WO2024041084A1 - 一种治疗银屑病的外用中药组合物及其制备方法与应用 - Google Patents
一种治疗银屑病的外用中药组合物及其制备方法与应用 Download PDFInfo
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- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/30—Boraginaceae (Borage family), e.g. comfrey, lungwort or forget-me-not
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- A—HUMAN NECESSITIES
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- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/48—Fabaceae or Leguminosae (Pea or Legume family); Caesalpiniaceae; Mimosaceae; Papilionaceae
- A61K36/484—Glycyrrhiza (licorice)
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- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/53—Lamiaceae or Labiatae (Mint family), e.g. thyme, rosemary or lavender
- A61K36/536—Prunella or Brunella (selfheal)
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- A—HUMAN NECESSITIES
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- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/44—Oils, fats or waxes according to two or more groups of A61K47/02-A61K47/42; Natural or modified natural oils, fats or waxes, e.g. castor oil, polyethoxylated castor oil, montan wax, lignite, shellac, rosin, beeswax or lanolin
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/0012—Galenical forms characterised by the site of application
- A61K9/0014—Skin, i.e. galenical aspects of topical compositions
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- A—HUMAN NECESSITIES
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- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P17/00—Drugs for dermatological disorders
- A61P17/06—Antipsoriatics
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
Definitions
- the invention belongs to the technical field of psoriasis treatment, and in particular relates to an external Chinese medicine composition for treating psoriasis and its preparation method and application.
- Psoriasis is a common chronic relapsing inflammatory skin disease.
- the basic skin lesions are erythema, papules, or plaques covered with multiple layers of silvery white scales.
- the site of occurrence is more common on the body surface and head. Multiple layers of silvery white dry scales often appear repeatedly on the erythema.
- the cause of psoriasis is not entirely clear and it is stubborn and difficult to treat. It is listed as one of the ten most stubborn diseases by the World Health Organization.
- topical drugs such as keratin enhancers, corticosteroid creams, etc.
- systemic drugs include immunosuppressants, retinoic acid, antibiotics, and corticosteroids. wait.
- these drugs have good short-term therapeutic effects on psoriasis, but long-term use has serious side effects and a high recurrence rate.
- methotrexate is the first-line systemic treatment drug for psoriasis.
- Some patients have mild adverse reactions after taking the drug, such as diarrhea, nausea, lack of appetite, ulcers, drowsiness, and skin erosion.
- Retinoic acid drugs are mostly used to treat moderate to severe psoriasis vulgaris, erythroderma type, and adenocarcinoma type. However, they are not very effective for joint psoriasis. Common adverse reactions include skin and mucosal symptoms, eye toxicity, and liver enzymes. Or elevated blood lipids, abnormal bone marrow development and teratogenicity, it is necessary to monitor indicators such as blood lipids and growth curves. Although biological agents (such as adalimumab, infliximab, efalizumab, etc.) are effective in treating psoriasis, they have many side effects, such as nodular vasculitis, severe infection, Heart failure and malignant tumor risk. Topical glucocorticoids may cause symptoms such as skin atrophy, telangiectasia, atrophic striae, purpura, and hirsutism.
- the object of the present invention is to provide a traditional Chinese medicine composition for treating psoriasis.
- Each medicinal flavor synergistically plays the role of clearing heat, detoxifying, cooling blood and dispersing blood stasis. It has significant curative effect in the external treatment of psoriasis, and It has the advantages of safety, economy and convenient operation.
- the invention provides an external Chinese medicine composition for treating psoriasis, which includes the following raw materials in parts by weight: 10 to 20 parts of madder, 10 to 20 parts of Lithospermum, 10 to 20 parts of Prunella vulgaris, and 30 to 30 parts of raw licorice. 50 servings.
- the preparation types of the external Chinese medicine composition include liquids, oils, powders, tinctures, lotions, creams, ointments, shower gels and gels.
- the invention also provides a method for preparing the above-mentioned external Chinese medicine composition, which includes the following steps: mixing madder, lithospermum, prunella vulgaris and raw licorice with oil, soaking, frying over low heat until brown, and removing the medicinal residue, to obtain Medicated oil.
- the oil agent includes sunflower oil or sesame oil.
- the material-to-liquid ratio of the sum of the four raw materials of madder, Lithospermum, Prunella vulgaris and raw licorice to the oil is 1g: 4.1ml to 9.2ml.
- the soaking time is 24h to 28h.
- the present invention also provides an application of the above-mentioned external Chinese medicine composition in preparing products for treating psoriasis.
- the external Chinese medicine composition exerts a therapeutic effect by affecting keratinocyte differentiation, cytokine activity and/or epidermal cell differentiation.
- the external Chinese medicine composition exerts a therapeutic effect by participating in cytokine receptor interactions, IL-17 signaling pathways, estrogen signaling pathways and/or protein digestion and absorption pathways.
- the products include pharmaceuticals.
- madder cools blood, promotes blood circulation, removes blood stasis and activates blood circulation, which is the monarch medicine
- Lithospermum chinensis clears away heat and detoxifies, cools blood and stops bleeding
- Prunella vulgaris clears away heat and detoxifies, and is a ministerial medicine together
- raw licorice serves as an auxiliary, detoxifying and harmonious medicine, making up the four medicines.
- FIG. 2 Histopathological changes in mice in each group, where A: Histopathological changes in the back and ears of mice in each group ( ⁇ 200), B: Quantification of back thickness of mice in each group on the 12th day, C: Mouse in each group on the 12th day Quantitative ear thickness at 12 days, note: compared with the model group, *p ⁇ 0.05, **p ⁇ 0.01; compared with the normal group, ### p ⁇ 0.001;
- Figure 4 The results of mRNA-seq analysis of back skin tissue of IMQ+SCY group and IMQ group.
- A Volcano plot of differentially expressed proteins (DEPs) between IMQ+SCY group and IMQ group. Green dots represent down-regulated DEPs, and red dots represent Up-regulated DEPs, gray points represent non-significant DEPs;
- B Heat map of 289 DEPs in IMQ+SCY group and IMQ group, cluster analysis based on gene expression, red represents highly expressed DEPs, green represents low expressed DEPs , each row represents the expression level of each DEPs in different groups;
- Figure 5 GO and KEGG enrichment analysis of differential genes in psoriasis skin lesions after SCY intervention, where A: GO analysis results of DEPs after SCY intervention, B: KEGG analysis results of DEPs after SCY intervention.
- the invention provides an external Chinese medicine composition for treating psoriasis, which includes the following raw materials in parts by weight: 10 to 20 parts of madder, 10 to 20 parts of Lithospermum, 10 to 20 parts of Prunella vulgaris, and 30 to 30 parts of raw licorice. 50 servings.
- the present invention has no special limitations on the specific sources of each of the above raw materials.
- the weight parts of each raw material are preferably 13 to 18 parts of Rubia cordata, 12 to 16 parts of Lithospermum vulgaris, 14 to 17 parts of Prunella vulgaris, and 35 to 45 parts of raw licorice, and more preferably 14 to 14 parts of Rubia cordata. 16 parts, comfrey 14-15 parts, prunella vulgaris 15-16 parts, raw licorice 38-42 parts.
- the external Chinese medicine composition for treating psoriasis of the present invention can preferably be made into liquids, oils, powders, tinctures, lotions, creams, ointments, bath agents and gels.
- the invention also provides a method for preparing the above-mentioned external Chinese medicine composition, which includes the following steps: mixing madder, lithospermum, prunella vulgaris and raw licorice with oil, soaking, frying over low heat until brown, and removing the medicinal residue, to obtain Medicated oil.
- the oil agent preferably includes sunflower seed oil or sesame oil
- the material-to-liquid ratio of the sum of the four raw materials of madder, lithospermum, prunella vulgaris and raw licorice to the oil agent is preferably 1g: 4.1ml to 9.2ml, more Preferably, it is 1 g: 5 ml to 8.5 ml.
- the present invention has no special limitation on the specific source of sunflower oil or sesame oil, and any conventional commercially available product in this field can be used.
- the soaking time is preferably 24h to 28h, more preferably 25h to 26h.
- the present invention has no special limitation on the specific method of removing drug residue, and any conventional removal method in this field can be used.
- the present invention also provides an application of the above-mentioned external Chinese medicine composition in preparing products for treating psoriasis.
- the products preferably include pharmaceuticals.
- the external Chinese medicine composition of the present invention preferably exerts a therapeutic effect by affecting keratinocyte differentiation, cytokine activity and/or epidermal cell differentiation, and more preferably by participating in cytokine receptor interaction, IL-17 signaling pathway, estrogen Hormone signaling pathways and/or protein digestion and absorption pathways play a therapeutic role.
- mice Male BABLc mice aged 6-8 weeks and weighing 20g ⁇ 2g were randomly divided into 3 groups, with 4 mice in each group: (1) Blank control group (Control group): Apply 62.5 mg of Vaseline cream daily Mouse back and inner and outer sides of both auricles. (2) Model group (IMQ group): 62.5 mg of 5% IMQ cream was applied to the back of mice and the inner and outer sides of both auricles respectively. 6 hours later, 62.5 mg of Vaseline cream was applied to the back of the mouse and the inner and outer sides of both auricles. (3) Sicao oil group (IMQ+SCY group): Apply 62.5 mg of 5% IMQ cream on the back of mice and the inner and outer sides of both auricles.
- Blank control group Control group
- IMQ group 62.5 mg of 5% IMQ cream was applied to the back of mice and the inner and outer sides of both auricles respectively. 6 hours later, 62.5 mg of Vaseline cream was applied to the back of the mouse and the inner and outer sides of both auricles.
- mice 6 hours later, smear the back of the mouse and the inner and outer sides of both auricles with 150 ul of the four-grass oil prepared in Example 1. All treatments of mice started from the day when IMQ was applied (day 0), once a day for 12 consecutive days.
- mice The severity of skin inflammation on the back and ears of mice was scored using the Psoriasis Clinical Area and Severity Index (PASI), including erythema, thickness, and scale. Scores indicate severity: 0 (none), 1 (mild), 2 (moderate), 3 (severe) and 4 (extremely severe). Mice were fasted before specimen collection and allowed to drink water for 12 h. At the end of the 13th day of the experiment, the mice were sacrificed with CO2 , and ear skin lesions and back skin lesions were collected for further experiments.
- Psoriasis Clinical Area and Severity Index including erythema, thickness, and scale. Scores indicate severity: 0 (none), 1 (mild), 2 (moderate), 3 (severe) and 4 (extremely severe). Mice were fasted before specimen collection and allowed to drink water for 12 h. At the end of the 13th day of the experiment, the mice were sacrificed with CO2 , and ear skin lesions and back skin
- Sicao oil can improve skin lesions on the back and ears of IMQ-induced psoriasis-like mice
- Sicao oil can improve the histopathology of IMQ-induced psoriasis-like mouse model
- RNA extraction from the mouse samples of the IMQ group and IMQ+SCY group in Example 4 according to the standard operating procedures provided by the manufacturer.
- the total RNA was quality inspected by NanoDrop ND-2000 spectrophotometer and Agilent Bioanalyzer 4200 (Agilent technologies, Santa Clara, CA, US). RNA that passed the quality inspection can be used for subsequent library construction.
- RNA-removed RNA was fragmented, double-stranded cDNA synthesized, second strand degraded, end repaired, A added to the 3' end, adapter ligated, and amplified.
- sequencing by synthesis put the flow cell with the cluster on the machine, and add four fluorescently labeled dNTPs, DNA polymerase and adapter primers to the flow cell for amplification.
- SBS sequencing by synthesis
- each sequencing cluster extends the complementary chain
- each fluorescently labeled dNTP is added to release corresponding fluorescence.
- the sequencer captures the fluorescent signal and converts the light signal into a sequencing peak through computer software to obtain the fragment to be tested. sequence information.
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- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Pharmacology & Pharmacy (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- General Health & Medical Sciences (AREA)
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Abstract
一种治疗银屑病的外用中药组合物,包括按照重量份数计的如下原料:茜草10~20份,紫草10~20份,夏枯草10~20份,生甘草30~50份。所述中药组合物中各药味共奏清热解毒、凉血散瘀之功,外用治疗银屑病有显著疗效。
Description
本发明属于银屑病治疗技术领域,尤其涉及一种治疗银屑病的外用中药组合物及其制备方法与应用。
银屑病是一种常见的慢性复发性炎症性皮肤病。基本皮损为红斑、丘疹,或斑块上覆有多层银白色鳞屑。发生部位多见于身体表面及头部,常是红斑上反复出现多层银白色干燥鳞屑。银屑病的病因并不完全清楚且顽固难治,被世界卫生组织列为十大顽症之一。目前对银屑病的治疗,多是采用局部外用药物,如角质促成剂、皮质类因醇霜剂等;而对全身使用的药物,则有免疫抑制剂、维甲酸、抗生素、皮质类因醇等。这些药物对银屑病是有较好的近期治疗效果,但长期使用副作用大,并且复发率较高。比如甲氨蝶呤是银屑病一线***治疗药物,部分患者用药后存在轻微不良反应,表现为腹泻、恶心、食欲缺乏、溃疡、嗜睡、皮肤糜烂等。维甲酸类药物多用于治疗中重度的寻常型、红皮病型及腺癌型银屑病,但对关节型银屑病效果欠佳,常见不良发应包括皮肤粘膜症状、眼毒性、肝酶或血脂升高、骨髓发育异常及致畸性,需监测血脂及生长曲线图等指标。生物制剂(如阿达木单抗、英夫利昔单抗、依法利珠单抗等)治疗银屑病虽见效快,但是存在很多副作用,如部分患者使用后出现结节性血管炎、严重感染、心衰及恶性肿瘤风险。外用糖皮质激素可能会出现皮肤萎缩、毛细血管扩展、萎缩纹、紫癜、多毛等症状。
中医药在改善轻中度银屑病患者临床症状、减少复发率方面表现出了一定的优势。但中药内服治疗周期长,故患者依从性不强,且部分药物存在潜在肝肾损伤可能,患者往往无法长期服用。随着人们对生活品质及身体健康的高水平要求,人们越来越要求更全面的治疗方法,银屑病的临床治疗方案应遵循更安全、有效、经济和便捷的原则。
发明内容
有鉴于此,本发明的目的在于提供一种治疗银屑病的中药组合物,各药味协同共奏清热解毒、凉血散瘀之功,在外用治疗银屑病上有显著疗效,且
具有安全、经济、操作便捷的优势。
为了实现上述发明目的,本发明提供了以下技术方案:
本发明提供了一种治疗银屑病的外用中药组合物,包括按照重量份数计的如下原料:茜草10~20份,紫草10~20份,夏枯草10~20份,生甘草30~50份。
优选的,所述外用中药组合物的制剂类型包括水剂、油剂、散剂、酊剂、洗剂、乳膏剂、软膏剂、沐浴剂和凝胶剂。
本发明还提供了一种上述外用中药组合物的制备方法,包括如下步骤:将茜草、紫草、夏枯草和生甘草与油剂混合,浸泡,小火煎至焦黄色,去除药渣,得药油。
优选的,所述油剂包括葵花籽油或芝麻油。
优选的,茜草、紫草、夏枯草和生甘草四种原料总和与油剂的料液比为1g:4.1ml~9.2ml。
优选的,所述浸泡的时间为24h~28h。
本发明还提供了一种上述外用中药组合物在制备治疗银屑病产品中的应用。
优选的,所述外用中药组合物通过影响角质形成细胞分化、细胞因子活性和/或表皮细胞分化,发挥治疗作用。
优选的,所述外用中药组合物通过参与细胞因子受体相互作用、IL-17信号通路、***信号通路和/或蛋白质消化吸收途径,发挥治疗作用。
优选的,所述产品包括药品。
本发明的有益效果:
本发明外用中药组合物中茜草凉血行血、祛瘀活血,为君药,紫草清热解毒、凉血止血,夏枯草清热解毒,共为臣药,生甘草为佐,解毒和药,四药配合共奏清热解毒,凉血润燥之功,能够很好的治疗银屑病,具有疗效明确显著、安全、经济且操作便捷的优势。
图1各组小鼠银屑病相关表型的变化(n=4),其中A:各组小鼠第12天背部与耳部皮损外观;B:各组小鼠背部皮肤PASI评分(红斑、鳞屑、厚度及总分);C:各组小鼠耳朵厚度及耳部皮肤PASI评分(红斑、鳞屑);注:与模型组相比,*p<0.05,**p<0.01,***p<0.001;与正常组相比,#p<0.05,##p<0.01,###p<0.001;
图2各组小鼠组织病理变化,其中A:各组小鼠背部与耳部组织病理学变化(×200),B:各组小鼠第12天背部厚度定量,C:各组小鼠第12天耳部厚度定量,注:与模型组相比,*p<0.05,**p<0.01;与正常组相比,###p<0.001;
图3数据分析流程;
图4IMQ+SCY组和IMQ组背部皮肤组织的mRNA-seq分析结果,其中A:IMQ+SCY组与IMQ组差异表达蛋白(DEPs)火山图,绿色的圆点表示下调的DEPs,红色的点表示上调的DEPs,灰色的点表示非显著的DEPs;B:IMQ+SCY组与IMQ组289个DEPs热图,根据基因表达量进行聚类分析,红色表示高表达的DEPs,绿色表示低表达的DEPs,每行表示每个DEPs在不同组中的表达水平;
图5SCY干预后银屑病皮损中差异基因的GO与KEGG富集分析,其中A:对SCY干预后的DEPs进行GO分析结果,B:对SCY干预后的DEPs进行KEGG分析结果。
本发明提供了一种治疗银屑病的外用中药组合物,包括按照重量份数计的如下原料:茜草10~20份,紫草10~20份,夏枯草10~20份,生甘草30~50份。
本发明对于上述各原料的具体来源没有特殊限定。在本发明中,所述各原料的重量份数优选的为茜草13~18份,紫草12~16份,夏枯草14~17份,生甘草35~45份,更优选的为茜草14~16份,紫草14~15份,夏枯草15~16份,生甘草38~42份。本发明所述治疗银屑病的外用中药组合物优选的可制成水剂、油剂、散剂、酊剂、洗剂、乳膏剂、软膏剂、沐浴剂和凝胶剂。
本发明还提供了一种上述外用中药组合物的制备方法,包括如下步骤:将茜草、紫草、夏枯草和生甘草与油剂混合,浸泡,小火煎至焦黄色,去除药渣,得药油。
在本发明中,所述油剂优选的包括葵花籽油或芝麻油,茜草、紫草、夏枯草和生甘草四种原料总和与油剂的料液比优选为1g:4.1ml~9.2ml,更优选为1g:5ml~8.5ml。本发明对于葵花籽油或芝麻油的具体来源没有特殊限定,采用本领域常规市售产品均可。在本发明中,所述浸泡的时间优选为24h~28h,更优选为25h~26h。本发明对于去除药渣的具体方式没有特殊限定,采用本领域常规去除方法均可。
本发明还提供了一种上述外用中药组合物在制备治疗银屑病产品中的应用。
在本发明中,所述产品优选的包括药品。本发明所述外用中药组合物优选的通过影响角质形成细胞分化、细胞因子活性和/或表皮细胞分化,发挥治疗作用,更优选的通过参与细胞因子受体相互作用、IL-17信号通路、***信号通路和/或蛋白质消化吸收途径,发挥治疗作用。
下面结合实施例对本发明提供的技术方案进行详细的说明,但是不能把它们理解为对本发明保护范围的限定。
下述实施例中,如无特殊说明,均为常规方法。
下述实施例中所用的材料、试剂等,如无特殊说明,均可从商业途径得到。
实施例1
取紫草10g,茜草10g,夏枯草10g,生甘草50g,混合后用500ml葵花籽油浸泡24h,小火煎至药材变成焦黄色,去除药渣,得四草油,置于4℃冰箱备用。
实施例2
取紫草20g,茜草20g,夏枯草20g,生甘草30g,混合后用550ml芝麻油浸泡24h,小火煎至药材变成焦黄色,去除药渣,得四草油,置于4℃冰箱备用。
实施例3
取紫草15g,茜草14g,夏枯草16g,生甘草40g,混合后用600ml葵花籽油浸泡24h,小火煎至药材变成焦黄色,去除药渣,得四草油,置于4℃冰箱备用。
实施例4
将雄性、6-8周龄、体重20g±2g的BABLc小鼠随机分为3组,每组4只小鼠:(1)空白对照组(Control组):用凡士林乳膏62.5mg每日涂抹小鼠背部与双耳廓内外侧。(2)模型组(IMQ组):将5%IMQ乳膏62.5mg分别涂抹小鼠背部与双耳廓内外侧。6h后用凡士林乳膏62.5mg涂抹小鼠背部与双耳廓内外侧。(3)四草油组(IMQ+SCY组):将5%IMQ乳膏62.5mg涂抹小鼠背部与双耳廓内外侧。6h后用实施例1制备所得的150ul四草油涂抹小鼠背部与双耳廓内外侧。小鼠所有治疗从应用IMQ之日起(第0天)开始,每天1次,连续12天。
采用银屑病临床面积和严重程度指数(PASI)对小鼠背部与耳部皮肤炎症的严重程度进行评分,包括红斑、厚度和鳞屑。分值表示严重程度:0(无),1(轻度),2(中度),3(重度)及4(极重度)。小鼠在采集标本前禁食,允许饮水12h。第13天实验结束后小鼠用CO2处死,分别采集耳朵皮损、背部皮损进行进一步实验。
实验结果
1)四草油可改善IMQ诱导的银屑病样小鼠背部与耳部皮损
结果如图1所示,IMQ造模第12天,Control组外用凡士林软膏,背部与耳部皮肤均无红斑、鳞屑、皮肤增厚现象;IMQ组小鼠背部与耳部均出现明显红斑、鳞屑、皮肤增厚现象。与Control组相比,IMQ组PASI评分均显著升高,耳朵肿胀明显。与IMQ组相比,IMQ+SCY组小鼠PASI评分及耳朵肿胀程度减轻。
2)四草油可改善IMQ诱导的银屑病样小鼠模型的组织病理
结果如图2所示:各组小鼠的背部与耳部的组织病理切片显示,IMQ造模第12天,Control组小鼠背部与耳部皮肤表皮层较薄,未出现炎性细胞浸润。与Control组相比,IMQ组小鼠背部与耳部皮肤表现为角化过度伴角化不全,角化不全区可见中性粒细胞微脓肿,颗粒层明显减少,基层增厚表皮突向下延伸。可观察到中性粒细胞、淋巴细胞等浸润(p<0.001)。IMQ+SCY组小鼠背部与耳部皮肤炎性细胞浸润和表皮增生减少(p<0.05)。
实施例5
为了探究本发明四草油治疗银屑病的作用机制,为四草油外用提供理论依据,特此进行下述mRNA-seq分析:
1.建库流程
(1)总RNA质检
采用RNeasy Mini Kit(Cat#74106,Qiagen)并且根据生产厂商提供的标准操作流程进行实施例4中IMQ组和IMQ+SCY组小鼠样品的total RNA抽提。总RNA经NanoDrop ND-2000分光光度计及Agilent Bioanalyzer 4200(Agilent technologies,Santa Clara,CA,US)进行质检,质检合格的RNA可进行后续的文库构建。
(2)文库构建与质检
对于特定真核物种,利用磁珠去除rRNA。接下来对去除rRNA的RNA进行片段化、双链cDNA合成、降解第二链、末端修复、3’末端加A、连接接头、扩增实验。构建的文库使用2.0 Fluorometer检测浓度,Agilent 2100检测大小。
(3)上机测序
质检合格的文库,准备进行Illumina测序,测序策略为PE150。测序基本原理为边合成边测序(SBS,Sequencing by Synthesis):将携有cluster的flow cell上机,并在flow cell中加入四种荧光标记的dNTP、DNA聚合酶以及接头引物进行扩增,在每一个测序簇延伸互补链时,每加入一个被荧光标记的dNTP就能释放出相对应的荧光,测序仪通过捕获荧光信号,并通过计算机软件将光信号转化为测序峰,从而获得待测片段的序列信息。
2.数据分析流程
如图3所示。
3.实验结果
(1)四草油干预后差异基因表达鉴定
通过mRNA-seq分析,共鉴定出718个差异基因。根据差异基因筛选条件:Foldchange=2倍且p-value<0.05,在IMQ+SCY组和IMQ组中筛选了160个上调基因和129个下调基因。结果见火山图(图4A)和聚类分析热图(图4B)。
(2)GO和KEGG富集分析揭示差异基因的功能和生物学途径
为了揭示差异表达蛋白(DEPs)的功能和生物学途径,对IMQ+SCY组SCY干预后小鼠银屑病皮损中的差异基因进行了GO和KEGG富集分析。GO分析表明IMQ+SCY组的DEPs主要与角质形成细胞分化、细胞因子活性、表皮细胞分化等有关(图5A)。KEGG富集分析表明,IMQ+SCY组的DEPs主要参与细胞因子受体相互作用、IL-17信号通路、***信号通路、蛋白质消化吸收等(图5B)。
以上所述仅是本发明的优选实施方式,应当指出,对于本技术领域的普通技术人员来说,在不脱离本发明原理的前提下,还可以做出若干改进和润饰,这些改进和润饰也应视为本发明的保护范围。
Claims (10)
- 一种治疗银屑病的外用中药组合物,其特征在于,包括按照重量份数计的如下原料:茜草10~20份,紫草10~20份,夏枯草10~20份,生甘草30~50份。
- 根据权利要求1所述的外用中药组合物,其特征在于,所述外用中药组合物的制剂类型包括水剂、油剂、散剂、酊剂、洗剂、乳膏剂、软膏剂、沐浴剂和凝胶剂。
- 权利要求1所述外用中药组合物的制备方法,其特征在于,包括如下步骤:将茜草、紫草、夏枯草和生甘草与油剂混合,浸泡,小火煎至焦黄色,去除药渣,得药油。
- 根据权利要求3所述的制备方法,其特征在于,所述油剂包括葵花籽油或芝麻油。
- 根据权利要求3所述的制备方法,其特征在于,茜草、紫草、夏枯草和生甘草四种原料总和与油剂的料液比为1g:4.1ml~9.2ml。
- 根据权利要求3所述的制备方法,其特征在于,所述浸泡的时间为24h~28h。
- 权利要求1所述外用中药组合物在制备治疗银屑病产品中的应用。
- 根据权利要求7所述的应用,其特征在于,所述外用中药组合物通过影响角质形成细胞分化、细胞因子活性和/或表皮细胞分化,发挥治疗作用。
- 根据权利要求7所述的应用,其特征在于,所述外用中药组合物通过参与细胞因子受体相互作用、IL-17信号通路、***信号通路和/或蛋白质消化吸收途径,发挥治疗作用。
- 根据权利要求7所述的应用,其特征在于,所述产品包括药品。
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CN104689132A (zh) * | 2015-04-01 | 2015-06-10 | 王惠兰 | 一种治疗银屑病的中药组合物 |
CN111000913A (zh) * | 2018-10-08 | 2020-04-14 | 国药集团健康产业研究院有限公司 | 一种治疗银屑病的中药组合物 |
CN115317538A (zh) * | 2022-08-23 | 2022-11-11 | 上海市皮肤病医院 | 一种治疗银屑病的外用中药组合物及其制备方法与应用 |
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CN103655814A (zh) * | 2013-11-26 | 2014-03-26 | 上海中医药大学附属岳阳中西医结合医院 | 一种治疗银屑病的中药组合物及其应用 |
CN104689132A (zh) * | 2015-04-01 | 2015-06-10 | 王惠兰 | 一种治疗银屑病的中药组合物 |
CN111000913A (zh) * | 2018-10-08 | 2020-04-14 | 国药集团健康产业研究院有限公司 | 一种治疗银屑病的中药组合物 |
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