TWI637747B - A composition comprising young watermelon fruit extract, young chiapao melon fruit extract or combination thereof having functionality to regulate the gene expression of interleukin-8, nitric oxide synthase 3, endothelin 1, tissue type-plasminogen activa - Google Patents

Abstract

The invention provides a composition containing watermelon young fruit extract, Gerber melon young fruit extract, or a combination thereof, which can regulate nitric oxide synthase 3 (NOS3) and von Willebrand factor (vWF) genes by upward regulation. And down-regulating interleukin-8 (IL-8), endothelin 1 (EDN1), tissue-type plasminogen initiator (PALT), human coagulation factor (F3), serine proteinase inhibitor 1 (SERPINE1) And the expression of the intercellular adhesion molecule 1 (ICAM1) gene, which in turn promotes keratinocyte metabolism and inhibits the secretion of interleukin-8, and finally achieves anti-inflammatory, anti-acne, and lower blood pressure and blood lipid effects.

Description

Used to regulate interleukin-8, nitric oxide synthase 3, endothelin 1, tissue fibrinolysis Compositions containing zymogen initiator, von Willebrand factor, human coagulation factor, serine proteinase inhibitor 1 and intercellular adhesion molecule 1 genes containing watermelon young fruit, Gerber melon young fruit extract or combination thereof

The invention provides a composition containing watermelon young fruit extract, Gerber melon young fruit extract, or a combination thereof, and particularly relates to a compound for regulating interleukin-8 (IL-8) and nitric oxide. Nitric oxide synthase 3 (NOS3), endothelin 1 (EDN1), tissue type-plasminogen activator (PALT), von Willebrand factor, vWF), human coagulation factor (F3), serine protease inhibitor 1 (SERPINE1) and intercellular adhesion molecule 1 (ICAM1) genes.

Modern people live in busy green, stressful environments, irregular meals, or eating processed foods. They face the problem that the largest sebaceous glands are too strong on the skin. Male skin often has more oil than female skin, especially Oily skin with large pores over 25 years of age, prolonged exposure to ultraviolet light, long-term travel, habits of sleeping late, drinking and coffee habits, smoking or exposure to second-hand smoke, and working in cold air for a long time , Often have problems with greasy skin, large pores clogged, and acne.

At the same time, with age, cholesterol will accumulate in blood vessels, causing endothelial cell function defects, changing the original anticoagulant and anti-inflammatory properties of endothelial cells, and narrowing blood vessels, which can easily lead to diseases such as hypertension and arteriosclerosis.

Therefore, in the face of the above problems, people must work hard to rely on the acquired care to delay the aging of the skin and body functions, and even return to young skin and healthy body. In recent years, in order to meet the needs of people to delay aging, various products on the market, whether by rubbing, wiping or oral, have been continuously developed. However, in order to achieve special effects, these products have added too many carcinogenic chemicals or environmental hormones, making consumers use the ingredients that will harm the skin and affect health without knowing it. Therefore, it is a gradual trend to add natural and effective ingredients to whitening products in modern times that emphasize natural health.

In view of this, the present invention provides a young fruit of Cucurbitaceae melon for preparing and regulating interleukin-8 (IL-8), nitric oxide synthase 3 (NOS3), and endothelin 1 (endothelin 1, EDN1), tissue type-plasminogen activator (PALT), von Willebrand factor (vWF), human coagulation factor (Homo sapiens coagulation factor III ( tissue factor), F3), serine protease inhibitor 1 (SERPINE1) and intercellular adhesion molecule 1 (ICAM1) gene expression levels of pharmaceutical compositions, wherein NOS3 and The VWF gene is up-regulated, and the EDN1, PLAT, F3, SERPINE1, IL-8 and ICAM1 genes are down-regulated; and the Cucurbita melon young fruit is selected from a watermelon young fruit extract and a Gerber melon A group of fruit extracts and combinations thereof.

The present invention also provides the use of a young fruit of Cucurbitaceae melon for preparing a medicinal composition for promoting keratinocyte metabolism, wherein the young fruit of Cucurbita melon is selected from the group consisting of a watermelon young fruit extract and a Gerber melon seed. A group of fruit extracts and combinations thereof.

The present invention further provides the use of a young fruit of Cucurbitaceae melon for preparing a pharmaceutical composition for lowering blood pressure and blood lipid, wherein the young fruit of Cucurbita melon is selected from a watermelon young fruit extract and a Gerber melon seed. A group of fruit extracts and combinations thereof.

In one embodiment of the present invention, the pharmaceutical composition has the effects of promoting keratinocyte metabolism, anti-inflammatory and anti-acne.

In one embodiment of the present invention, the pharmaceutical composition inhibits interleukin-8 (IL-8) secretion.

In one embodiment of the present invention, the pharmaceutical composition has the effect of reducing blood pressure and blood lipid.

In one embodiment of the present invention, the Cucurbita melon young fruit extract uses water, alcohols, hydroalcohols or a combination thereof as an extraction solvent, and the extraction solvent is respectively young with Cucurbita melon young fruit. The volume ratio of the fruit is 2 to 20: 1 to 5.

In one embodiment of the present invention, the pharmaceutical composition is used for regulating interleukin-8 (IL-8), nitric oxide synthase 3 (NOS3), endothelin 1 (endothelin 1, EDN1), tissue type-plasminogen activator (PALT), von Willebrand factor (vWF), human coagulation factor (Homo sapiens coagulation factor III ( tissue factor), F3), serine protease inhibitor 1 (SERPINE1), and intercellular adhesion molecule 1 (intercellular adhesion molecule 1, ICAM1) gene expression, and NOS3 and VWF genes are up-regulated, EDN1, PLAT, F3, SERPINE1, IL-8 and ICAM1 genes are down-regulated.

In one embodiment of the present invention, the pharmaceutical composition further comprises a pharmaceutically acceptable carrier.

In one embodiment of the present invention, the pharmaceutical composition is administered by oral or skin application.

Therefore, the composition containing the watermelon young fruit extract, the Gerber melon young fruit extract, or a combination thereof can regulate nitric oxide synthase 3 (NOS3) and vascular hemophilia through upward regulation. Factor (von Willebrand factor, vWF) gene, and down-regulation of interleukin-8 (IL-8), endothelin 1 (EDN1), tissue type plasminogen initiator (tissue type -plasminogen activator (PALT), Homo sapiens coagulation factor III (tissue factor) (F3), serine protease inhibitor 1 (SERPINE1), and intercellular adhesion molecule 1, ICAM1) gene expression, which in turn promotes keratinocyte metabolism and inhibits interleukin-8 secretion, and finally achieves anti-inflammatory, anti-acne, and lower blood pressure and blood lipid effects. Therefore, the composition containing watermelon young fruit extract, Gerber melon young fruit extract, or a combination thereof according to the present invention can be used for preparing a pharmaceutical composition.

The embodiments of the present invention will be further described below with reference to the drawings. The examples listed below are intended to clarify the present invention and are not intended to limit the scope of the present invention. Any person skilled in the art will not depart from the spirit and scope of the present invention. Within the scope, when some changes and retouching can be done, the protection scope of the present invention shall be determined by the scope of the attached patent application scope.

FIG. 1 is a data chart of keratinocyte proliferation promoted by watermelon young fruit extract or Gerber melon young fruit extract of the present invention. Compared with the control group, * p <0.05, ** p <0.01, and *** p <0.001.

FIG. 2 is a data chart of the reduction of proinflammatory cytokine IL-8 production by watermelon young fruit extract or Gerber melon young fruit extract of the present invention. Compared to the control group * p <0.05, ** p <0.01.

FIG. 3 shows nitric oxide synthase 3 (NOS3) which regulates vasodilation and endothelin 1 which regulates vasoconstriction by the watermelon young fruit extract or Gerber melon young fruit extract of the present invention. , EDN1) gene performance data. Compared to the control group * p <0.05, ** p <0.01.

FIG. 4 shows the von Willebrand factor (vWF) that regulates thrombolysis and the tissue-type plasminogen that inhibits angiogenesis by the watermelon young fruit extract or Gerber melon young fruit extract of the present invention. Gene map of gene expression of tissue type-plasminogen activator (PALT), Homo sapiens coagulation factor III (tissue factor), and serine protease inhibitor 1 (SERPINE1). Compared to the control group * p <0.05, ** p <0.01.

FIG. 5 shows that the watermelon young fruit extract or the Gerber melon young fruit extract of the present invention regulates the pro-inflammatory factors interleukin-8 (IL-8) and intercellular adhesion molecule 1 , ICAM1) gene performance data. Compared to the control group * p <0.05, ** p <0.01.

The invention provides a composition containing young fruits of cucurbitaceae, which is extracted at a temperature of 50 to 100 ° C., and is obtained by extracting and centrifuging a watermelon young fruit and a Gerber melon young fruit with a solvent, respectively. The extraction solvent is water, alcohols, aqueous alcohols, or a combination thereof. The present invention utilizes a cell proliferation reagent kit to carry out the analysis of promoting the keratinocyte metabolism of the watermelon young fruit extract or Gerber melon young fruit extract of the present invention, and using human interleukin 8 (IL-8 / CXCL8) enzyme immunoassay ( enzyme-linked immunosorbent assay (ELISA) analysis and detection reagent kit, to analyze the inhibitory ability of the watermelon young fruit extract or Gerber melon young fruit extract of the present invention on the pro-inflammatory cytokine IL-8, and carry out the test through gene expression detection Analysis of the regulation of vascular endothelial cell genes by the invention of watermelon young fruit extract or Gerber melon young fruit extract to confirm that the composition containing watermelon young fruit extract or Gerber melon young fruit extract of the present invention can promote keratin Cell metabolism and inhibition of interleukin-8 secretion ultimately achieve anti-inflammatory, anti-acne, and lower blood pressure and blood lipid effects.

Example 1 Preparation method of cucurbitaceous melon young fruit extract of the present invention

Watermelon ( Citrullus lanatus ), anciently known as cold gourd, is a plant or fruit of the genus Watermelon in the Cucurbitaceae family. Native to Africa, it is a dicotyledon flowering plant shaped like a vine with feathery leaves. The fruit it produces is a false fruit, and belongs to a category that botanists call fake berries. The skin of the fruit is smooth with green or yellow and dark green patterns, and the fruit is juicy and red or yellow.

Citrullus lanatus (Thunb.) Matsum et Nakai is one of the early watermelon varieties in Taiwan. It was originally called Garbo Papaya, a pioneer of orange flesh. Only Penghu County in Taiwan is economically cultivated and has a long history. The special flavor is deeply favored by residents and tourists in Penghu. Therefore, its output value and farmers' willingness to cultivate are both high. It has already become a special agricultural product with high economic value in Penghu.

In one embodiment of the present invention, after the young watermelon fruit or the young Garbo melon is washed, the water, alcohol, or an aqueous alcohol or a combination of extraction solvents thereof are used to separate the young watermelon fruit or the young Garbo melon fruit, respectively. The volume ratio is from 2 to 20: 1 to 5. After homogenization, extraction is performed at 50 to 100 ° C for 0.5 to 3 hours to obtain a crude extract of watermelon young fruit and Jiabao melon young fruit respectively; after extraction, cool to room temperature, and The crude extract is centrifuged, and the supernatant after centrifugation is taken. The supernatant is filtered through a 400mech filter to obtain a filtrate. Finally, the filtrate is concentrated under reduced pressure at 45 to 70 ° C to obtain the watermelon young fruit extract of the present invention. Or Gerber melon young fruit extract. among them The watermelon young fruit is defined as a young fruit within 3 weeks and the flesh is white; the Gerber melon is defined as a young fruit within 20 days and the flesh is white; and a whole melon including a peel is used in the embodiment of the present invention. Extraction is performed simultaneously.

Example 2 The effect of the watermelon young fruit extract or Gerber melon young fruit extract of the present invention in promoting keratinocyte metabolism

The present invention analyzes the promotion of keratinocytes metabolism. Human Keratinocytes (PCS-200-010 ^™ ) were purchased from the American Type Culture Collection (ATCC). The cells were cultured in a dermal cell basal medium (ATCC, USA, PCS-200-030 ^TM ), and keratinocyte growth factor (ATCC, USA, PCS-200-040 ^TM ) was added, which contained : 2.0 mL of 0.4% bovine pituitary extract (BPE), 0.5 mL of 0.5 ng / mL recombinant human transforming growth factor α (rh TGF-α), and 15.0 mL of 6 mM L-glutamine (L -glutamine), 0.5 mL of 100 ng / mL hydrocortisone hemisuccinate, 0.5 mL of 5 mg / mL recombinant human insulin (rh insulin), 0.5 mL of 1.0 mM, and 0.5 mL of 5 mg / mL transferrin ( apo-Transferrin).

The analysis was performed using a cell proliferation reagent kit (Roche, Switzerland, No. 116472229001). First, 6,000 keratinocytes were inoculated into each hole in a 96-well culture plate, and cultured at 37 ° C for 2 hours in the above medium; 100 μL of 0.25 mg / mL Gerber melon young fruit extract and Gerber melon ripe fruit were added to the holes, respectively. The extract (prepared in the same manner as in Example 1) and watermelon young fruit extract were used as the experimental group; 100 μL of 50 ng / mL Epidermal growth factor (EGF) was added as a control group; and the medium was added without any ingredients As a control group, 10 μL of 100 μM Bromodeoxyuridine (BrdU) was added to each hole, and the culture was continued for 24 hours under the same conditions. BrdU mosaic DNA content was used to detect cell proliferation. Remove the culture medium, add 200 μL of Fixing Solution (FixDenat) to each well and react at room temperature for 30 minutes. Remove the fixing solution and wash it once with Phosphate buffered saline (PBS). Add 100 μL of anti-BrdU-POD to each well and let it stand at room temperature for 90 minutes, so that anti-BrdU labeled with peroxidase can identify BrdU. After removing the anti-BrdU-POD, wash with 200 to 300 μL PBS three times, and finally add matrix solution (TMB) to each hole to make the reaction color. After standing at room temperature for about 5 to 30 minutes, add 25 μL of 1M H ₂ The SO ₄ reaction was terminated, and the absorbance at 450 nm was measured with an ELISA reader (Bio Tek, USA), and statistical analysis was performed by Excel.

The experimental results of the effect of the watermelon young fruit extract or Gerber melon young fruit extract on promoting keratinocyte metabolism are shown in Fig. 1. After the watermelon young fruit extract, Gerber melon young fruit extract, and Gerber melon ripe fruit, After treatment, keratinocyte proliferation increased significantly. Compared to keratinocyte proliferation results of epidermal growth factors known to promote keratinocyte proliferation, watermelon young fruit extract increased by approximately 10%, Gerber melon young fruit extract increased by approximately 5%, and Gerber melon mature fruit extract and Epidermal growth factor has similar keratinocyte proliferation. Therefore, the watermelon young fruit extract, Gerber melon young fruit extract, or a combination thereof according to the present invention has an excellent effect of promoting keratinocyte proliferation and can effectively metabolize keratinocytes.

Example 3 The watermelon young fruit extract or Gerber melon young fruit extract of the present invention inhibits interleukin-8 (IL-8) secretion

Since interleukin-8 (IL-8) is known to have a function to regulate the inflammatory response, the present invention further tests the watermelon young fruit extract or Gerber melon young fruit extract on the pro-inflammatory cytokine IL- 8's ability to inhibit.

Using keratinocytes as in Example 2 described above, the present invention was analyzed using a human interleukin 8 (IL-8 / CXCL8) ELISA assay detection reagent kit (R & D systems, USA). First, in a 24-well culture plate, each well containing 500 μL of the above medium was seeded with 5 × 10 ⁴ keratinocytes and cultured at 37 ° C. for 24 hours; carefully remove the medium without disturbing the attached cells, and then Add 120 μL of 0.25mg / mL Gerber melon young fruit extract, Gerber melon ripe fruit extract, or watermelon young fruit extract as experimental groups in each hole; and use medium without adding any ingredients and not exposed to ultraviolet rays As a control group, another group was exposed to ultraviolet radiation as a control group, and each group was repeated three times. The cells were then exposed to 300 mJ ultraviolet (UVB) radiation at 37 ° C for 24 hours, and 120 μL of the cell suspension was collected.

Capture antibody diluted in PBS was added to a 96-well ELISA plate, and the volume of each well was 100 μL. The samples were placed at 4 ° C. until overnight to adsorb the antibodies to the 96-well ELISA plate. Then wash the 96-well ELISA plate three times with 200 μL of washing buffer (0.05% Tween 20 and PBS mixed solution), add 300 μL of blocking buffer (BSA and PBS mixed solution) and place at 37 ° C for 2 hours to block non-specificity Adsorption. Then, the 96-well ELISA plate was washed three times with 200 μL of washing buffer, and 100 μL of detection antibody was added and left at 37 ° C. for 2 hours. Then, the 96-well ELISA plate was washed three times with 200 μL of washing buffer, and 100 μL of streptavidin-HRP (streptavidin-HRP) was added and left at room temperature for 20 minutes. Then, the 96-well ELISA plate was washed three times with 200 μL of washing buffer, 100 μL of the matrix solution was added and left at room temperature for 20 minutes, and 50 μL of the stop solution was added to stop the reaction. ) Measure the absorbance at 450nm and perform t- test statistical analysis with Excel.

The experimental results of the effect of the watermelon young fruit extract, Gerber melon young fruit extract, or a combination thereof on inhibiting the secretion of interleukin-8 (IL-8) are shown in FIG. 2. Inflammation caused by external stimuli. Compared to the amount of IL-8 secreted by cells exposed to 300mJ ultraviolet (UVB) radiation, Gerber melon young fruit extract can reduce the production of proinflammatory cytokines IL-8 by about 60%. Melon Fruit Extract It can reduce the production of pro-inflammatory cytokine IL-8 by about 33%, and watermelon young fruit extract can reduce the production of pro-inflammatory cytokine IL-8 by about 26%. Therefore, the watermelon young fruit extract or Gerber melon young fruit extract of the present invention can reduce the production of proinflammatory cytokine IL-8 and achieve anti-inflammatory effect.

Example 4 Regulatory effect of vascular endothelial cell genes of watermelon young fruit extract or Gerber melon young fruit extract of the present invention

According to the above embodiments, the watermelon young fruit extract or Gerber melon young fruit extract of the present invention can promote keratin metabolism and anti-inflammation, and finally achieve the effect of preventing acne and facial blisters. With the increase of age, abnormal diet, poor work and rest, etc., cholesterol will accumulate in blood vessels, causing endothelial cell dysfunction, changing the original anticoagulant and anti-inflammatory properties of endothelial cells, and narrowing blood vessels can easily lead to hypertension. , Arteriosclerosis and other diseases. The main functions of vascular endothelial cells are to regulate vasoconstriction, the production of cytokines, and the role of immune cells in the blood. Therefore, the present invention further examines the effect of watermelon young fruit extract or Gerber melon young fruit extract on the expression of related genes in vascular endothelial cells.

In the present invention, human umbilical vein endothelial cells (HUVEC) (obtained from the Center for Biological Resources Preservation and Research, BCRC number H-UV001) are used to perform experiments on the expression levels of vascular endothelial cells. First, inoculate 1 × 10 ⁵ human umbilical vein endothelial cells in each hole in a 6-well culture plate, add 2 mL of EC medium (Gibico, USA, No. M-200-500), and add 100 μL of 0.03125 mg / mL Jia respectively. Gourd young fruit extract, Gerber melon mature fruit extract, and watermelon young fruit extract were used as the experimental group; and a medium without any added ingredients was used as the control group, and cultured for 6 hours; then human umbilical vein endothelial cells were collected. RNA was extracted from the above cells using an RNA extraction reagent kit (Genemark, USA), and then reverse-transcribed into cDNA using reverse script (SuperScript® III Reverse Transcriptase) (Invitrogen, USA), followed by KAPA SYBR® FAST qPCR reagent Group (KAPA Biosystems, USA) measured target genes, including interleukin-8 (IL-8), nitric oxide synthase 3 (NOS3), and endothelin 1 , EDN1), tissue type-plasminogen activator (PALT), von Willebrand factor (vWF), Homo sapiens coagulation factor III (tissue factor), F3), serine protease inhibitor 1 (SERPINE1) and intercellular adhesion molecule 1 (ICAM1) genes were analyzed by quantitative PCR.

The main function of vascular endothelial cells is to regulate vasoconstriction, cytokine production, and immune cells in the blood. The results are shown in Figure 3. After the action of watermelon young fruit extract, it has been shown that the expression of genes that regulate vasodilation NOS3 can be improved And it can reduce the expression of genes that regulate vasoconstriction EDN1, indicating that the watermelon fruit extract of the invention has the effect of lowering blood pressure; as shown in Figure 4, the watermelon fruit extract can improve the gene expression of thrombolytic vWF, and can Achieving gene expression that inhibits angiogenesis PLAT, F3, and SERPINE1, indicating that watermelon young fruit extract can increase thrombolysis and reduce thrombosis; as shown in Figure 5, watermelon young fruit extract can reduce proinflammatory factors IL-8 and ICAM1 The gene expression indicates that the watermelon young fruit extract can achieve anti-inflammatory effect.

In summary, the composition containing watermelon young fruit extract, Gerber melon young fruit extract, or a combination thereof according to the present invention can enhance the gene expression of good factors of vascular endothelial cells, such as NOS3 and VWF, and inhibit endothelial cells The expression of vascular necrosis factor genes, such as EDN1, PLAT, F3, SERPINE1, IL-8, and ICAM1, is thus achieved, thereby promoting keratinocyte metabolism and inhibiting interleukin-8 secretion, and finally reaching anti-inflammatory, anti-acne and lowering blood pressure, Effect of blood lipids. Therefore, the composition containing watermelon young fruit extract or Gerber melon young fruit extract according to the present invention is a natural ingredient and does not produce traditional artificial inhibition The side effects brought by the agent can be used to regulate skin keratin metabolism, anti-inflammatory, anti-acne, and lower blood pressure and blood lipids. The composition is in the form of powder, granule, liquid, gel or paste. And its dosage form is provided in the form of food, drink, medicine, reagent or nutritional supplement, and can be administered orally or smeared.

Claims (10)

An extract of cucurbit fruit young fruit is used for preparing and regulating interleukin-8 (IL-8), nitric oxide synthase 3 (NOS3), endothelin 1, EDN1), tissue type-plasminogen activator (PALT), von Willebrand factor (vWF), Homo sapiens coagulation factor III (tissue factor), F3 ), Serine protease inhibitor 1 (SERPINE1) and intercellular adhesion molecule 1 (ICAM1) gene expression of pharmaceutical compositions, wherein NOS3 and VWF genes are up-regulated, EDN1, PLAT, F3, SERPINE1, IL-8 and ICAM1 genes are down-regulated; and the Cucurbita melon young fruit extract is selected from the group consisting of a watermelon young fruit extract, a Gerber melon young fruit extract, and A group consisting of the combination; the Cucurbita melon young fruit extract uses water as an extraction solvent. The use as described in item 1 of the scope of patent application, wherein the pharmaceutical composition has the effects of promoting keratinocyte metabolism, anti-inflammatory and anti-acne effects. The use according to item 1 of the scope of patent application, wherein the pharmaceutical composition inhibits interleukin-8 (IL-8) secretion. The use as described in item 1 of the scope of patent application, wherein the pharmaceutical composition has the effect of lowering blood pressure and blood lipids. A cucurbit melon young fruit extract is used for preparing a medicinal composition for promoting keratinocyte metabolism. The cucurbit melon young fruit extract is selected from the group consisting of a watermelon young fruit extract and a Gerber melon seed. Fruit extracts and combinations thereof; the squash gourd young fruit extract uses water as an extraction solvent. A squash melon young fruit extract for preparing a pharmaceutical composition for lowering blood pressure and blood lipid, wherein the squash melon young fruit extract is selected from the group consisting of a watermelon young fruit extract and a Jiabao melon seed. Fruit extracts and combinations thereof; the squash gourd young fruit extract uses water as an extraction solvent. The use as described in any one of claims 1, 5 and 6, wherein the volume ratio of the extraction solvent to the young fruits of the Cucurbita melon fruits is 2 to 20: 1 to 5. The use according to item 5 or 6 of the scope of the patent application, wherein the pharmaceutical composition is used for regulating interleukin-8 (IL-8), nitric oxide synthase 3, NOS3), endothelin 1 (EDN1), tissue type-plasminogen activator (PALT), von Willebrand factor (vWF), human coagulation factor (Homo Expression of sapiens coagulation factor III (tissue factor) (F3), serine protease inhibitor 1 (SERPINEI) and intercellular adhesion molecule 1 (ICAM1) genes, and NOS3 and VWF genes Up regulation, EDN1, PLAT, F3, SERPINE1, IL-8 and ICAM1 genes are down regulation. The use as described in any one of claims 1, 5 and 6, wherein the pharmaceutical composition further comprises a pharmaceutically acceptable carrier. The use according to any one of claims 1, 5 and 6, wherein the pharmaceutical composition is administered orally or by skin application.