined Relationships of more than 1 are diagnosed with bronchial asthma, and with a relationship of less than 1, obliterating bronchiolitis. The disadvantage of this method is its lack of accuracy in predicting the course of bronchial obstructive syndrome in combination with morbidity (obtaining fluid bronchoalveolar lavage by bronchoscopy.

 The aim of the invention is to improve the accuracy of predicting the course of obstructive bronchitis in young children, which will prevent the development of bronchial asthma.

 This goal is achieved by the study of blood serum, in which the content of dopamine, serotonin and cortisol is determined, and with the following indicators: an increase in the concentration of dopamine to 3.6 = 1.2 μg / l compared with the age norm 1.2 ± 0.7 μg / l , a decrease in the concentration of serotonin to 8.7 ± 2.1 μg / L compared with the age norm of 19 ± 2.3 μg / L and an increase in cortisol to 1000 ± 80.6 nmol / L compared with the norm of 700 ± 50.6 nmol / L predict an unfavorable outcome of the disease.

The proposed method is as follows:
The determination of dopamine is carried out by the fluorometric method (B. M. Kogan, I. V. Nechaev, J. "Laboratory" 1, 1979). 1 ml of whole blood is shaken for 5 min with 5 ml of N-butanol.

 Then the sample is centrifuged for 10 minutes at 3000 rpm. The organic phase is transferred to a test tube containing 0.4 ml of 0.1% cysteine hydrochloride prepared in a 0.1 N HC1 solution. 6 ml of heptane are poured into the same tube. The sample is shaken for 3 minutes, then centrifuged for 5 minutes at 2000 rpm. The organic phase is discarded. Then, 0.4 ml of the inorganic phase are taken into 2 tubes (control and experimental). 0.4 ml of 0.1 mol of EDTA sodium acetate buffer, pH 6.9, is also poured here. In a control tube, 50 μl of a 0.2 mol solution of iodine prepared with absolute alcohol is added.

 0.2 ml of 0.1 N alkaline sulfite prepared in a 5 N NaOH solution is added to the test tube. After 2 minutes, an alkaline sulfite solution is poured into the test tube, and iodine into the control solution. After 3 minutes, 0.4 ml of a 5 N solution of acetic acid is poured into both tubes. All samples are placed in a boiling bath for 6 minutes. It is then cooled on ice and the fluorescence readings are taken on a Hittachi 3000 spectrofluometer. For dopamine, the excitation wavelength is 320 nm and the absorption wavelength is 375 nm.

 The determination of serotonin is also carried out by a fluorometric method. To determine the content of serotonin, 0.4 ml of the inorganic phase is taken into a test tube, 0.1 ml of 0.1 N orthophthalic aldehyde prepared on methanol is added. Then, 6 ml of a 10 N HC1 solution is added to the sample. The sample is placed in a boiling bath for 10 minutes. Then, the fluorescence intensity is also recorded on a Hittachi spectrofluometer with respect to a control sample. For serotonin, the excitation wavelength is 357 nm, and the absorption wavelength is 470 nm.

To calculate the concentration of serotonin and dopamine, the formula is used, where the intensity of a standard concentration solution of 0.001 μg / l is taken as the unit of fluorescence:
0.01x1000x1 ml of fluorescence of an experimental sample
0.4 ml (volume of oxidizing agents) x standard fluorescence index.

 Determination of cortisol is carried out by enzyme immunoassay using standard kits for the determination of cortisol in serum. To do this, 50 μl of blood is poured into the cell of the plate and incubated in an incubator for 30 minutes with constant shaking. Then the serum was taken, the cell plate was washed three times with phosphate buffer and 50 μl of conjugate was added. 15 minutes after the development of staining in the dark, 20 μl of stabilizer (concentrated HCl) was added. The staining intensity of the sample is measured on a vertical photometer at a wavelength of 460 nm. Calculation of the concentration of cortisol is carried out according to the calibration curve attached to the device.

Clinical examples
1. Patient Golda M., 11 months old, was admitted to Children's Hospital 2 with a diagnosis of structural bronchitis, respiratory failure 2. The complex of clinical and laboratory studies included the evaluation of indicators by the proposed method. Revealed: the content of serotonin 6.52 μg / l, dopamine 3.75 μg / l, cortisol 1100 nmol / l, which indicated an unfavorable prognosis of the course of the disease. Further observation of the patient for two years revealed the manifestation of bronchial obstructive syndrome, namely the formation of bronchial asthma.

 2. Patient L. Chinoian, 8 months old, was admitted to Children's Hospital 2 with a diagnosis of obstructive bronchitis, respiratory failure 2. The child underwent a comprehensive clinical and laboratory study according to the proposed method. Revealed: the content of serotonin 20.7 μg / l, dopamine 1.3 μg / l, cortisol 670 nmol / l. A favorable variant of the course of the disease was predicted, which was subsequently confirmed in the process of dynamic observation for two years.

 Our proposed method was tested on large clinical material (238 patients). The data are presented in the table.

 The proposed method in comparison with the prototype allows more accurately predict the adverse outcome of obstructive bronchitis - the transition to bronchial asthma.

Claims (1)

A method for predicting the course of obstructive bronchitis in young children by the method of studying a biological fluid, characterized in that dopamine, serotonin, cortisol are determined in the blood serum and with an increase in dopamine to 3.6 ± 1.2 μg / l (norm 1.2 ± 0 , 7 μg / L), a decrease in serotonin to 8.7 ± 2.1 μg / L (normal 19 ± 2.3 μg / L), an increase in cortisol to 1000 ± 80.6 nmol / L (normal 700 ± 50.6 nmol / l) predict an unfavorable outcome of obstructive bronchitis - a transition to bronchial asthma.

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