Application of Phanginin A in the preparation of drugs for the prevention and / or treatment of skin diseases

Abstract

The application belongs to the technical field of medicines, and specifically provides application of Phanginin A in preparation of medicines for preventing and / or treating skin diseases. The skin diseases include atopic dermatitis and skin epidermal barrier damage. The application finds that Phanginin A can reduce atopic dermatitis lesion scores, improve atopic dermatitis skin pathological thickening and inflammatory cell infiltration, reduce spleen inflammatory response, inhibit volume increase, promote thymus function recovery, down-regulate expression of atopic dermatitis related inflammatory factors IL-6 and TNF-alpha, and can be used for preventing and / or treating atopic dermatitis. In addition, the application also finds that Phanginin A can promote keratinocyte scratch healing, and can be used for repairing skin epidermal barrier damage.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical technology, specifically relating to the application of Phanginin A in the preparation of drugs for the prevention and / or treatment of skin diseases, particularly in the preparation of drugs for the repair of atopic dermatitis and skin epidermal barrier damage. Background Technology

[0002] The skin, the largest organ in the human body, plays a vital role in protection, excretion, temperature regulation, and sensing external stimuli. However, the skin is also susceptible to various factors, leading to a variety of skin diseases, such as atopic dermatitis and damage to the skin's epidermal barrier. Atopic dermatitis (AD) is a chronic, relapsing, pruritic inflammatory skin disease characterized by impaired skin barrier function, immune dysregulation, and abnormal neurosensory function. Its typical clinical manifestations include polymorphic skin lesions such as erythema, papules, exudative eczematous lesions, lichenification, and intense itching, often accompanied by dry skin and hyperresponsiveness (a vicious cycle of itching and scratching). AD shows a significant bimodal age distribution, with 60% of cases beginning in infancy and 30% persisting into adulthood. Its global incidence has increased 2-3 times in the past 20 years, and it has become one of the leading health problems in terms of disease burden among non-fatal diseases. The pathogenesis of AD is complex, involving multidimensional interactions. The causes of atopic dermatitis include: ① Genetic susceptibility: Approximately 30% of patients have filaggrin gene mutations leading to filaggrin deficiency and disruption of the skin's physical barrier; polymorphisms in genes related to the Th2 / Th22 / Th17 (Th helper T cells) immune pathway drive immune imbalance. ② Disruption of the immune-barrier-microbe axis: Damage to the epidermal barrier allows allergens to penetrate, triggering type 2 inflammation (interleukin-4 / interleukin-13) and neuroimmune activation (interleukin-33 and nerve growth factor mediate pruritus signals); Staphylococcus aureus colonization further exacerbates inflammation. ③ Environmental and neuropsychological factors: Climate change, pollutants, and psychological stress affect disease progression through epigenetic modifications. AD patients often have comorbid allergic diseases (asthma, allergic rhinitis, food allergies) and immunodeficiency, and chronic itching significantly increases the risk of anxiety, depression, and sleep disorders, constituting a multidimensional health burden. Therefore, there is an urgent need for safe and effective drug formulations for the treatment of atopic dermatitis.

[0003] Phanginin A is a kazonesane diterpenoid compound that has been shown to possess a variety of biological activities. It is an effective orally active salt-induced kinase 1 (SIK1) activator that can promote skeletal muscle glucose uptake, lower blood glucose levels, improve glucose tolerance and dyslipidemia by inhibiting gluconeogenesis, increasing p-SIK1 expression and decreasing p-CREB expression, thus showing potential for the treatment of type 2 diabetes.

[0004] Currently, there are no reports of SIK1 as a drug target for atopic dermatitis, no reports of SIK1 activators inhibiting atopic dermatitis, and no reports of Phanginin A in the treatment of atopic dermatitis and skin epidermal barrier damage. Summary of the Invention

[0005] To address the problems of existing technologies, this invention provides the use of Phanginin A in the preparation of drugs for the prevention and / or treatment of skin diseases, particularly in the preparation of drugs for the repair of atopic dermatitis and skin epidermal barrier damage.

[0006] This invention is achieved through the following technical solution:

[0007] First, the present invention provides the use of Phanginin A or a pharmaceutically acceptable salt thereof in the preparation of medicaments for the prevention and / or treatment of skin diseases.

[0008] Phanginin A is derived from *Caesalpinia sappan*, a plant belonging to the genus *Caesalpinia* in the legume family. It is a kazoline diterpenoid compound with the CAS number 1011528-58-7 and the molecular formula C2. 21 H 28 O5, its chemical structural formula is shown below:

[0009]

[0010] Preferably, the pharmaceutically acceptable salt includes at least one of the following: lithium, potassium, and sodium salts formed with alkali metals; calcium and magnesium salts formed with alkaline earth metals; hydrochlorides, sulfates, and phosphates formed with inorganic acids; and tartrates, maleates, succinates, citrates, p-toluenesulfonates, butyrates, camphorsulfonates, and methanesulfonates formed with organic acids.

[0011] Preferably, the skin disease includes atopic dermatitis and damage to the skin's epidermal barrier.

[0012] Preferably, the drug can reduce the skin lesion score of atopic dermatitis, improve the pathological thickening of the skin and the infiltration of inflammatory cells in the skin, reduce the inflammatory response of the spleen, inhibit its enlargement, and promote the recovery of thymus function.

[0013] Preferably, the drug can downregulate the expression of atopic dermatitis-related inflammatory factors IL-6 and TNF-α.

[0014] Preferably, the drug can promote the healing of keratinocyte scratches.

[0015] Preferably, the dosage of Phanginin A or its pharmaceutically acceptable salt is 6.09–12.19 mg / kg based on body weight. This dosage is the human equivalent based on mouse experimental doses.

[0016] Preferably, the drug further includes pharmaceutically acceptable excipients.

[0017] Preferably, the pharmaceutically acceptable excipients include any one or a combination of at least two of the following: sustained-release agents, excipients, fillers, binders, wetting agents, disintegrants, absorption promoters, surfactants, and lubricants. The combination of at least two is, for example, a combination of binders and excipients, a combination of binders and flavoring agents, a combination of binders and fillers, etc. Other combinations are also possible and will not be elaborated here.

[0018] Preferably, the drug dosage form is a tablet, capsule, granule, injection, oral liquid, pill, ointment, suspension, dispersant, syrup, suppository, gel, aerosol, or patch.

[0019] Preferably, the drug is administered orally, via non-gastrointestinal route, via local administration, or via an implanted drug storage device.

[0020] Secondly, the present invention provides a pharmaceutical composition for the prevention or treatment of skin diseases, comprising Phanginin A or a pharmaceutically acceptable salt thereof and pharmaceutical excipients that can be used in dermatological drugs.

[0021] Preferably, the pharmaceutical composition contains 0.001 to 99 wt% of Phanginin A or a pharmaceutically acceptable salt thereof.

[0022] Beneficial effects of this invention:

[0023] This invention marks the first discovery of a novel application of Phanginin A and its pharmaceutically acceptable salts in the preparation of drugs for the prevention and / or treatment of skin diseases, particularly in the prevention and / or treatment of atopic dermatitis and damage to the skin's epidermal barrier. Experiments conducted in this invention have shown that Phanginin A can improve 2,4-dinitrochlorobenzene (DNCB)-induced atopic dermatitis in mice, reduce atopic dermatitis lesion scores, improve pathological thickening and inflammatory cell infiltration in the skin, alleviate splenic inflammation and inhibit its enlargement, promote thymus function recovery, and downregulate the expression of atopic dermatitis-related inflammatory factors IL-6 and TNF-α, demonstrating the potential of Phanginin A in improving inflammatory skin diseases such as atopic dermatitis. Furthermore, Phanginin A can promote keratinocyte scratch fusion, indicating its potential for repairing skin barrier damage. Attached Figure Description

[0024] Figure 1 The effect of Phanginin A on skin lesions in AD model mice.

[0025] Figure 2 The effect of Phanginin A on the histopathological changes of skin tissue in AD model mice (HE staining).

[0026] Figure 3 The results of the effect of Phanginin A on the expression of the related inflammatory factor IL-6 in the skin tissue of AD model mice (ELISA method). "" indicates that compared with the model group, P < 0.05. "" indicates that compared with the model group, P < 0.01. "P < 0.001" indicates that compared to the model group.

[0027] Figure 4 The results of the effect of Phanginin A on the expression of the related inflammatory factor TNF-α in the skin tissue of AD model mice (ELISA method). "" indicates that compared with the model group, P < 0.05. "P < 0.01" indicates that compared to the model group.

[0028] Figure 5 The effect of Phanginin A on the relative expression level of IL-6 mRNA in skin tissue of AD model mice (RT-qPCR method, "#" indicates P<0.05 compared with the control group; "" indicates P<0.05. "" indicates that compared with the model group, P < 0.01. "P < 0.001" indicates that compared to the model group.

[0029] Figure 6 The effect of Phanginin A on the relative expression level of TNF-α mRNA in skin tissue of AD model mice (RT-qPCR method, "#" indicates P<0.05 compared with the control group; "" indicates P<0.05. "" indicates that compared with the model group, P < 0.05. "" indicates that compared with the model group, P < 0.01. "P < 0.001" indicates that compared to the model group.

[0030] Figure 7 The results of the cell scratch assay of Phanginin A on HaCaT cells. Detailed Implementation

[0031] The present invention will now be described in further detail with reference to the accompanying drawings and embodiments, but this does not limit the present invention in any way. Any modifications or improvements made based on the teachings of the present invention shall fall within the protection scope of the present invention.

[0032] The processes, conditions, reagents, and experimental methods used in implementing this invention, except as specifically mentioned below, are all common knowledge and general knowledge in the field, and this invention does not have any particular limitations. Experimental methods in the embodiments that do not specify specific conditions are generally performed under conventional conditions or as recommended by the manufacturer.

[0033] Unless otherwise stated, all technical terms and scientific terms used in this specification have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. However, in the event of any conflict, the specification containing the definitions shall prevail.

[0034] The following are some of the sources of reagents and consumables in the examples:

[0035] Sappanwood ( Biancaea sappan Seeds of *L.) Tod.* were collected in Baise, Guangxi in July 2014 and identified by the Kunming Institute of Botany, Chinese Academy of Sciences. Specimen (20140712c01) is deposited at the State Key Laboratory of Phytochemistry and Sustainable Utilization of Western Resources, Kunming Institute of Botany. 2,4-Dinitrochlorobenzene (DNCB) (Sigma-Aldrich, USA); Dexamethasone acetate (Shanghai Aomei Biotechnology Co., Ltd.); BALB / c female mice (Beijing Spefol Biotechnology Co., Ltd.); HaCaT cells (Wuhan Pronosei Biotechnology Co., Ltd.); Strong RIPA lysis buffer (Shanghai Beyotime Biotechnology Co., Ltd.); ELISA kit (Hangzhou Lianke Biotechnology Co., Ltd.); Trizol reagent (Abixin Shanghai Biotechnology Co., Ltd.); Real-time quantitative PCR premixed reagent (Beijing Kangrun Chengye Biotechnology Co., Ltd.).

[0036] Example 1: Preparation of compound Phanginin A

[0037] Sappanwood ( Biancaea sappan(L.) Tod.) seeds (11 kg) were air-dried and pulverized, then extracted three times at room temperature with 95% ethanol (3 × 50 L, v / v, 48 h each time), and filtered. The filtrate was concentrated under reduced pressure to obtain a crude extract (2.1 kg), which was then suspended in water and extracted sequentially with ethyl acetate (3 × 6 L). The ethyl acetate phase (1.2 kg) was subjected to 100-200 mesh silica gel column chromatography, eluted with a petroleum ether-acetone gradient (9:1 → 1:1, v / v) to obtain five fractions (AE). Fraction C (120 g) was separated by medium-pressure preparative liquid chromatography (MCI gel column) with a methanol-water gradient (50:50 → 100:0) to obtain six subfractions (C1-C6). Fraction C2 (35 g) was repeatedly purified by silica gel column chromatography (eluent: petroleum ether-ethyl acetate 9.5:0.5 → 7:3) to obtain four secondary fractions (C2.1-C2.4). Secondary component C2.2 (15 g) was recrystallized from methanol to obtain Phanginin A (10 g) with a purity of 97%.

[0038] Experimental Example 2: The therapeutic effect of Phanginin A on 2,4-dinitrochlorobenzene (DNCB)-induced atopic dermatitis in mice.

[0039] 1 Experimental Methods

[0040] 1.1 Animal model establishment and drug administration method

[0041] Female BALB / c mice (6-8 weeks old, 20±2 g) were acclimatized for one week in an SPF-grade animal laboratory and then randomly divided into 5 groups (n=5) according to body weight: blank control group, AD model group, positive control group (dexamethasone acetate, 1 mg / kg), low-dose Phanginin A group (PA-L, 75 mg / kg), and high-dose Phanginin A group (PA-H, 150 mg / kg). Before modeling, a bare area of ​​approximately 2.5 cm × 2.5 cm was prepared on the back of the mice using a shaving tool and depilatory cream. Except for the blank control group, the other groups were induced to establish AD models using DNCB. The specific steps are as follows: First, sensitization was performed for 2 consecutive days using 2% DNCB solution (dissolved in a blank matrix solution of acetone:olite = 4:1, 100 μL / mouse); from day 7, stimulation was performed using 0.5% DNCB solution (100 μL / mouse) every 2 days until the end of the experiment on day 15, at which time samples were collected. During the same period, mice in the blank control group were treated with an equal amount (100 μL / mouse) of blank matrix solution at the same site.

[0042] From day 1 of modeling, each group was administered the corresponding drug solution by gavage at a volume of 0.1 mL / 10 g (drug volume / mouse body weight). The blank control group and AD model group were administered 0.25% (w / v) sodium carboxymethyl cellulose (CMC-Na) solution by gavage. The low-dose Phanginin A group (dose, 75 mg / kg) was administered 7.5 mg / mL Phanginin A (prepared with 0.25% CMC-Na) by gavage. The high-dose Phanginin A group (dose, 150 mg / kg) was administered 15 mg / mL Phanginin A (prepared with 0.25% CMC-Na) by gavage. The positive control group (dose, 1 mg / kg) was administered 0.1 mg / mL dexamethasone acetate (prepared with 0.25% CMC-Na) by gavage. The administration frequency was once daily for 14 consecutive days.

[0043] 1.3 Photographs and scoring of skin lesions

[0044] Photographs of skin lesions: On days 7 and 14 after the establishment of the AD mouse model by DNCB, photographs of skin lesions on the back of the mice were taken and recorded.

[0045] Skin lesion scoring: Starting from day 4 of modeling, the inflammation of the skin on the back of mice was observed and scored every two days. The sum of the scores for erythema, edema, epidermal abrasion, and scaling was the total score. The specific scoring criteria are shown in Table 1.

[0046] Table 1 Scoring Indicators and Grading Standards

[0047]

[0048] 1.4 Animal sampling and preservation

[0049] Animal tissue was harvested on day 15 of modeling. Mice were fasted for 12 hours after the last drug administration, but water was permitted. After euthanizing the mice, they were fixed on a sterile animal-specific operating table for tissue harvesting. Skin tissue of approximately 1 cm × 1 cm from the hair-removed area was harvested, trimmed neatly, and placed in pre-labeled sterile centrifuge tubes containing 4% paraformaldehyde. The tubes were stored at room temperature for subsequent tissue sectioning. The remaining skin tissue from the hair-removed area was placed in cryopreservation tubes, sealed, flash-frozen in liquid nitrogen, and stored at -80°C.

[0050] Remove the spleen and thymus, and remove any excess surrounding tissue. Weigh them using a precision balance, record the weight, and calculate their organ index (organ index (mg / g) = organ weight (mg) / body weight (g)). ±s).

[0051] 1.5 HE staining

[0052] ①Dewaxing and rehydration of paraffin sections: Mouse skin tissue was fixed, embedded in paraffin, and sectioned. It was then immersed in environmentally friendly dewaxing solution I and environmentally friendly dewaxing solution II for 20 min each. Subsequently, it was treated with anhydrous ethanol I, anhydrous ethanol II and 75% ethanol for 5 min each. Finally, it was rinsed with running tap water to complete the hydration.

[0053] ② Frozen sections were warmed and fixed: Frozen sections were taken out of the -20℃ freezer and brought to room temperature. They were fixed with tissue fixative for 15 min and then rinsed with running water.

[0054] ③ Hematoxylin staining: Immerse the sections in hematoxylin staining solution for 3-5 min, wash with tap water, differentiate with differentiation solution, wash with tap water, blue back solution, and rinse with running water.

[0055] ④ Eosin staining: The sections were dehydrated in 85% and 95% graded alcohol for 5 min each, and then stained in eosin dye for 5 min.

[0056] ⑤ Dehydration, clearing and mounting: The sections were dehydrated in anhydrous ethanol I, anhydrous ethanol II and anhydrous ethanol III for 5 min each; then cleared in xylene I and xylene II for 5 min each, and finally mounted with neutral resin.

[0057] ⑥ Microscopic examination: Images were acquired and analyzed to examine the pathological changes in each group of mice. The cell nucleus appeared blue, and the cytoplasm appeared red.

[0058] 1.6 ELISA method for detecting the expression of inflammatory factors IL-6 and TNF-α in skin lesions

[0059] Skin tissue samples were taken from -80℃, treated with pre-cooled 1× PBS buffer, washed to remove blood, and excess water was absorbed with absorbent paper. The tissue was placed in a 1.5 mL sterile centrifuge tube, and the skin tissue was thoroughly minced. Lysis buffer was added (100 μL of strong RIPA lysis buffer per 10 mg of tissue). A grinding bead was added to the centrifuge tube, and the tissue was ground at low temperature (70 Hz, 60 s / time, 4–5 times). The homogenized tissue was allowed to stand at low temperature for 30 min for lysis, then centrifuged at 12000 rpm for 15 min. The protein supernatant was collected, and the protein concentration was determined by BCA method. The expression of related inflammatory factors IL-6 and TNF-α in the skin tissue was detected by ELISA.

[0060] 1.7 Real-time quantitative PCR detection of relative expression levels of IL-6 and TNF-α mRNA in skin tissue

[0061] RNA extraction: Weigh 20 mg of tissue, treat with pre-chilled 1× PBS buffer, and place in a 1.5 mL sterile centrifuge tube. Mince thoroughly, add 1 mL of Trizol, and grind thoroughly (70 Hz, 60 s / cycle, 3-5 times). Incubate on ice for 10 min, add 20% chloroform, mix well, incubate on ice for 10 min, centrifuge at 12000 rpm / 15 min / 4℃, collect the supernatant, add an equal volume of isopropanol, and incubate for 10 min. Centrifuge at 12000 rpm / 10 min / 4℃, discard the supernatant, and retain the precipitate. Add 1 mL of 75% ethanol to each tube, invert once to allow the RNA to float. Centrifuge at 12000 rpm / 5 min / 4℃. Repeat twice. Aspirate the ethanol and evaporate to dryness, then add enzyme-free water (20-50 μL) to dissolve the RNA.

[0062] Reverse transcription: The reaction system (20 μL) consisted of Starscript III All-in-one RT Mix (1 μL), 5×Starscript III All-in-one RT Buffer (4 μL), sample RNA (1 μg), and enzyme-free water. The reaction program was set sequentially as follows: 37℃ for 2 min, 50℃ for 15 min, and 85℃ for 2 min. cDNA was stored at -20℃.

[0063] Real-time quantitative PCR: The reaction system (20 μL) consisted of cDNA (2 μL), 2×RealStar Fast SYBR qPCRMix (10 μL), each of the forward and reverse primers (0.5 μL each) (primer nucleic acid sequences are shown in Table 2), and enzyme-free water (7 μL). The PCR reaction cycling conditions were as follows: pre-reaction (95℃ for 2 min), 40 cycles (95℃ for 15 s, 60℃ for 30 s), and melting phase (95℃ for 15 s, 60℃ for 1 min, 95℃ for 15 s).

[0064] Table 2 Primers for Real-Time Quantitative PCR

[0065]

[0066] 1.8 Statistical Analysis

[0067] All data are expressed as mean ± standard deviation and follow a normal distribution. Data analysis was performed using SPSS software, and one-way ANOVA was used for statistical analysis. A p-value < 0.05 was considered statistically significant.

[0068] 2. Experimental Results

[0069] 2.1 Effects of Phanginin A on skin lesions in AD model mice

[0070] The effects of Phanginin A on skin lesions in AD model mice are as follows: Figure 1 As shown, this experiment used DNCB to induce AD ​​mice. During the experimental sensitization period, the mice in the model group showed severe edema, erythema, scaling, and epidermal erosion on their backs. From the time of drug administration to the final sampling stage, the AD model group mice showed obvious inflammatory manifestations such as skin erythema, edema, scaling, erosion, and epidermal shedding. Compared with the AD group, the above symptoms were significantly improved in the treatment groups with different doses of drugs. The area of ​​erythema crusting was significantly reduced, and the new hair growth was significantly increased. By day 12-14 of modeling, only some scales remained on the skin of the mice in the drug-treated group.

[0071] 2.2 Effect of Phanginin A on the skin lesion severity score in AD model mice

[0072] Table 3 shows the severity scores of skin lesions in AD model mice after Phanginin A intervention. The scores were taken on day 4 after the sensitization period of days 1 and 2, and on day 8 after the first stimulation on day 7. The results showed that the skin lesions in the model group showed a tendency to heal naturally on day 12. The improvement in skin lesion scores in the Phanginin A intervention group was more significant and statistically significant.

[0073] Table 3. Skin lesion scores on the backs of mice in each experimental group ( ±s, n=5)

[0074]

[0075] Note: Compared with the control group, ###P<0.001; compared with the model group, P<0.05, P<0.01, P<0.001.

[0076] 2.3 Effects of Phanginin A on organ indices in AD model mice

[0077] The effects of Phanginin A on organ indices in AD model mice are shown in Table 4. After continuous DNCB stimulation, the spleen index of AD model mice increased significantly compared with the blank control group (P<0.01), while the spleen index decreased in the Phanginin A group. The spleen is an important organ involved in the immune response; its weight increases during inflammation. Phanginin A can reduce the spleen index, indicating that it may have anti-inflammatory effects. Compared with the blank control group, the thymus index of AD model mice decreased significantly (P<0.05). After Phanginin A intervention, the thymus index of mice recovered. The thymus index is an important indicator for assessing immune imbalance in AD. The fact that Phanginin A can increase the thymus index in AD mice indicates that Phanginin A promotes the recovery of thymic function in atopic dermatitis.

[0078] Table 4 Comparison of organ indices in mice of different experimental groups ( ±s, n=5)

[0079]

[0080] Note: Compared with the control group, #P<0.05, ##P<0.01; compared with the model group, P<0.05, P<0.01.

[0081] 2.4 Effects of Phanginin A on histopathological changes in skin tissue of AD model mice

[0082] The effects of Phanginin A on the histopathological changes of skin tissue in AD model mice (HE staining) are as follows: Figure 2 As shown, the results indicated that the skin tissue of mice in the blank control group was smooth and without inflammatory infiltration. Compared with the blank control group, the AD model group showed significant thickening of the epidermis and dermis, hyperkeratosis, and significant inflammatory cell infiltration in the dermis. Compared with the AD model group, the epidermis and dermis of mice in the low-dose Phanginin A group and the high-dose Phanginin A group were significantly thinner, and the inflammatory cell infiltration in the dermis was reduced. This demonstrates that Phanginin A can improve pathological thickening of the skin and inflammatory cell infiltration in the skin, thus playing a role in improving AD.

[0083] 2.5 Effects of Phanginin A on the expression levels of inflammatory factors IL-6 and TNF-α

[0084] The results of ELISA detection of Phanginin A's effect on IL-6 and TNF-α expression in skin tissue of AD model mice are as follows: Figure 3 and Figure 4 As shown, the results indicated that compared with the blank control group, the levels of IL-6 and TNF-α were increased in the model group; compared with the model group, the levels of IL-6 and TNF-α were decreased in the Phanginin A dosage group. These results suggest that Phanginin A can improve inflammation by downregulating the expression of AD-related inflammatory factors.

[0085] 2.6 Effects of Phanginin A on IL-6 and TNF-α mRNA expression

[0086] The results of RT-qPCR detection of IL-6 and TNF-α mRNA expression in skin tissue of AD model mice by Phanginin A are as follows: Figure 5 and Figure 6 As shown, the results of reverse transcription and real-time quantitative PCR detection of RNA extracted from skin tissue showed that, compared with the blank control group, the relative expression levels of IL-6 and TNF-α mRNA in the model group were increased; compared with the model group, the relative expression levels of IL-6 and TNF-α mRNA in the Phanginin A dose group were decreased, suggesting that the drug has a therapeutic effect on atopic dermatitis.

[0087] Example 3: Experiment on Phanginin A promoting keratinocyte scratch healing

[0088] HaCaT cells are immortalized human keratinocytes that retain the differentiation and barrier function characteristics of epidermal cells, and are widely used in studies simulating skin barrier repair and wound healing. The HaCaT cell scratch assay simulates skin barrier damage, and the scratch closure rate is observed under a microscope to reflect the barrier repair capacity.

[0089] 1. Experimental Procedure

[0090] HaCaT cells with good morphology and in the logarithmic growth phase were prepared in complete culture medium (containing 10% FBS and 1% penicillin antibiotics) at a concentration of 1.25 × 10⁶ cells / mL. 5 Cell suspensions of 10 cells / mL were seeded into scratching inserts of 24-well plates. After the cells adhered and formed scratches, the inserts were removed and the cells were treated with different concentrations (18.75 μmol / mL and 37 μmol / mL) of Phanginin A for 12 h. The cells were then observed and photographed under an inverted microscope and processed using ImageJ software.

[0091] 2 Experimental Results

[0092] The results are as follows Figure 7As shown, the results indicate that Phanginin A treatment can promote keratinocyte scratch healing, suggesting that Phanginin A has the potential to prevent and / or treat skin barrier damage.

[0093] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. The use of Phanginin A or a pharmaceutically acceptable salt thereof in the preparation of medicaments for the treatment of atopic dermatitis.

2. The application according to claim 1, characterized in that, The drug can repair damage to the skin's epidermal barrier.

3. The application according to claim 1, characterized in that, The drug can reduce the skin lesion score of atopic dermatitis, improve the pathological thickening of the skin and the infiltration of inflammatory cells in the skin, reduce the inflammatory response of the spleen, inhibit its enlargement, and promote the recovery of thymus function.

4. The application according to claim 1, characterized in that, The drug can downregulate the expression of atopic dermatitis-related inflammatory factors IL-6 and TNF-α.

5. The application according to claim 1, characterized in that, The drug can promote the healing of keratinocyte scratches.

6. The application according to claim 1, characterized in that, The dosage of Phanginin A or its pharmaceutically acceptable saline solution is 6.09–12.19 mg / kg by body weight.

7. The application according to any one of claims 1-6, characterized in that, The drug also includes pharmaceutically acceptable excipients.

8. The application according to any one of claims 1-6, characterized in that, The drug dosage forms are tablets, capsules, granules, injections, oral liquids, pills, ointments, suspensions, dispersants, syrups, suppositories, gels, aerosols, or patches.

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