Use of disopyramide in the preparation of a medicament for treating a disease associated with lacrimal gland secretory dysfunction

Abstract

The application is a divisional application of 2025116336025, and provides application of desipramine in preparation of a medicine for treating diseases related to lacrimal gland secretion dysfunction, and the technical key points are as follows: in the application, a dry eye mouse model with lacrimal gland dysfunction is constructed, and the liposome-encapsulated desipramine is used for intervention. Through comprehensive analysis of multiple physiological and histological indexes, it is found that the desipramine treatment significantly improves the ocular surface function and structure of the model animal. Specifically, compared with the model control group, the fluorescein sodium staining score of the desipramine treatment group of mice is significantly reduced, indicating that the corneal epithelial injury is effectively repaired; at the same time, the epithelial layer structure in the corneal tissue tends to be complete, and the inflammatory cell infiltration in the stroma layer is significantly reduced; the density of goblet cells in the conjunctiva also rises significantly, indicating that the mucus layer secretion function of the ocular surface is restored. And the desipramine treatment has an improvement trend on the tear film break-up time and tear secretion volume, indicating its potential role in maintaining the stability of the tear film and promoting the secretion of the basal tear.

Description

[0001] This application is a divisional application of Chinese invention application filed on November 10, 2025, application number 2025116336025, entitled "A coordinate axis organoid chip for high-throughput in-situ tracking drug screening and its construction method". Technical Field

[0002] This invention relates to the field of biomedical technology, and in particular to the application of disepine in the preparation of drugs for treating diseases related to lacrimal gland secretion dysfunction. Background Technology

[0003] Dry eye syndrome is the most common eye disease seen in ophthalmology consultations besides refractive errors. It is mainly characterized by insufficient tear production or excessive tear evaporation, leading to dryness, irritation, and inflammation of the ocular surface. Traditional treatments (such as artificial tears and anti-inflammatory drugs) can relieve symptoms, but they often fail to fundamentally repair damaged ocular surface tissues and require frequent use, resulting in poor patient compliance. Therefore, exploring effective and alternative treatment interventions is crucial to meeting urgent medical needs. In drug screening for dry eye syndrome, traditional 2D cell and animal models are far from meeting practical requirements. Establishing new experimental medical models and research paradigms is of great significance for a deeper understanding of the pathogenesis of dry eye syndrome and the development of effective treatments. Summary of the Invention

[0004] The purpose of this invention is to address the lack of effective treatments for dry eye syndrome in the existing technology.

[0005] To achieve the above objectives, the present invention adopts the following technical solution:

[0006] Application of disepine in the preparation of drugs for treating diseases related to lacrimal gland secretion dysfunction.

[0007] Preferably, the lacrimal gland secretion dysfunction-related diseases include dry eye syndrome.

[0008] This application also provides a medicament for treating diseases related to lacrimal gland secretion dysfunction, the medicament comprising diserpine.

[0009] Preferably, the drug also includes a pharmaceutically acceptable carrier or excipient.

[0010] Compared with the prior art, this application has the following beneficial effects:

[0011] This study established a mouse model of dry eye due to lacrimal gland dysfunction and treated it with liposome-encapsulated diserpine. Comprehensive analysis of multiple physiological and histological indicators showed that diserpine treatment significantly improved ocular surface function and structure in the model animals. Attached Figure Description

[0012] Figure 1 This is a bright-field image of the lacrimal gland organoids after stimulation with the traditional Chinese medicine monomer disheping according to one embodiment of the present invention, showing its morphological changes and secretion-related responses.

[0013] Figure 2 This is an image of corneal fluorescein-stained mice after treatment with liposome-coated diserine in one embodiment of the present invention, showing the improvement of corneal epithelial damage.

[0014] Figure 3 This is a representative image of the conjunctival sac and corneal region of a dry eye mouse after treatment with liposome-coated diserpine, according to one embodiment of the present invention, to further verify the protective effect of the drug on ocular surface tissues.

[0015] Figure 4 This is a comparison chart of tear film breakup time and tear secretion after disepine treatment according to one embodiment of the present invention, wherein the left chart is the tear film breakup time and the right chart is the tear secretion. Detailed Implementation

[0016] The present invention will be further described in detail below with reference to specific embodiments.

[0017] I. Experimental Materials and Sources

[0018] 1. Preparation of engineered liposomes

[0019]

[0020] 2. Animal experiments

[0021]

[0022] II. Example 1: Verification of the application of disepped in the preparation of a drug for treating diseases related to lacrimal gland secretion dysfunction, wherein the chemical formula of disepped is shown below:

[0023]

[0024] (I) Preparation of liposomes (LNP)

[0025] 1. Preparation of lipid films:

[0026] Accurately weigh D-Lin-MC3-DMA, distearate-sn-glycerol-3-phosphorylcholine (DSPC), polyethylene glycol 2000 (PEG2000), and cholesterol according to a molar ratio of 50:10:1.5:38.5, ensuring a total mass of 1 mg. Dissolve the above lipid material in 500 μL of chloroform, an organic solvent, and stir until the lipids are completely dissolved.

[0027] The dissolved lipid solution was transferred to a rotary evaporator and subjected to rotary evaporation at 40°C to remove the organic solvent and form a uniform lipid film. The rotary-evaporated lipid film was then placed in a vacuum drying oven and further dried for 2 hours to ensure complete removal of any residual organic solvent.

[0028] 2. Hydration treatment:

[0029] The dried lipid film was placed in an appropriate amount of double-distilled water and hydrated at 65°C for 30 minutes to ensure complete hydration and the formation of primitive liposomes. During the hydration process, a magnetic stirrer can be used to gently stir the film to promote liposome formation.

[0030] 3. Adjust particle size:

[0031] The hydrated liposomes were extruded sequentially through polycarbonate filters of 1.0 μm, 0.8 μm, 0.4 μm, and 0.1 μm, 10 times each time, to gradually reduce the liposome particle size. After extrusion, the particle size distribution of the liposomes was measured using dynamic light scattering (DLS) to ensure that the particle size reached the required nanoscale size (e.g., 100-200 nm).

[0032] If the particle size still does not meet the requirements, the extrusion steps can be repeated until the ideal particle size range is achieved.

[0033] (II) Liposome-coated traditional Chinese medicine monomer disheping

[0034] 1. Solubility of lipids and drugs:

[0035] Accurately weigh D-Lin-MC3-DMA, distearate-sn-glycerol-3-phosphorylcholine (DSPC), polyethylene glycol 2000 (PEG2000), and cholesterol according to a molar ratio of 50:10:1.5:38.5, ensuring a total mass of 1 mg.

[0036] The above-mentioned lipid material and the traditional Chinese medicine monomer disheping were dissolved in 500 μL of organic solvent chloroform. The total lipid mass ratio of the traditional Chinese medicine monomer to the carrier LNP was 1:10. The mixture was stirred evenly to completely dissolve the lipids and drugs, forming a homogeneous lipid-drug solution.

[0037] 2. Formation of a thin lipid membrane:

[0038] The lipid-drug solution was transferred to a rotary evaporator and rotary evaporated at 40°C to remove the organic solvent chloroform, forming a uniform thin lipid film.

[0039] After evaporation, the flask is placed in a vacuum drying oven for 2 hours to ensure complete removal of residual organic solvents.

[0040] 3. Hydrated lipid membrane:

[0041] After drying, place the thin lipid membrane in an appropriate amount of aqueous medium (such as physiological saline or phosphate buffer). The temperature of the aqueous medium should be higher than the phase transition temperature (Tc) of the lipid to promote lipid hydration.

[0042] By gently stirring or oscillating, the lipid membrane is brought into full contact with the aqueous phase, triggering the self-assembly process of lipids to form multilayer liposomes (MLVs).

[0043] 4. Particle size control:

[0044] To obtain smaller and more uniform liposome particle sizes, the formed liposomes were further processed using an extrusion method. The liposomes were extruded through polycarbonate filter membranes of 1 μm, 0.8 μm, 0.4 μm, and 0.1 μm, 10 times each time, until the desired particle size range (e.g., 100-200 nm) was achieved.

[0045] The particle size distribution of liposomes was measured using dynamic light scattering (DLS) to ensure uniform and stable particle size.

[0046] 5. Purification and post-processing:

[0047] The purity and stability of liposomes can be improved by removing unencapsulated drugs, residual organic solvents, and excess aqueous media components through methods such as dialysis, ultrafiltration, or centrifugation.

[0048] Liposomes can also be sterilized as needed to meet the requirements of clinical applications.

[0049] (III) Construction of a dry eye mouse model

[0050] 1. Preparation of laboratory animals:

[0051] Select 6-8 week old C57BL / 6 mice, weighing between 20-25 grams, and acclimatize them for 1 week to ensure they are in good physiological condition before the experiment begins.

[0052] Mice were randomly divided into experimental and control groups, with at least 5 mice in each group, to ensure the reliability and reproducibility of the experimental results.

[0053] 2. Dry eye model induction:

[0054] A dry eye model was induced using a 0.2% benzalkonium chloride (BAK) solution, as follows:

[0055] Twice daily (9:00 AM and 9:00 PM), 5 μL of 0.2% BAK solution was instilled into both eyes of each mouse. This was repeated for 7 consecutive days to simulate the pathological process of dry eye syndrome.

[0056] 3. Model Evaluation:

[0057] After induction, the success of the dry eye mouse model was assessed using the following methods:

[0058] Tear secretion measurement: Tear secretion in mice was measured using the phenol red cotton thread method (CFT). A phenol red cotton thread was inserted into the inner side of the lower eyelid of the mouse, removed after 30 seconds, and the length of the thread wetted by tears was measured to assess tear secretion.

[0059] Tear film breakup time (TBUT) determination: Tear film breakup time (TBUT) was measured using the sodium fluorescein staining method. A 0.5% sodium fluorescein solution was instilled into the conjunctival sac of mouse eyes, and the integrity of the tear film was observed using a cobalt blue slit-lamp microscope. The time when the first tear film breakup point appeared was recorded.

[0060] Corneal fluorescein staining: The integrity of the corneal epithelium was assessed using the fluorescein staining method. A 0.5% fluorescein solution was instilled into the conjunctival sac of the mouse eye, and the corneal staining was observed using a cobalt blue slit-lamp microscope. The area and intensity of the corneal staining were scored.

[0061] (iv) Treatment and efficacy evaluation

[0062] To verify the effectiveness of the traditional Chinese medicine monomer disheping in treating dry eye syndrome, this invention treated a constructed mouse model of dry eye and comprehensively evaluated the efficacy using multiple methods.

[0063] The specific steps are as follows:

[0064] 1. Experimental Groups:

[0065] The constructed dry eye mice were randomly divided into the following groups, with at least 5 mice in each group, to ensure the reliability and reproducibility of the experimental results:

[0066] Control group: treated with physiological saline (PBS).

[0067] Positive control group: treated with 0.05% cyclosporine solution.

[0068] Liposome (LNP) group: treated with drug-free liposomes.

[0069] Drug group: treated with liposomes (LNP) coated with the herbal monomer disheping.

[0070] Healthy control group: A group of age-matched mice without induced dry eye model were introduced as the healthy control group.

[0071] 2. Treatment plan:

[0072] The treatment period was 7 days, twice a day (9 am and 9 pm), with 5 μL of the corresponding treatment solution instilled into both eyes of each mouse.

[0073] Ensure the accuracy and consistency of each eye drop procedure to avoid the influence of human factors on the experimental results.

[0074] 3. Efficacy evaluation indicators:

[0075] After treatment, the efficacy of the treatment in each group of mice was evaluated using the following methods:

[0076] Tear secretion measurement: Tear secretion in mice was measured using the phenol red cotton thread method (CFT). A phenol red cotton thread was inserted into the inner side of the lower eyelid of the mouse, removed after 30 seconds, and the length of the thread wetted by tears was measured to assess tear secretion. Changes in tear secretion in each group were analyzed by comparing with pre-treatment levels and a healthy control group.

[0077] Tear film breakup time (TBUT) was measured using the sodium fluorescein staining method. A 0.5% sodium fluorescein solution was instilled into the conjunctival sac of mice, and the integrity of the tear film was observed using a cobalt blue slit-lamp microscope. The time of the first tear film breakup was recorded. Changes in tear film stability were assessed by comparing the TBUT of different groups of mice.

[0078] Corneal fluorescein staining: The integrity of the corneal epithelium was assessed using fluorescein staining. A 0.5% fluorescein solution was instilled into the conjunctival sac of mice, and the corneal staining was observed using a cobalt blue slit-lamp microscope. The area and intensity of the corneal staining were scored. A lower corneal staining score indicated less corneal epithelial damage and a better treatment outcome.

[0079] Histological analysis: Histological sections and staining (such as hematoxylin-eosin staining) were performed on the eyeballs of some mice to observe pathological changes in tissues such as cornea, conjunctiva and lacrimal gland, and to further evaluate the protective effect of treatment on ocular surface tissues.

[0080] 4. Data Analysis:

[0081] Perform statistical analysis on the collected data and use appropriate statistical methods (such as one-way ANOVA, t-test, etc.) to compare the differences between groups.

[0082] The efficacy of the traditional Chinese medicine monomer disheping in improving dry eye symptoms was analyzed using a p-value less than 0.05 as the criterion for statistical significance.

[0083] To systematically evaluate the therapeutic effect of the traditional Chinese medicine monomer diseping on a dry eye model, this application constructed a mouse model of dry eye due to lacrimal gland dysfunction and intervened with liposome-encapsulated diseping. Comprehensive analysis of multiple physiological and histological indicators showed that diseping treatment significantly improved ocular surface function and structure in the model animals. Please refer to [link to relevant documentation]. Figures 1-4 Specifically, compared with the model control group, the corneal fluorescein staining score of mice in the diserpine treatment group was significantly reduced ( Figure 2 This indicates that corneal epithelial damage has been effectively repaired; simultaneously, the epithelial structure in the corneal tissue is becoming more intact, and inflammatory cell infiltration in the stromal layer is significantly reduced. Figure 3 In addition, the density of goblet cells in the conjunctiva also showed a significant increase. Figure 3 This indicates that the ocular surface mucus secretion function has recovered to some extent. Preliminary observations also found that diserpine treatment tends to improve tear film breakup time and tear secretion volume (e.g., Figure 4 As shown in the figure, this further supports its potential role in maintaining tear film stability and promoting basal tear secretion. These results, from multiple perspectives, confirm that disepine has good therapeutic potential in intervening in dry eye diseases related to lacrimal gland secretion dysfunction.

Claims

1. Application of disepine in the preparation of drugs for treating dry eye syndrome.

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