Application of IL-33 in preparation of anti-inflammatory drugs

Abstract

The invention discloses application of IL-33 in preparation of anti-inflammatory drugs, and relates to the technical field of biological medicines, and the key points of the technical scheme are as follows: it is found through mouse experiments that injection of IL-33 in a Zusanli acupoint area can effectively improve the survival rate of mice with systemic inflammation induced by LPS and effectively reduce the levels of serum inflammatory factors IL-1beta, IL-6 and TNF-a. It is proved that IL-33 has the anti-inflammatory effect, technical support is provided for research and development of new anti-inflammatory drugs, and a new application approach is provided for IL-33. Meanwhile, a new treatment mode is provided for treating the LPS-induced systemic inflammation, and a theoretical basis is provided for researching and treating the LPS-induced systemic inflammation in the future.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, and more specifically, to the application of IL-33 in the preparation of anti-inflammatory drugs. Background Technology

[0002] IL-33 (interleukin-33) is an important member of the IL-1 family, playing a crucial role in the immune system by regulating various cell types and inflammatory responses. IL-33 primarily promotes Th2 immune responses by binding to its receptor ST2, activating eosinophils, mast cells, natural killer cells (NK cells), and innate lymphocyte type 2 (ILC2). For example, in allergic reactions, IL-33 exacerbates inflammatory responses in diseases such as asthma by inducing Th2 cell differentiation and the secretion of factors such as IL-4 and IL-5.

[0003] Existing technologies have demonstrated that IL-33 can induce skin inflammation and trigger inflammatory responses in the nasal mucosa. For example, IL-33 activates ILC2 and Th2 cells, triggering mast cell degranulation and leading to allergy symptoms such as sneezing and runny nose. Furthermore, IL-33 is associated with the inflammatory process of diseases such as rheumatoid arthritis and may influence disease progression by regulating obesity-related metabolic processes. In addition, in immunologically sensitive tumors such as pancreatic cancer, IL-33 enhances anti-tumor immune responses by promoting the formation of tertiary lymphoid structures (TLSs). Recent studies show that its optimized protein can significantly improve anti-tumor effects in mouse models.

[0004] IL-33, located in the cell nucleus in its resting state, is a cytokine that plays multiple key roles in the immune system, which can be summarized as an "alarm" and a "coordinator." It is mainly released when tissues are damaged or infected, thereby initiating, coordinating, and amplifying the immune response. Summary of the Invention

[0005] The purpose of this invention is to provide the application of IL-33 in the preparation of anti-inflammatory drugs. This invention found that injecting IL-33 into the Zusanli acupoint can effectively improve the survival rate of mice with lipopolysaccharide (LPS)-induced systemic inflammation and effectively reduce the levels of serum inflammatory factors IL-1β, IL-6 and TNF-α, thus exhibiting a certain anti-inflammatory effect.

[0006] The above-mentioned technical objective of the present invention is achieved through the following technical solution: the application of IL-33 in the preparation of drugs that improve the survival rate of mice with LPS-induced systemic inflammation and anti-inflammatory drugs.

[0007] The present invention is further configured such that the effective component of the drug for improving the survival rate of mice with LPS-induced systemic inflammation and the anti-inflammatory drug is IL-33 cytokine.

[0008] The present invention is further configured such that the drug and anti-inflammatory drug for improving the survival rate of mice with LPS-induced systemic inflammation include pharmaceutically acceptable excipients.

[0009] The present invention is further configured such that: the pharmaceutically acceptable excipient is selected from at least one of the following pharmaceutically acceptable solvents, solubilizers, cosolvents, emulsifiers, osmotic pressure regulators, stabilizers, suspending agents, anti-adhesives, binding agents, penetration enhancers, pH adjusters, buffers, surfactants, absorbents, diluents, filter aids, and sustained-release materials.

[0010] The present invention is further configured such that the drug and anti-inflammatory drug for improving the survival rate of mice with LPS-induced systemic inflammation include pharmaceutically acceptable dosage forms.

[0011] The present invention is further configured such that the pharmaceutically acceptable dosage form is any one of powder, granule, or aqueous solution.

[0012] The present invention is further configured such that the drug for improving the survival rate of mice with LPS-induced systemic inflammation and the anti-inflammatory drug need to be prepared as an injection solution when used.

[0013] The present invention is further configured such that the injection solution is applied to the Zusanli acupoint.

[0014] In summary, this invention has the following beneficial effects: Mouse experiments showed that injecting IL-33 into the Zusanli acupoint effectively improved the survival rate of mice with LPS-induced systemic inflammation and effectively reduced the levels of serum inflammatory factors IL-1β, IL-6, and TNF-α. This demonstrates that IL-33 possesses anti-inflammatory effects, providing technical support for the development of new anti-inflammatory drugs and offering a novel application pathway for IL-33. Simultaneously, it provides a new treatment method for LPS-induced systemic inflammation and lays a theoretical foundation for future research and development of treatments for LPS-induced systemic inflammation. Attached Figure Description

[0015] Figure 1 This is an analysis diagram of the experimental results of the mouse experiment in the embodiments of the present invention.

[0016] A. Effect of IL-33 on survival rate in LPS-induced systemic inflammation mice. Ctrl group: injected with 5% trehalose; MA group: injected with PBS followed by manual acupuncture; IL-33 group: injected with IL-33 (n = 20). B. Effect of IL-33 on IL-1β, IL-6, and TNF-α levels in LPS-induced systemic inflammation mice. Grouping same as A (n = 5). C. Effect of sST2 on survival rate in LPS-induced systemic inflammation mice during acupuncture. Ctrl group: injected with PBS; MA group: injected with PBS followed by manual acupuncture; sST2 + MA group: injected with sST2 followed by manual acupuncture (n = 20). D. Effect of sST2 on IL-1β, IL-6, and TNF-α levels in LPS-induced systemic inflammation mice during acupuncture. Grouping same as C (n = 5). *, **, ***, and **** represent p < 0.05, 0.01, 0.001, and 0.0001, respectively. Detailed Implementation

[0017] The following is in conjunction with the appendix Figure 1 The present invention will be described in further detail below.

[0018] The IL-33 used in this embodiment of the invention was purchased from Thermo Fisher, catalog number: Cat#210-33-10μg. sST2 was purchased from R&D Company, catalog number: Cat#1004-MR.

[0019] Example 1: Inhibitory effect of IL-33 on LPS-induced systemic inflammation in mice

[0020] Preparation procedure for IL-33 working solution: Dissolve 10 μg of IL-33 powder in 1 mL of 5% trehalose buffer to prepare a stock solution with a concentration of 10 μg / mL. Before use, dilute the stock solution to a concentration of 4 μg / mL for IL-33 working solution.

[0021] Experimental grouping: Eight-week-old C57 / BL6J mice were selected for the experiment and divided into a control group (Ctrl), a manual acupuncture group (MA), and an IL-33 group, with 25 mice in each group. The acupoint ST36 (located approximately 4 mm below the knee joint and 2 mm lateral to the anterior tibial tuberosity) was selected for injection and acupuncture. A standard Hamilton microsyringe was used for subcutaneous injection at the ST36 acupoint. After inserting the syringe into the acupoint, aspiration was performed. If no blood return was observed, the liquid was slowly injected into the acupoint, with a standard injection volume of 5 μL per acupoint.

[0022] in:

[0023] Control group mice were injected with 5% trehalose buffer, and 25 minutes after the injection, LPS (8 mg / kg per mouse) was injected intraperitoneally to induce systemic inflammatory response in mice. Twenty mice / groups were subjected to a 7-day survival observation experiment, and blood was collected from the eyes of the other 5 mice / groups 90 minutes after the LPS injection.

[0024] After injection of 5% trehalose buffer, the mice in the manual acupuncture group underwent manual acupuncture. Acupuncture needles (0.25 × 13 mm, Hua Tuo brand) were inserted into the bilateral ST36 of the mice, and a rotating manipulation was performed with a balanced tonifying and reducing technique. The needles were manipulated once every 5 minutes for 1 minute each time, with a rotation amplitude of 180° and a frequency of 2 Hz. The total acupuncture duration was 25 minutes. Immediately afterwards, LPS (8 mg / kg per mouse) was injected intraperitoneally to induce a systemic inflammatory response in the mice. Twenty mice in one group were used for a 7-day survival observation experiment, while blood was collected from the eyeballs of the other 5 mice in one group 90 minutes after the LPS injection.

[0025] The IL-33 group was injected with IL-33 working solution, and 25 minutes after the injection, LPS (8 mg / kg per mouse) was injected intraperitoneally. Twenty mice / groups were observed for survival for 7 days, while the other 5 mice / groups had their eyeballs enucleated and blood collected 90 minutes after the LPS injection.

[0026] Survival rate observation over 7 days revealed that IL-33 effectively improved the survival rate of mice, comparable to the effect of acupuncture. Blood samples collected from mice 90 minutes after injection were analyzed. The collected whole blood was allowed to stand at room temperature for 2 hours, then centrifuged at 4000 rpm for 10 minutes at 4 °C. Serum was collected for inflammatory factor level detection. The results showed that IL-33 effectively reduced the levels of IL-1β, IL-6, and TNF-α, indicating that IL-33 possesses a certain anti-inflammatory effect.

[0027] Example 2: Effect of sST2 on IL-33's inhibition of LPS-induced systemic inflammation in mice

[0028] Preparation procedure for IL-33 working solution: Dissolve 10 μg of IL-33 powder in 1 mL of 5% trehalose buffer to prepare a stock solution with a concentration of 10 μg / mL. Before use, dilute the stock solution to a concentration of 4 μg / mL for IL-33 working solution.

[0029] Preparation procedure for sST2 working solution: Dissolve sST2 powder in PBS buffer to prepare a stock solution with a concentration of 100 μg / mL. Before use, dilute the stock solution to a concentration of 30 μg / mL.

[0030] Experimental Groups: Eight-week-old C57 / BL6J mice were selected for the experiment and divided into a control group (Ctrl), a manual acupuncture group (MA), and a sST2+ manual acupuncture group (sST2+MA), with 25 mice in each group. The Zusanli (ST36) acupoint was selected for injection and acupuncture. ST36 is located approximately 4 mm below the knee joint and approximately 2 mm lateral to the anterior tibial tuberosity. A standard Hamilton microsyringe was used for subcutaneous injection at the ST36 acupoint. After inserting the syringe into the acupoint, aspiration was performed. If no blood return was observed, the liquid was slowly injected into the acupoint, with a standard injection volume of 5 μL per acupoint.

[0031] The control group was injected with 5% trehalose buffer. 25 minutes after the injection, LPS (8 mg / kg per mouse) was injected intraperitoneally to induce a systemic inflammatory response in the mice. 20 mice / groups were used for a 7-day survival observation experiment. The other 5 mice / groups had their eyeballs enucleated and blood collected 90 minutes after the LPS injection.

[0032] After injection of 5% trehalose buffer, the mice underwent manual acupuncture. Acupuncture needles (0.25 × 13 mm, Hua Tuo brand) were inserted into the bilateral ST36 fossa of the mice using a rotating manipulation technique, applying a balanced tonifying and reducing method. The needles were manipulated once every 5 minutes for 1 minute each time, with a rotation amplitude of 180° and a frequency of 2 Hz, for a total acupuncture duration of 25 minutes. Immediately afterwards, LPS (8 mg / kg per mouse) was injected intraperitoneally to induce a systemic inflammatory response. Twenty mice in each group underwent a 7-day survival observation experiment, while blood was collected from the eyeballs of the other five mice in each group 90 minutes after LPS injection.

[0033] After injection of sST2 working solution, the sST2+ hand acupuncture group underwent hand acupuncture. Acupuncture needles (0.25×13 mm, Huatuo brand) were inserted into the bilateral ST36 of mice, and a rotating manipulation was performed with a balanced tonifying and reducing technique. The needles were manipulated once every 5 minutes for 1 minute each time, with a rotation amplitude of 180° and a frequency of 2 Hz. The total acupuncture time was 25 minutes. Immediately afterwards, LPS (8 mg / kg per mouse) was injected intraperitoneally to induce a systemic inflammatory response in the mice. Twenty mice / groups were used for a 7-day survival observation experiment, while the other 5 mice / groups had their eyeballs enucleated and blood collected 90 minutes after the LPS injection.

[0034] The results showed that injecting the free IL-33 receptor sST2 into the acupoints before acupuncture significantly reduced the acupuncture effect and decreased the survival rate of mice, showing a significant difference compared to the acupuncture group (n=20). Although it still reduced serum inflammatory markers, the difference was still significant compared to the acupuncture group (n=5). This indicates that IL-33 can effectively inhibit LPS-induced systemic inflammation in mice.

[0035] This specific embodiment is merely an explanation of the present invention and is not intended to limit the invention. After reading this specification, those skilled in the art can make modifications to this embodiment without contributing any inventive step, but such modifications are protected by patent law as long as they are within the scope of the claims of the present invention.

Claims

1. Use of IL-33 for the manufacture of an anti-inflammatory medicament, characterized in that: The medicine can reduce the levels of serum inflammatory factors IL-1β, IL-6 and TNF-a.

2. Use according to claim 1, characterized in that: The medicine can improve the survival rate of mice with LPS-induced systemic inflammation.

3. Use according to claim 1 or 2, characterized in that: The effective components of the medicine are all IL-33 cytokines.

4. Use according to claim 1 or 2, characterized in that: The medicine comprises pharmaceutically acceptable excipients.

5. Use according to claim 4, characterized in that: The pharmaceutically acceptable excipients are selected from at least one of pharmaceutically acceptable solvents, solubilizers, cosolvents, emulsifiers, osmotic pressure regulators, stabilizers, suspending agents, anti-adhesion agents, integrating agents, penetration enhancers, pH value regulators, buffers, surfactants, absorbents, diluents, filtration aids, controlled-release materials.

6. Use according to claim 1 or 2, characterized in that: The medicine comprises pharmaceutically acceptable dosage forms.

7. Use according to claim 6, characterized in that: The pharmaceutically acceptable dosage forms are any one of powders, powders or water.

8. Use according to claim 7, characterized in that: The medicine needs to be prepared into an injection solution when used.

9. Use according to claim 8, characterized in that: The injection solution acts on the Zusanli acupoint.

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