RU2375758C1 - Method for simulation of experimental malignant tumours of lungs - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: invention refers to experimental medicine, namely to experimental oncology, and can be used for simulation of malignant tumour growing in lung tissue. That is ensured by puncture blood sampling in amount 0.3 ml from rat's subclavian vein in sterile conditions in a syringe washed with heparin solution. A needle is left in a wound and covered with a cottonwool tampon with heparin. The same syringe is filled with overoxidised olive oil 0.2 ml preheated to 38°C with dissolved 3,4-benzpyrene 1 mg, all to be quickly mixed and introduced in subclavian vein. A puncture point is tightly pressed with a cottonwool tampon with alcohol within 1 minute. Then 3-3.5 months later a rat manifest tumour of lung, specifically round cell sarcoma.

EFFECT: method provides adequate reproduction of model.

Description

The invention relates to experimental medicine, namely to experimental oncology, and can be used to reproduce a malignant tumor growing in lung tissue.

Animal lung tumors are an important experimental model for research aimed at improving early diagnosis, developing effective prevention and drug therapy for lung cancer. It is known that induced tumors, like spontaneous ones, are a more adequate model of human tumors. It is believed that the only animal on which the reproduction of malignant lung tumors adequate to those in humans is possible is a rat. This is due to the fact that lungs of rats are more similar to human lungs than in mice. In rats, in particular, as in humans, there are mucous glands in the bronchi that are absent in mice.

After oncologist experimenters got the opportunity to work with pure carcinogens, the idea arose of reproducing a tumor in the lung by introducing them directly into the tissue of this organ. Feit et al. after the cotton fiber impregnated with methylcholanthrene was introduced into the lung, squamous metaplasia, proliferation, cell atypia, and even isolated cases of squamous cell carcinoma were detected (Feit, Sveyda, Hoffman "Experimentaly Produced Precancerosis of the Lung in Rats". - "Neoplasma", 1967, 14, 3).

Stanton Blackwell (1961) developed an original method (Stanton MF, Blackwell R. Induction of epidermoid carcinoma in lungs of rats: a "New" method bases upon deposition of methylcholanthrene in areas of pulmonary infraction. - "J. Nat. Cancer Inst." , 1961, 27, 2, 375), we have chosen as a prototype. The authors administered methylcholanthrene dissolved in tricaprilin and in the form of crystals mixed in a solvent intravenously to rats. At the same time, a halogenated hydrocarbon, hexachloro-tetrafluorobutane, was injected intravenously, which caused multiple heart attacks in the lungs associated with damage to small vessels. Against the background of these changes, squamous cell carcinomas were observed, resembling bronchogenic tumors in general appearance and cellular composition, but appearing mainly from small bronchi and therefore pathogenetically corresponding to only some, mainly peripheral, human carcinomas. In this case, according to the authors, the tumors arose from the regenerating epithelium of the bronchus, often turning into a flat keratinizing. If a carcinogenic hydrocarbon was injected with a thrombotic substance, not carcinomas but sarcomas appeared in the lungs. The physicochemical state of carcinogenesis was also important for the histogenesis of tumors. So, after the introduction of fully dissolved methylcholanthrene, in some cases, carcinosarcomas appeared. With the introduction of a carcinogenic substance in the form of crystals mixed in a solvent, only carcinomas appeared. Then the authors revealed that in cases where 7,12-dimethylbenz (a) anthracene or benz (a) pyrene were added to fluorocarbon, fibrosarcomas arose. Tumors did not appear when N-2-fluorenylacetamide, diethylnitrosamine, chromium oxide, asbestos, and tobacco smoke condensate were delayed in places of a heart attack.

However, the low percentage of malignant tumors and their heterogeneous histological structure (from adenomas to squamous cell carcinoma and sarcomas), the inability to systematically obtain material in the predicted amount, makes this method difficult for studies related to the identification of pathognomonic signs of malignant growth in the lung and the development of experimental therapy methods .

An object of the present invention is to provide a method for producing a standard induced malignant tumor growing in rat lung tissue.

This goal is achieved by the fact that 0.5 ml of a mixture containing 1 mg of 3,4-benzpyrene in 0.2 ml of peroxidized olive oil and 0.3 ml of rat blood is injected into the subclavian vein of males of outbred rats. Tumors in the lung appear after 3-3.5 months and morphologically represent squamous sarcoma.

The invention "A method for producing experimental malignant lung tumors" is new, since it is unknown from the level of medicine, namely in experimental oncology, when the malignant process is induced in the lungs of experimental animals.

Analysis of known methods for reproducing an induced experimental malignant process in the lungs gives reason to talk about the novelty of the proposed method, which is represented by morphologically confirmed round-cell sarcoma and reproducibility in the range of 95-96%.

In contrast to the prototype, the proposed "Method for the production of experimental malignant tumors in the lung" is carried out without prior administration to the animals of sufficiently toxic substances that can cause their death, is performed using white outbred rats and obeys the requirements for experimental tumors, one of which postulates that the induced tumors should reproduce in at least 90% of cases, grow and cause animals to die.

The developed method has an inventive step, as it obviously does not follow for a specialist from the known level of development of experimental oncology and reproduction of induced malignant tumors in the lungs of rats. In well-known sources of information in Russia, the CIS countries and abroad, a similar method was not found.

The invention is industrially applicable, since it can be repeatedly reproduced in research institutions of oncological profile with vivariums.

The method is as follows.

All manipulations with animals are made in boxing. Tools, dishes, hands are sanitized in the usual way. The assistant fixes the rat in a supine position, lubricates the supraclavicular region with 70% alcohol with a small content of iodine solution. Then the experimenter in sterile conditions from the subclavian vein by means of a puncture draws 0.3 ml of blood into a syringe washed with a heparin solution. The needle is left in the wound, covered with a cotton ball with heparin, 0.2 ml of pre-heated to 38 ° C preoxidized olive oil with 1 mg of 3,4-benzpyrene dissolved in it is collected into the same syringe, quickly mixed and injected into the subclavian vein. The puncture site is tightly clamped with a cotton swab with alcohol and held for 1 minute.

The procedure was performed on 130 animals. Induced tumors appeared in 124 rats, i.e. at 95.4%.

After opening the animals in the lungs, as a rule, 1-2 focal tumor growth was detected. On histological preparations, a significant blood supply to the lungs with foci of hemorrhages is visualized. The walls of the vessels are significantly thickened, sclerosed. In them, an accumulation of blood cells is noted. Against the background of blood supply and swelling of the interalviolar septa, a destruction of the bronchiolar-alveolar apparatus is observed. The reaction of connective tissue elements that form growths in the form of free cords, as well as perivascular and peribronchial growths, as a manifestation of sclerotization processes, is especially intensely detected.

The picture of destructive changes in both lungs is supplemented by an abundance of homogeneous cells, most often rounded with hyperchromic nuclei resembling lymphocytes. This kind of undifferentiated cells can be attributed to the immature form of sarcoma, namely to squamous cell sarcoma. The stroma in such a tumor is small and it is represented mainly by thin-walled capillaries, but they are not identified on lung preparations.

In 1/3 of the animals, signs of swelling and hemorrhage were determined in the pericardial tissue. Large foci of infiltration with rounded sarcoma-like cells with pronounced hyperchromic nuclei were noted.

The technical and economic efficiency of the “Method for the production of experimental malignant lung tumors” lies in the fact that the resulting induced malignant tumors are morphologically confirmed squamous cell sarcoma, reproduce in 90% of cases, grow and cause animals to die. The method is carried out on white outbred rats, in which the lungs are similar to the lungs of a person, and tumors reproduce without prior administration to animals of toxic substances that can cause their death. The method is economical, affordable, provides the reproduction of tumors in a predicted amount.

Claims (1)

A method of obtaining experimental malignant lung tumors, including the induction of an experimental malignant process, characterized in that from the subclavian vein under sterile conditions, 0.3 ml of blood is drawn into a syringe by a puncture with a heparin solution, the needle is left in the wound, covered with a cotton ball with heparin, 0.2 ml of pre-heated to 38 ° C pre-oxidized olive oil with 1 mg of 3,4-benzpyrene dissolved in it is drawn into the same syringe, mix rapidly and injected into the subclavian vein, place okola tightly clamped with a cotton swab with alcohol for 1 min, after 3-3.5 months a tumor appears in the rat lung - squamous sarcoma.