Method of modelling iron deficiency anaemia

FIELD: medicine.

SUBSTANCE: method includes the introduction of deferoxaminum to mice. Deferoxaminum is introduced subcutaneously in a dose of 0.5 g/kg 2 times with a 3 day interval.

EFFECT: method makes it possible to simply and efficiently model iron deficiency anaemia in mice.

3 tbl, 1 ex

 

The invention relates to medicine, specifically to experimental Hematology, and relates to a method of modeling of iron deficiency anemia.

Iron deficiency and iron deficiency anemia are a national problem of health systems in different countries, as it is the most common pathology in the world after respiratory viral infections. Iron deficiency anemia is considered as a clinico-hematological syndrome characterized by violation of hemoglobin due to iron deficiency in the serum and bone marrow and the development of trophic disorders in all organs and tissues. Changes in anemia and even latent iron deficiency, leading to metabolic, volemic, trophic disorders, will lead to reduced efficiency, increased maternal mortality, negative impact on children's development[3, 6, 7].

In addition, anemia is one of the most common complications of pregnancy. Anemia in pregnant women increases the frequency of premature labor, threatened abortion, fetal malnutrition, labor abnormalities, infectious complications and hypogalactia in parturients, as well as many other complications. In the presence of severe anemia may develop such obstetric pathology, as ej�Menno detachment of the placenta, bleeding during labor and the postpartum period. In women with anemia most children born with low birth weight, signs of fetal malnutrition [4].

Various methods for the modeling of iron-deficiency anaemia, which allows to investigate the mechanisms of development of anemia syndromes and methods of their correction. However, all these methods have drawbacks such as the need for multiple invasive intervention that results in excessive trauma of the animal, as well as the long duration simulation of iron deficiency anemia, so the development of new models of iron deficiency anemia continues to be an urgent task of modern pathophysiology [5, 8].

The closest (prototype) is a method of producing experimental iron deficiency anemia by daily intramuscular injection outbred rats-females deferoxamine at a dose of 150 mg/kg prolongation of the injection timing to 8 weeks [5].

However, the application of this method requires multiple invasive interventions over a long period of time, which complicates the process and leads to unnecessary trauma of the animal and given the short gestation period in animals is not suitable for modeling of iron deficiency anemia complicating pregnancy.

The problem solved by the present invention yavl�is a simplification of the method of modeling of iron deficiency anemia and improve its efficiency.

The task is achieved by the fact that laboratory animals (mice) deferoxamine is administered subcutaneously at 0.5 g/kg 2 times with an interval of 3 days.

New in the present invention is that iron deficiency anemia simulate subcutaneous administration of deferoxamine 0.5 g/kg 2 times with an interval of 3 days.

In the reviewed literature was not found modeling method of iron deficiency anemia by subcutaneous injection of deferoxamine 0.5 g/kg 2 times with an interval of 3 days.

Comparison of the invention with existing methods of modeling of iron deficiency anemia showed that for the first time proposed to simulate iron-deficiency anemia twice by subcutaneous administration of deferoxamine at a dose of 0.5 g/kg with an interval of 3 days. Although in the literature it is known creating a model of iron deficiency anemia by daily intramuscular injection to rats deferoxamine at a dose of 150 mg/kg prolongation of the injection timing to 8 weeks, the introduction of deferoxamine in mice subcutaneously at 0.5 g/kg 2 times with an interval of 3 days of obtaining the desired result, for the specialist is not obvious. The experiment showed unpredictable results.

Thus, the claimed invention meets the criteria of the invention of "Novelty" and "Inventive step", as it is I�mainly not to a person skilled in the art. The present invention meets the criterion of "Industrial applicability" because it can be used in experimental medicine (Hematology, physiology, pathological physiology, pharmacology) for modeling pathological conditions of humans and animals.

The method is as follows.

Laboratory animal (mouse) administered deferoxamine at a dose of 0.5 g/kg subcutaneously, three days later, repeat the introduction of deferoxamine at a dose of 0.5 g/kg subcutaneously.

The proposed method was studied in experiments on animals mice female line F1 (Swahs/6) 54 pieces, weighing 18-20 g. the Animals category 1, the conventional linear mouse, obtained from the nursery of the Department of experimental biomedical modeling, Institute of pharmacology named after E. D. Goldberg, Moscow (certificate available).

Example 1.

The method is as follows. Iron deficiency anemia is modeled using a sequestering agent deferoxamine (Desferal, NOVARTIS PHARMA). The drug is administered subcutaneously at a dose of 0.5 g/kg twice at 1 and 4 days of experiment. Control animals on a similar scheme introduced into the water for injection.

Example 2.

Method prototype. Iron deficiency anemia is modeled using complexing of the drug substance deferoxamine (Desferal, NOVARTIS PHARMA) by daily �nutrimedical administration in the dose of 150 mg/kg prolongation of the injection timing to 8 weeks. Control animals on a similar scheme introduced into the water for injection.

Example 3.

The method of simulation of iron deficiency anemia used in pregnant female mice. To achieve pregnancy for female mice hoisted male. Pregnancy confirmed by the presence of vaginal plugs. 1 day after confirmation of pregnancy, the mice by subcutaneous injection is administered a solution of deferoxamine (Desferal, NOVARTIS PHARMA). The drug is administered subcutaneously at a dose of 0.5 g/kg twice in 3 days. The control group are pregnant females receiving water for injection according to the same scheme.

To confirm the development of anemia in mice subjected to modeling pathological conditions for 56 days by the method prototype and 8 and day 11 of the proposed method, determine the peripheral blood. Blood sampling is carried out from vessels of the tail and on the Hematology analyzer veterinary mode determine the level of hemoglobin, erythrocytes and hematocrit [2]. Processing of results is carried out by the method of variation statistics using t-test and non-parametric U-test of Wilcoxon - Mann-Whitney [1].

The results of the study. In the case of using the prototype method on the eighth week in experimental animals revealed anemia, characterized by the reduction of hemoglobin 31.7% from the beginning of�the diesel values (p< 0,001), decreased red blood cell count (p<0.001) and hematocrit (p<0,001) (table 1).

Table 1.
Indices of peripheral blood of mice of the F1 (Swahs/6) after daily intramuscular administration of deferoxamine at a dose of 150 mg/kg (1) or water for injection (2), (X±m)
The time of the studyHemoglobin, g/lErythrocytes X1012/lHematocrit, %
intact15,96±0,2911,27±0,1235,89±0,81
8 weeks111,10±0,35*8,83±0,11*27,43±0,56*
216,25±0,3811,73±0,0936,58±0,74
Note: here and below * - the difference is significant in comparison with indicators of intact mice; the difference is significant compared to
performance of mice in the control group

Experimental mo�melirovanie iron deficiency anemia according to the proposed method showed that on the 7th day after receiving the full dose (day 11 of the experiment) in mice observed the signs of anemia - a decrease in the level of hemoglobin, erythrocytes and hematocrit compared to non-treated and the control group. The reduction of hemoglobin relative to the control group occurred 36.6% in the group of non-pregnant animals and 12.1% in the group of pregnant females (p<0,001). In the group of pregnant females, the reduction of hemoglobin relative to the intact group was 18.9%. The level of red blood cells in the group of non-pregnant mice decreased by 38% compared to the control group, in the group of pregnant animals - 18.4% (p<0,001). In the group of pregnant females reduced level of red blood cells relatively intact group was 23%. The hematocrit decreased by 32.6% in the group of non-pregnant mice compared with that in the control group and 13.4% in the group of pregnant females (p<0,001). In the group of pregnant females, the decrease in the level of hematocrit relatively intact group was 16.8% (tables 2, 3).

Table 2.
Indices of peripheral blood of mice of the F1 (Swahs/6) after subcutaneous administration of deferoxamine at 0.5 mg/kg 2 times with an interval of 3 days (1) or water for injection (2), (X±m)
The time of the studyHemoglobin, g/lErythrocytes X1012/lHematocrit, %
intact16,17±0,5411,61±0,1535,30±1,14
8 days116,24±0,5310,54±0,3433,88±1,1
216,67±0,3411,52±0,1037,00±0,78
11 days110,45±0,52*7,54±0,45*24,20±0,86*
216,86±0,2511,9±0,0735,90±0,87
Table 3.
The peripheral blood of pregnant female mice of the F1 (Swahs/6) after subcutaneous administration of deferoxamine at 0.5 mg/kg 2 times with an interval of 3 days (1) or water for injection (2), (X±m)
The time of the study Hemoglobin, g/lErythrocytes X1012/lHematocrit, %
intact15,35±0,3611,73±0,0835,06±0,61
8 days112,47±0,3*10,26±0,22*30,87±0,67*
215,8±0,2511,47±0,0836,37±0,58
11 days111,73±0,32*9,51±0,29*Of 29.17±0,8*
214,37±0,3410,83±0,3133,67±0,87

Thus, the introduction of deferoxamine subcutaneously at 0.5 g/kg 2 times with an interval of 3 days leads to the development of iron deficiency anemia in experimental animals (mice). The present invention allows to simplify the method of simulation of iron deficiency anemia and improve its efficiency.

Literature

1) Glants S. Mediko-biological statistics / per. s angl. Danilov Yu. a. / under the editorship of N.�. Busikashvili, D. V. Samoilov. M.: Practice, 1999. 459 S.

2) Clinical laboratory diagnostics: national leadership in 2 t. T. I./ ed. by V. V. Dolgov, V. V. Menshikov. Moscow: GEOTAR-Media, 2012. 928 S.

3) Kovalev L. Iron deficiency anemia. M. Doctor, No. 12, 2002, pp. 4-9.

4) Konovalova E. N., Burlev V. A. Iron status in pregnant and postpartum women // Obstetrics and gynecology. 2012. No. 1. P. 137-142.

5) I. Krasnikova, M. Pathogenetic substantiation of efficiency of application ferrogel in experimental anemia: abstract of Cand. Biol. Sciences. Irkutsk, 2003

6) Pathological physiology / ed. by V. V. Novitsky, E. D. Goldberg. Tomsk, 2001. 716 S.

7) A. L. Tikhomirov, sarsaniia S. I. Rational therapy and modern principles of diagnosis of iron-deficiency States in obstetric practice // Farmateka. 2009; 1. C32-39.

8) Hubbard, A. S., Bandyopadhyay S., et all. Effect of dietary iron on fetal growth in pregnant mice. // Comp Med. 2013 Apr; 63 (2): P. 127-35.

The method of simulation of iron deficiency anemia, which consists in the mice injected deferoxamine, characterized in that deferoxamine is administered subcutaneously at 0.5 g/kg 2 times with an interval of 3 days.



 

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