New application of pentagalloylglucose in preparation of product for intervening iron overload liver injury

Abstract

The invention relates to a novel application of pentagalloyl glucose as an iron overload liver injury protective agent. It is revealed for the first time that pentagalloyl glucose can reduce excessive iron accumulation in iron overload liver cells and animal liver injury models and inhibit cell death, and has the effect of resisting iron overload liver injury; the invention provides a scientific basis for developing liver protection products for resisting iron overload damage based on pentagalloylglucose, and has important application value.

Description

Technical Field

[0001] The present invention belongs to the field of medical technology, and specifically relates to a new use of pentagalloyl glucose as a liver protection ingredient in the preparation of a product for intervening in liver damage caused by iron overload. Background Art

[0002] Iron is one of the essential trace elements for the human body. The iron in the body transfers electrons through changes in valence and participates in a variety of physiological activities. However, studies have shown that long-term intake of large amounts of red meat will lead to an increase in the iron content in the liver. The increased consumption of iron nutritional supplements and fortified foods has also led to an increasing risk of iron overnutrition in the population. In addition, excessive intake of vitamin C, drinking water with excessive iron, long-term excessive drinking, and aging of the body may all cause excessive iron storage in the body. People with iron overload are at a higher risk of abnormal blood sugar metabolism and liver function. The risk of abnormal liver function in the high ferritin group is about 2.96 times that of the low ferritin group. It can be seen that the harm of iron load disorder to health cannot be ignored. Like iron deficiency, iron overload can also have an adverse effect on physical health.

[0003] Galloyl glucose compounds belong to the hydrolyzable tannins among polyphenols. Their basic structure is a D-glucose core connected to multiple gallic groups. They are widely found in fruits and vegetables such as Chinese toon, sea buckthorn, emblica, and southern jujube, as well as mango pits. Among them, pentagalloylglucose (1,2,3,4,6-pentagalloylglucose, PGG) is formed by connecting one gallic acid to each of the C1, C2, C3, C4, and C6 hydroxyl groups of glucose. It has multiple active functions such as antioxidant, antiviral, hypoglycemic and lipid-reducing, and anti-cancer, and has a wide range of uses and high biosafety.

[0004] At present, the existing technology has not yet studied the mechanism of intervention of galloyl glucose compounds in iron overload liver damage. The existing technology CN105193833A discloses the use of the herbaceous chrysanthemum monomer in the preparation of liver protection drugs, but the liver protection mechanism of the compound is not disclosed. And the herbaceous chrysanthemum monomer is also quite different from the galloyl glucose compounds. The glucose is connected to the chalcone dihydrochalcone, which is quite different from the single galloyl structure, and the properties of the compound are obviously different.

[0005] Prior art CN110354135A discloses the application of 1,2,3,6-tetragalloyl glucose ester in the preparation of lipase inhibitor drugs or health foods, which mentions the application of 1,2,3,6-tetragalloyl glucose ester in the preparation of drugs for the treatment of non-alcoholic fatty liver disease. Experiments have shown that 1,2,3,6-tetragalloyl glucose ester can restore ALT levels, relieve hepatocyte swelling, and reduce the volume and number of lipid droplets in obese mice induced by HFD. The mouse model used is C57BL / 6 mice fed with a high-fat diet (HFD).

[0006] Prior art WO2024103215A1 discloses a pharmaceutical combination for treating fatty liver disease and its use, which uses metformin and 1,2,3,4,6-O-pentagalloylglucose. Through the synergistic effect of the two, the performance of glycine N-methyltransferase (GNMT) in liver cells can be greatly improved, while reducing the accumulation of oil droplets in liver cells, and can overcome the problems caused by metformin alone. It can be used for the treatment of fatty liver disease. The mouse model it targets also uses C57BL / 6 mice fed with a high-fat diet (HFD) to model.

[0007] In summary, the prior art has not yet studied and verified the mechanism of liver damage caused by iron overload by galloyl glucose compounds. Based on the above technical problems, the present invention is proposed. Summary of the invention

[0008] The first aspect of the present invention is to provide a new use of pentagalloyl glucose in the development of liver protection products. The new use of pentagalloyl glucose in the development of liver protection products provided by the present invention is: the use of pentagalloyl glucose as a liver protection ingredient in the preparation of products for intervention of iron overload liver damage. The product can be a medicine, food, health food, etc., and the liver protection can be used to intervene in iron overload liver damage. When pentagalloyl glucose is used as an ingredient for intervention of iron overload liver damage, it can be added to the basic formula for intervention of iron overload damage, made into powders, tablets, capsules, etc., and can also be added to food.

[0009] The second aspect of the present invention provides a medicine or health food capable of intervening in liver damage caused by iron overload. The medicine or health food capable of intervening in liver damage caused by iron overload provided by the present invention contains pentagalloyl glucose as an effective ingredient. The damage may specifically be liver damage induced by excessive iron accumulation. The medicine or health food capable of intervening in liver damage caused by iron overload containing pentagalloyl glucose as an effective ingredient is used after the occurrence of liver damage and is administered once a day.

[0010] The third aspect of the present invention is to provide a method for intervening in liver damage caused by iron overload. The method comprises taking pentagalloyl glucose or the above-mentioned medicine or health food with pentagalloyl glucose as an effective ingredient capable of intervening in iron overload damage once a day after liver damage caused by iron accumulation after excessive iron intake.

[0011] In another aspect, the present invention provides the use of pentagalloylglucose in the preparation of a product for protecting liver damage caused by iron overload. Preferably, the product is a medicine or a food.

[0012] On the other hand, the present invention provides the use of pentagalloyl glucose in the preparation of a protective agent for cell damage caused by an ferroptosis inducer. Preferably, the ferroptosis inducer is a glutathione peroxidase 4 inhibitor. More preferably, the ferroptosis inducer is RSL3 and / or dextran iron. The cell damage is hepatocyte damage, and preferably, the hepatocyte is L02 cell.

[0013] The protective agent is in the form of powder, tablet or capsule. Other preparable drug carriers and other drug components may also be added, and the drug components are not limited to cross-linking agents, disintegrants, sweeteners, taste masking agents, dispersants, etc.

[0014] The beneficial effects of the present invention include:

[0015] (1) Screening of 1,3,4,6-tetragalloylglucose (TeGG) and PGG among the galloylglucose compounds that have significant protective effects against liver damage.

[0016] (2) The dual liver injury models induced by RSL3 and dextran iron verified that PGG had a good protective effect on animal models of liver injury caused by iron overload and could inhibit liver damage.

[0017] (3) PGG can reduce the level of iron in the liver and restore iron homeostasis in animal models of iron overload.

[0018] (4) Compared with its structural analogue, chamoside A, it has a better protective effect. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1Gallic acid (GA), 1-Galloyl-beta-glucose (MGG), trigalloyl-glucose (TGG), tetragalloyl-glucose (TeGG) and pentagalloyl-glucose (PGG) inhibit iron overload injury of hepatocytes induced by ferroptosis inducers in vitro (AC) and pentagalloyl-glucose can reduce intracellular iron levels (DE).

[0020] Figure 2 Pentagalloylglucose can significantly reduce the levels of AST and ALT in the serum of iron-overloaded animals (AC) and reduce liver damage induced by iron overload.

[0021] Figure 3 Pentagalloylglucose can reduce iron accumulation in the serum (A) and tissues (BD) of iron-overloaded mice, thereby playing a protective role in intervening iron-overload liver damage.

[0022] Figure 4 .Comparison of the protective effect of pentagalloylglucose with that of thonningianin A (TGA), a structural analogue known to have protective effects against liver damage (AC). DETAILED DESCRIPTION

[0023] The following is a further detailed description of the specific implementation methods and technical solutions of the present invention in conjunction with the accompanying drawings and specific embodiments. It should be made clear that those skilled in the art can refer to the content of this article and appropriately improve the process parameters for implementation. It is particularly important to point out that all similar replacements and modifications are obvious to those skilled in the art, and they are all deemed to be included in the present invention. The methods and applications of the present invention have been described through preferred embodiments, and relevant personnel can obviously modify or appropriately change and combine the methods and applications described herein without departing from the content, spirit and scope of the present invention to implement and apply the technology of the present invention.

[0024] The present invention is described below by using specific embodiments, but the present invention is not limited thereto.

[0025] Unless otherwise specified, the experimental methods used in the following examples are all conventional methods; the reagents, biological materials, etc. used in the following examples are all commercially available unless otherwise specified.

[0026] Normal human liver cells L02 are from the American type culture collection (ATCC cell bank).

[0027] Example 1 Pentagalloylglucose inhibits hepatocyte injury and iron accumulation induced by ferroptosis inducers in vitro.

[0028] Experimental method: In vitro, 0-8 μM gallic acid (GA), 1-Galloyl-beta-glucose (MGG), 1,3,6-Tri-O-galloylglucose (TGG), 1,3,4,6-Tetragalloylglucose (TeGG) and 1,2,3,4,6-O-Pentagalloylglucose (PGG) were used to act on L02 cells induced by 0.5 μM RSL3 for 24 hours, and the cell survival rate was detected by crystal violet staining. Figure 1 AB results showed that gallic acid (GA), 1-galloyl glucose (MGG) and 1,3,6-trigalloyl glucose (TGG) could not protect L02 cells from ferroptosis induced by RSL3 at concentrations of 0-8 μM, while 1,3,4,6-tetragalloyl glucose (TeGG) and PGG had significant protective effects, and PGG had the strongest effect. 1 μM could increase the cell survival rate from about 35% to about 90%, while 1 μM TeGG could only increase it to about 50%, indicating that the number of galloyl groups may affect the inhibitory effect on ferroptosis. After RSL3 treatment of L02 cells, the cells shrank into a spherical shape and floated in the culture medium, and the number of cells decreased. After PGG, the cell morphology recovered, the adhesion state was good, and the number increased significantly ( Figure 1 C). The intracellular iron level was detected, and the results showed that PGG treatment could reduce the intracellular iron accumulation caused by RSL3 ( Figure 1 D). Ferritin FTH1 is a storage form of intracellular iron. When the intracellular iron level increases, it will induce the expression of FTH1. The expression of cellular FTH1 was detected by Western blot. The results also showed that RSL3 can increase the accumulation of cellular iron, and PGG treatment can reduce the expression of FTH1 ( Figure 1 E).

[0029] Example 2 Pentagalloylated glucose inhibits liver damage induced by dextran iron in an animal model of iron overload.

[0030] Experimental methods: Fifteen 4-week-old male KM mice (Beijing Weitong Lihua Experimental Technology Co., Ltd.) were randomly divided into three groups after one week of adaptive feeding, namely blank control group, iron overload model group and PGG intervention group. Both the model group and the intervention group were injected with 25 mg / kg bw dextran iron (INFeD) intraperitoneally 8 times to induce liver iron overload injury, while the control group was injected with an equal amount of normal saline once every two days. After 16 days, the intervention group was gavaged with 50 mg / kg bw PGG solution, and the model group and the control group were gavaged with an equal amount of solvent. The mice were killed after 6 weeks for specimen collection and testing. The experimental process is shown in the figure. Figure 2 A. The serum liver injury indicators AST and ALT increased significantly in the model group, but decreased after PGG intervention compared with the model group, indicating that iron dextran injection induced liver injury and PGG had the effect of inhibiting this liver injury ( Figure 2 B, 2C).

[0031] Example 3 Pentagalloylglucose reduces iron accumulation in an in vivo iron overload model.

[0032] Experimental method: The animal experimental method is the same as in Example 2. The iron levels in serum and liver tissue were detected using a kit. The results showed that PGG intervention significantly reduced the iron accumulation in the serum and liver of iron-overloaded mice ( Figure 3 A, 3B), the spleen is one of the organs of iron metabolism, and the change of its iron level can also represent the state of iron load in the body. Consistent with the liver and serum, the iron content in the spleen also decreases ( Figure 3 C). In the PGG-treated group, the expression of FTH1 in the liver decreased to a certain extent, and the expression of the iron absorption protein TfR1 was restored, indicating that PGG reduced the level of iron in the liver and restored iron homeostasis ( Figure 3 D).

[0033] Example 4 Comparison of the liver protective effects of pentagallocalloyl glucose and cypermethrin a.

[0034] Test method: Thonningianin A (TGA), also known as thonningianin dihydrochalcone-7-O-[3”-O-galloyl-4”,6”-hexahydroxybiphenyl dicarboxyl]-β-D-glucose, is mainly derived from thonningianin extract and has a similar glucose nucleus and galloyl group to PGG. Its structural formula is shown in Figure 4 A. Studies have reported that TGA has a good hepatoprotective effect, can prevent and treat liver damage, and can be used to prepare liver protection drugs. In order to compare the difference in the protective effect of TGA and PGG against iron overload liver damage, L02 cells were treated with 0.5μM RSL3 to induce the same ferroptosis model, and 1μM PPGG and TGA were used for intervention at the same time. The cell survival rate was observed after 24 hours. The results showed that at the same concentration, PGG protected the cells by 100%, while TGA had no significant protective effect ( Figure 4 B, 4C), indicating that PGG has a better protective effect than TGA against iron overload liver injury.

Claims

1. Application of pentagalloylglucose in the preparation of products for protecting liver damage caused by iron overload.

2. Application of pentagalloylglucose in the preparation of drugs for protecting liver damage caused by iron overload.

3. Application of pentagalloylglucose in the preparation of protective agents for cell damage caused by ferroptosis inducers.

4. The use according to claim 3, characterized in that The ferroptosis inducer is a glutathione peroxidase 4 inhibitor.

5. The use according to claim 4, characterized in that The ferroptosis inducing agent is RSL3 and / or iron dextran.

6. The use according to any one of claims 3 to 5, characterized in that: The cell damage is liver cell damage.

7. The use according to claim 6, characterized in that The hepatocytes are L02 cells.

8. The use according to any one of claims 1 to 3, characterized in that: The pentagalloyl glucose structural formula is shown in formula (I):

9. The use according to any one of claims 1 to 3, characterized in that: The protective agent is in the form of powder, tablet or capsule.

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