首页|特应性皮炎寒湿证病证结合小鼠模型的构建研究

特应性皮炎寒湿证病证结合小鼠模型的构建研究

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目的 构建特应性皮炎(atopic dermatitis,AD)寒湿证病证结合小鼠模型,并对模型进行宏观表征及微观指标评价.方法 将Balb/c小鼠随机分为正常对照组、疾病模型组、寒湿证组、病证结合组,每组 6 只.其中正常对照组小鼠外涂基质液;疾病模型组小鼠外涂二硝基氯苯(dinitrochlorobenzene,DNCB),模拟特应性皮炎疾病模型;寒湿证组小鼠在外涂基质液基础上,采用人工气候箱模拟寒湿环境[温度(4±2)℃、相对湿度(90±5)%],并结合高脂饲料喂养模拟寒湿证;病证结合组小鼠在DNCB造模基础上,结合寒湿证证候模拟方法进行复合造模.采用经皮水分流失测量仪对动物经皮水分流失(transepidermal water loss,TEWL)进行测量和评估;对小鼠皮炎和寒湿证证候情况进行观察评价;对搔抓行为进行记录分析;HE染色观察小鼠背部皮肤组织病理改变,并对其表皮厚度和炎症细胞浸润情况进行分析;Western blot法检测皮肤中紧密连接蛋白 1(Claudin-1)、瞬时受体电位香草酸 3(transient receptor potential vanilloid 3,TRPV3)和TRPV4 蛋白的表达水平;利用流式细胞术分析淋巴结辅助性T细胞17(helper T cells 17,Th17)/调节性T细胞(regulatory T cells,Treg)比值.结果 与疾病模型组比较,病证结合组小鼠的证候评分明显升高(P<0.001),TEWL值明显升高、背部皮肤表皮厚度明显增加(P<0.05),背部皮肤的Claudin-1、TRPV3 蛋白表达量显著降低(P<0.01),而TRPV4 蛋白表达显著升高(P<0.05).结论 采用DNCB进行特应性皮炎疾病造模,并结合寒湿环境及高脂饮食的寒湿证造模方式,能够成功构建特应性皮炎寒湿证病证结合小鼠模型.该研究为后续深入探索特应性皮炎寒湿证病证结合动物模型构建及中药复方对AD的药效学研究提供了较理想的动物模型.
Construction Research of A Disease Syndrome Combination Mouse Model of Cold Dampness Syndrome with Atopic Dermatitis
Objective To establish a mouse model of atopic dermatitis(AD)with cold dampness syndrome,and evaluate the model by detecting macro and micro indicators.Methods Balb/c mice were randomly divided into control group(CON),atopic dermatitis model group(AD),cold dampness syndrome model group(CDS),and a disease-syndrome combination model group(DSC),with 6 mice in each group.The mice in control group were treated with matrix liquid.In AD group,mice were treated with Dinitrochlorobenzene(DNCB)to simulate AD disease model.In CDS group,the mice were treated with matrix liquid,and combined with artificial climate box to mimic the cold-dampness environment[temperature(4±2)℃,relative humidity(90±5)%],and fed with high-fat feed to simulate the cold dampness syndrome.The mice in the DSC group were modeled by using a combination of DNCB and cold dampness syndrome simulation methods.The transepidermal water loss(TEWL),cold dampness syndrome,and scratching behavior of animals were measured.HE staining was used to observe the pathological changes of skin tissue on the back of mice,and the epidermal thickness and inflammatory cell infiltration were analyzed.Western blot was used to detect the expression levels of Claudin-1,TRPV3 and TRPV4.The ratio of Th17/Treg in lymph nodes was analyzed by flow cytometry.Results Compared with the AD group,the syndrome score of mice was significantly increased(P<0.001),the TEWL value and the epidermal thickness of the back skin of mice in the DSC group were significantly increased(P<0.05).The expression of Claudin-1 and TRPV3 proteins in the back skin decreased significantly(P<0.01),while the expression of TRPV4 protein increased significantly(P<0.05)in mice of the DSC group compared with those in the AD group.Conclusion Using DNCB to establish the model of AD,combined with the cold dampness environment and high-fat diet,the mouse model of AD with cold dampness syndrome can be successfully established.This study provides an ideal animal model for further exploring the establishment of animal model of AD combined with cold dampness syndrome,and the efficacy of TCM compound on AD syndrome mouse model.

Atopic dermatitisCold dampness syndromeCombination with disease and syndromeClaudin-1TRPV3TRPV4Th17/Treg cell ratio

吴汶丰、余彦栩、吴清、叶思褀、张瑜、刘俊峰、莫秀梅、陈达灿、晏烽根

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广州中医药大学第二临床医学院,广州 510405

广州中医药大学第二附属医院,广州 510120

粤港澳中医药与免疫疾病研究联合实验室,广州 510120

省部共建中医湿证国家重点实验室,广州 510120

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特应性皮炎 寒湿证 病证结合 Claudin-1 TRPV3 TRPV4 Th17/Treg细胞比值

国家自然科学基金青年基金广东省科技部共建中医湿证国家重点实验室项目广东省科技部共建中医湿证国家重点实验室项目广东省重点领域研发计划广东省科技厅中医药防治难治性慢病重点实验室项目市校(院)企联合资助项目

82004364SZ2021ZZ17SZ2021ZZ392020B11111000102018B0303220122024A03J0544

2024

中国中医基础医学杂志
中国中医研究院基础理论研究所

中国中医基础医学杂志

CSTPCD
影响因子：0.779
ISSN：1006-3250
年,卷(期)：2024.30(9)