Method for predicting leprosy patient treatment effectiveness

FIELD: medicine.

SUBSTANCE: method involves determining mean cytochemical coefficient of lipid accumulation in peripheral blood leukocytes in conditional units before beginning therapy application (MCC1) and in 2-3 or 5-6, or 10-12, or 20-24 months of therapy application (MCC2). Therapy effectiveness coefficient is calculated in conditional units from formula K= MCC2/MCC1. The value being equal to or greater than 1, leprosy therapy is predicted to be effective.

EFFECT: simplified prognosis method.

1 dwg, 1 tbl

 

The invention relates to medicine, namely to laboratory methods, and can be, in particular, used to predict the effectiveness of therapy in patients with leprosy.

From the practice of medicine known method for predicting the effectiveness of therapy in patients with leprosy laboratory investigation, namely, that determine the number solyusulfona in a daily urine sample and calculate the rate of excretion of the drug from the body of the patient, based on which predict the effectiveness of sulfonic therapy in patients with leprosy (Ryzhova NA, Evstratova, VA Determination of the speed of removing solyusulfona from the body leprosy patients to assess the effectiveness of sulfonic therapy. Methodical recommendations. Astrakhan, 1975, 7 C.). The disadvantages of this method are: a preliminary introduction to the patient a standard test dose solyusulfona that is not suitable for drug drug intolerance, reduction informative way in patients with disorders vadovedyelare and concentration of the kidneys, shall not be conducted during implementation of the method other laboratory tests of urine, the need for implementing the method of stationary laboratory equipment (photoelectrocolorimeter) and pre-build CALIB ovocny curve solyusulfona; the presence of toxic and not readily available reagents, and the long duration of the process (1 to 3 days), which does not allow to obtain a specific technical result is a simplification of the method.

Also known is a method for predicting the effectiveness of therapy in patients with leprosy laboratory tests that identify antibodies to different antigenic determinants .leprae using complex serological reactions in the serum of patients with leprosy (Decina M.N., Yushchenko A., Degtyarev O.V., Lukin Y., Ibraimov M.I. Serological monitoring the effectiveness of chemotherapy for leprosy. Methodical recommendations. Astrakhan, 1994, 16 S.). The disadvantages of this method are: the need for parallel execution 5 serological tests, which requires the implementation of the method in the immunological laboratory, prior receipt of blood serum of the patient as a biological material, the presence of a large number (at least 19), including expensive and not readily available chemical reagents, pooled sera from donors and patients with active stage of leprosy, sets of antibodies of different classes of human immunoglobulins, test antigens USD-.leprae and ultrasound-.avium, and long duration (not less than 20 hours) and high demands on technical equipment method (p is a flat-bottomed anxiety polystyrene, the pipettes-the pipettes variable volume, microphotometer, pH meter, a thermostat, a device for carrying out counter immunoelectrophoresis), which ultimately does not allow to obtain a specific technical result is a simplification of the method.

Closest to the present invention is a method for predicting the effectiveness of therapy in patients with leprosy laboratory research technique lies in the fact that in cells of hematopoietic origin, tissue macrophages, and epithelial cells to determine the total activity of esterases, and the ratio of activity (efficiency ratio) determine the effectiveness of therapy in patients with leprosy (Osipov A.A. Esterna activity in skin lesions in leprosy: the dissertation on competition of a scientific degree of candidate of medical Sciences. - Moscow, 1983. - 23 S.).

The similarity of this method to offer is that they both relate to laboratory research methods based on the study of the metabolism of cells of hematopoietic origin. The disadvantages of this method are:

is a multistage process consisting of a collection of material from the patient using biopsy, a special method of fixing material in a solution of calcium - formula at the fixed temperature mode (0-4°C)prepare on m is Crotone - the cryostat of the material several histological sections with subsequent incubation in a buffer solution with a limited range of temperature (+37° (C) and pH 7.4, estimation of the intensity of histochemical reactions on the scanning microscope - photometer, which ultimately determines the large complexity and technical complexity of the method;

most traumatic way as a result of repeated physical and psychological trauma patient due to the sampling of the material by the method of minor surgery - biopsy of the area of the skin that requires prior anesthesia, observance of the rules of asepsis and the optimal choice of location, depth and size byobserving part of the skin that affects the outcome of the study;

- need to use at least 5 of expensive and toxic chemicals;

- long duration of the method - not less than 18 hours;

- expensive technical equipment necessary to provide a consistent way, namely, - microtome - cryostat MK-25, the stationary thermostat, refrigerator and microscope - photometer SMP-01 company “Opton”, allows you to zoom in study of histological sections not less than 800 times, and cutting tools (scissors, scalpel), which ultimately makes possible the implementation of the method only in the us is the conditions of specialized pathology laboratory.

Thus, these shortcomings do not allow to obtain a specific technical result is a simplification of the method.

The present invention solves the main task is to facilitate the method. The essence of the invention is expressed by a set of essential features that are sufficient to provide the technical invention of the result.

The problem is solved in the invention by the fact that determine the average cytochemical the coefficient of accumulation of lipids in peripheral blood leukocytes in conventional units before therapy (CCS1and after 2-3 or 5-6, 10-12, 20 to 24 months of therapy (CCS2), then calculate the coefficient of effectiveness of therapy in conventional units (K) by the formula:

and when it is equal to 1.20 conventional units and above, predict the effectiveness of therapy in patients with leprosy.

Simplification of the method makes it possible to completely eliminate the two and to simplify the three phases of the prototype method. The duration of the method is reduced to not less than 35 times, and the number of reagents is not less than 1.5 times. In addition, there is no need to use a microtome - cryostat MK-25, the stationary thermostat, cutting tools, fridge and scanning microscope - photometer SMP-01 company “Opton”. The implementation of the separation method does not require special training or perhaps in any laboratory and field conditions.

The problem of predicting the effectiveness of therapy in patients with leprosy remains one of the most important in modern leprology. Its most important aspect is the search for new therapies and individualized treatment regimens for individual patients with leprosy, which is impossible without use in everyday practice physician-leprology reliable and available laboratory methods for predicting the effectiveness of treatment of patients with leprosy. The use of these methods contributes to the timely detection of the .leprae drug resistance, is connected with further progression, the development of complications of the disease result in ineffective therapy of a patient with leprosy.

The proposed method is as follows.

Determine the coefficient of effectiveness of therapy by the formula:

where K - the coefficient of effectiveness of therapy in conventional units;

The CCS1- the calculated value of the average cytochemical coefficient accumulation of lipids in peripheral blood leukocytes (in arbitrary units)as defined before the start of therapy;

The CCS2- the calculated value of the average cytochemical coefficient accumulation of lipids in peripheral blood leukocytes (in arbitrary units)as defined in 2-3 or 5-6, 10-12, 20-24 months tera is AI.

The coefficient of effectiveness of therapy (To) define, respectively, by 2-3 or 5-6, 10-12, 20-24 months of therapy.

Lipids in peripheral blood leukocytes is determined by known methods (Raskin I.M., “oil containing leukocytes in ischemic heart disease”, Moscow, “Medicine”, 1977, p.42-47). To do this, prepare the drug - peripheral blood smear of a patient on a glass slide according to the standard technique (Won, 1999). After conducting a specific color dried in the air looking at the drug under the microscope (oil immersion system, lens X 90, eyepiece X 7). Viewed 100 peripheral blood leukocytes, depending on the intensity of the specific color is conventionally divided into four groups. Groups, depending on the intensity of the specific color included in them of peripheral blood leukocytes, conventionally denoted as“1+”, “2+”, “3+ and “4+”. The first group (1+) leukocytes slabopolojitelen specific color, the cytoplasm of which is painted on the periphery in the form of a thin pale blue rim; the kernel contractno or weak contrast. The second group (2+) - leukocytes with positive specific color, cytoplasm which diffuse painted in pale blue, and the core has a fuzzy contours. The third group (3+) leukocytes with dramatically positive specific color, cytoplas is and are intensely colored in blue color; the core has sharp contours. The fourth group (4+) leukocytes with a maximum specific color, cytoplasm which diffuse painted blue, contains concentrations of dye in the form of individual granules; the kernel has a clear contours. The degree of accumulation of lipids in peripheral blood leukocytes Express the value of the average cytochemical coefficient (CCS), which is calculated by summing the products of the quantities of pluses (1 to 4) and peripheral blood leukocytes of each group and divide this amount by 100 (G.Astaldi et L.Verga, 1957).

where CCS is the calculated value of the average cytochemical coefficient accumulation of lipids in peripheral blood leukocytes (in arbitrary units)as defined before and after the start of therapy;

A, B, C, D - the number of peripheral blood leukocytes in each of the four groups of 100, viewed in the product;

1, 2, 3, 4 - the number of pluses corresponding to each of the four groups is seen in the product 100 peripheral blood leukocytes according to their specific coloring;

100 - the total number reviewed in the preparation of peripheral blood leukocytes.

All used reagents were valid for laboratory research category of quality. The work was carried out using a microscope domestic is about the production of “Micmed-1, option 2”.

Statistical processing of the obtained results was carried out using Microsoft Excel spreadsheets with generally accepted definition of statistical parameters: arithmetic mean (M)mean-square deviation (S) and student test (t).

Summary of the invention is illustrated by the graph and the table.

From the graph it is seen that in patients with leprosy average value of the CCS1reflecting the degree of accumulation of lipids in peripheral blood leukocytes, has the lowest value (1,72 conventional units) prior to the beginning of therapy. Subsequently, after 2-3 months of treatment, the degree of accumulation of lipids in peripheral blood leukocytes (the value of the CCS2) significantly (p<0,01) increases in comparison with the initial value (the value of the CCS1) and becomes equal 2,04 conventional units. In addition, parallel to the emergence of clinical and laboratory signs of effectiveness of therapy and duration of treatment, namely, 5-6 and 10-12 months of treatment, experienced a further statistically significant (p<0.01) increase in the average value of the CCS2that is 2.2 and 2,28 conventional units, respectively. The maximum value of this indicator (CCS2equal 2,39 conventional units) reaches after 20-24 months of therapy on the background of the formation of the ball the majority of patients with signs of regression of the disease. Thus, in patients with leprosy as a result of treatment and in parallel to the emergence of clinical and laboratory signs of its effectiveness increased accumulation of lipids in peripheral blood leukocytes (the value of the CCS2compared with the same measure defined before the start of therapy (the value of the CCS1).

This allows you to use the value of this index (before and at different times of treatment) for the calculation of the effectiveness of therapy (To) in patients with leprosy.

Table
The coefficient of effectiveness of therapy (To) after the start of therapy in conventional units
After 2-3 monthsAfter 5-6 months10-12 monthsAfter 20-24 months
1,201,281,331,40

The table shows that after the start of therapy as signs of regression of the disease, an increase in the value of the coefficient of effectiveness of therapy (To), which is equal to 1.20, 1,28, of 1.33 and 1.40 conventional units, respectively, 2-3, 5-6, 10-12 and 20-24 months of treatment. Moreover, the maximum value of this ratio (1,40 conventional units) coincides with the period of the regression of clinical and histological signs of illness evania after 20-24 months of therapy. In addition, after 2-3 months of therapy the value of the coefficient of effectiveness of therapy (To) is equal to 1.20 conventional units. This value was adopted as the minimum value of the coefficient of effectiveness of therapy (K), the value of which is equal to 1.20 units, and higher predicted the effectiveness of therapy in patients with leprosy. Thus, in the leprosy therapy and parallel to the emergence of clinical and laboratory signs of regression of the disease, an increase in the value of the coefficient of effectiveness of therapy (To), allowing you to use this indicator as a laboratory test for predicting the effectiveness of therapy in patients with leprosy.

The proposed method was successfully tested in the clinic of the research Institute for the study of leprosy health Ministry on 15 patients with leprosy during 1999-2002. All examined patients at the time of the study had clinical manifestations activity leprosula process, which were confirmed by laboratory methods, namely bakteriostaticheski and histologically. The total number of studies have 54 of analysis.

Below are the results of testing.

Example 1.

Patient Mr, born in 1928, (history No. 3875). Sick with leprosy since 1993. In February 2001 for the first time hospitalized in the clinic of the research Institute for the study of leprosy health Ministry with the diagnosis of the m: leprosy, lepromatosis type, LLp, active stage. Clinically the disease is characterized by widespread leprosum process of the skin, namely, diffuse deep infiltration and laromana, facies leonina, as well as lesions of the upper respiratory tract and peripheral nervous system. Bacterioscopic index (BIN) is equal to 1.83+. Histologically admission: Infiltrates lepromatosis structures containing a large number of homogeneous and granular forms .leprae.

To predict the effectiveness of therapy was determined by the coefficient of effectiveness of therapy (K) by the formula:

where K - the coefficient of effectiveness of therapy in conventional units;

The CCS1- the calculated value of the average cytochemical coefficient accumulation of lipids in peripheral blood leukocytes (in arbitrary units)as defined before the start of therapy;

The CCS2- the calculated value of the average cytochemical coefficient accumulation of lipids in peripheral blood leukocytes (in arbitrary units)as defined in 2 months of therapy.

The coefficient of effectiveness of therapy (K) were determined, respectively, by 2 months of therapy.

Preparing the drug - peripheral blood smear of a patient on a glass slide according to the standard technique. After conducting specific OCD the ski according to the method Imersion (1977) reviewed the preparation under the microscope using the oil immersion system. 100 viewed peripheral blood leukocytes, depending on the intensity of the specific color is conventionally divided into four groups: with slabopolojitelen (1+), positive (2+)and strongly positive (3+) and maximum (4+) color. The degree of accumulation of lipids in peripheral blood leukocytes expressing the calculated value of the average cytochemical coefficient (CCS) in arbitrary units.

The value of the CCS1(before treatment) was equal to 1.70 conventional units. The value of the CCS2defined after 2 months of therapy - 2,05 conventional units. The coefficient of effectiveness of therapy (the ratio of the CCS1and the CCS2) after 2 months of treatment, respectively, was 1.20 conventional units. Thus, the coefficient of effectiveness of therapy (K)equal to 1.20 conventional units, which corresponds to the established authors of the lower bounds for the values of the coefficient of effectiveness of therapy (1,20 conventional units), it is possible to reliably predict the effectiveness of treatment after 2 months of treatment.

Example 2.

Patient-N., born in 1936, history No. 3866). Sick with leprosy since 1984. In October 1999 for the first time hospitalized in the clinic of the research Institute for the study of leprosy health Ministry with a diagnosis of leprosy, lepromatous type, LLp active stage. Clinically the disease is characterized by diffuse deep infiltration and laromme on the skin is, lesions of the upper respiratory tract and peripheral nervous system. Bacterioscopic index (BIN) equal 2,33+. Histologically: Infiltrates lepromatosis structures containing a large number .leprae. Predicting the effectiveness of therapy was carried out by the method described in example 1. The value of the CCS1(before treatment) was equal to 1.55 conventional units. The value of the CCS2defined after 5 months of therapy - 2,47 conventional units. The coefficient of effectiveness of therapy (the ratio of the CCS1and the CCS2after 5 months of therapy, respectively, amounted to 1.59 conventional units. Thus, the coefficient of effectiveness of therapy (To), 1.59 conventional units, which is higher than set by the authors of the lower bounds for the values of the coefficient of effectiveness of therapy (1,20 conventional units), it is possible to reliably predict the effectiveness of therapy after 5 months of treatment.

Example 3.

Patient M., born in 1936 history No. 3120). Sore of leprosy since 1963. In November 2000, hospitalized in the clinic of the research Institute for the study of leprosy health Ministry with recurrent disease: leprosy, lepromatous type, LLs, active stage. Clinically the disease is characterized by diffuse deep infiltration and laromme on the skin, lesions of the peripheral nervous system. Bacterioscopic index (BIN) equal to 2.50+. Histologically: Actively almyroudis infiltrate lepromatosis patterns, contains a large number of homogeneous and granular forms .leprae. Predicting the effectiveness of therapy was carried out by the method described in example 1. The value of the CCS1(before treatment) was 2,21 conventional units. The value of the CCS2defined after 20 months of therapy - 2,39 conventional units. The coefficient of effectiveness of therapy (the ratio of the CCS1and the CCS2) after 20 months of therapy, respectively 1,08 conventional units. Thus, the coefficient of effectiveness of therapy (K)equal 1,08 conventional units, which is lower than the authors of the lower bounds for the values of the coefficient of effectiveness of therapy (1,20 conventional units), cannot reliably predict the effectiveness of treatment after 20 months of treatment.

The application of the proposed method compared with the method of the prototype achieved a positive result, which consists in simplifying the prediction of the effectiveness of therapy in patients with leprosy. Simplification of the way full with the exception of two laborious and lengthy stages of the implementation of the prototype method, namely the preparation of the material the patient several histological sections and their incubation in a buffer solution with a limited interval and temperature and pH. This, in turn, eliminates the need for implementing the method of the microtome - cryostat MK-25, stationary thermostat and preliminary preparation of buffer solution. In addition, simplified stage sampling of material from the patient, because instead used for this purpose in the method-prototype method minor surgery - biopsy of skin, in the proposed method uses conventional and low-impact method of sampling peripheral blood of the patient through a puncture in the flesh of the finger. This avoids multiple physical and psychological trauma patient, does not require pre-anesthesia, cutting tools, and does not affect the outcome of the study. Simplified the commit phase of the material, as there is no need for the preservation of the activity of intracellular esterases to apply a special method of fixation of a material at a specific temperature (0-4° (C)that eliminates the need in the refrigerator for technical implementation of the proposed method and gives the opportunity to replace the solution of calcium formula for affordable and inexpensive solution of formalin. Simplifying the evaluation phase, the intensity of the histochemical reaction allows to replace expensive and technically difficult to work scanning microscope - photometer SMP-01 company “Opton" available in any laboratory and easy to use light microscope domestic production. In addition, the COI is whether the light microscope, the proposed method allows to reduce the required size of the larger image is not less than 1.3 times in comparison with the method of the prototype. Thus, simplification of the method allows for a shorter period of time (approximately 35 times smaller in comparison with the method of the prototype) to carry out the prediction of the effectiveness of therapy in patients with leprosy. In turn, this will give an opportunity to establish the presence or formation of drug resistance in the causative agent of leprosy, therefore, to estimate the probability of disease progression and development of complications. This will allow you to adequately plan anti-epidemic activities in relation to the patient, and contact with him faces. the Amount of chemicals required for the implementation of the proposed method is reduced to not less than 1.5 times in comparison with the method of the prototype. The implementation of the method is possible in the field without requiring special training, stationary equipment, expensive and toxic reagents.

Thus, the authors proposed a new, never before do not offer a method for predicting the effectiveness of treatment of patients with leprosy.

The proposed method can be recommended for clinical use in leprosy institutions.

A method for predicting the effectiveness of therapy in patients with leprosy, consisting in the study of the metabolism of cells of hematopoietic about the convergence, characterized in that an average cytochemical the coefficient of accumulation of lipids in peripheral blood leukocytes in conventional units before therapy (CCS1and after 2-3 or 5-6, 10-12, 20 to 24 months of therapy (CCS2), then calculate the coefficient of effectiveness of therapy in conventional units (K) by the formula

and when it is equal to 1.20 conventional units and above, predict the effectiveness of therapy in patients with leprosy.



 

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EFFECT: higher accuracy of detection.

FIELD: medicine, psychiatry, neurology.

SUBSTANCE: the present innovation deals with the ways to detect latent epileptogenesis that provides the chance to evaluate the degree of its compensation or mobility and enables to improve diagnostics of epilepsy at its pre-clinical stage. One should carry out electroencephalographic monitoring by calculating fractal dimensionality of spectral power fluctuation α and biochemical testing paroxysmal cerebral activity due to a test system that enables to detect blood level of autoantibodies to glutamate-binding membranous cerebral protein, moreover, additionally, one should evaluate immune homeostasis, detect the level of total thiolic groups of proteins (SH-groups) in blood serum due to calorimetric technique based upon interaction of molecular iodine with free sulfhydrylic groups of proteins, and spectrophotometrically evaluate the content of average-molecular oligopeptides (AMP), moreover, the values of SH-groups being equal to 2.87-3.42 mM/l and AMP values being equal to 2.89-3.38 g/l in combination with other values prove the availability of pre-clinical stage of epilepsy in patients.

EFFECT: higher accuracy of diagnostics.

3 tbl

FIELD: medicine.

SUBSTANCE: method involves carrying out bacteriological examination of sputum for availability of tuberculosis mycobacterium drug resistance. X-ray and hematological examination is carried out at the disease detection stage. Primary tuberculosis mycobacterium drug resistance being available, large focused shadow of diameter greater than 3.6 cm being found on lung X-picture, leukocytosis and segmented neutrophils availability being found greater than 68% in blood analysis, surgical treatment necessity on the background of short term chemotherapy course before and after operation is to be predicted.

EFFECT: high accuracy of prognosis.

2 dwg

FIELD: medicine, biology.

SUBSTANCE: invention relates to nutrient medium used for accumulation of cells for the following cytological and/or immunocytochemical analysis carrying out. Invention relates to medium containing salts NaCl, KCl, anhydrous CaCl2, MgSO4 x 6 H2O, MgCl2 x 6 H2O, Na2HPO4 x 2 H2O, KHPO4, NaHCO3, and also glucose and Henx's solution, 10% albumin solution and polyglucin taken in the ratio 1:1:1. Invention provides enhancing the preservation of cells.

EFFECT: improved an valuable properties of nutrient medium.

3 ex

FIELD: medicine, urology.

SUBSTANCE: the present innovation deals with differential diagnostics of prostatic cancer and other prostatic diseases at the stage of primary inspection. The method includes the detection of PCA and calculation of probability coefficient for prostatic cancer (PCC) by the following formula: where e - the foundation of natural logarithm (e=2.718…), PCA - the level of total blood PCA in ng/ml, V - patient's age in years. At PCC value being above 0.2 one should diagnose prostatic cancer and to establish final diagnosis one should perform polyfocal prostatic biopsy. The method enables to increase accuracy of diagnostics at decreased number of unjustified prostatic biopsies.

EFFECT: higher efficiency of diagnostics.

2 ex

FIELD: medicine, juvenile clinical nephrology.

SUBSTANCE: disease duration in case of obstructive pyelonephritis should be detected by two ways: either by detecting the value of NADPH-diaphorase activity, as the marker of nitroxide synthase activity in different renal department and comparing it to established norm, or by detecting clinico-laboratory values, such as: hemoglobin, leukocytes, eosinophils, urea, beta-lipoproteides, lymphocytes, neutrophils, the level of glomerular filtration, that of canalicular reabsorption, urinary specific weight, daily excretion of oxalates, arterial pressure, and estimating their deviation against average statistical values by taking into account a child's age.

EFFECT: higher efficiency of detection.

7 dwg, 1 ex, 6 tbl

FIELD: medicine, psychiatry.

SUBSTANCE: one should isolate DNA out of lymphocytes of peripheral venous blood, then due to the method of polymerase chain reaction of DNA synthesis one should amplify the fragments of hSERT locus of serotonin carrier gene and at detecting genotype 12/10 one should predict the risk for the development of hallucino-delirious forms of psychoses of cerebro-atherosclerotic genesis.

EFFECT: more objective prediction of disease development.

3 ex

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