Method of predicting beginning of chronic obstructive lung disease

FIELD: medicine.

SUBSTANCE: in order to realise method of chronic obstructive lung disease prediction content of CD3-CD16+, CD4+CD25+, CD95+, CD45RA+CD4+/CD45RO+CD4+ in blood is defined by method of flow cytofluorometry, Interleukin-6 (IL-6) in blood serum on analyser. If values of CD3-CD16+ content are over 0.18*103, CD4+CD25+ over 0.12*103, CD95+ over 0.03*103, CD45RA+CD4+/CD45RO+CD4+ less than 1.3, IL-6 over 2.4 Pkg/ml beginning of chronic obstructive lung disease is predicted.

EFFECT: method allows to increase accuracy of prediction of disease beginning, is safe, technically realisable.

2 ex

 

The invention relates to medicine, namely to pulmonology, and can be used to predict the onset of chronic obstructive pulmonary disease (COPD).

COPD is one of the most important causes of ill health and death worldwide. Over the years many people suffer from COPD, prematurely dying from it or from its complications. COPD is the fourth cause of death in the world, and, as predicted in the coming decades will see an increase in the incidence and mortality from it.

COPD is a disease characterized by irreversible obstruction of the Airways. Restricting air flow rate is progressive and associated with abnormal inflammatory response of the lungs to pathogenic particles or gases. (Global strategy for diagnosis, treatment and prevention of chronic obstructive pulmonary disease. Revision 2007)

A known method for predicting the development of chronic obstructive pulmonary disease in persons who do not have functional symptoms of bronchial obstruction (RF patent No. 2303956, 2007), including the identification of the examined functional spirometric indices. As well as the identification of the life history of certain clinical symptoms and risk factors: signs of dyspnea, passive Smoking, productive cough, wheezing, availability is pistov cough for over 3 weeks the floor, snoring. Installed features assign grades and numeric values, on the basis of which determine the prognostic factors. Then numerical characteristics between a compare and assess the risk of developing COPD.

The prototype of the invention is a method for predicting the development of chronic obstructive pulmonary disease among smokers entities (patent RF №2303957, 2007). The examined determine the functional spirometric indices, with no signs of bronchial obstruction, determine clinical symptoms and risk factors of the history of life: the experience of Smoking, number of cigarettes smoked per day, the presence of dyspnea or wheezing, the degree of intensity of Smoking and other indicators. Installed features assign grades and numeric values, on the basis of which determine the prognostic factors. Then numerical characteristics between a compare and assess the risk of developing COPD.

It is known that Smoking leads to changes in both its structure and functioning of the Central and peripheral Airways, alveoli and capillaries, as well as the immune system of the lungs. Caused by Smoking changes in the epithelium of the respiratory tract include loss of cilia, hypertrophy of mucous glands, increased number of goblet cells and increased permeability, l is to carry through the development of respiratory symptoms.

The mechanism of occurrence of obstructive lung disease due to Smoking is that Smoking causes an imbalance of proteases and antiproteolytic lung function, possibly manifested destruction of the parenchyma and obstruction of the respiratory tract (Chuchalin A.G. Chronic obstructive pulmonary disease, Moscow, 2000. - S-365).

The objective of the invention is to develop a method for predicting the development of chronic obstructive pulmonary disease on immunological parameters.

The technical result - getting pathogenetic markers-criteria for the development of chronic obstructive pulmonary disease, improving the accuracy of the method.

The proposed method is as follows: in the blood taken on an empty stomach from Vienna, determine the content of CD3-CD16+, CD4+CD25+, CD95+, CD45RA+CD4+/CD45RO+CD4+, Interleukin-6 (IL-6). The study of populations and subpopulations of lymphocytes was carried out using flow cytofluorimetry using monoclonal antibodies labeled with different fluorochromes (Karpischenko A.I. Medical laboratory technology. - St. Petersburg: Intermedica, 2002).

Cell staining performed according to the standard technique (Lugovskaya S.A., the Postman M.E. Ter-Minassian, Tupitsyn NN. Immunophenotyping in the diagnosis of hematological malignancies. - Tver: Triad, 2005; haidouks SV, Shurochka A.V. Questions modern protoc the ow cytometry. Clinical application. - Chelyabinsk, CGMA, 2008).

The procedure is performed in a standard disposable polypropylene tubes for flow cytofluorimetry. At the bottom of each test tube add 5 ál of monoclonal antibodies, add 100 ál of well-mixed sample (whole blood), mix. Incubated for 20 min in the dark at room temperature (20-25°C). After incubation for lysis of red blood cells in test tubes make 1 ml of the working lytic reagent Cal-lyse (Caltag, Austria), mixed and incubated for 10 min at room temperature in the dark. The tubes centrifuged for 5 min in mode 1500 rpm Remove supernatant, add 2 ml of Cell-WASH to launder sample from the destroyed cells. Carefully resuspended precipitate and centrifuged for 5 min in mode 1500 rpm Remove supernatant, the precipitated cells resuspended in 0.5 ml Cell-WASH. The samples analyzed by collecting at least 10 LLC events in the flowing cytofluorimetry using research program CellQuest. The results are expressed in percentage of cells bearing the antigen. Given the evidence of the number of leukocytes and the leukocyte composition formula obtained previously on the Hematology analyzer, calculate the absolute number of cells bearing a particular antigen. The level of IL-6 in serum were determined met the house enzyme immunoassay analyzer ACCESS 2 (Beckman Coulter).

When values of the content of CD3-CD16+above 0,18·103, CD4+CD25+ above 0,12·103, CD95+ above 0,03·103, CD45RA+CD4+/CD45RO+CD4+ below 1.3, IL-6 above 2.4 PCG/ml predict the development of COPD.

Produced a study in the blood of healthy people (control group - 30 people) and patients with chronic obstructive pulmonary disease (30 persons), taken on an empty stomach from Vienna, determined the content of cells, the absolute number of lymphocytes (ACHL), neutrophils (young, stab, segmented), lymphocytes, monocytes, eosinophils, basophils, circulating immune complexes (CIC), interleukin-6 (IL-6), subpopulation structure of peripheral blood lymphocytes (CD3+CD16-, CD3+CD4+, CD3+CD8+, CD4+/CD8+, CD19+, CD3-CD16+, CD3+CD16+, CD3+CD8+, CD4+CD25+, CD25+, CD4+95+, CD95+, CD4+HLADR+, HLADR+, CD45RO+CD4+, CD45RA+CD4+, CD45RA+CD4+/CD45RO+CD4+, CD19+CD23+), we investigated the functional activity of lymphocytes (spontaneous proliferation, response to PWM, the answer to Konvalin And (Sopa), response to phytohemagglutinin (PHA)), apoptosis of lymphocytes, phagocytosis, metabolic activity in the test recovery narasinga of tetrazole (NBT-test), the level of serum immunoglobulins.

Revealed a significant (p<0,05) differences between the control group and patients with chronic obstructive pulmonary disease according to the following criteria: CD3-CD16+, CD4+CD25+, CD95+, CD45RA+CD4+/CD45RO+CD4+and IL-6.

The results serve as a basis for recommendations measurement level is I CD3-CD16+, CD4+CD25+, CD95+, CD45RA+CD4+/CD45RO+CD4+and IL-6 in healthy individuals in the presence of the risk of COPD and values: CD3-CD16+ above 0,18·103, CD4+CD25+ above 0,12·103, CD95+ above 0,03·103, CD45RA+CD4+/CD45RO+CD4+ below 1.3, IL-6 above 2.4 PCG/ml to predict the occurrence of COPD.

The proposed method is illustrated by the following examples.

Example 1.

25.02.2004 to the district physician turned patient A., 35 years old complaining of a persistent cough with little sputum.

Objectively: skin and visible mucous physiological color.

Thorax correct form.

Palpation intercostal space expanded, chest excursion limited, voice shaky and weak. Percussion of the chest and lung sounds, borders lungs expanded.

Auscultation: the weakening of respiratory noise, scattered, single dry rales. Respiratory rate 20 per minute.

Cardiovascular system: heart sounds are muffled. Pulse and heart rate of 80 per minute.

Liver at the edge of the costal arch, megaelosia sensitive to palpation, smooth.

Laboratory data: CD3-CD16+1.5·103, CD4+CD25+-0,32·103, CD95+-0,2·103, CD45RA+CD4+/CD45RO+CD4+0,14, IL-6 - 2.43 PCG/ml Fluorography OGK: Chronic bronchitis.

Diagnosis: Chronic non-obstructive bronchitis. On the basis of changes of immunological indicators is her we predict the occurrence of chronic obstructive pulmonary disease in the future.

After 5 years this patient comes to our clinic complaining of persistent cough with yellow sputum, increased shortness of breath with little exertion, increased body temperature.

Objectively: skin and visible mucous cyanotic.

The jugular veins. The chest is barrel-shaped.

Palpation intercostal space expanded, chest excursion limited, voice shaky and weak. Percussion of the chest - sound box, the border of the lungs are expanded.

Auscultation: the weakening of respiratory noise, mixed dry rales. Respiratory rate is 26 per minute.

Cardiovascular system: heart sounds are muffled, accent 2 tone of the pulmonary trunk. Pulse and heart rate is 84 per minute.

Liver at the edge of the costal arch, megaelosia sensitive to palpation, smooth.

Laboratory data: CD3-CD16+2.85·103, CD4+CD25+-0,42·103, CD95+-0,4·103, CD45RA+CD4+/CD45RO+CD4+-1,14, IL-6 - 2.46 PCG/ml

Radiography OGK: Chronic obstructive process. Emphysema of the lungs.

Spirography: Significant decrease in VC. Moderate obstruction distal bronchi. FEV1 - 55,44%, VC - 45%, FVC - 61%. A significant disruption in the status of the valve is AI.

FBC: Subtropically endobronchitis.

Diagnosis: Chronic obstructive pulmonary disease, moderate, acute stage.

Our prediction was confirmed

Example 2. 12.10.2002 in the clinic asked the Patient P., 45 years old complaining of a persistent cough, more in the morning, with a small amount of sputum.

Objectively: the skin is pale pink color.

Relative to the horizontal direction of the ribs, barrel-shaped rib cage.

The number of respiratory movements alone 20 per minute.

Palpation of the chest - resistance, expansion of the intercostal spaces. Percussion of the lung - sound box, voice shaky and weak.

Auscultation: the weakening of the respiratory noise, dry wheezing during forced exhalation.

Heart tones are rhythmic, muffled.

Palpation of abdominal organs: liver stands out from the edge of a costal arch on 1.5 cm, sensitive, with a smooth surface.

Laboratory data: CD3-CD16+to 1.4·103, CD4+CD25+-0,27·103, CD95+-0,29·103, CD45RA+CD4+/CD45RO+CD4+0,2, IL-6 - 2.51 PCG/ml).

Radiography: Chronic bronchitis.

Spirography FEV1 87%, VC 90%.

The experience of Smoking for 20 years.

Diagnosis: Chronic non-obstructive bronchitis.

We predict the appearance of the patient after a few years COPD.

In 2009, the same will patientportal to the medical Department with complaints of cough with white sputum, shortness of breath with little exertion, feeling of stuffiness in the chest.

1 month ago, increased cough, increased sputum, increased shortness of breath (with little exercise), General weakness, feeling of tightness in the chest that occurs during physical activity.

Objective: cyanotic staining mucous membranes.

Relative to the horizontal direction of the ribs, barrel-shaped rib cage.

The number of respiratory movements alone 22 min.

Swelling up to the upper third of the tibia (the sign of right ventricular failure).

Palpation of the chest - resistance, expansion of the intercostal spaces, uniform but limited participation in the act of breathing. Percussion of the lung - sound box, voice shaky and weak.

Auscultation: the weakening of the respiratory noise, dry wheezing during quiet breathing (remote wheezing).

Heart tones are weakened.

Palpation of abdominal organs: liver stands out from the edge of a costal arch on 1.5 cm, sensitive, with a smooth surface.

Laboratory data: CD3-CD16+2.36·103, CD4+CD25+-0,57·103, CD95+-0,79·103, CD45RA+CD4+/CD45RO+CD4+-1,2, IL-6 - 2.51 PCG/ml).

X-ray: pulmonary pattern is reinforced deformed. The fibrous roots of the lungs modified tyazhistye. Aperture is limited in podnizhnecheljustnye: Chronic bronchitis. Pneumosclerosis.

Spirography - extremely sharp decline in VC, abrupt disturbance of the airway. VC - 50%, FVC - 55%, FEV1 OF 50%.

Diagnosis: Chronic obstructive pulmonary disease severe stage, the stage of exacerbation.

Our prediction was confirmed.

A method for predicting the onset of chronic obstructive pulmonary disease, characterized in that in the blood determines the content of CD3-CD16+, CD4+CD25+, CD95+, CD45RA+CD4+/CD45RO+CD4+ method flow cytofluorimetry, Interleukin-6 (IL-6) in blood serum analyzer and the values of the content of CD3-CD16+ above 0,18·103, CD4+CD25+ above 0,12·103, CD95+ above 0,03·103, CD45RA+CD4+/CD45RO+CD4+ below 1.3, IL-6 above 2.4 PCG/ml predict occurrence of the disease.



 

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