Differential diagnostic technique for degree of rheumatoid arthritis activity

FIELD: medicine.

SUBSTANCE: patient's peripheral blood is examined for the content of cytotoxic lymphocytes, %: (CD8/CD16), (CP8/GranzymeB), (CP16/GranzymeB), (CDS), (CD16/CD56) and (CD8/HLA DR). It is conmbined with determining cytotoxic activity (CTA), %. It is followed by calculating a RA activity index (AI) by formula: If observing the condition 11.8%<AI≤18.2%, a moderate degree of rheumatoid arthritis activity is diagnosed. If observing the condition AI>18.2%, a high degree of rheumatoid arthritis activity is diagnosed.

EFFECT: using the technique enables high-reliability differential diagnosis of the degree of rheumatoid arthritis activity.

2 ex

 

The invention relates to medicine, namely to rheumatology, and can be used for differential diagnosis of the degree of activity of rheumatoid arthritis, namely medium and high RA activity.

Rheumatoid arthritis (RA) is the most common chronic inflammatory disease that causes early to violate the congruence of the joints due to the erosive process, formation of pannus, hyperproductive synovial fluid, inflammation of the muscular, that already in the first two years of the disease leads to the violation, and then to the loss of function of the joints with subsequent disability of the patient (Ardis W., Folomeev O.M. Rheumatic diseases and disabilities of the adult population of the Russian Federation // Scientific and practical rheumatology, 2007, No. 4, p.4).

There are medium and high activity RA (Karataev CU, Olyunin Y.A. ON the classification of rheumatoid arthritis // Scientific and practical rheumatology, 2008, No. 1, p.13).

Today there are many ways the differential diagnosis of the degree of activity of RA. However, due to the fact that existing methods do not take into account the immunopathogenesis of RA, and also because not enough high accuracy differential diagnosis of RA, they do not meet the requirements of modern medicine (IBIS styles Gdynia Reda MSM. Comorbid conditions in patient with rheumatic diseases: an update. Curr Opin Rheumatol. - 2004. Vol. No. 16. - P.109-113).

Thus, development of new methods of differential diagnostics of the degree of activity of rheumatoid arthritis is an actual problem of modern rheumatology (Nasonov EL Rheumatoid arthritis as a medical problem // Therapeutic archive, 2004, №5, p.5-7).

Conducted research on the medical-scientific and patent literature found various methods of differential diagnostics of the degree of activity of rheumatoid arthritis.

There is a method of differential diagnosis of the degree of activity of RA using the index the index DAS28 (Van Riel P.L., van Gestel A.M. Clinical outcome measures in rheumatoid arthritis. - 2000. Vol. No. 59. - P.28-31).

In the calculation of the DAS28 index take into account the number of swollen and tender joints out of 28 to be assessed, the General condition of the patient and ESR (Provo ML, van't Hof MA, Kipper HH, van Leuven MA, van de Putte LB, van Riel PL. Modified disease activity scores that include twenty-eight-joint counts. Development and validation in a prospective longitudinal study of patients with rheumatoid arthritis. Arthritis Rheum. - 1995. Vol. No. 56. -P.44-48). When the index value of DAS28 in the range of 3.2 to 5.1 diagnose average degree of activity of RA, when the value of DAS28>5,1 diagnosed with a high degree of activity of PA (Prevoo ML, van't Hof MA, Kuper HH, van Leeuwen MA, van de Putte LB, van Riel PL. Modified disease activity scores that include twenty-eight-joint counts. Development and validation in a prospective longitudinal study of patients with rheumatoid arthritis. Arthritis Rheum. - 1995. Vol. No. 38. - P.44-48).

The disadvantage of this method is the tsya his lack of credibility, as the degree of activity of the RA does not always correlate directly with anatomical changes in the joints that are used in the calculation of the DAS28 index.

RF patent №2222259 (2004 BIPM No. 3), secure "Method of assessing the activity of rheumatoid arthritis". Ultrasonic studies assess synovial effusion and proliferation of the synovial membrane. When the bag acquiring an oval shape, a moderate quantity of synovial effusion heterogeneous echostructure due to the presence of fine suspended solids in combination with fuzzy contours of the synovial membrane with signs of focal proliferative changes diagnosed with moderate activity. When the bag acquiring a rounded shape, a large number of synovial effusion heterogeneous echostructure in the form of a hyperechogenic coarse flocculent suspensions in combination with irregular thickening of the synovial membrane and the presence of focal or diffuse proliferative changes in the form of a hyperechogenic structures with fuzzy blurred contours or irregular growth diagnosed with a high degree of activity of RA.

The disadvantage of this method is the lack of accuracy related to the fact that RA can occur without lesions of the knee joint, and considering the systemic disease is, not accurate enough to carry out differential diagnostics of the degree of activity of RA only one joint.

The closest technical solution adopted for the prototype, is protected by the RF patent №2184967 (2002, BIPM No. 19) "Method of evaluating the degree of activity of RA", which consists in the study of peripheral blood of the patient. Register viscosity characteristics collapsing the blood of the period of reaction r, the constant of thrombin k, the maximum amplitude AM, the time of formation perintalmanna structure of the bunch, T, the total rate of retraction and spontaneous lysis of the clot F. Additionally determine the intensity of spontaneous platelet aggregation AG and the rate of activity of thrombin Kk angiography in the range of 3±0.24 Rel. units, r in the range of 3.8±0,17 min, AM in the interval 752±17.2 Rel. unit T in the interval of 63.7±2,05 min, F in the range of 10.6±0,92 Rel. unit, in the absence of statistically significant changes in k and Kk determine the average degree of activity of rheumatoid arthritis, when AG is in the range of 3.1±0,28 Rel. units, r in the range of 2.9±0,10 min, Kk in the interval 32,4±0,62 Rel. unit k in the range of 3.8±0,13 min, AM in the interval 844±19,4 Rel. unit T in the interval 47,7±1,12 min, in the absence of a statistically significant change F determines a high degree of activity of rheumatoid arthritis.

The disadvantages of the prototype is the lack of reliability differentialdiagnosis RA, related to the fact that the parameters of the blood coagulation system may change due to concomitant diseases, such as vasculitis, antiphospholipid syndrome, hepatitis, lymphoproliferative diseases.

The aim of the invention is to improve the accuracy of differential diagnosis of the degree of activity of rheumatoid arthritis.

This goal is achieved by the fact that examine the peripheral blood of the patient, which determine the content of cytotoxic lymphocytes, %: (CD8/CD16), CD8 (t/Grazing), (CD16/Grazing), CD8 (t), (CD16/CD56) and (CD8/HLA-DR). Also determine their cytotoxic activity (CTA), %. Then calculate the index of the degree of activity of RA (IA) according to the formula:

If the condition of 11.8%<IA≤18,2%, diagnose average degree of activity of rheumatoid arthritis. If the condition IA>18,2% diagnosed high degree of activity of rheumatoid arthritis.

Detailed description of the method.

The patient fasting taking venous blood volume of 5 ml Lymphocytes secrete on the density gradient urografin-ficoll (Kheifets LB separation of the formed elements of human blood in the density gradient urografin-ficoll // Laboratory business, 1973, No. 10, page S-581).

Determine the content of cytotoxic lymphocytes (CD8/CD16), CD8 (t/Grazing), (CD16/Grazing), CD8 (t), (CD16/CD56) and (CD8/HLADR) using the indirect immunofluorescence reaction, for example, the flowing laser cytofluorimetry "FC 500" (Becman Coulter, USA) using the mouse monoclonal antibodies provided by the firm CHM "Sorbent" Institute of immunology MZ the Russian Federation. Cyto-toxic activity (CTA) is determined using target cells: leukemia K562 cells sensitive to lysis by natural killer (Pinegin B.V., Yarylo A.A., A. Simonov et al. Application of flow cytometry to assess the functional activity of the human immune system. / Manual for laboratory doctors. - Moscow: Federal state unitary enterprise "Medservice", - 2001. P.48-53).

For diagnosing the degree of activity of rheumatoid arthritis activity index IA (IA), is calculated by the formula:

If the condition of 11.8%<IA≤18,2%, diagnose average degree of activity of rheumatoid arthritis. If the condition IA>18,2% diagnosed high degree of activity of RA.

The practical implementation of the proposed method is illustrated by examples from clinical practice.

Example 1: patient N. 38 years old, medical history, No. 261, was admitted to the cardiological Department of the Rostov state medical University, Rostov state medical University) for specification of the diagnosis due to the ineffectiveness of therapy. The patient survey was conducted, according to the claimed method.

The pain is Oh prandial implemented a sampling of venous blood in a volume of 5 ml. Lymphocytes were isolated by density gradient urografin-ficoll (Kheifets LB separation of the formed elements of human blood in the density gradient urografin-ficoll // Laboratory business, 1973, No. 10, page S-581).

The contents of cytotoxic lymphocytes (CD8/CD16), CD8 (t/Grazing), (CD16/Grazing), CD8 (t), (CD16/CD56) and (CD8/HLA-DR) were determined using the indirect immunofluorescence reaction on flow laser cytofluorimetry "FC 500" (Becman Coulter, USA) using the mouse monoclonal antibodies provided by the firm CHM "Sorbent" Institute of immunology MZ the Russian Federation. Cytotoxic activity was determined using target cells: leukemia K562 cells sensitive to lysis by natural killer (Pinegin B.V., Yarylo A.A., A. Simonov et al. Application of flow cytometry to assess the functional activity of the human immune system. / Manual for laboratory doctors. - Moscow: Federal state unitary enterprise "Medservice", - 2001. P.48-53).

Results were obtained: (CD8/CD16)=23.1 per cent, (CD8/Granting)=7%, (CD16/Granting)=3,53%, (CTA)=14,1%, CD8 (t)=24%, (CD16/CD56)=2,27%, CD8/HLA-DR)=5,17%.

Determined the activity index RA patients, substituting the obtained results into the formula:

It was obtained the value of IA=28.57 percent. As it was made a condition IA=to 28.57>18,2 (%), the patient was diagnosed with a high degree of activity of RA. Hurt who was appointed as an adequate therapy, the results of which confirmed the validity of the claimed method of differential diagnostics of the degree of activity of RA, and an adequate therapy, the effectiveness of which has proved a reliable method of differential diagnosis of the degree of activity of RA.

Example 2: patient M 29 years old, medical history, No. 287, was admitted to the cardiological Department of the Rostov state medical University, Rostov state medical University) for specification of the diagnosis due to the ineffectiveness of therapy. The patient survey was conducted, according to the claimed method.

The patient fasting has carried out the sampling of venous blood in a volume of 5 ml Lymphocytes were isolated by density gradient urografin-ficoll (Kheifets LB separation of the formed elements of human blood in the density gradient urografin-ficoll // Laboratory business, 1973, No. 10, page S-581).

The contents of cytotoxic lymphocytes (CD8/CD16), CD8 (t/Grazing), (CD16/Grazing), CD8 (t), (CD16/CD56) and (CD8/HLA-DR) were determined using the indirect immunofluorescence reaction on flow laser cytofluorimetry "FC 500" (Becman Coulter, USA) using the mouse monoclonal antibodies provided by the firm CHM "Sorbent" Institute of immunology MZ the Russian Federation. Cytotoxic activity was determined using target cells: leukemia K562 cells, the feeling is sustained fashion to lysis by natural killer (Pinegin B.V., The Yarylo A.A., A. Simonov et al. Application of flow cytometry to assess the functional activity of the human immune system/ Manual for laboratory doctors. - Moscow: Federal state unitary enterprise "Medservice", - 2001. P.48-53).

Results were obtained: (CD8/CD16)=19,8%, CD8/Granting)=6,9%, (CD16/Granting)=1,159%, (CTA)=13,7%, CD8 (t)=21%, (CD16/CD56)=1,47%, CD8/HLA-DR)=5,02%.

Determined the activity index RA patients, substituting the obtained results into the formula:

It was obtained the value of IA=13,99%. As it was made a condition that 11,8%<IA=13,99%≤18,2%, the patient was diagnosed with a moderate degree of activity of rheumatoid arthritis. The patient was conducted adequate therapy, the results of which confirmed the validity of the claimed method of differential diagnostics of the degree of activity of RA.

We examined 25 patients with a diagnosis of RA according to the claimed method. 11 patients were diagnosed with a moderate degree of activity of RA, 14 patients had a high degree of activity of RA. The results of treatment of all patients confirmed the reliability of the delivered according to the claimed method of diagnosis the differential diagnosis of the degree of activity of rheumatoid arthritis.

Thus, the proposed method allows high accuracy differential diagnosis of the degree of activity of rheumatoid and is TRITA.

The method of differential diagnosis of the degree of activity of rheumatoid arthritis (RA), providing for the investigation of peripheral blood of a patient, characterized in that determine the content of cytotoxic lymphocytes, %: (CD8/CD16), CD8 (t/Grazing), (CD16/Grazing), CD8 (t), (CD16/CD56), (CD8/HLA-DR), as well as their cytotoxic activity (CTA)%, then calculate the index of the degree of disease activity (IA), %, according to the formula

and when 11,8%<IA≤18,2% diagnose average degree of activity of rheumatoid arthritis, the condition of the IA>18,2% - high level of activity of RA.



 

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