Method of modelling autoimmune gastritis in rats

FIELD: medicine.

SUBSTANCE: invention relates to experimental medicine, namely to experimental gastroenterology and can be used for modelling autoimmune gastritis in rats. For this purpose water-salt antigen, containing mixture of homogenate of allogenic mucous membrane of stomach in physiological solution and complete Freund's adjuvant, with component ratio 1:4, is introduced to animal. Method ensures possibility of obtaining model on rats, what results in decrease of research work cost.

EFFECT: ensuring possibility of obtaining model on rats, what results in decrease of research work cost.

2 dwg, 6 cl

 

The invention relates to medicine, in particular for experimental surgery, and is intended for simulation of autoimmune gastritis in rats.

There are various ways of modeling gastritis, for example, by ligation of the gastro-epiploic, gastro-splenic and other veins of the stomach (Norkunas P.I. and others, 1967, 1975) or by cryosurgery over - or subphrenic cut the main trunk of the vagus nerve (Shalimov S. and others, 1989). The disadvantage of the first method is the inability to create the necessary time and power compression ligatures, as long and sharp clamping of the veins of the stomach leads to venous hyperemia of the mucous membrane, up to the formation of erosions and ulcers of the stomach. The disadvantages of the second method are the invasiveness of the method, which should always be accompanied by piloroplasty, prevent the development of stagnant and putrefactive processes in the stomach, as well as heavy portability his animals, resulting in significant part to their death. Thus, these methods do not provide an adequate model of experimental gastritis, especially its chronic course.

In addition, there is a method of modeling of chronic gastritis with symptoms of intestinal metaplasia glands (Wroblewski P.K., 1983), based on a systematic introduction to the animals (dogs) Hello the military or xenogenic antigens stomach, obtained from all walks its walls.

This method is much more effective than the previous one, but also has some significant drawbacks.

1. Extremely long periods of injection of the antigen.

2. In 60% of cases accompanied by the formation of erosions in the mucosa.

3. The introduction of antigens stomach obtained from all walks its walls, in some cases leads to perforation of the stomach with the development of peritonitis and death.

4. The simulation method is carried out on dogs, which increases the cost of the animals and their research works.

The closest analogue of the proposed method is a method of modeling of chronic atrophic gastritis autoimmune Genesis on dogs (Helfman A.E., and others, 1969), which is based on the intake to 5 g of the gastric mucosa by pre-laparotomy and gastrotomy. Mucous membrane prepared water-salt antigen in the amount of 12 ml, mixed with 4 ml of adjuvant's adjuvant and injected subcutaneously in 8 ml twice with an interval of 7 days. The pathological process develops in about a month.

This method in comparison with the previous has the advantage of that is that immunization of dogs can be autologous antigen from the mucous membrane of the stomach, which prevents the development of Grozny is slojnenia - perforation of the organ with the development of peritonitis. In addition, the use of adjuvant's adjuvant allows not systematic introduction of antigen, and 2-3-fold.

However, this method has several disadvantages.

1. This method as an experimental animal to create the model involves the use of dogs, which extremely affects the cost of all the research work and the care of large animals.

2. When modeling diseases involving the immune component should be considered different species sensitivity of laboratory animals to the effects of antigens. In this regard, the dog belongs to one of the most sensitive animals that accompanied 30-40% of their destruction during experimental work.

3. Modeling diseases with dogs requires additional personnel for the anesthesia and surgical intervention.

4. The method is traumatic, accompanied by high rates of mortality.

The aim of the invention is the ability to use as an experimental animal to create a model of autoimmune gastritis rats as the least sensitive to the effects of antigens for modeling diseases involving the immune component, as well as significant is the reduction of the cost of all works (from animal care to research).

Description of the invention.

The method of modelling of autoimmune gastritis in rats as follows.

The first step is preparation of animals (rats) for fence material (stomach).

For 10-12 hours prior to surgery and biopsy specimens of animals exclude food, leaving the drink. For immunization 15-20 rats enough to make a fence material in 5 animals.

In the second stage, taking material for the preparation of water-salt antigen.

Under ether anesthesia after treatment of the surgical field by Piloncillo-Grossao produce laparotomy, the gastrectomy. Material (stomach) is transferred into a sterile Petri dish. Then open the stomach along the greater curvature, 2-3 times lower in a sterile container with saline solution to remove food residue. Then transferred into a sterile H. Petri, which produces a sampling of the gastric mucosa and homogenized.

At the third stage, the preparation of water-salt antigen.

The obtained homogenate of the gastric mucosa in the ratio of 1:10 is mixed with saline solution (1 part homogenate to 10 parts saline) and passed through a cartridge filter (filter) to get rid of large particles of gastric tissue. The resulting solution in a ratio of 1:4 mixed with complete adjuvant the Blockers is (1 part solution to 4 parts complete adjuvant's adjuvant), resuspending until smooth. Then consider water-salt antigen is ready to use. Before using water-salt antigen necessarily within 1 minute resuspending and in the form delivered to the operating room for the introduction of animals or storage at a temperature of 4-6C until use.

At the fourth stage, the introduction of animal water-salt antigen.

The animal is prepared for immunization, for this after a routine inspection of animals under ether anesthesia handle the injection of antigen (paws animal) on Piloncillo-Grossao. Then carry out the introduction of water-salt antigen. In a volume of 0.2 ml antigen is taken in a syringe and injected into the subcutaneous tissue of the tabs on its inner surface. Introduction antigen carried out three times with an interval between doses of 5 days.

This invention is illustrated by the following example.

For modeling of autoimmune gastritis took the rat. After a routine inspection of the animal under ether anesthesia was treated with the injection of antigen (paws animal) on Piloncillo-Grossao. The introduction of water-salt antigen produced after careful resuspendable with complete adjuvant's adjuvant in a dilution of 1:4. In a volume of 0.2 ml was taken in a syringe and injected into the subcutaneous tissue three times with an interval of 5 d is it. 25-30 days after injection of antigen in these intervals occur autoimmune gastritis. Macroscopically it is characterized by redness, swelling of the mucous membranes, hemorrhage (figure 1).

Histologically observed moderate stasis microvasculature, infiltration of the gastric mucosa by neutrophils and lymphocytes, plasmatic cells. Destruction of the gastric mucosa is absent (figure 2).

The present invention was tested on 30 rats and achieved a similar result.

Thus, the present invention allows it to be used as an experimental animal to create a model of autoimmune gastritis rat as the least sensitive to the effects of antigens for modeling diseases involving the immune component, and to achieve a significant reduction in the cost of all works (from animal care to research).

1. The method of modelling of autoimmune gastritis in rats, including the introduction of animal water-salt antigen containing a mixture of homogenate allogeneic gastric mucosa in physiological solution and complete adjuvant's adjuvant at the ratio of components of the mixture 1:4.

2. The method according to claim 1, characterized in that the dilution of the homogenate allogeneic gastric mucosa physiologist in the logical solution is 1:10.

3. The method according to claim 1, characterized in that a single injection of water-salt antigen is carried out in a volume of 0.2 ml.

4. The method according to claim 1, characterized in that the water-salt antigen before the introduction resuspending until smooth.

5. The method according to claim 1, characterized in that the water-salt antigen injected into the subcutaneous tissue of the rat paw on its inner surface.

6. The method according to claim 1, characterized in that the water-salt antigen injected three times with an interval between doses for 5 days.



 

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