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Establishment of mouse tuberculosis natural dormant infection model preparation and drug evaluation methods

A technology of latent infection and mice, which is applied in the field of mouse tuberculosis natural latent infection model preparation and drug evaluation method establishment, can solve the problems of inability to study the immune mechanism, long incubation period, long research cycle, etc., and achieve convenient latent and relapse mechanism , the effect of stabilizing the acute phase and shortening the incubation period

Inactive Publication Date: 2016-02-03
INST OF LAB ANIMAL SCI CHINESE ACAD OF MEDICAL SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although the infection process of the low-dose natural infection model is similar to that of human latent infection, the existing models have a high load of bacteria in the lung and spleen (4-5 log10CFU) and a long incubation period (up to 107 weeks), so the mechanism of latent recurrence can be studied , and the evaluation of drug vaccines using recurrence is too long, which is not conducive to the development of research
However, the latent infection model obtained after the intervention factor is different from the natural latent infection process of humans. The early immune mechanism cannot be studied, the incubation period is long and varied, and the recurrence varies greatly. It is difficult to standardize the evaluation of research drugs and vaccines.

Method used

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  • Establishment of mouse tuberculosis natural dormant infection model preparation and drug evaluation methods
  • Establishment of mouse tuberculosis natural dormant infection model preparation and drug evaluation methods
  • Establishment of mouse tuberculosis natural dormant infection model preparation and drug evaluation methods

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] The construction of embodiment 1 mouse tuberculosis latent infection model

[0036] SPF-grade C57 mice (6-8 weeks old, female, 450 mice) were purchased from Victoria Lihua Company and adapted to ABSL-3 for 1 week. All mice were raised in accordance with the ethical requirements of animal welfare, and obtained the approval number ILAS-PC-2013-015 from the Animal Welfare Ethics Committee of the Institute of Medical Experimental Animals, Chinese Academy of Medical Sciences. After the mice entered the ABSL-3 laboratory, 6 mice per cage, free access to water and food, laboratory humidity (50±10%), light (12 / 12h light / dark cycle), temperature (23±2°C). At the end of the experiment, mice were fixed and euthanized.

[0037] Mycobacterium tuberculosis H37Rv grows on the L-J medium for 3 weeks to the logarithmic growth phase, add 2ml of 0.9% NaCl, scrape off the tuberculosis colonies with a sterile L glass rod, blow and mix them repeatedly with a 1ml pipette, and pass the bacter...

Embodiment 2

[0040] The detection of the lung of embodiment 2 mouse latent infection model of tuberculosis, spleen tissue bacteria load

[0041] 1, 3, 5, 8, 12, 16, 20 and 24 weeks after the infection of Example 1, 6 mice were dissected respectively, and the lung and spleen tissues were put in 4% H 2 SO 4 Remove possible miscellaneous bacteria, put the tissue in a 2ml glass grinder, add 1ml 0.9% NaCl for grinding. The homogenate was serially diluted 10 times, and 100 μL of the homogenate was inoculated on the L-J slope, and placed at 37°C, 5% CO 2 cultured in an incubator. After 21 days counting, 6 mice per time point were subjected to statistical analysis. (T-test)

[0042] The bacterial loads of lung and spleen tissues at 1, 3, 5, 8, 12, 16, 20 and 24 weeks after infection are shown in Table 1:

[0043] Table 1 Bacteria load in mouse lung and spleen tissue

[0044]

[0045] Note: *** shows significant difference in the number of sclerotia compared with the blank control group (P...

Embodiment 3

[0048] Lung and spleen histopathological detection of embodiment 3 mouse tuberculosis latent infection model

[0049] Example 1 Dissect 6 mice at 1, 3, 5, 8, 12, 16, 20, and 24 weeks after infection, and dissect 6 mice at each time point at 34, 38, 44, and 52 weeks after infection in the first experiment , while the second and third experiments, the remaining mice after 24 weeks were dissected at 52 weeks. The lung, spleen and liver tissues of the mice were dissected, checked for gross pathology, and soaked in formalin. According to the routine pathological embedding and repair method, HE staining was performed after sectioning, and histopathological lesions were diagnosed by senior animal pathology experts, and pathological scoring was performed. According to the size of the lesion and the degree of inflammation, it was divided into: +, 25%; ++ ,50%; +++,75% of the 3 grades.

[0050] Throughout the experimental period, there were no visible lesions in the lung and spleen ti...

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Abstract

The invention relates to the technical field of animal model building, and in particular to a mouse tuberculosis dormant infection model establishment method. An incubation period of a mouse tuberculosis dormant infection model built by the method provided by the invention can be controlled to be within 20 weeks. In addition, the model built by the method provided by the invention accords with natural dormant infection characteristics of human. The condition that in the early phase of incubation (within 8 weeks), the mouse tuberculosis dormant infection model can be used for quickly and effectively evaluating antituberculosis drugs and vaccines without the need of using relapse in a later period of incubation is discovered for the first time. In addition, a model preparation technology in the invention can shorten the incubation period, stabilization in the incubation period is realized, and relapse conditions are relatively coincident, thus facilitating the research on incubation and relapse mechanisms. In addition, any intervention factor is not used, and an infection process of the model is the same as the natural dormant infection process of human, thus facilitating the research on an immunologic mechanism in the early phase.

Description

technical field [0001] The invention relates to the technical field of animal model construction and drug evaluation, in particular to a method for preparing a mouse natural latent infection model of tuberculosis and establishing a drug evaluation method. Background technique [0002] Tuberculosis is a chronic infectious disease caused by Mycobacterium tuberculosis, which can invade many organs, with pulmonary tuberculosis infection being the most common. Tuberculosis is a chronic infectious disease that seriously endangers people's health. At present, about 2 billion people are infected in the world. There are about 8-10 million new tuberculosis patients every year, and about 2-3 million deaths due to tuberculosis every year. At present, the annual incidence of tuberculosis in my country is about 1.3 million, and the number of deaths due to tuberculosis reaches 130,000 per year, exceeding the total number of deaths from other infectious diseases. [0003] The latent infect...

Claims

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Application Information

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IPC IPC(8): A61K35/74A61K49/00A01K67/02
Inventor 占玲俊唐军林树柱秦川
Owner INST OF LAB ANIMAL SCI CHINESE ACAD OF MEDICAL SCI
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