RU2585385C2 - Method for assessing tuberculoma activity - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: whole blood CD19+ percentage of total leukocyte count is measured by flow cytofluorometry. If the CD19+ cell count is 9 or more per cent, tuberculoma is considered to be inactive, whereas the value of less than 9 per cent shows active tuberculoma.

EFFECT: simplifying a procedure of tuberculoma activity assessment, providing more instant diagnosis.

5 ex

Description

The invention relates to medicine, namely to laboratory clinical diagnosis, and can be used to assess the activity of tuberculoma without invasive research methods (puncture, intraoperative biopsy), which subsequently affects the decision on the choice of tactics of patient management, in particular the use of surgical or conservative treatment method.

The tension of the epidemiological situation of tuberculosis is currently maintained, and, as a result, tuberculosis alertness is becoming increasingly important for the timely detection of tuberculosis (http://www.medicinform.net/tuberculos/bookytbbook4.htm). At the same time, clinical observations show that patients with tuberculoma often do not know about its existence. In half of them, changes in the lungs are detected only with a preventive x-ray examination. Some patients have mild symptoms of intoxication and changes in laboratory data. In particular, during the period of remission of the disease, the indicators are within normal values, and with an exacerbation of ESR it can increase to 20-25, less often 35-40 mm / h, leukocytosis is noted up to 8-11 × 10 ⁹ / l, a shift of the leukocyte formula to the left is observed . These changes are inconsistent and may be absent, despite the activity of tuberculomas. Mycobacterium tuberculosis is usually found in the sputum or lavage water of the bronchi usually during the breakdown of tuberculoma. The danger of the situation lies in the fact that tuberculoma constantly contains Mycobactérium tuberculosis, which are especially active in the areas of molten caseosis, which poses a threat to the health of both the patient and others (http://osop.ru/krovoobrashhenie/499-tuberkulema-legkix.html )

X-ray tuberculomas are divided into small (up to 2 cm), medium (up to 4 cm), large (up to 6 cm) and giant (more than 6 cm) (Tuberculosis in children and adolescents: a guide for doctors / E.N. Yanchenko. - Hippocrates, 1999 .-- 335 p.). There are no vessels in tuberculomas, and therefore antituberculosis drugs penetrate casease with great difficulty, which complicates the treatment (http://osop.ru/krovoobrashhenie/499-tuberkulema-legkix.html).

The potential activity of tuberculosis is usually not very dynamic, however, it should be borne in mind that the integrity of the capsule is quite easily violated in various physiological and pathological conditions at almost any time, even very distant from its formation (Kalabukha I.A. Pulmonary tuberculoma - see a surgeon / I. A. Kalabukha, V.E. Ivashchenko // Health of Ukraine. - 2010. - No. 6. - P. 40-41). The latter can lead to the progression of the pathological process, which is observed in approximately 10% of patients with prolonged stabilization. In this regard, the assessment of tuberculoma activity is important in practice. In addition, in the absence of the effect of antibacterial treatment for 4-6 months, patients, especially with large tuberculomas (more than 4 cm in diameter), are indicated for surgical treatment (http://vse-zabolevaniya.ru/bolezni-ftiziatrii/tuberkulema- leg; kih.html).

Thus, it is not enough to detect tuberculoma, it is necessary to establish the degree of its activity. Currently, a rough assessment of the activity of tuberculosis can be carried out, based on generally accepted research methods.

1. Clinical data. The age criterion is important, due to the high potential activity of tuberculosis more often in adolescence. There is also evidence of the important role of the state of the intrathoracic lymph nodes (http://www.blackpantera.ru/pediatrics/27943/). The disadvantage of these indicators is the inaccuracy of the criteria.

2. Radiographic studies. The activity of tuberculoma radiologically manifests itself in an increase in its size, the formation of a destruction site, new foci of bronchogenic dissemination (mainly in the lower parts of the lungs) and daughter tuberculomas. Often, a “pathway” can be identified that connects tuberculoma with the root of the lung due to perivascular and peribronchial infiltration (http://www.radiomed.ru/publications/tuberkulez-tuberkuloma-0, Tuberculosis in children and adolescents: a guide for doctors / E.N. Yanchenko. - Hippocrates, 1999.- 335 p., Http://pulmonolog.com/content/ klinicheskaya-kartina-i-techenie-tuberkulomy). The disadvantage of this method is the qualitative characteristics of the assessment and, as a consequence, subjectivity, the need to observe the patient in dynamics, radiation exposure to complete the study.

3. Computed tomography. Computed tomography reveals tuberculomas and the phases of their activity. The detection efficiency of tuberculosis in the progression phase by this method is 36.0 ± 9.6% (http://www.dissercat.com/content/kompyuternaya-tomografiya-organov-grudnoi-kletki-v-khirurgicheskom-lechenii-tuberkuleza-legk# ixzz31kyGDx8v). The disadvantage of this method is radiation exposure during the study, the low efficiency of detecting activity of tuberculosis.

4. Clinical diagnostic studies. It is described that with active tuberculoma in half of the patients there is a slight increase in the level of fibrinogen in the blood plasma, dysproteinemia increases with the progression of the disease (http://pulmonolog.com/content/klinicheskaya-kartina-i-techenie-tuberkulomy). The disadvantage of this method is the low information content of the method (50%), the absence of an exact value of the laboratory criterion, the need for dynamic observation (repeated research).

There is a method for determining the activity of tuberculous changes in the lungs (RF Patent No. 2161313), including a blood test, characterized in that the activity of leukocyte elastase and elastase-like activity are recorded in blood serum, and the content of cationic proteins is measured in plasma neutrophils, and with leukocyte elastase activity up to 3.1 Units / ml, elastase-like activity of blood serum up to 183 IU / ml and cationic protein content of not more than 7 conventional units determine inactive pulmonary tuberculosis, with higher values of these indicators or at least one of them determine active pulmonary tuberculosis. The disadvantage of this method is the need to simultaneously determine three laboratory parameters, the preparation of a large number of reagents (Tris buffer, phosphate buffer and others). In addition, there is no exact indication that the method can be used to assess the activity of tuberculoma.

There is also a laboratory method for determining the activity of the tuberculosis process (RF Patent No. 2238564), characterized in that mucin (3EG5, u / ml), immunoglobulin A (IgA, g / l), phagocytosis (%), lysozyme, ea are determined. , fibronectin, ng / ml, T-cells (abs), T-helpers (abs), T-suppressors (abs), B-cells (abs), T-killers (abs), two classification functions are calculated: f1 - value functions for healthy: f1 = 3EG5 0.056 + IgA8.72 + phagocytosis 2.48 + lysozyme 2.25 + fibronectin 0.246 + T-cells 32.1 + T-helpers 4.3 + T-suppressors 8.2 + B-cells 15.5 + T-killers 46.57 - 188.226; f2 - function value for patients: f2 = 3EG5 (-0.12) + IgA13.4 + phagocytosis 0.9 + lysozyme 1.025 + fibronectin 0.74 + T cells 39.15 + T-helpers 0.059 + T-suppressors 5 65 + B cells 29.76 + T-killers 29.25 - 371.09; and with f1 <f2, the activity of the tuberculosis process is established. The disadvantage of this method is the need to perform a large number of tests and the uncertainty of its use to assess the activity of tuberculoma.

5. Bacteriological studies. According to reports, a bacteriological study of sputum in patients with tuberculoma to assess its activity is uninformative. It allows only about half of active tuberculomas to be detected, and bacilli in sputum are occasionally found in tuberculomas that lose their activity or even completely lose it, which is later confirmed on the basis of histopathological findings (http://meduniver.com/Medical/Xirurgia/ 2504.html, http: //pulmonolog.com/content/klinicheskaya-kartina-i-techenie-tuberkulomy).

There is also evidence that inactive tuberculomas usually detect L-forms of mycobacteria, however, the accuracy of this prognosis is not very high (http://medkarta.com/?cat=article&id=26267).

There is a method for predicting the course of the tuberculosis process in the lungs by microbiological isolation of L-forms of tuberculosis mycobacteria from the sputum of patients, characterized in that the selected L-forms of the pathogen determine antilysocyme activity and, with the severity of this symptom equal to or greater than 4 μg / ml, exacerbation is predicted tuberculosis (Application for invention No. 96124291). The disadvantage of this method is the wide interpretation of the concept of "exacerbation of tuberculosis", as well as the need to work with a culture of Mycobactérium tuberculosis, which is dangerous for the health of the researcher.

In general, the disadvantages of bacteriological studies are low information content, the need to work with the culture of Mycobactérium tuberculosis, which unreasonably puts the researcher at risk, as well as significant material costs for conducting microbiological studies due to the fact that the work is carried out in compliance with SANPIN 1.3.2322-08 providing a whole range of conditions for conducting research (the presence and processing of rooms in the "clean" and "dirty" zones, ventilation, compliance with the "flow" of the procedures performed, the presence of a cabinet in biological protection, disposable material for working with biological material, etc.).

6. Skin test. A hyperergic reaction to tuberculin is often observed in newly diagnosed patients with a progressive course of pulmonary tuberculoma (with an inactive process, tuberculin tests are hypo- and normergic) (http://www.radiomed.ru/publications/tuberkulez-tuberkuloma-0). The disadvantage of this method is the low sensitivity of the method, low specificity (many cross-positive reactions, ranging from other diseases caused by the presence of Mycobactérium tuberculosis, to allergic reactions to food, livelihoods, drugs), the need for a double visit to a medical institution (for making a sample and for taking results), the introduction of restrictions for the patient for the period of observation of the reaction (you can not take a bath, go to the pool, etc.)

7. Assessment of activity based on clinical, radiological, and microbiological signs. Includes detection of decay, fresh infiltrative and focal shadows around tuberculoma, instability of x-ray shadows, intoxication and bacillus excretion. Each of these signs allows you to identify only a quarter of the active tuberculosis (http: //meduniver.corn/Medical/Xirurgia/2504.html). The disadvantage of this method is the “fuzzy” criteria, the need for the simultaneous use of several types of research (radiological, microbiological), the high cost of the study when involving all types of research, labor costs, the need to use a large number of personnel (phthisiatrist, radiologist, radiologist, laboratory assistant, Clinical laboratory diagnostics doctor, medical laboratory technician, nurses), duration (crops are performed for at least 7 days).

8. Closest to the proposed is a morphological study. It is known that the morphological analysis of tuberculoma based on the structural study of the capsule, caseosis and the state of the surrounding lung tissue allows the most reliable assessment of its activity (Kholodok O.A. Morphological aspects of the activity of pulmonary tuberculosis // Bulletin of physiology and respiratory pathology / O.A. Kholodok, M.I. Cheremkin. - 2013. - No. 49. - S. 51-54). However, the disadvantage of this method is its invasiveness. There is an increased risk of infection for the researcher (tuberculoma always contains Mycobactérium tuberculosis) and the potential for the spread of infection in the patient. Another significant drawback of the method is the need to use expensive additional equipment to ensure the safety of the procedure (disposable materials), its painlessness (painkillers with a possible risk of allergic reactions, as well as material costs for their acquisition and use) and material costs for the study itself (preliminary processing of biomaterial, preparation of slices, staining, visualization). The disadvantage is the duration of the study - it can not be performed earlier than after 3 days.

The task is to simplify the method for determining the activity of tuberculoma, reduce the invasiveness of manipulations, increase the speed of diagnosis by taking a small amount (3 ml) of venous blood once to determine the number of B-lymphocytes using flow cytometry.

To solve the problem when determining the activity of tuberculoma, 3 ml of blood is taken from a patient in a test tube containing EDTA as an anticoagulant, from the ulnar vein or vein of the rear of the hand. The determination of CD 19 + cells is carried out using a flow cytofluorimeter of any manufacturer and monoclonal antibodies against CD19 + cells. The content of the number of cells is expressed as a percentage of the number of all white blood cells. With a CD 19 + cell count of 9% or more, tuberculoma is considered inactive, and with an indicator value of less than 9%, tuberculoma is considered active. Based on the data obtained, a conclusion is drawn about the need to maintain or change the method of treatment of the patient.

Example 1. Patient K., 30 years old, IB No. 510. Ds: pulmonary tuberculoma, confirmed by the growth of culture, multidrug resistance, comorbidity - astigmatism. The diagnosis was first established in 1987, since that time the patient is under constant supervision. In 2000, when he was admitted to the hospital for the next examination, the number of CD 19 + cells was determined by flow cytofluorimetry, which amounted to 11.2%. Based on this, it was concluded that tuberculoma is inactive, a decision was made to continue conservative treatment. The patient was given a course of taking the following drugs: capreomycin, pyrazinamide, para-aminosalicylic acid, cycloserine, protionamide, levofloxacin. The patient was not subjected to surgical treatment. According to the results of the study, a morphological conclusion was obtained: conglomerate tuberculoma in the phase of subsiding activity with foci of deposition of lime salts in caseous masses and a moderate epithelioid-cell reaction, which confirmed the evidence that the patient was admitted to hospital that tuberculoma is inactive, and the correctness of the prescribed treatment .

Example 2. Patient P., 21 years old, IB No. 389, Ds: pulmonary tuberculoma, confirmed histologically, there is no concomitant pathology. The diagnosis was first established in 2010. Upon admission after 5 months, to determine the choice of treatment method by flow cytofluorimetry, the number of CD 19 + cells was determined, the number of which was 2.9%. Based on this, it was concluded that the tuberculoma was active, the patient was operated on 03/31/2011, the duration of treatment in the hospital was 40 days, the therapy had no complications, and the strain Mycobacterium tuberculosis was sensitive. The subsequent conclusion of the morphologist on the results of the study of surgical material at discharge from the hospital was as follows: a small homogeneous active tuberculoma with massive deposits of lime salts in caseous masses, which confirmed the correctness of the conclusion made when the patient was admitted to the clinic for treatment based on the proposed method.

Example 3. Patient C., 40 years old, IB No. 82354. Ds: cerebral tuberculoma, confirmed histologically, concomitant pathology - peptic ulcer of the duodenum in remission. The diagnosis was established in 2012. In 2013, upon admission to the hospital to address the issue of treatment tactics, the number of CD 19 + cells was determined by flow cytometry. Their number was 6.5%. Based on this, it was concluded that the tuberculoma is active, the patient was operated on 04/18/2013. The duration of treatment in the hospital was 29 days. The histological conclusion of the study confirmed the activity of tuberculoma. Active homogeneous tuberculoma with marginal decay of caseous masses was detected.

Example 4. Patient K., 30 years old, IB No. 85723. Ds: tuberculoma of the liver. At the time of detection of the disease (05/11/2011), the patient was 29 years old. For inpatient treatment, she was admitted 11 months after diagnosis, concomitant pathology - no. When determining the number of CD 19 + lymphocytes, 16.2% of them were detected, which is associated with inactive tuberculoma. A decision was made on the conservative tactics of treating the patient, which amounted to 58 days. In the future, according to the results of a biopsy study, the conclusion about the phase of tuberculoma activity was confirmed: homogeneous tuberculoma in the phase of subsiding activity with a small deposition of lime salts in caseous masses.

Example 5. Patient Ch., 50 years old, IS No. 81415. Ds: tuberculoma of the kidney. The diagnosis was first made at the age of 32. During the examination, it was found that the number of CD 19 + cells was 2.5%, a conclusion was made about the active phase of tuberculoma, a decision was made on surgical treatment of the patient. The length of hospital stay was 54 days. According to the results of a histological examination, it was subsequently confirmed that excised conglomerate tuberculoma with decay was in the active phase, contained multiple fibrous-caseous foci of various stages of activity.

The method was tested at the Federal State Budgetary Institution "Ural Research Institute of Phthisiopulmonology" of the Ministry of Health of Russia on 31 patients with a diagnosis of "tuberculoma", of which 15 were active, the rest did not. The predictive value of the proposed test or its predictive score is 83.9%. It was calculated by the authors according to the formula: 26 (correctly differentiated cases) * 100/31 (total cases) = 83.9%. To assess the possibility of using this test in clinical practice, the authors calculated standard statistical indicators, sensitivity and specificity, according to generally accepted formulas (http://pubhealth.spb.ru/E BM / Epid / EpidRusG04.htm). Calculated by the authors

the sensitivity was 13 * 100/15 = 86.7%,

specificity - 13 * 100/16 = 81.3%.

Thus, the application of the proposed method on the basis of determining the number of CD19 + cells of whole blood by flow cytofluorimetry allows us to accurately predict the activity of tuberculoma. Determining the activity of tuberculoma by the proposed method will allow, without much effort, quickly and safely for the patient to adjust the tactics of patient management, which in modern conditions reduces material costs for a medical institution to provide medical care. The proposed method is necessary for practical use in the work of clinical diagnostic laboratories and phthisiatric clinics. The method is highly informative, reproducible.

Claims (1)

A method for evaluating the activity of tuberculoma, including a blood test, characterized in that in whole blood the number of CD 19+ cells is determined as a percentage of the number of all leukocytes by flow cytofluorimetry, and if their number is less than 9 percent, they are evaluated as active, and at a value equal to or more than 9 percent, rate it as inactive.