Application of tautomycin in regulating immunity and resisting infection

Abstract

The invention discloses application of tautomycin in preparation of a medicine for preventing or treating infectious diseases. Experiments show that intraperitoneal injection of Tautomycin can promote migration of mononuclear cells in a mouse to an abdominal cavity, so that an abdominal cavity mononuclear-macrophage component is transformed from mature macrophages to precursor mononuclear cells, the abdominal cavity mononuclear-macrophage component is transformed into mature macrophages more quickly under the infection condition, the effect of swallowing pathogenic microorganisms to remove infection is achieved, and the effect of removing infection is achieved. The survival rate of the E.coli infected mouse model is improved, and the effect is achieved by adjusting immune-related signal channels instead of directly killing pathogenic microorganisms. According to the application, the application range of the Tautomycin is widened, and the potential of the Tautomycin for preventing and treating infectious diseases of mammals is also prompted.

Description

Technical Field

[0001] The invention belongs to the field of biotechnology, and in particular relates to the application of tautomycin in regulating immunity and resisting infection. Background Art

[0002] Tautomycin is a fungal antibiotic isolated from Streptomyces spiroverticillatus broth in China in the late 1980s when screening antibiotics against rapeseed sclerotinia. 41 H 66 O 13 , molecular weight is 766.96. Tautomycin significantly inhibited the growth of microorganisms (MIC = 6.25 × 10 -4 mM), has strong antifungal activity against Streptococcus sclerotiorum, and is a potent and specific inhibitor of protein phosphatase 1 and 2A subunits (PP1 and PP2A) in the absence of Ca 2+ In the case of inducing smooth muscle contraction, the Kiapp values ​​were 0.16nM and 0.4nM, respectively.

[0003] In terms of plant diseases, Tautomycin has good antifungal activity, especially against Sclerotinia sclerotiorum of rapeseed. Tautomycin can cause morphological abnormalities such as swelling and abnormal branching of Sclerotinia sclerotiorum hyphae; the activity of protein phosphatase in Sclerotia sclerotiorum treated with Tautomycin is reduced, and the cell membrane permeability changes, thereby inhibiting the stem rot of rapeseed caused by it.

[0004] There are few studies on the effects of tautomycin in mammalian cells. Some studies have shown that tautomycin (250-1000nM) can inhibit the proliferation of medullary thyroid cancer (MTC) cells and reduce the expression of neuroendocrine markers in MTC cells by inhibiting GSK-3β. Other studies have shown that the combination of tautomycin and enzalutamide can synergistically inhibit the cell proliferation and AR signaling of prostate cancer LNCaP cells and C4-2 cells, as well as androgen receptor (AR) variant-positive 22RV1 cells.

[0005] Until now, the role of Tautomycin in immunity has remained unknown.

[0006] Summary of the invention

[0007] Purpose of the invention: The technical problem to be solved by the present invention is to provide an application of tautomycin in the preparation of drugs for preventing or treating infectious diseases, which mainly plays a role in preventing and treating infectious diseases by regulating innate immune cells rather than directly killing pathogenic microorganisms.

[0008] Technical solution: In order to solve the above technical problems, the present invention provides the use of tautomycin in the preparation of drugs for preventing or treating infectious diseases.

[0009] The infectious diseases include diseases caused by bacteria, fungi or other pathogenic microorganisms.

[0010] The bacteria include but are not limited to Escherichia coli, and the fungi include but are not limited to Candida albicans.

[0011] Wherein, the drug includes a single component or its analogs, derivatives or composite preparations.

[0012] The application is to promote monocyte migration, thereby increasing the proportion of mature macrophages during infection and thus exerting an anti-infection effect.

[0013] The application activates immune signaling pathways such as NF-kappa B signaling pathway, Chemokine signaling pathway, and MAPK signaling pathway of cells, thereby upregulating the levels of cytokines such as RANTES and CCL28 and reducing the levels of cytokines such as Epigen and CCL6.

[0014] Wherein, the application is to fight infection by regulating the conversion of innate immune monocytes-macrophages.

[0015] Wherein, the in vivo experimental concentration of the tautomycin is 0.1-100 ng / g.

[0016] Beneficial effects: The present invention has found through experiments that intraperitoneal injection of Tautomycin can promote the migration of monocytes in mice to the peritoneal cavity, transform the peritoneal monocyte-macrophage components from mature macrophages to promonocytes, and transform them into mature macrophages faster and more under infection conditions, play the role of phagocytizing pathogenic microorganisms to eliminate infection, and improve the survival rate of E. coli infected mouse models. This effect is achieved by regulating immune-related signal pathways rather than directly killing pathogenic microorganisms. The present invention not only broadens the application scope of Tautomycin, but also suggests its potential for preventing and treating infectious diseases in mammals. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1The cell counts of peritoneal lavage fluid of mice in the control group and Tautomycin-treated group;

[0018] Figure 2 The ratio of monocytes to macrophages in the peritoneal lavage fluid of mice in the control group and the Tautomycin-treated group;

[0019] Figure 3 The survival rate of the E. coli infected mouse model in the control group and the Tautomycin-treated group;

[0020] Figure 4 The ratio of monocytes to macrophages in the peritoneal lavage fluid of the E. coli infected mouse model in the control group and the Tautomycin treated group;

[0021] Figure 5 This is the heat map result of the difference of cytokine chip in peritoneal lavage fluid of mice in the control group and Tautomycin treatment group;

[0022] Figure 6 The KEGG enrichment results of Top30 differential cytokines in the peritoneal lavage fluid of mice in the control group and Tautomycin treatment group;

[0023] Figure 7 Tautomycin has no direct killing effect on E. coli and Candida albicans. DETAILED DESCRIPTION

[0024] In order to enable those skilled in the art to better understand the solution of the present application, the technical solution in the embodiments of the present application will be clearly and completely described below in conjunction with the drawings in the embodiments of the present application. Obviously, the described embodiments are only part of the embodiments of the present application, not all of the embodiments. Based on the embodiments in the present application, all other embodiments obtained by ordinary technicians in this field without creative work should fall within the scope of protection of the present application.

[0025] Example 1

[0026] 6-8 week old male C57BL / 6J mice (Wuhan Cloud-Clone Technology Co., Ltd., 8 mice, 18-20g) were intraperitoneally injected with Tautomycin (Millipore, CAS109946-35-2, 1ng / g body weight / day), and the control group (Ctrl) (Wuhan Cloud-Clone Technology Co., Ltd., 8 mice, 18-20g) were intraperitoneally injected with an equal amount of saline at the same time. Two days later, the mice were anesthetized and the peritoneal lavage fluid was obtained by flushing the peritoneal cavity with saline, and the cells in the peritoneal lavage fluid were counted. Compared with the control group, the number of cells in the peritoneal lavage fluid of mice treated with Tautomycin increased significantly ( Figure 1The peritoneal lavage fluid was centrifuged at 1000 rpm for 5 min to obtain the cell pellet, and the cell pellet was resuspended in an appropriate volume of 1×PBS (0.01 M, pH 7.4) and then the cell surface antibodies F4 / 80-FITC (ebioscience, catalog number 11-4801-82) and Ly-6C-PE (ebioscience, catalog number 12-5932-82) were added to label the cells. The monocyte-macrophage ratio in the peritoneal lavage fluid was analyzed by flow cytometry. It was found that compared with the control group, Tautomycin treatment could increase the number of monocytes (Ly6c + F4 / 80 - , 6.87% vs 45.16%) migrated more to the peritoneal cavity, making the peritoneal monocyte-macrophage component composed of mature macrophages (Ly6c - F4 / 80 + ) to pro-monocytes (Ly6c + F4 / 80 - )Conversion( Figure 2 ).

[0027] Example 2

[0028] 6-8 week old male C57BL / 6J mice were intraperitoneally injected with Tautomycin (1 ng / g body weight / day), and the control group (Ctrl) was intraperitoneally injected with an equal volume of saline at the same time. 7 CFUs / mouse E. coli (E.coli ATCC8739) was used to establish an infection mouse model. After 24 hours, the survival of the mice was recorded. It was found that 100% of the mice in the Tautomycin-treated group survived, which was significantly higher than that in the control group injected with normal saline ( Figure 3 ). After the mice were anesthetized, the peritoneal cavity was flushed with normal saline to obtain peritoneal lavage fluid. The peritoneal lavage fluid was centrifuged at 1000 rpm for 5 min to obtain the cell pellet, and the cell pellet was resuspended with an appropriate volume of 1×PBS (0.01M, pH 7.4) and then labeled with cell surface antibodies F4 / 80-FITC (ebioscience, catalog number 11-4801-82) and Ly-6C-PE (ebioscience, catalog number 12-5932-82). The ratio of monocytes to macrophages in the peritoneal lavage fluid of the two groups of mice was analyzed by flow cytometry. It was found that compared with the control group, the mature macrophages (Ly6c - F4 / 80 + ) was significantly higher (33% vs 53.77%) ( Figure 4), indicating that monocytes induced by Tautomycin treatment to migrate to the peritoneal cavity can be transformed into mature macrophages more quickly under infection conditions, playing the role of phagocytizing pathogenic microorganisms to eliminate infection.

[0029] Example 3

[0030] 6-8 week old male C57BL / 6J mice were intraperitoneally injected with Tautomycin (1 ng / g body weight / day), and the control group (Ctrl) was intraperitoneally injected with an equal amount of saline at the same time. Two days later, the mice were anesthetized and the peritoneal cavity was flushed with saline to obtain peritoneal lavage fluid. The peritoneal lavage fluid was centrifuged at 1200 rpm for 10 min to remove the cell precipitate and the supernatant was collected. After aliquoting, it was frozen at -80 degrees for testing. The mouse cytokine chip (RayBiotech, catalog number GSM-CAA-4000-1) was used to detect the levels of cytokines in the peritoneal lavage fluid of the control group and the treatment group, including inflammatory factors, chemokines, etc. It was found that the levels of cytokines in the peritoneal lavage fluid of mice were significantly changed after Tautomycin treatment (calibrated p value <0.05, |logFC|>0.263), for example, the levels of factors such as RANTES and CCL28 increased significantly, and the levels of factors such as Epigen and CCL6 decreased significantly ( Figure 5 ). Signaling pathway enrichment analysis found that the KEGG signaling pathways that were significantly enriched in cytokines after Tautomycin treatment were closely related to immune response, including Cytokine-cytokine receptor interaction, TNFsignaling pathway, NF-kappa B signaling pathway, Chemokine signaling pathway, MAPK signaling pathway, etc. ( Figure 6 ), indicating that Tautomycin can regulate immune-related signaling pathways.

[0031] Example 4

[0032] In order to further verify that the anti-infection effect of Tautomycin is exerted by regulating the innate immune monocyte-macrophage conversion rather than directly killing microorganisms, we used the paper diffusion method drug sensitivity test to detect the antibacterial effect of Tautomycin. Tautomycin was diluted to the corresponding concentration with 1×PBS (0.01M, pH 7.4) and then dropped on the blank drug sensitivity test paper. The paper with different concentrations of Tautomycin (1nM, 10nM, 100nM, 1μM) was pasted on the TSA plate coated with the following bacterial solution. The plate with the paper was placed at 37℃ for 18-24h, and the size of the inhibition zone was used to determine whether the drug had an antibacterial effect. The results showed that Tautomycin of different concentrations (1nM, 10nM, 100nM, 1μM) had no inhibitory effect on the growth of E.coliATCC8739. In addition, based on its known antifungal activity against Sclerotinia sclerotiorum, we tested the effect of Tautomycin on Candida albicans (CMCC(F)98001), a common pathogenic fungus in the human digestive and urogenital tracts, and found that different concentrations of Tautomycin (1nM, 10nM, 100nM, 1μM) had no inhibitory effect on the growth of Candida albicans ( Figure 7 ). Therefore, the anti-infection effect of Tautomycin is through enhancing the innate immune response, regulating the monocyte-macrophage conversion, phagocytosis and elimination of pathogenic microorganisms, rather than directly killing pathogenic microorganisms.

Claims

1. The use of mutamicin in the preparation of drugs for preventing or treating infectious diseases.

2. The use according to claim 1, characterized in that: The infectious diseases include diseases caused by bacteria, fungi or other pathogenic microorganisms.

3. The use according to claim 2, characterized in that: The bacteria include Escherichia coli, and the fungi include Candida albicans.

4. The use according to claim 1, characterized in that: The medicine includes a single component or its analogs, derivatives or composite preparations.

5. The use according to claim 1, characterized in that: The application is to promote the migration of monocytes, thereby increasing the proportion of mature macrophages during infection and thus exerting an anti-infection effect.

6. The use according to claim 1, characterized in that: The application activates the NF-kappa B signaling pathway, the Chemokine signaling pathway, and the MAPK signaling pathway immune signaling pathways of the cells, thereby upregulating the levels of RANTES and CCL28 cytokines and reducing the levels of Epigen and CCL6 cytokines.

7. The use according to claim 1, characterized in that: The application is to fight infection by regulating the innate immune monocyte-macrophage switch.

8. The use according to claim 1, characterized in that: The in vivo experimental concentration of the tautomycin is 0.1-100 ng / g.

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