Use of metformin for the preparation of a medicament for the treatment of interstitial cystitis

Abstract

The application discloses application of metformin in preparation of a medicine for treating interstitial cystitis, and finds that metformin can reduce pain sensitivity of an interstitial cystitis (IC) rat, reduce edema degree of a bladder of the IC rat, restore bladder capacity of the IC rat, repair the bladder mucosa of the IC rat and reduce submucosal hemorrhage, reduce mast cell infiltration, and has a significant curative effect on interstitial cystitis, and can provide a new medicine for treating interstitial cystitis.

Description

Technical Field

[0001] This invention relates to a novel use of metformin, and more particularly to the use of metformin in the preparation of a medicament for treating interstitial cystitis. Background Technology

[0002] Interstitial cystitis / bladder pain syndrome (IC / BPS) is a multifactorial chronic bladder disease characterized by bladder and pelvic pain, increased pain sensitivity, and lower urinary tract symptoms such as urinary frequency, urgency, or nocturia, significantly reducing patients' quality of life. Existing studies on bladder wall morphology in BPS / IC most commonly report reduced urothelial thickness, glycosaminoglycan (GAG) layer disorder, abnormal smooth muscle structure, upregulated microvascular and nerve density, mast cell infiltration, and fibrosis. However, its pathogenesis, natural course, and pathological mechanisms remain incompletely understood.

[0003] Currently, the treatment of IC / BPS emphasizes individualized and comprehensive treatment strategies. Treatment methods include non-pharmacological treatment, pharmacological treatment, and surgical treatment. Among them, the drugs used in pharmacological treatment are divided into oral drugs (alpha receptor blockers, m receptor blockers, antidepressants, and muscle relaxants, etc.) and intravesical instillation drugs (dimethyl sulfoxide, hyaluronic acid, heparin, pentosan polysulfate, lidocaine, etc.).

[0004] Metformin (Met) has been designated as one of the most important first-line treatments for type 2 diabetes. It primarily works by activating AMPK (adenosine monophosphate-activated protein kinase) in cells, thereby reducing hepatic glucose output. Metformin can also regulate glucose and lipid metabolism in cardiomyocytes, reducing the production of advanced glycation end products (AGEs) and reactive oxygen species in endothelial cells, thus minimizing cardiovascular risk. Studies have found that it has inhibitory effects on tumor cell proliferation and induces apoptosis, showing inhibitory effects on malignant tumors of organs such as the breast, kidney, brain, ovary, lung, and endometrium. Preclinical studies have also demonstrated the neuroprotective effects of metformin in Parkinson's disease, Alzheimer's disease, multiple sclerosis, and Huntington's disease. Currently, there are no reports of metformin being used to treat interstitial cystitis. Summary of the Invention

[0005] This invention is based on the discovery that metformin can significantly improve the symptoms of interstitial cystitis, and thus invented an application of metformin in the preparation of drugs for treating interstitial cystitis.

[0006] The technical solution of the present invention is: the application of metformin in the preparation of a drug for treating interstitial cystitis.

[0007] This invention discovers that metformin can reduce pain sensitivity in rats with interstitial cystitis (IC), reduce bladder edema, restore bladder capacity, repair bladder mucosa, reduce submucosal hemorrhage, and reduce mast cell infiltration. In other words, metformin has significant therapeutic effects on interstitial cystitis and can provide a new drug for the treatment of interstitial cystitis. Attached Figure Description

[0008] Figure 1 These are the experimental results of the Von Frey pain threshold test in rats for each experimental group of this invention.

[0009] Figure 2 These are ultraviolet light images of rat urine spots from various experimental groups of this invention.

[0010] Figure 3 This is a statistical chart showing the analysis of the area of ​​rat urinary spots in the experimental groups of this invention.

[0011] Figure 4 This is a comparison chart of the wet weight of rat bladders in each experimental group of this invention.

[0012] Figure 5 These are HE staining images of rat bladder tissue from various experimental groups in this invention.

[0013] Figure 6 These are toluidine blue staining images of rat bladder tissue from various experimental groups in this invention.

[0014] Figure 7 This is a schematic diagram of mast cell counts in rat bladder tissue from each experimental group of the present invention. Detailed Implementation

[0015] I. Animal Material Sources and Grouping Experimental Methods

[0016] Twelve SPF-grade SD rats (220-250g) were housed under pathogen-free conditions (room temperature 22℃, humidity 50%), with free access to food and water and 12 hours of light per day. All rats were paired with similar weights and then randomly assigned to a control group (control group), an inflammation group (IC group), a gavage treatment group (Met 100mg / kg group), and a bladder instillation treatment group (Met 10mg / ml group), with three rats in each group. On day 1, the inflammation group, gavage treatment group, and bladder instillation treatment group were administered protamine sulfate (PS) 10mg / ml and lipopolysaccharide (LPS) 1mg / ml respectively to establish an interstitial cystitis model via bladder instillation. The control group received saline bladder instillation concurrently. After successful model establishment, the gavage treatment group received metformin hydrochloride 100mg / kg via gavage on days 2, 4, and 6, and the bladder instillation treatment group received metformin hydrochloride 10mg / ml bladder instillation on days 2, 4, and 6. The control group received saline bladder instillation concurrently. Metformin is sourced from Merck & Co., Ltd.

[0017] II. Verification methods and results for the treatment of interstitial cystitis

[0018] On the 7th day of the group experiment, the following experiments were conducted and the corresponding results were obtained.

[0019] 1. Von Frey pain threshold test

[0020] Experimental Method: Each group was placed on a wire mesh, and their hind paws were tested. Using a specific specification of Von Frey filament (e.g., 0.6g as the starting specification), the filament was gently touched to the sole of the animal's hind paw from under the mesh, maintaining contact for 1-2 seconds. The filament specification was adjusted based on the rat's response. If no withdrawal response (e.g., paw retraction or licking behavior) was observed, the filament specification was increased; if a response was observed, the filament specification was decreased until a filament that elicited the withdrawal reflex was found. This process was repeated multiple times (e.g., 5 times), with at least 2 minutes between each repetition to ensure the rat returned to its baseline state.

[0021] The results are as follows Figure 1 As shown, Figure 1 *: p < 0.05, **: p < 0.01. The results showed that compared with the control group, the pain threshold of rats in the IC group was significantly lower (p < 0.05), indicating that their pain perception was more pronounced. The pain threshold of rats in both groups treated with metformin was significantly increased (p < 0.05), but after the third treatment, it was no different from the control group, indicating that metformin restored the pain sensitivity of the interstitial cystitis model rats to normal.

[0022] 2. Urine stain test

[0023] During the experiment, a single SD rat was placed in an experimental cage and on filter paper. The cage containing the rat was placed in a quiet environment to allow the rat to urinate. The test lasted for 4 hours. No water was provided during the test, only food was provided. After 4 hours, the urine stains on the filter paper were collected, imaged with ultraviolet light, and analyzed using ImageJ software to measure the size of the urine stains.

[0024] Ultraviolet imaging, such as Figure 2 As shown, the statistical results analyzed using ImageJ software are as follows: Figure 3 As shown, Figure 3 **: p<0.01.** The results showed that compared with the control group, the urinary spot area in the IC group was significantly reduced (p<0.05), indicating decreased urine output, i.e., reduced bladder capacity and impaired bladder function. After metformin treatment, the urinary spot area in rats significantly increased (p<0.05), especially in rats treated with bladder instillation, demonstrating that metformin treatment increased bladder capacity and restored bladder function in the interstitial cystitis model rats.

[0025] 3. Heavy bladder due to dampness

[0026] Rat bladder samples were collected, rinsed on ice with PBS buffer, and then the surface moisture was blotted dry with filter paper before weighing.

[0027] The results are as follows Figure 4 As shown, Figure 4 *: p<0.05, indicating that compared with the control group, the wet weight of the bladder in IC rats was significantly increased, indicating that the degree of bladder edema was obvious. After treatment with metformin, the wet weight of the bladder in the interstitial cystitis model rats was significantly reduced, especially for rats treated by bladder instillation, which showed a more significant reduction in wet weight, proving that the degree of bladder edema in the interstitial cystitis model rats was reduced after metformin treatment.

[0028] 4. HE staining

[0029] Bladder tissue was fixed in 4% paraformaldehyde and embedded in paraffin. 5μm thick sections were cut using a microtome, dewaxed, rehydrated, cleared, and stained with hematoxylin and eosin.

[0030] HE staining results are as follows Figure 5 As shown in the figure. The results indicated that the bladders of rats in the control group exhibited normal histological morphology, with intact bladder mucosa and no edema or hemorrhage. However, in IC rats, the bladder mucosa showed significant abrasion, edema, and hemorrhage. After treatment with metformin, the bladder mucosa of the interstitial cystitis model rats recovered to its original state without edema or hemorrhage, with the bladder instillation treatment group showing a more significant effect.

[0031] 5. Toluidine blue staining

[0032] Bladder tissue was fixed in 4% paraformaldehyde and embedded in paraffin. 5μm thick sections were cut using a microtome, dewaxed, rehydrated, cleared, stained with toluidine blue, and mast cell counted.

[0033] The results of bladder histamine blue staining are as follows Figure 6 As shown in the diagram, the mast cell counting diagram is as follows: Figure 7 As shown, Figure 7 *: p < 0.05, **: p < 0.01. The results showed that compared with the control group, the number of mast cells in the IC group was significantly increased (p < 0.05), while the number of mast cells in rats with interstitial cystitis treated with metformin was significantly reduced (p < 0.05), with the reduction in mast cell number being particularly significant in the bladder instillation treatment group.

[0034] In summary, the experimental results demonstrate that metformin is significantly effective in treating interstitial cystitis and can be used in the preparation of drugs for treating interstitial cystitis. The drug dosage form can be an inhaler, tablet, capsule, powder, pill, granule, oral liquid, or injection. The route of administration can be oral, intravenous, intraperitoneal, or bladder instillation.

Claims

1. The use of metformin in the preparation of a drug for treating interstitial cystitis.

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