Application of cajanin in preparation of medicine for treating psoriasis and acne

Abstract

The invention provides an application of cajanin in preparation of a medicine for treating psoriasis and acne, and belongs to the technical field of traditional Chinese medicines. The invention finds that the longistyline can significantly inhibit proliferation of keratinocytes and secretion of inflammatory factors IL-6 and IL-8 in vitro, significantly inhibit psoriasis skin lesion, significantly reduce psoriasis injury area and severity index score, and significantly reduce epidermis thickness and Ki67 expression in psoriasis skin tissue; in addition, the longistyline can also significantly inhibit the formation of acnes, and can reduce the mRNA expression of acne-related inflammatory factors IL-6, IL-1alpha, IL-1beta, TNF-alpha and IL-23 in skin tissues. The cajanin is used as an effective component, the psoriasis and the acne can be effectively treated in an external application mode, and a new thought is provided for treatment of the psoriasis and the acne.

Description

Technical Field

[0001] The invention belongs to the technical field of traditional Chinese medicine, and in particular relates to the application of a traditional Chinese medicine monomer, cajanin, in preparing drugs for treating psoriasis and acne. Background Art

[0002] Psoriasis is a common chronic, inflammatory skin disease characterized by obvious erythema, scaling and itching on the skin, which seriously affects the quality of life of patients. Although there are many treatment options, such as traditional topical medications, oral medications, phototherapy and various biological agents in recent years, these treatments are often accompanied by poor treatment effects, side effects or high prices, and recurrence after drug discontinuation. Therefore, more new and effective drugs for the treatment of psoriasis are still urgently needed in clinical practice.

[0003] Acne is a chronic inflammatory skin disease of the hair follicles and sebaceous glands, which mainly occurs in adolescents and has a significant impact on the patient's appearance and mental health. Its pathogenesis is relatively complex, involving multiple links such as increased androgen levels, vigorous sebaceous gland secretion, abnormal keratinization of the hair follicles and sebaceous gland ducts, the reproduction of Propionibacterium acnes, and inflammatory reactions. Common clinical manifestations include a variety of skin lesions such as acne, papules, pustules, nodules, and cysts. Although there are currently a variety of treatment methods, such as retinoic acid drugs, antibiotics, and anti-androgen drugs, these drugs also have many limitations. For example, retinoic acid drugs may cause adverse reactions such as skin irritation, dryness, and desquamation, and long-term use of antibiotics can easily lead to bacterial resistance. Moreover, some patients are still prone to relapse after treatment, and existing treatment options are difficult to achieve long-term and stable treatment effects. The treatment of acne also faces huge challenges, and there is an urgent need to explore safer and more effective treatment methods.

[0004] Traditional Chinese medicine has a long history in the treatment of psoriasis and acne. Cajanine, as a natural phenolic compound, has antiviral, anti-tumor, osteoclast differentiation and natural killer cell function regulation effects. So far, there are no reports on the research of cajanine and its related preparations in the treatment of psoriasis and acne. Summary of the invention

[0005] In view of this, the purpose of the present invention is to provide the use of the Chinese medicine monomer caesalpin in the preparation of psoriasis and acne treatment drugs, so as to provide a new idea for the treatment of psoriasis and acne.

[0006] The present invention provides the use of a Chinese medicine monomer, caesalpin, in the preparation of a psoriasis treatment drug. Preferably, the caesalpin can significantly inhibit the proliferation of keratinocytes and the secretion of inflammatory factors IL-6 and IL-8 in vitro, significantly inhibit psoriasis skin lesions, significantly reduce the psoriasis lesion area and reduce the severity index score, and further analysis shows that caesalpin can significantly reduce the thickness of the epidermis and the expression of Ki67 in psoriasis skin tissue.

[0007] The present invention also provides the use of the Chinese medicine monomer caesalpin in the preparation of acne treatment drugs. Preferably, the caesalpin can significantly inhibit the formation of acne and reduce the mRNA expression of acne-related inflammatory factors IL-6, IL-1α, IL-1β, TNF-α and IL-23 in skin tissue.

[0008] The invention provides an external preparation for treating psoriasis and acne, wherein the active ingredient of the external preparation comprises cajanin.

[0009] Preferably, the content of cajanin in the external preparation is 5 mg / mL.

[0010] Preferably, the external preparation further comprises a solvent, a dispersant, an emulsifier and water.

[0011] Preferably, the external preparation is any one of a cream, an ointment, a lotion, a gel or a spray.

[0012] Compared with the prior art, the present invention has the following beneficial effects:

[0013] The present invention is the first to discover that caesalpinol has significant effects in treating psoriasis and acne, providing a new drug option for the treatment of these two common skin diseases. Especially for patients with poor results or more side effects from existing treatments, caesalpinol topical preparations are expected to become a potential, safe and effective alternative treatment method.

[0014] In the treatment of psoriasis, caesalpin can prevent the chemotaxis and aggregation of inflammatory cells, further reducing the pathological changes at the site of inflammation. In addition, the inhibitory effect of caesalpin on Ki67 expression can effectively control the excessive proliferation of keratinocytes, improve the symptoms of abnormal skin keratinization in patients with psoriasis from the root, and promote normal repair and regeneration of the skin. For the treatment of acne, caesalpin can reduce the expression of a variety of acne-related inflammatory factors (such as IL-6, IL-8, TNF-α, IL-1β), and comprehensively inhibit the inflammatory response during the onset of acne. By regulating the secretion of inflammatory factors, caesalpin helps to reduce inflammation caused by Propionibacterium acnes infection, reduce sebaceous gland secretion, and improve abnormal keratinization of hair follicles and sebaceous gland ducts, thereby effectively preventing and treating various skin lesions of acne, such as comedones, papules, pustules, etc.

[0015] As a natural phenolic compound, caesalpinol has the advantages of wide sources and relatively low cost. Its natural properties also make it easier for patients to accept psychologically, while reducing the potential risks of chemical synthetic drugs, which is in line with the modern trend of people's pursuit of natural drugs. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 The figure shows the inhibitory effect of caesalpin on the proliferation of M5-induced psoriasis cell model cells. In the figure, A is cultured for 24 hours, and B is cultured for 48 hours;

[0017] Figure 2 The figure shows the inhibitory effect of caesalpin on the secretion of inflammatory factors in the M5-induced psoriasis cell model. In the figure, A is the content of IL-6, and B is the content of IL-8;

[0018] Figure 3 The figure shows the therapeutic effect of caesalpinol on psoriasis model mice, where A is the skin lesions of mice in each group, and B is the Psoriasis area and severity index (PASI) score of mice in each group;

[0019] Figure 4 The effect of caesalpin on the skin thickness and keratinocyte proliferation of psoriasis model mice, where A is HE staining of skin tissue of mice in each group, B is the thickness of epidermis of the back skin of mice, C is Ki67 staining of skin tissue of mice, and D is the proportion of Ki67 positive cells;

[0020] Figure 5 The effect of caesalpin on acne formation in the skin of acne model mice, where A is the skin appearance of mice in each group, and B is HE staining of the skin tissue of mice in each group;

[0021] Figure 6 The effect of cajanin on the expression of skin-related inflammatory factors (IL-6, IL-1β, TNF-α, IL-1α and IL-23) in acne model mice. DETAILED DESCRIPTION

[0022] The present invention provides the use of caesalpin in the preparation of a psoriasis treatment drug. Caesalpin can significantly inhibit the proliferation of keratinocytes and the secretion of inflammatory factors IL-6 and IL-8 in vitro, significantly inhibit psoriasis skin lesions, significantly reduce the psoriasis lesion area and reduce the severity index score. Further analysis shows that caesalpin can significantly reduce the thickness of the epidermis and the expression of Ki67 in psoriasis skin tissue.

[0023] The present invention also provides the use of caesalpin in the preparation of acne treatment drugs. Caesalpin can significantly inhibit the formation of acne and reduce the mRNA expression of acne-related inflammatory factors IL-6, IL-1α, IL-1β, TNF-α and IL-23 in skin tissue.

[0024] Cajanine is a monomer compound extracted from the leaves of the natural plant pigeon pea. Its structure is similar to diethylstilbestrol. It is a phenolic hydroxyl compound with the molecular formula: C 21 H 22 O4, which includes a benzene ring structure and multiple hydroxyl groups (-OH) and a keto group (C=O).

[0025]

[0026] The present invention also provides an external preparation for treating psoriasis and acne, wherein the active ingredient of the external preparation includes cajanin. Preferably, the content of cajanin in the external preparation is 5 mg / mL.

[0027] The external preparation of the present invention preferably further comprises a solvent, a dispersant, an emulsifier and water; further preferably, the solvent is selected from one or more of dimethyl sulfoxide, ethanol and methanol, the dispersant is selected from one or more of polyethylene glycol, polyvinyl pyrrolidone, sodium polyacrylate and lignin sulfonate, and the emulsifier is selected from one or more of Tween series (Tween 20, Tween 60, Tween 80), sodium lauryl sulfate and lecithin.

[0028] The present invention also provides a method for preparing the topical preparation, wherein the caesin is weighed and dissolved in a solvent to prepare a mother liquor, a dispersant, an emulsifier and water are sequentially added to the mother liquor, and the topical preparation is obtained by mixing and clarifying. As an implementation method, the caesin powder is dissolved in a solvent DMSO solution to prepare a mother liquor, and a dispersant PEG300, an emulsifier Tween 80 and water are sequentially added to the mother liquor, and the mixture is mixed and clarified.

[0029] In the present invention, the external preparation is preferably any one of a cream, an ointment, a lotion, a gel or a spray. The present invention has no particular limitation on the specific preparation process of the above dosage form, and the commonly used preparation process in the art can be used.

[0030] The technical solutions provided by the present invention are described in detail below in conjunction with the embodiments, but they should not be construed as limiting the protection scope of the present invention.

[0031] Example 1

[0032] Effects of cajanin on proliferation of keratinocytes in M5-induced psoriasis cell model

[0033] Experimental method: HaCaT cells in the logarithmic growth phase were taken and pipetted to form a single cell suspension. After cell counting, 6.0×10 3 The cells were seeded into a 96-well plate at a density of 100 μg / well and placed in a cell culture incubator for 24 hours to adhere to the wall. The cells were then replaced with a culture medium containing 5 μM, 10 μM, 20 μM, 40 μM, and 50 μM quinolin, and a control group without drugs was set up. After 3 hours of culture, the culture medium in the M5 group was added with M5 cytokines (including IL-17A, IL-22, oncostatin M (OSM), TNF-α, IL-1α) to make the final concentration 10 ng / mL to establish a psoriasis cell model. After the above cells were cultured in a 37°C incubator for 24 hours and 48 hours, 20 μL of MTT solution was added to each well of the cells, and the OD value of each well at 450 nm was measured using an enzyme marker after incubation for 2 hours in a cell culture incubator. The cell viability was calculated according to the following formula:

[0034] Cell viability (%) = (OD value of drug administration group - OD value of blank well) / (OD value of control group - OD value of blank well) × 100%, wherein the OD value of the blank well is the OD value of the cell-free well in DMEM medium containing 20uL MTT test solution at 450nm.

[0035] Test results: Figure 1 As shown, 50 μM cajanin could significantly inhibit the proliferation of HaCaT cells induced by M5.

[0036] Example 2

[0037] Effect of cajanin on the secretion of inflammatory factors in M5-induced psoriasis cell model

[0038] Experimental method: The cell treatment method was the same as in Example 1. After 24 h of culture, the cell culture supernatant was taken and the contents of IL-6 and IL-8 in the supernatant were determined by IL-6 ELISA kit (Solerbo, Human IL-6 ELISA KIT, SEKH-0013) and IL-8 ELISA kit (Solerbo, Human IL-8 ELISA KIT, SEKH-0016).

[0039] Test results: Figure 2 As shown, 20 μM cajanin significantly inhibited the secretion of IL-6 and IL-8 by HaCaT cells induced by M5, suggesting the inhibitory effect of cajanin on psoriasis.

[0040] Example 3

[0041] Therapeutic effect of cajanin solution on psoriasis model mice

[0042] Preparation of cajuna solution: Dissolve 50 mg cajuna powder in 1000 μL DMSO solution to prepare a mother solution (mother solution concentration is 50 mg / mL). Take 160 μL DMSO mother solution, add 640 μL PEG300, 160 μL Tween80 and 640 μL saline in sequence, and mix well to prepare a 5 mg / mL cajuna solution.

[0043] Experimental method: The mouse skin psoriasis model was established by imiquimod induction method, and the specific method is as follows: 8-week-old BALB / c mice were shaved on the back skin of 2 cm × 2 cm, and from the second day after shaving, each mouse was applied with imiquimod ointment (imiquimod content 5%, produced by Sichuan Mingxin Pharmaceutical Co., Ltd.) 62.5 mg for modeling stimulation at 10-11 am, and 5 mg / mL Example 3 pigeon arsenic solution was applied once at 14-15 pm (100 μL each time) for 6 days. After 6 days, the PASI score was performed on the lesions in the drug-applied area on the back of the mice to evaluate whether the psoriasis modeling was successful and the severity of psoriasis.

[0044] The PASI score includes the lesion area score and the lesion severity score.

[0045] The lesion area score is based on the percentage of surface area: 0 = no rash, 1 = 1%-9%, 2 = 10%-29%, 3 = 30%-49%, 4 = 50%-69%, 5 = 70%-89%, 6 = 90%-100%.

[0046] The severity of skin lesions was evaluated based on erythema, infiltration, and scaling, with each feature scored on a 0-4 scale: 0 = none; 1 = mild; 2 = moderate; 3 = severe; 4 = very severe.

[0047] The lesion area score and lesion severity score were added together to obtain a total score ranging from 0 to 18 points.

[0048] Test results: Figure 3 As shown in Figure 2, pigeon argan significantly inhibited the skin lesions of psoriasis model mice ( Figure 3 A), PASI scores of psoriasis mice treated with quinolin were significantly reduced ( Figure 3 B), indicating the effective therapeutic effect of caesalpinin on psoriasis.

[0049] Example 4

[0050] Effects of cajanin on skin thickness and keratinocyte proliferation in psoriasis model mice

[0051] Experimental methods: (1) Sampling: Anesthetize the mice and ensure that they are completely sedated. Use sterile scissors to cut the back skin of blank control mice, imiquimod-induced psoriasis mice, and caesalpino-treated psoriasis mice, with a size of approximately 0.5 cm × 1 cm. Immediately place the collected skin tissue in a 4% neutral paraformaldehyde solution and fix it at room temperature for 24-48 hours. (2) Paraffin embedding: The fixed tissue is dehydrated with ethanol, permeabilized with xylene, and then embedded in paraffin to form a solid paraffin block. Use a microtome to cut the paraffin block into continuous thin slices with a thickness of approximately 4-6 μm. Flatten the slices in a 40°C warm water bath and then transfer them to a glass slide. Place the slices in a 60°C constant temperature oven and bake for about 2 hours to allow the tissue to adhere tightly. (3) HE staining: Dewaxing was performed before staining. The sections were placed in environmentally friendly dewaxing solution I for 20 min, environmentally friendly dewaxing solution II for 20 min, anhydrous ethanol I for 5 min, anhydrous ethanol II for 5 min, and 75% alcohol for 5 min, and then washed with tap water. After that, the sections were placed in high-definition constant staining pretreatment solution for 1 min. Staining was performed according to the following steps: ① First, the sections were placed in hematoxylin staining solution for 3-5 min, and then washed with tap water; ② Differentiation solution was used for differentiation, and then washed with tap water, and then blueing solution was used for blueing, and then rinsed with running water; ③ Eosin staining was performed, and the sections were placed in 95% alcohol for 1 min for dehydration, and then stained in eosin staining solution for 15 s; ④ Dehydration and sealing: The sections were placed in anhydrous ethanol I for 2 min, anhydrous ethanol II for 2 min, anhydrous ethanol III for 2 min, n-butanol I for 2 min, n-butanol II for 2 min, xylene I for 2 min, and xylene II for 2 min to make them transparent, and then sealed with neutral gum. (4) Microscope examination, image acquisition and analysis. Under a 40× microscope, HE staining photos of the mouse back skin were observed and taken. The thickness of the epidermis was measured on the vertical cross section of each slice using Image J software. The average of the epidermis thickness at 5 random points for each mouse was calculated, and the independent sample t test was used for statistics. (5) Immunohistochemical staining: The dewaxing method was the same as that of HE staining. After dewaxing, antigen retrieval was performed using sodium citrate antigen retrieval solution with pH 6.0 at 95°C. After natural cooling, the slides were placed in PBS (pH 7.4) and washed on a decolorizing shaker for 3 times, each time for 5 minutes. The slices were placed in 3% hydrogen peroxide solution and incubated at room temperature in the dark for 25 minutes. The slides were placed in PBS (pH 7.4) and washed on a decolorizing shaker for 3 times, each time for 5 minutes. 3% BSA was added to the tissue to evenly cover the tissue and blocked at room temperature for 30 minutes. The blocking solution was gently shaken off, and Ki67 primary antibody diluted in PBS 1:500 was added to the slices. The slices were placed flat in a humidified box and incubated overnight at 4°C. On the second day, wash the slides in PBS (pH 7.4) on a decolorizing shaker for 3 times, 5 min each time. After the sections are dried slightly, add a secondary antibody (HRP labeled) of the same species as the primary antibody to cover the tissue and incubate at room temperature for 50 min. Wash the slides in PBS (pH 7.4) on a decolorizing shaker for 3 times, 5 min each time.After the slices were slightly dried, freshly prepared DAB coloring solution was added to the circle. The coloring time was controlled under a microscope. The positive color was brown-yellow. The slices were rinsed with tap water to stop the coloring. Re-staining of cell nuclei: Re-staining with hematoxylin for about 3 minutes, washing with tap water, differentiation with hematoxylin differentiation solution for a few seconds, washing with tap water, blueing with hematoxylin blueing solution, and washing with running water. After staining, dehydrate and seal the slices: put the slices in 75% alcohol for 5 minutes-85% alcohol for 5 minutes-anhydrous ethanol I for 5 minutes-anhydrous ethanol II for 5 minutes-n-butanol for 5 minutes-xylene I for 5 minutes to dehydrate and make them transparent. Take the slices out of xylene and dry them slightly, and seal them with sealing glue. The results were interpreted under a white light microscope. The number of Ki67-positive cells in the field of view was counted using the software Image J, and the average value of Ki67-positive cells in 3 random fields of view for each mouse was calculated, and statistics were performed using the independent sample t test.

[0052] Test results: Figure 4 As shown in A, compared with the control group, the epidermal lesions in the psoriasis model group showed hyperkeratosis, incomplete keratinization, thickening of the spinous layer, hyperplasia of the granular layer, and a large number of inflammatory cells and eosinophils infiltrating the dermis. Compared with the pathological sections of the psoriasis model group, the spinous layer thickening of the mice in the pigeon argan group was significantly reduced, and the inflammatory cells and eosinophils in the dermis were greatly reduced; Figure 4 B shows that the back epidermis thickness of psoriasis mice treated with caesalpinin was significantly lower than that of psoriasis mice not treated with caesalpinin; Figure 4 C and 4D show that cajanin significantly inhibited the expression of Ki67 in the skin tissue of psoriasis mice. The above results further show that cajanin can significantly reduce the abnormal proliferation of epidermal cells in psoriasis mice and thus play a therapeutic role.

[0053] Example 5

[0054] Therapeutic effect of cajanin on acne model mice

[0055] Experimental method: The mouse skin acne model was established by intradermal injection of Propionibacterium acnes. The specific method was as follows: 2 cm × 2 cm of the back skin of 8-week-old ICR mice was shaved, and Propionibacterium acnes (1 × 10 7 CFU / mL) and oleic acid was applied. On the third day, each mouse was applied with oleic acid at 10-11 am and the pigeon arsenic solution of Example 3 was applied at 14-15 pm, and continued for 5 days. Photos were taken every day. On the 7th day, skin tissue samples were taken from the mice and HE staining was performed, and the experimental procedures were the same as in Example 5.

[0056] Test results: Figure 5 As shown in A, the mice in the acne model group developed obvious acne on the back on the third day after modeling, and the acne on the back of the mice in the pigeon argan oil group was significantly weaker than that in the acne model group; Figure 5B shows that the skin inflammation of the mice in the caesalpin group was significantly reduced compared with the acne model group, the thickening of the stratum spinosum was significantly reduced, and the inflammatory cells and eosinophils in the dermis were greatly reduced. These results indicate that caesalpin has the effect of inhibiting the formation of acne.

[0057] Example 6

[0058] Effects of cajanin on the expression of skin-related inflammatory factors in acne model mice

[0059] Experimental method: The control group, acne model group and pigeon argan group mice were sampled, RNA extracted and inflammatory factor expression detected. The experimental steps were the same as those in Example 5.

[0060] Test results: Figure 6 As shown, caesalpin significantly inhibited the mRNA expression of inflammatory factors IL-6, IL-1α, IL-1β, TNF-α and IL-23 in the skin of acne model mice, indicating that caesalpin can significantly inhibit the inflammatory response in the skin tissue of acne mice.

[0061] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. Application of quinoa in the preparation of drugs for the treatment of psoriasis.

2. The use according to claim 1, characterized in that: The cajanin can significantly inhibit the proliferation of keratinocytes and the secretion of inflammatory factors IL-6 and IL-8 in vitro.

3. The use according to claim 1, characterized in that: The cajanin can significantly inhibit psoriasis skin lesions, significantly reduce the psoriasis lesion area and reduce the severity index score.

4. The use according to claim 1, characterized in that: The cajanin can significantly reduce the thickness of the epidermis and the expression of Ki67 in psoriasis skin tissue.

5. Application of caesalpinol in the preparation of acne treatment drugs.

6. The use according to claim 5, characterized in that: The cajanin can obviously inhibit the formation of acne.

7. The use according to claim 5, characterized in that: The cajanin can significantly inhibit the mRNA expression of inflammatory factors IL-6, IL-1α, IL-1β, TNF-α and IL-23 in acne skin.

8. An external preparation for treating psoriasis and acne, characterized in that: The active ingredient of the external preparation includes cajanin.

9. The external preparation according to claim 8, characterized in that The external preparation further includes a solvent, a dispersant, an emulsifier and water.

10. The external preparation according to claim 8, characterized in that The external preparation is any one of a cream, an ointment, a lotion, a gel or a spray.

Images