Use of matrine in preparation of medicine for preventing or treating radiation damage

Abstract

The present application belongs to the field of medicine and biotechnology, and particularly relates to the use of matrine in the preparation of a medicine for preventing or treating radiation damage. The radiation damage includes skin damage, lung damage, intestinal damage, hematopoietic damage, reproductive function damage, and delaying aging caused by radiation. The radiation includes non-ionizing radiation and ionizing radiation. The medicine further comprises other medicines for preventing or treating radiation damage. In the screening of anti-radiation medicines based on various cell models, it is found that matrine has a repair-promoting effect on small intestinal mucosal epithelial cell damage caused by radiation; through a mouse radiation damage model, it is found that matrine can effectively protect the body, reduce the death caused by a lethal dose of Co-60 gamma rays, and delay aging caused by radiation. Matrine has a good clinical application prospect in preventing or treating radiation damage, and has no obvious toxic side effects.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical biotechnology, specifically relating to the use of echinacoside in the preparation of drugs for the prevention or treatment of radiation damage. Background Technology

[0002] With the continuous development of economy and technology, radiation is widely used in various fields. While bringing convenience to mankind, this also poses a significant threat to human health. In particular, the rapid development of the nuclear industry has exposed more and more people to the threat of nuclear radiation. Furthermore, the incidence of malignant tumors is constantly increasing, and radiotherapy, as a conventional anti-tumor treatment, damages normal cells while killing tumor cells.

[0003] Currently, there is a lack of effective radiation protectants in clinical practice. Traditional chemical anti-radiation drugs, such as amifostine, while effective and approved by the FDA, cause toxic side effects such as nausea, vomiting, and low blood pressure. Therefore, there is a need to find safer and more effective anti-radiation drugs to meet clinical needs. In addition, it is also necessary to protect against radiation-related diseases, such as aging and other conditions. In recent years, researchers have turned their attention to the effective anti-radiation active ingredients in traditional Chinese medicine, which have the characteristics of high safety, low toxicity, few side effects, and good patient tolerance, and hold promise as a new avenue for discovering anti-radiation drugs.

[0004] Echinacoside was first discovered in extracts of *Echinacea*, a plant native to North America (family Asteraceae). It is also a major component of *Cistanche deserticola*, a desert plant (family Orobanchaceae) known for its kidney-tonifying, blood-nourishing, and bowel-moistening properties. Previous studies have found that echinacoside possesses antioxidant, anti-inflammatory, anti-tumor, liver-protective, nerve-protective, wound-healing, and memory-improving effects.

[0005] However, there are currently no research reports on the anti-radiation effects of echinacoside. Summary of the Invention

[0006] To address the current lack of effective radiation protection agents in clinical practice, the purpose of this invention is to provide a safe, effective, convenient, and economical drug for the prevention or treatment of radiation damage, offering a new approach to the clinical prevention or treatment of radiation damage.

[0007] The technical solution for achieving the objective of this invention is as follows:

[0008] This invention provides the use of echinacoside in the preparation of medicaments for the prevention or treatment of radiation damage in subjects.

[0009] Preferably, the echinacoside can be extracted and isolated from the flowers, leaves, and fruits of plants containing echinacoside using a biological purification method, or it can be purchased from commercially available natural product monomers.

[0010] Alternatively, in the above-described uses, the radiation damage may be skin damage, lung damage, intestinal damage, hematopoietic damage, reproductive function damage, or radiation-induced aging.

[0011] In addition, the drug prolongs the lifespan of subjects with radiation damage.

[0012] Preferably, the intestinal injury is damage to the small intestinal mucosal epithelial cells.

[0013] More preferably, the echinacoside can upregulate the relative abundance of Lachnoclostridium, Roseburia, Prevotella and Tyzzerella microorganisms.

[0014] Preferably, the subject is a mammal.

[0015] More preferably, the subject is a human, companion animal, farm animal, or laboratory animal.

[0016] Alternatively, in the above-described applications, the radiation may be non-ionizing or ionizing.

[0017] Alternatively, in the above-described applications, the radiation may be ultraviolet radiation, X-ray radiation, gamma-ray radiation, alpha-ray radiation, beta-ray radiation, neutron radiation, or laser radiation.

[0018] Preferably, the radiation is X-ray radiation, gamma-ray radiation, alpha-ray radiation, or beta-ray radiation.

[0019] Alternatively, in the above-described uses, the radiation may be caused by cancer treatment, diagnostic procedures, occupational exposure, nuclear and radiation emergencies, or terrorist attacks.

[0020] Preferably, the diagnostic procedure is a dental or bone X-ray.

[0021] Preferably, the diagnostic process includes PET or CT scans.

[0022] Preferably, the occupational exposure is the occupational exposure of laboratory technicians or nuclear power plant workers.

[0023] Alternatively, in the above-described uses, the drug may also include other drugs for the prevention or treatment of radiation damage.

[0024] Alternatively, in the above-described uses, the drug is a chemical anti-radiation drug or an anti-radiation drug of natural origin.

[0025] Alternatively, in the above-mentioned uses, the chemical anti-radiation drug is amifostine, and the naturally derived anti-radiation drug is astragaloside A, ginsenoside Rg1, gypenosides, angelica polysaccharide, wolfberry polysaccharide, astragalus polysaccharide, ganoderma polysaccharide, polygonatum polysaccharide, silymarin, quercetin, baicalin, or hesperidin.

[0026] Preferably, the drug comprises a pharmaceutically active ingredient and a pharmaceutically acceptable carrier.

[0027] Preferably, the pharmaceutically acceptable carrier refers to a conventional pharmaceutical carrier in the field of pharmaceutical preparations, selected from one or more of fillers, binders, disintegrants, lubricants, suspending agents, wetting agents, pigments, flavoring agents, solvents, and surfactants.

[0028] More preferably, the active pharmaceutical ingredient is echinacoside as the sole active ingredient or is composed of echinacoside and Polygonatum polysaccharide.

[0029] More preferably, in the active pharmaceutical ingredient, the weight ratio of echinacoside to polygonatum polysaccharide is 10:1 to 1:1.

[0030] For example, the weight ratio of echinacoside to polygonatum polysaccharide is 10:1, 9:1, 8:1, 7:1, 6:1, 5:1, 4:1, 3:1, 2:1 or 1:1.

[0031] Alternatively, in the above-described uses, the drug may be an oral or topical preparation.

[0032] Alternatively, in the above-described uses, the oral preparation is a tablet, capsule, granule, or oral liquid, and the topical preparation is a gel, ointment, aerosol, or spray.

[0033] Compared with the prior art, the present invention has the following beneficial effects:

[0034] (1) This invention provides a safe, effective, convenient and economical drug for the prevention or treatment of radiation damage, providing a new approach for the clinical prevention or treatment of radiation damage.

[0035] (2) Pharmacological experiments have shown that in the screening of anti-radiation drugs based on multiple cell models, echinacoside has a promoting effect on the repair of radiation-induced small intestinal mucosal epithelial cell damage; through mouse radiation damage model, it was found that echinacoside can effectively protect the body, reduce death caused by lethal dose of cobalt-60 γ-rays, alleviate radiation-induced intestinal damage, reduce radiation-induced damage to the male mouse reproductive system, and delay radiation-induced aging.

[0036] (3) The natural product monomer echinacoside used in this invention is widely available and can be produced through mature industrial methods, enabling the drug of this invention to be produced on a large industrial scale. In addition, by adding a pharmaceutically acceptable carrier, the drug of this invention can be formulated into conventional oral or topical preparations, making the drug treatment stable, improving the efficacy, and making it convenient to use. Attached Figure Description

[0037] Figure 1 Effects of echinacoside on HIEC-6 human small intestinal mucosal epithelial cells irradiated with 4 Gy cobalt-60 γ rays.

[0038] Figure 2 Effects of echinacoside on human lymphoblast AHH-1 cells irradiated with 4 Gy cobalt-60 γ rays.

[0039] Figure 3 Effects of echinacoside on human vascular endothelial cells (HUVECs) irradiated with 3 Gy cobalt-60 γ rays.

[0040] Figure 4 The dosage range of echinacoside for alleviating radiation damage to small intestinal mucosal epithelial cells.

[0041] Figure 5 Echinacoside increased the 30-day survival rate of mice after a single 6.5 Gy whole-body irradiation.

[0042] Figure 6 Echinacoside increased the 30-day survival rate of mice after a single 7.0 Gy whole-body irradiation.

[0043] Figure 7 Echinacoside increased the 30-day survival rate of mice after a single 7.5 Gy whole-body irradiation.

[0044] Figure 8 : Species diversity of each group of samples.

[0045] Figure 9 Differences and changes in the abundance of fecal microorganisms in different mouse species.

[0046] Figure 10 Echinacoside inhibited the decrease in peripheral blood leukocyte count in irradiated mice.

[0047] Figure 11 Echinacoside combined with polysaccharides inhibited the decrease in peripheral blood leukocyte count in irradiated mice.

[0048] Figure 12 Echinacoside combined with polysaccharides reduced the risk of death in mice irradiated with 8 Gy cobalt-60.

[0049] Figure 13Echinacoside combined with polysaccharides increased the median survival time of mice irradiated with 8.5 Gy cobalt-60. Detailed Implementation

[0050] To facilitate understanding of the present invention by those skilled in the art, the main terms used in the present invention are explained below:

[0051] The drugs described herein may be used at a “therapeutic effective amount.” A “therapeutic effective amount” is the amount that effectively achieves the desired therapeutic outcome within the necessary dose and time period. Therapeutic effective amounts can vary depending on factors such as an individual’s disease state, age, sex, weight, and the ability of the composition to elicit the desired response in an individual. A therapeutic effective amount is also the amount in which the beneficial therapeutic effect outweighs the toxic or harmful effects of the molecule.

[0052] As used herein, the phrase “pharmaceutically acceptable” means compounds, materials, compositions, carriers, and / or dosage forms that are suitable for contact with human and animal tissues within the limits of reasonable medical judgment without excessive toxicity, irritation, allergic reactions, or other problems or complications, and that are commensurate with a reasonable benefit / risk ratio.

[0053] "Pharmaceutically acceptable carrier" means an excipient that can be used to prepare pharmaceutical compositions that are generally safe, non-toxic, and not biologically or otherwise adverse, and includes carriers acceptable for both veterinary and human use. As used herein, "pharmaceutically acceptable carrier" includes one or more such carriers.

[0054] The medicament of this invention can be administered to a subject by any suitable method known to those skilled in the art, such as oral, parenteral, transmucosal, transdermal, intramuscular, intravenous, intradermal, subcutaneous, intraperitoneal, intravenous, intracranial, intravaginal, intratumoral, intravaginal, intratumoral, intratumoral, intratumoral, intratumoral, intratumoral, or intraoral administration. It can also be administered via controlled release by encapsulating the active ingredient in a suitable polymer, which can then be inserted subcutaneously, intratumorally, intraorally, as a skin patch, or intravaginally. The invention also includes coating a medical device with the active ingredient.

[0055] In some embodiments, the pharmaceutical ingredients of the present invention are preferably administered orally and are therefore formulated in a form suitable for oral administration, i.e., as solid or liquid dosage forms. Suitable solid oral dosage forms include tablets, capsules, pills, granules, and pellets. Suitable liquid oral dosage forms include solutions, suspensions, dispersants, emulsions, and oils. In some embodiments, the active ingredients are formulated into capsules. According to this embodiment, the compositions of the present invention also contain a desiccant in addition to the active compound and an inert carrier or diluent, and a gelatin capsule in addition to other excipients.

[0056] In some embodiments, the medicament of the present invention is applied topically to the body surface and is thus formulated into a form suitable for topical application. Suitable topical formulations include gels, ointments, creams, lotions, drops, and controlled-release polymers. For topical application, the echinacoside active ingredient is prepared and applied as a solution, suspension, or emulsion in a physiologically acceptable diluent, with or without a drug carrier.

[0057] Diseases or conditions that can be treated or prevented by the drugs of the present invention include, for example, but not limited to, radiation damage, aging phenotypes, aging-related diseases or conditions, radiation-related aging diseases or conditions, and signs of aging.

[0058] In one instance, the subject requiring radiation protection or radiation mitigation according to the methods provided herein is a subject who is about to, has, or has already been exposed to potentially harmful amounts of radiation. It should be understood that such exposure can be a single exposure, periodic exposure, sporadic exposure, or continuous exposure to radiation. It should also be understood that such radiation exposure includes accidental, incidental, or intentional exposure.

[0059] Examples of subjects for whom the method according to the invention may require radiation protection or radiation mitigation include, but are not limited to, patients exposed to radiation (e.g., proton radiation, photon radiation) as part of a treatment regimen (e.g., cancer patients requiring radiotherapy), subjects exposed to radiation for the diagnosis of a disease or condition (e.g., subjects undergoing dental or bone X-rays, patients undergoing PET scans, CT scans, etc.). Examples of subjects for whom the method according to the invention may require radiation protection or radiation mitigation also include people who may be exposed to radiation due to their occupational or lifestyle choices (e.g., aircraft crew members or other frequent air travelers, even space travelers exposed to above-average radiation levels, laboratory technicians and other workers, or those exposed through the use of electronic devices) or those exposed to radon accumulation (e.g., accumulation in residences or mines), or outdoor workers or sunbathers exposed to natural radiation from the sun. Other subjects for whom the method according to the invention may require radiation protection include those accidentally exposed to radiation (e.g., leaks or spills) (e.g., nuclear reactor leaks or accidents or laboratory spills). Those exposed to radiation from exploding nuclear warheads due to war or terrorism are also considered. Other subjects include those exposed to conventional explosives containing radioactive material detonated by terrorists.

[0060] The term "subject" includes mammals such as humans, companion animals (e.g., dogs, cats, birds, etc.), farm animals (e.g., cattle, sheep, pigs, horses, poultry, etc.), and laboratory animals (e.g., rats, mice, guinea pigs, birds, etc.). In addition to humans, subjects may include dogs, cats, pigs, cattle, sheep, goats, horses, buffalo, ostriches, guinea pigs, rats, mice, birds (e.g., long-tailed parrots), and other wild, domesticated, or commercially useful animals (e.g., chickens, geese, turkeys, fish). The term "subject" does not exclude individuals who are normal in all respects. The term "subject" includes, but is not limited to, people who require treatment or are susceptible to the disease or its sequelae.

[0061] Any patents, patent application publications, or scientific publications cited in this article are incorporated herein in their entirety by reference.

[0062] In the following embodiments, numerous specific details are set forth in order to provide a thorough understanding of the invention. However, those skilled in the art will understand that the invention can be practiced without these specific details. In other instances, well-known methods, procedures, and components have not been described in detail so as not to obscure the invention. Therefore, these embodiments should in no way be construed as limiting the broad scope of the invention.

[0063] Example 1: Echinacoside has a protective effect against radiation-induced damage to human small intestinal mucosal epithelial cells.

[0064] The hematopoietic, circulatory, and digestive systems are highly sensitive to ionizing radiation. In patients exposed to low to medium doses of ionizing radiation, a decrease in peripheral blood leukocytes, lymphocytes, and platelets can be detected, along with petechial hemorrhages, abdominal pain, diarrhea, bloody stools, and other gastrointestinal reactions.

[0065] In this embodiment, human peripheral blood lymphoblastic blast cell line AHH-1, human umbilical vein endothelial cell line HUVEC, and human small intestinal mucosal epithelial cell line HIEC-6 were selected to conduct an in vitro evaluation study on the radiation damage protection effect of echinacoside.

[0066] Human peripheral blood lymphoblastic blast cell line AHH-1 was purchased from ATCC. One hour before irradiation, echinacoside (China National Institutes for Food and Drug Control) solution was prepared using 1640 culture medium (Hyclone) containing 10% fetal bovine serum (Gibco). This solution was serially diluted and added to 96-well plates for prophylactic intervention of AHH-1 cells. The final echinacoside concentrations were 0.39, 0.78, 1.56, 3.12, 6.25, 12.5, 25, 50, and 100 μg / ml. For the sham irradiation group, the irradiation model group (IR), and the cell-free blank culture medium group, each well contained an equal volume of 1640 culture medium containing 10% fetal bovine serum. Radiation damage to AHH-1 cells was induced by irradiation with 4 Gy of cobalt-60 gamma rays. The sham irradiation group was shielded with a 6 cm thick lead brick. Forty-eight hours after irradiation, 10 μL of CCK-8 was added to each well, and the absorbance of each well was read at a wavelength of 450 nm using a microplate reader to calculate the cell viability of each group.

[0067] Human umbilical vein endothelial cell (HUVEC) cells were purchased from ScienCell as primary cells. HUVEC cells in the logarithmic growth phase (passage 6 or less) were trypsinized and resuspended, then the cell concentration was adjusted to 2 × 10⁶ cells / year using ECM medium (ScienCell). 4 100 μL of HUVEC cell suspension was added to each well of a 96-well plate. After 12 hours, HUVECs were fully adhered to the plate, and echinacoside intervention and irradiation were initiated. One hour before irradiation, echinacoside solution was prepared using ECM culture medium and serially diluted to achieve final concentrations of 0.39, 0.78, 1.56, 3.12, 6.25, 12.5, 25, 50, and 100 μg / mL. The culture medium in each well of the 96-well plate was replaced according to these groups for preventative intervention. An equal volume of ECM culture medium was added to each well of the sham irradiation group, the irradiation model group (IR), and the cell-free blank culture group. HUVECs were irradiated with 3 Gy of cobalt-60 gamma rays to induce radiation damage. The sham irradiation group was shielded with a 6 cm thick lead brick. Five days after irradiation, the supernatant was aspirated, and 100 μL of serum-free and cytokine-free culture medium containing 10% CCK-8 was added to each well. The absorbance of each well was read at a wavelength of 450 nm using an ELISA reader, and the cell viability of each group was calculated.

[0068] The human small intestinal mucosal epithelial cell line HIEC-6 was purchased from Shanghai Jining Industrial Co., Ltd. After trypsin digestion and resuspending of HIEC-6 cells in logarithmic growth phase, the cell concentration was adjusted to 1×10⁶ cells / year using Opti-MEM medium (Gibco) containing 5% fetal bovine serum (FBS). 4100 μL of HIEC-6 cell suspension was added to each well of a 96-well plate. After 12 hours, HIEC-6 cells were fully adhered to the plate, and echinacoside intervention and irradiation were initiated. One hour before irradiation, echinacoside solution was prepared using Opti-MEM medium containing 5% fetal bovine serum and serially diluted to final concentrations of 6.25, 12.5, 25, 50, 100, and 200 μg / mL. The medium in each well of the 96-well plate was replaced according to the group for preventative intervention. For the sham group, irradiation model group (IR), and cell-free blank medium group, an equal volume of Opti-MEM medium containing 5% fetal bovine serum was added to each well. HIEC-6 cells were irradiated with 4 Gy of cobalt-60 gamma rays to induce radiation damage. The sham group was shielded with a 6 cm thick lead brick. Five days after irradiation, the supernatant was aspirated, and 100 μL of serum-free Opti-MEM culture medium containing 10% CCK-8 was added to each well. The absorbance of each well was read at a wavelength of 450 nm using a microplate reader, and the cell viability of each group was calculated.

[0069] Cell viability = (OD450 of test group - OD450 of blank culture medium group) ÷ (OD450 of sham irradiation group - OD450 of blank culture medium group)

[0070] For the cell viability assays described above, each group had six replicates. The Student's T-test was used to compare the cell viability of each concentration of echinacea glycoside intervention group with that of the irradiated model group; P < 0.05 was considered statistically significant.

[0071] like Figures 1 to 3 As shown in the results, administration of echinacea glycoside one hour before irradiation resulted in significantly higher cell viability in HIEC-6 radiation-damaged cells at the 100 and 200 μg / mL echinacea glycoside intervention groups compared to the irradiation model group (P < 0.05). However, in AHH-1 and HUVEC cells, cell viability in the 0.39–100 μg / mL echinacea glycoside intervention groups did not increase and even showed a decreasing trend. This suggests that echinacea glycoside can effectively alleviate small intestinal mucosal epithelial cell damage caused by ionizing radiation. However, echinacea glycoside did not show any protective effect against radiation damage at the cell viability level in peripheral blood lymphoblasts (AHH-1) and vascular endothelial cells (HUVEC).

[0072] Example 2: Dosage range of echinacoside in alleviating radiation damage to small intestinal mucosal epithelial cells

[0073] The human small intestinal mucosal epithelial cell line HIEC-6 was purchased from Shanghai Jining Industrial Co., Ltd. After trypsin digestion and resuspending of HIEC-6 cells in logarithmic growth phase, the cell concentration was adjusted to 1×10⁶ cells / year using Opti-MEM medium (Gibco) containing 5% fetal bovine serum (FBS). 4100 μL of HIEC-6 cell suspension was added to each well of a 96-well plate. After 12 hours, HIEC-6 cells were fully adhered to the plate, and echinacoside intervention and irradiation were initiated. One hour before irradiation, echinacoside solution was prepared using Opti-MEM medium containing 5% fetal bovine serum and serially diluted to a final concentration of 50, 60, 70, 80, 90, and 100 μg / mL. The medium in each well of the 96-well plate was replaced according to the group for preventative intervention. An equal volume of Opti-MEM medium containing 5% fetal bovine serum was added to each well of the sham irradiation group, the irradiation model group (IR), and the cell-free blank medium group. HIEC-6 cells were irradiated with 4 Gy of cobalt-60 gamma rays to induce radiation damage. The sham irradiation group was shielded with a 6 cm thick lead brick. Five days after irradiation, the supernatant was aspirated, and 100 μL of serum-free Opti-MEM culture medium containing 10% CCK-8 was added to each well. Two hours later, the absorbance of each well was read at a wavelength of 450 nm using a microplate reader, and the cell viability of each group was calculated.

[0074] Cell viability = (OD450 of test group - OD450 of blank culture medium group) ÷ (OD450 of sham irradiation group - OD450 of blank culture medium group)

[0075] For the cell viability assays described above, each group had six replicates. The Student's T-test was used to compare the cell viability of each concentration of echinacea glycoside intervention group with that of the irradiated model group; P < 0.05 was considered statistically significant.

[0076] like Figure 4 As shown, the experimental results indicate that 60-100 μg / mL echinacoside can effectively alleviate the damage to the small intestinal mucosal epithelium caused by 4 Gy cobalt-60 γ-ray irradiation, significantly improve its cell viability (P<0.0001), and enhance the level of cell viability of normal small intestinal mucosal epithelial cells.

[0077] Example 3: Echinacoside improves 30-day survival rate of mice irradiated with lethal doses of cobalt-60 gamma rays.

[0078] The 30-day survival rate is the most direct indicator for evaluating the efficacy of anti-radiation drugs. SPF-grade male Balb / c mice, 6 weeks old and weighing 20±1g, were purchased from Spiford (Beijing) Biotechnology Co., Ltd. After being housed in IVC cages in an SPF-grade environment for 3 days, cobalt-60 gamma irradiation induced damage to hematopoiesis, immunity, and the intestines, leading to death. It was observed that 50% of mice died within 30 days after a single 6.0 Gy whole-body irradiation, 70% died within 30 days after a single 6.5 Gy whole-body irradiation, and all mice died within 30 days after a single 6.8 Gy or higher whole-body irradiation.

[0079] In this embodiment, mice were irradiated with a 70% lethal dose of 6.5 Gy. Figure 5 Mice irradiated with a 100% lethal dose of 7.0 Gy ( Figure 6 This study investigated the effect of prophylactic gavage administration of echinacoside on radiation-induced mortality in mice. The 30-day survival records showed that oral administration of 60-120 mg / kg echinacoside one hour before irradiation effectively protected mice exposed to lethal doses, prolonging their lifespan and increasing their chances of receiving treatment. Furthermore, oral administration of 60-120 mg / kg echinacoside one hour before irradiation effectively increased the survival rate of mice exposed to lethal doses by 70%, raising the 30-day survival rate of Balb / c mice irradiated with 6.5 Gy from 30% to over 80%. In particular, 60 mg / kg echinacoside showed superior effects in prolonging lifespan and increasing survival rate in mice irradiated with 6.5–7.0 Gy compared to amifostine, the only FDA-approved anti-radiation drug. When mice were irradiated with a higher total lethal dose of 7.5 Gy, no effect was observed on survival time and survival rate from a single prophylactic gavage administration of echinacoside before irradiation, but a single therapeutic gavage administration after irradiation increased the survival rate by 40%. Figure 7 The above examples suggest that echinacoside is effective for both prevention and treatment in radiation-induced fatal emergencies. Furthermore, echinacoside is preferably administered orally and can be stored at room temperature away from light; while WR2721 (amifostine) is administered intravenously, and its lyophilized powder must be stored at 4°C and prepared immediately before use. Based on the above results and analysis, it is evident that echinacoside, as an anti-radiation drug, can be formulated into tablets or granules, making it easier to store, carry, distribute, and use, and significantly improving patient compliance.

[0080] Example 4: Echinacoside-regulated gut microbiome of mice irradiated with lethal dose of cobalt-60 γ-rays

[0081] The gut microbiota is the largest and most complex micro-ecosystem in the human body, comprising approximately 10 times the number of somatic cells, and its impact on metabolic capacity and overall health is significant. Existing literature suggests that gut microbiota play a beneficial role in mitigating intestinal radiation damage and improving the survival rate of mice exposed to lethal doses, making them a reliable target for radiation damage prevention and treatment.

[0082] This embodiment uses Balb / c mice, at a dose of 6.5 Gy 60 Seven days after a single whole-body irradiation with Co-γ rays, fecal samples were collected from mice in a sterile environment. The effects of the anti-radiation drug echinacoside (ECH) on the species and relative abundance of gut microbiota in mice were analyzed using 16S rRNA detection technology. Bioinformatics analysis of the 16S rRNA detection results showed that the species diversity of fecal microorganisms in mice killed by ionizing radiation was significantly different from the other four groups. Furthermore, the microbial diversity of mice treated with echinacoside tended to approach that of normal mice. Figure 8Further analysis revealed that echinacoside could alleviate radiation-induced damage and allow irradiated mice to survive by regulating the relative abundance of microorganisms such as Lachnoclostridium, Roseburia, Prevotella, and Tyzzerella. Figure 9 ).

[0083] Example 5: Echinacoside effectively delays aging and prolongs lifespan in mice surviving high-dose cobalt-60 gamma ray irradiation.

[0084] Male Balb / c mice were divided into three groups: a sham irradiation group, an irradiation model group (IR), and intervention groups containing echinacoside at concentrations of 30, 60, and 90 mg / kg, with 10 mice in each group. After being housed in IVC cages in an SPF-grade animal facility for 3 days, mice were administered a single oral gavage one hour before irradiation. The sham and IR groups were given 0.2 mL of distilled water per mouse, while the intervention groups were administered 0.2 mL of different concentrations of echinacoside solution via gavage to achieve oral intervention doses of 30, 60, and 90 mg / kg echinacoside. The mice were then transported to the cobalt source at Beijing Normal University in SPF-grade irradiation cages and irradiated for 6 minutes at a radiation dose rate of 1 Gy / min. Immediately after irradiation, the mice were returned to the SPF-grade animal facility and housed in IVC cages as usual. After receiving 6 Gy... 60 At week 52 following a single whole-body Co-γ ray irradiation, of the five surviving mice in the irradiation model group, one died, and the remaining four exhibited significantly more signs of aging than the mice in the 60 mg / kg echinacea glycoside prophylaxis group. All four showed symptoms of hair loss, dermatitis, and diarrhea; three (3 / 4, 75%) showed lens opacity, and two (2 / 4, 50%) showed anal prolapse. These four irradiation survivors subsequently died at week 75. In contrast, the mice orally administered 60 mg / kg echinacea glycoside showed less hair loss at week 52 compared to the irradiation-only survivors, with only one showing mild cataracts. All five mice survived at week 75, and four remained alive at week 85. Therefore, prophylactic oral echinacea glycoside can effectively delay the onset of aging symptoms and prolong the lifespan of mice surviving high-dose Co-60 γ-ray irradiation.

[0085] Example 6: Protective effect of echinacoside on peripheral blood counts in mice irradiated with cobalt-60 γ-rays

[0086] Male Balb / c mice were housed in IVC cages in an SPF-grade animal facility for 3 days. One hour before irradiation, each mouse was orally administered 0.2 mL / g orally. The sham-irradiation group and the irradiation model group were given distilled water, while the 30, 60, and 90 mg / kg echinacoside intervention groups were administered the corresponding concentrations of echinacoside aqueous solution by gavage. After exposure to 3.5 Gy... 60 On day 7 following a single whole-body irradiation with Coγ rays, the peripheral blood leukocyte count reached its lowest point. For example... Figure 10As shown, a single prophylactic oral gavage of 60 mg / kg echinacoside one hour before irradiation increased the peripheral blood leukocyte count in irradiated mice. The average peripheral blood leukocyte counts in the 30, 60, and 90 mg / kg echinacoside groups were all increased compared with those in the irradiated model group. Student's T test analysis revealed that the 60 mg / kg echinacoside group showed a statistically significant increase in leukocyte count compared with the irradiated control group (P<0.05).

[0087] Example 7: Combined use of echinacoside and plant polysaccharides enhances the anti-radiation effect.

[0088] Balb / c male mice were subjected to 3.5 Gy and 8 Gy 60 One hour before a single whole-body Co-γ irradiation, mice were orally administered 0.2 mL of the corresponding solution per mouse. The normal control group was administered distilled water by gavage, the echinacoside group was administered echinacoside aqueous solution (dose 50 mg / kg) by gavage, the Polygonatum polysaccharide group was administered Polygonatum polysaccharide aqueous solution (dose 12.5 mg / kg) by gavage, and the combined echinacoside + Polygonatum polysaccharide group was administered a mixed aqueous solution of the two (dose 50 mg / kg echinacoside + 12.5 mg / kg Polygonatum polysaccharide).

[0089] like Figure 11 As shown, the number of peripheral blood leukocytes in mice decreased significantly on day 3 after 3.5 Gy irradiation. Both echinacoside and polygonatum polysaccharide alone could effectively increase the number of peripheral blood leukocytes (P<0.05). The number of peripheral blood leukocytes in the combined echinacoside and polygonatum polysaccharide group was further increased compared with the groups treated with either one alone, and there were statistically significant differences compared with the echinacoside group and the polygonatum polysaccharide group (P<0.05).

[0090] like Figure 12 As shown in the results, the study found that at a completely lethal dose of 8 Gy 60 Under single whole-body irradiation with Coγ rays, the median survival time of mice was 4.5 days, and the 30-day survival rate was zero. One hour before irradiation, a single oral administration of echinacea glycoside or polygonatum polysaccharide effectively prolonged the median survival time and average survival duration of irradiated mice, although the 30-day survival rate remained zero (P<0.05). The median survival time of mice in both the echinacea glycoside and polygonatum polysaccharide groups was greater than or equal to 10 days. Mice that were administered echinacea glycoside and polygonatum polysaccharide together by gavage one hour before irradiation showed a significantly increased median survival time compared to the 8 Gy irradiation group (P<0.05), increasing from 4.5 days to 10.5 days. Although there was no statistically significant difference in median survival time between the combined echinacea glycoside and polygonatum polysaccharide groups and the groups treated with either treatment alone, the 30-day survival rate increased from zero to 20%.

[0091] like Figure 13 As shown in the survival curve of the research results, when60 When the single whole-body dose of Coγ-rays increased to 8.5 Gy, there was no difference in survival rate among the groups. Log rank test showed that the median survival time of mice in the echinacea glycoside and polygonatum polysaccharide intervention groups was no different from that of the irradiation model group. However, the median survival time of mice in the combined intervention group was significantly increased compared with the irradiation model group, the echinacea glycoside intervention group, and the polygonatum polysaccharide intervention group (P<0.05).

[0092] The study suggests that echinacoside alone has anti-radiation effects, which can alleviate hematopoietic damage in irradiated subjects, inhibit peripheral blood leukopenia, and improve the survival of mice. However, it cannot reduce the risk of radiation-induced death when irradiated subjects (such as mice) are exposed to 8.5 Gy or higher doses. The combined use of echinacoside and Polygonatum polysaccharide increases the anti-radiation effect, further inhibits peripheral blood leukopenia in irradiated subjects, reduces the risk of death caused by higher irradiation doses, or effectively increases the median survival time of subjects exposed to higher lethal doses of irradiation.

[0093] Furthermore, in the aforementioned acute and long-term animal experiments, no significant adverse reactions were observed with the drug of the present invention, and the experimental animals all showed good tolerance to it.

[0094] Obviously, those skilled in the art can make various modifications and variations to this invention without departing from its spirit and scope. Therefore, if these modifications and variations fall within the scope of the claims of this invention and their equivalents, this invention also intends to include these modifications and variations.

Claims

1. The use of the active pharmaceutical ingredient composed of echinacoside and polygonatum polysaccharide in the preparation of a drug for preventing or treating gamma-ray radiation damage in subjects, characterized in that: The weight ratio of echinacoside to polygonatum polysaccharide is 4:1, the radiation damage is hematopoietic damage, and the drug prolongs the lifespan of radiation-damaged subjects.

2. The use according to claim 1, characterized in that: The subjects were mammals.

3. The use according to claim 2, characterized in that: The subjects were either humans or laboratory animals.

4. The use according to any one of claims 1 to 3, characterized in that: The drug is an oral preparation.

5. The use according to claim 4, characterized in that: The oral preparation is a tablet, capsule, granule, or oral liquid.

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