Application of genkwanin in preparation of medicine for treating diabetic ulcer

Abstract

The invention relates to the technical field of drugs for treating diabetic ulcer, in particular to application of genkwanin in preparation of drugs for treating diabetic ulcer. It is found that genkwanin has the effect of treating diabetic ulcer, a remarkable treatment effect is achieved, healing and reepithelization of diabetic ulcer wounds can be promoted, the content of active oxygen in the diabetic ulcer wounds can be reduced, and technical support is provided for clinically developing efficient strategies to treat diabetic ulcer in the future.

Description

Technical Field

[0001] The invention relates to the technical field of diabetic ulcer drugs, and in particular to the application of genkwain in preparing drugs for treating diabetic ulcers. Background Art

[0002] Diabetes is becoming more common worldwide. As the course of diabetes increases, the risk of diabetic ulcers in patients increases significantly. Diabetic ulcers are usually difficult to heal and are prone to infection. In severe cases, they can lead to gangrene and even require amputation, which not only causes great pain to patients, but also seriously affects their quality of life and places a heavy burden on the medical system. Modern medicine mainly treats diabetic ulcers by controlling blood sugar, debridement of wounds, anti-infection treatment, and local application of dressings. However, these methods are not ideal for some complex and difficult-to-heal diabetic ulcers. Although simple blood sugar control is the basis, it cannot directly promote wound healing; wound debridement can only remove necrotic tissue and has limited effect on promoting the growth of new tissue; anti-infection treatment is necessary when infection exists, but long-term use of antibiotics may lead to problems such as drug resistance; and ordinary dressings mainly protect the wound surface, but lack the ability to actively promote healing. Therefore, finding an efficient and simple treatment method for diabetic ulcers is a key problem that needs to be solved in clinical practice.

[0003] Genkwanin (GKA) is a flavonoid compound extracted from the plant Genkwanin. It belongs to the alkaloid component and is widely used in traditional medicine and modern pharmacology research. At present, Genkwanin has been studied in the treatment of intervertebral disc degeneration, breast cancer, colorectal cancer, and Parkinson's disease, reflecting its potential and application prospects in the treatment of various diseases. However, there is no report on the correlation between Genkwanin and the treatment of diabetic ulcers in this field. Summary of the invention

[0004] In order to solve the above problems, the present invention provides the use of Genkwanin in preparing a drug for treating diabetic ulcers. The present invention finds that Genkwanin has a significant therapeutic effect in treating diabetic ulcers.

[0005] In order to achieve the above object, the present invention provides the following technical solutions:

[0006] The present invention provides application of genkwain in preparing medicine for treating diabetic ulcer.

[0007] Preferably, the drug for treating diabetic ulcer is a drug that promotes wound healing and re-epithelialization of diabetic ulcer.

[0008] Preferably, the drug for treating diabetic ulcer is a drug for reducing the content of active oxygen in the wound surface of diabetic ulcer.

[0009] The invention provides a medicine for treating diabetic ulcer, wherein the effective component of the medicine comprises genkwain.

[0010] Preferably, the concentration of Genkandrae Chinensis per unit of drug is 10 mM.

[0011] Preferably, the drug is an external preparation.

[0012] Preferably, the auxiliary materials of the drug include: dimethyl sulfoxide, polyethylene glycol 300, Tween-80 and physiological saline.

[0013] Preferably, the volume ratio of dimethyl sulfoxide to physiological saline is 10:45; the volume ratio of polyethylene glycol 300 to physiological saline is 40:45; and the volume ratio of Tween-80 to physiological saline is 5:45.

[0014] Beneficial effects:

[0015] The present invention provides the use of Genkwanin in the preparation of a drug for treating diabetic ulcers. The present invention finds that Genkwanin has the effect of treating diabetic ulcers and has achieved significant therapeutic effects, can promote the healing and re-epithelialization of diabetic ulcer wounds, and reduce the content of active oxygen in diabetic ulcer wounds, providing technical support for the future clinical development of efficient strategies for treating diabetic ulcers. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required to be used in the embodiments are briefly introduced below.

[0017] Figure 1 The effect of Genkandrin on wound healing in diabetic ulcer mice; A is the wound healing photography result and wound healing rate calculation result, B is the HE staining result and HE quantitative calculation result, scale bar = 1mm; * indicates P < 0.05, ** indicates P < 0.01, *** indicates P < 0.001;

[0018] Figure 2 The effect of genkwain on ROS expression in the wounds of mice with diabetic ulcers; A is the result of ROS detection in the wounds by fluorescence microscopy, and B is the quantitative calculation result of ROS detection in the wounds by fluorescence microscopy; * indicates P < 0.05, and *** indicates P < 0.001. DETAILED DESCRIPTION

[0019] The present invention provides application of genkwain in preparing medicine for treating diabetic ulcer.

[0020] The molecular formula of the genkwain of the present invention is C 16 H 12 O5, chemical structure is shown in formula Ⅰ:

[0021]

[0022] The present invention has no particular limitation on the specific source of genkwain, which can be obtained by self-extraction and preparation, or purchased from the market.

[0023] As an embodiment, the drug for treating diabetic ulcer is a drug that promotes healing and re-epithelialization of diabetic ulcer wounds.

[0024] As an embodiment, the drug for treating diabetic ulcer is a drug for reducing the content of active oxygen in the wound surface of diabetic ulcer.

[0025] The present invention finds that Genkwanin has the effect of treating diabetic ulcers and has achieved significant therapeutic effects. It can promote the healing and re-epithelialization of diabetic ulcer wounds and reduce the content of active oxygen in diabetic ulcer wounds, providing technical support for the future clinical development of efficient strategies for the treatment of diabetic ulcers.

[0026] Based on the above advantages, the present invention provides a drug for treating diabetic ulcers, wherein the active ingredient of the drug includes genkwain.

[0027] As an embodiment, the concentration of Genkandrae Chinensis in each unit of the drug is 10 mM.

[0028] As an embodiment, the drug is an external preparation.

[0029] As an embodiment, the auxiliary materials of the drug include: dimethyl sulfoxide, polyethylene glycol 300, Tween-80 and physiological saline.

[0030] As an embodiment, the volume ratio of dimethyl sulfoxide to physiological saline is 10:45; the volume ratio of polyethylene glycol 300 to physiological saline is 40:45; and the volume ratio of Tween-80 to physiological saline is 5:45.

[0031] The drug provided by the present invention has significant therapeutic effect in treating diabetic ulcers, can reduce the ROS expression in the wound surface of diabetic ulcer mice, and has more significant therapeutic effect than the positive control Beifuji.

[0032] In order to further illustrate the present invention, the application of Genkandrae Chinensis provided by the present invention in the preparation of a drug for treating diabetic ulcer is described in detail below in conjunction with the accompanying drawings and examples, but they should not be construed as limiting the scope of protection of the present invention.

[0033] Example 1 Effect of Genkwanin on Wound Healing in Mice with Diabetic Ulcer

[0034] 1. Animals: Animal experiments used m / m control mice and db / db diabetic mice, male, 8 weeks old, weighing 22-24 g, provided by the Experimental Animal Center of Yueyang Hospital of Integrated Traditional Chinese and Western Medicine Affiliated to Shanghai University of Traditional Chinese Medicine, and purchased from Jiangsu GemPharmatech Co., Ltd. All animals were kept at a standard temperature of 23±2°C, with SPF grade 12h light / 12h dark treatment, and the animals had free access to high-fat or standard diet and water.

[0035] Animal model: 5 m / m mice were used as normal control group, and 15 db / db diabetic mice were randomly divided into diabetic group, diabetic genkwain treatment group and diabetic positive treatment group, with 5 mice in each group. The hair on the back of m / m normal control and db / db diabetic mice was shaved, and the shaved area was about 4×4cm; before the ulcer model was established, the mice were anesthetized with isoflurane, and then a wound model with an area of ​​6×6mm was created with a puncher, with 4 wounds on the back of each mouse, and the experiment was carried out under sterile conditions.

[0036] 2. Grouping and treatment:

[0037] (1) Normal group (m / m): ulcer model was established and treated with the drug base solvent. The ulcer of the mouse was wet-compressed at a dosage of 80 μl / mouse / day for 9 consecutive days; the base solvent was composed of the following components in volume concentrations: 10% dimethyl sulfoxide (DMSO), 40% polyethylene glycol 300 (PEG300), 5% Tween-80 (Tween-80), and 45% saline (Saline); (2) Diabetes group (db / db): ulcer model was established and treated with the drug base solvent at a dosage of 80 μl / mouse / day The ulcers of mice were wet-compressed for 9 consecutive days; (3) Diabetes genkwain treatment group (db / db+GKA): genkwain monomer was dissolved in the drug base solvent at a concentration of 10mM and applied to the ulcers of mice at a dosage of 80μl / mouse / day for 9 consecutive days; (4) Diabetes positive treatment group (db / db+rb-bFGF): bovine basic fibroblast growth factor solution (rb-bFGF) with a specification of 63000IU / bottle was topically applied to the ulcers of mice at a dosage of 80μl / mouse / day for 9 consecutive days.

[0038] 3. Experimental methods:

[0039] (1) Wound healing analysis

[0040] Macroscopic observation: Pictures were taken on the 1st, 3rd, 5th, 7th and 9th days after treatment, and the pictures were processed with Image-J software to calculate the wound ulcer area. The wound closure was calculated using the following formula:

[0041] Wound healing rate = [(initial wound area - residual wound area) / initial wound area] × 100%.

[0042] (2) Histological hematoxylin-eosin staining (HE staining)

[0043] On the 9th day after induction, mice were killed and skin specimens were fixed in 4% neutral formaldehyde buffer for 24 hours. Routine histological examination, paraffin embedding, sectioning, and HE staining were performed. The sections were observed under a digital section scanner.

[0044] 4. Experimental Results

[0045] Genkwanin improves wound healing in diabetic ulcer mice

[0046] Wound healing analysis and HE staining were used to observe the effect of Genkwain on diabetic ulcer wound healing in vivo. Figure 1 As shown in A, compared with the diabetic ulcer group, the diabetic Genkandrin treatment group had more significant wound healing effects on days 5, 7, and 9 after injury, and the effect was better than that of the positive treatment group. HE staining results ( Figure 1 Middle B) shows that on day 9, the wound of the diabetic genkwain treatment group was significantly smaller than that of the diabetic ulcer group, and the effect was better than that of the diabetic positive treatment group. This result shows that the wound healing of db / db diabetic mice is slowed down, and the intervention of genkwain can effectively promote the healing and re-epithelialization of diabetic wounds, and the effect is more significant than that of the positive treatment group.

[0047] Example 2 Effect of Genkandrin on ROS Expression in Wounds of Mice with Diabetic Ulcer

[0048] 1. Animal model: m / m control mice and db / db diabetic mice, sourced and treated as in Example 1.

[0049] 2. Grouping and treatment:

[0050] (1) Normal group (m / m): ulcer model was established, and the drug base solvent (same as in Example 1) was used for treatment, and the ulcer of the mouse was wet-applied at a dosage of 80 μl / mouse / day for 9 consecutive days; (2) Diabetes group (db / db): ulcer model was established, and the drug base solvent was used for treatment, and the ulcer of the mouse was wet-applied at a dosage of 80 μl / mouse / day for 9 consecutive days; (3) Diabetes genkwain treatment group (db / db+GKA): genkwain monomer was dissolved in the drug base solvent at a concentration of 10 mM, and the ulcer of the mouse was wet-applied at a dosage of 80 μl / mouse / day for 9 consecutive days; (4) Diabetes positive treatment group (db / db+rb-bFGF): bovine basic fibroblast growth factor external solution (Beifuji, rb-bFGF) with a specification of 63000 IU / bottle was topically applied to the ulcer of the mouse at a dosage of 80 μl / mouse / day for 9 consecutive days.

[0051] 3. Experimental methods:

[0052] (1) Fluorescence staining and semi-quantitative analysis of ROS in wound surface

[0053] On the 9th day after induction, mice were killed, wound skin specimens were taken, and the expression of ROS in the wound was detected using a dihydroethidium ROS detection kit. 200 μl of cleaning solution was added to 10 μm frozen sections for 5 min, and the cleaning solution was removed; 50 μM dihydroethidium fluorescent probe (purchased from Sigma, USA, catalog number 37291) was diluted 1000 times with PBS, and then added to the frozen sections, protected from light, and the wet box was placed in a 37°C water bath for incubation for 30 min; the dihydroethidium fluorescent probe was removed, washed 3 times with PBS, counterstained with DAPI, and washed 3 times with PBS; the sections were sealed with anti-fluorescence quenching sealing agent; ROS was detected under a fluorescence microscope. The mean fluorescence intensity of the fluorescence images was semi-quantitatively analyzed using Image-J software, and the mean fluorescence intensity was calculated using the following formula:

[0054] Average fluorescence intensity = total fluorescence intensity of the region / area of ​​the region.

[0055] 4. Experimental results:

[0056] Genkwanin can reduce the expression of ROS in the wound of diabetic ulcer mice

[0057] The dihydroethidium ROS detection kit was used to detect the ROS content and observe the effect of Genkandrin on the expression of ROS in the wound surface of diabetic ulcer mice. Figure 2 As shown in the figure, compared with the diabetic ulcer group, the ROS content in the wound of the diabetic genkwain treatment group was significantly reduced and lower than that of the diabetic positive treatment group. This result shows that the expression of ROS in the wound of db / db diabetic mice is abnormally increased, and the intervention of genkwain can effectively reduce the expression of ROS in diabetic wounds, and the effect is more significant than that of the positive control Beifuji.

[0058] In summary, Genkwain has significant therapeutic effect in the treatment of diabetic ulcers and can reduce the expression of ROS in the wound surface of diabetic ulcer mice.

[0059] Although the above embodiment describes the present invention in detail, it is only a part of the embodiments of the present invention, not all of the embodiments. People can also obtain other embodiments based on this embodiment without creativity, and these embodiments all fall within the protection scope of the present invention.

Claims

1. Application of Genkwanin in the preparation of drugs for treating diabetic ulcers.

2. The use according to claim 1, characterized in that: The drug for treating diabetic ulcer is a drug for promoting the healing and re-epithelialization of diabetic ulcer wounds.

3. The use according to claim 1, characterized in that: The drug for treating diabetic ulcer is a drug for reducing the content of active oxygen in the wound surface of diabetic ulcer.

4. A drug for treating diabetic ulcer, characterized in that: The active ingredients of the medicine include genkwain.

5. The drug according to claim 4, characterized in that The concentration of Genkandrin in each unit of drug was 10 mM.

6. The drug according to claim 4 or 5, characterized in that The medicine is an external preparation.

7. The drug according to claim 6, characterized in that The auxiliary materials of the medicine include: dimethyl sulfoxide, polyethylene glycol 300, Tween-80 and normal saline.

8. The drug according to claim 7, characterized in that The volume ratio of the dimethyl sulfoxide to the physiological saline is 10:45; the volume ratio of the polyethylene glycol 300 to the physiological saline is 40:45; and the volume ratio of the Tween-80 to the physiological saline is 5:45.

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