黄连素调节PI3K/AKT/NF-κB信号通路对慢性湿疹大鼠皮肤损伤的影响

doi:10.11958/20221627

摘要/Abstract

摘要：

目的探讨黄连素对慢性湿疹大鼠皮肤损伤及磷脂酰肌醇-3-激酶（PI3K）/蛋白激酶B（AKT）/核因子-κB（NF-κB）信号通路的影响。方法 60只SD大鼠分为对照组，慢性湿疹组，黄连素低、中、高剂量组和泼尼松组，每组10只。除对照组外，其余组大鼠背部涂抹2，4-二硝基氯苯（DNCB）构建慢性湿疹模型。进行湿疹面积及严重度指数（EASI）评分；酶联免疫吸附试验（ELISA）检测血清组胺、胃泌素释放肽（GRP）、免疫球蛋白E（IgE）、白细胞介素（IL）-4、IL-6、肿瘤坏死因子α（TNF-α）、γ干扰素（IFN-γ）水平；苏木素-伊红（HE）染色观察皮损组织病理学改变；Western blot法检测皮损组织PI3K/AKT/NF-κB信号通路相关蛋白表达。结果与对照组比较，慢性湿疹组大鼠皮损组织受损严重，血清组胺、GRP、IgE、IL-4、IL-6、TNF-α水平以及皮损组织IL-4、IL-6、TNF-α蛋白表达和p-PI3K/PI3K、p-AKT/AKT、p-NF-κB p65/NF-κB p65、p-NF-κB抑制蛋白α（IκBα）/IκBα比值升高，血清和皮损组织IFN-γ降低（P<0.05）。与慢性湿疹组比较，黄连素各剂量组和泼尼松组大鼠皮损组织病理损伤有所改善，EASI评分下降，血清组胺、GRP、IL-4、IL-6、TNF-α水平以及皮损组织IL-6、TNF-α蛋白表达和p-PI3K/PI3K、p-AKT/AKT、p-NF-κB p65/NF-κB p65、p-IκBα/IκBα比值降低，血清和皮损组织IFN-γ升高（P<0.05），同时黄连素中、高组和泼尼松组大鼠血清IgE和皮损组织IL-4降低（P<0.05），且黄连素高剂量组效果更好；黄连素高剂量组和泼尼松组上述指标比较差异无统计学意义（P>0.05）。结论黄连素尤其是高剂量黄连素可抑制PI3K/AKT/NF-κB信号通路激活，减轻免疫失衡和炎症反应，改善慢性湿疹大鼠皮肤损伤。

关键词: 小檗碱, 皮肤疾病, 湿疹性, 免疫, 细胞, 炎症, 疾病模型, 动物, PI3K/AKT/NF-κB信号通路

Abstract:

Objective To investigate the effect of berberine on skin damage and phosphatidylinositol-3-kinase (PI3K)/protein kinase B (AKT)/nuclear factor-κB (NF-κB) signaling pathway in rats with chronic eczema. Methods Sixty SD rats were divided into the control group, the chronic eczema group, the berberine (low, medium and high dose) groups and the prednisone group (n=10 for each group). Except for the control group, the other groups of rats were treated with 2,4-dinitrochlorobenzene (DNCB) on back to establish chronic eczema model. Eczema area and severity index (EASI) scores were measured in groups. Serum levels of histamine, gastrin-releasing peptide (GRP), immunoglobulin E (IgE), interleukin (IL)-4, IL-6, tumor necrosis factor α (TNF-α) and γ interferon (IFN-γ) were detected by enzyme-linked immunosorbent assay (ELISA). Histopathological changes of skin lesions were observed by hematoxylin-eosin (HE) staining. Expression levels of PI3K/AKT/NF-κB signaling pathway-related proteins in skin lesions were detected by Western blot assay. Results Compared with the control group, skin lesions were seriously damaged in the chronic eczema group. Serum levels of histamine, GRP, IgE, IL-4, IL-6 and TNF-α, and protein expression levels of IL-4, IL-6 and TNF-α and p-PI3K/PI3K, p-AKT/AKT, p-NF-κB p65/NF-κB p65 and p-NF-κB inhibitor protein α (IκBα)/IκBα ratio in skin lesions were increased, serum and skin lesion IFN-γ decreased (P<0.05). Compared with the chronic eczema group, the pathological damage of skin lesions of rats was improved in the berberine groups and the prednisone group. EASI score, serum levels of histamine, GRP, IgE, IL-4, IL-6 and TNF-α levels, and protein expression levels of IL-4, IL-6 and TNF-α and p-PI3K/PI3K, p-AKT/AKT, p-NF-κB p65/NF-κB p65, p-IκBα/ IκBα ratio in skin lesions decreased, serum and skin lesion tissue IFN-γ increased (P<0.05). Serum IgE and IL-4 in skin lesions of rats decreased in the berberine groups (medium and high dose groups) and the prednisone group (P<0.05). The effect was better in the high dose berberine group. There were no significant differences in the above indexes between the high dose berberine group and the prednisone group (P>0.05). Conclusion Berberine, especially high dose berberine, can inhibit the activation of PI3K/AKT/NF-κB signaling pathway, reduce immune imbalance and inflammatory response, and improve skin damage in rats with chronic eczema.

Key words: berberine, skin diseases, eczematous, immunity, cellular, inflammation, disease models, animal, PI3K/AKT/NF-κB signal path

中图分类号:

R758.23

图/表 8

参考文献 26

[1]	BARRETT A, HAHN-PEDERSEN J, KRAGH N, et al. Patient-reported outcome measures in atopic dermatitis and chronic hand eczema in adults[J]. Patient, 2019, 12(5):445-459. doi:10.1007/s40271-019-00373-y.
[2]	DUBIN C, DEL DUCA E, GUTTMAN-YASSKY E. Drugs for the treatment of chronic hand eczema: successes and key challenges[J]. Ther Clin Risk Manag, 2020, 16:1319-1332. doi:10.2147/TCRM.S292504.
[3]	陈兴, 黄丽霞, 李忻红, 等. 中药外治湿疹用药规律的数据挖掘研究[J]. 辽宁中医杂志, 2021, 9(1):86-90.
	CHEN X, HUANG L X, LI Q H, et al. Data mining research on medication laws of external treatment of eczema with traditional Chinese medicine[J]. Liaoning J Tradit Chin Med, 2021, 9(1):86-90. doi:10.13192/j.issn.1000-1719.2021.09.023.
[4]	赖亭吉. 消银颗粒联合卤米松乳膏治疗慢性湿疹疗效观察[J]. 东南大学学报:医学版, 2020, 39(2):190-192.
	LAI T J. Curative effect of Xiaoyin granules combined with halometasone cream on chronic eczema[J]. J Southeast Univ(Med Sci Edi), 2020, 39(2):190-192. doi:CNKI:SUN:NJTD.0.2020-02-011.
[5]	WANG J, WANG L, LOU G H, et al. Coptidis Rhizoma:a comprehensive review of its traditional uses,botany,phytochemistry,pharmacology and toxicology[J]. Pharm Biol, 2019, 57(1):193-225. doi:10.1080/13880209.2019.1577466.
[6]	ANDOH T, YOSHIHISA Y, REHMAN M U, et al. Berberine induces anti-atopic dermatitis effects through the downregulation of cutaneous EIF3F and MALT1 in NC/Nga mice with atopy-like dermatitis[J]. Biochem Pharmacol, 2021, 185:114439. doi:10.1016/j.bcp.2021.114439.
[7]	郭斐斐, 杜雪洋, 李丹, 等. 小檗碱乳膏对大鼠耳廓复合痤疮模型皮损及IGF-1,DHEA的影响[J]. 中国皮肤性病学杂志, 2021, 35(10):1103-1110.
	GUO F F, DU X Y, LI D, et al. Effects of berberine cream on skin lesions,IGF-1 and DHEA in rats with auricle acne[J]. Chin J Dermatovenereol, 2021, 35(10):1103-1110. doi:10.13735/j.cjdv.1001-7089.202012139.
[8]	张荣榕, 李宁, 邵帅, 等. 基于P38MAPK信号通路探讨加味黄连膏对小鼠湿疹模型的影响[J]. 中国医院药学杂志, 2020, 40(16):1728-1732.
	ZHANG R R, LI N, SHAO S, et al. Effect of modified huanglian ointment on mouse eczema model based on p38MAPK signal pathway[J]. Chin J Hosp Pharm, 2020, 40(16):1728-1732. doi:10.13286/j.1001-5213.2020.16.08.
[9]	邹效. 黄连素在皮肤疾病中的应用[J]. 科学养生, 2021, 2(3):294.
	ZOU X. Application of berberine in skin diseases[J]. Regimen, 2021, 2(3):294.
[10]	胡志平, 何绍前, 王传明, 等. 甘草酸抑制PI3K/AKT/NF-κB减轻胶原诱导型类风湿关节炎大鼠炎症反应[J]. 中国老年学杂志, 2020, 40(13):2852-2856.
	HU Z P, HE S Q, WANG C M, et al. Glycyrrhizic acid inhibits PI3K/AKT/NF-κB alleviates the inflammatory response in rats with collagen induced rheumatoid arthritis[J]. Chin J Gerontol, 2020, 40(13):2852-2856. doi:10.3969/j.issn.1005-9202.2020.13.054.
[11]	谢婷妃, 袁树珍, 隋晓露, 等. 尿酸诱导人肾小管上皮细胞炎症损伤中PI3K/AKT/NF-κB信号通路的调控机制[J]. 中华肾脏病杂志, 2021, 37(1):36-42.
	XIE T F, YUAN S Z, SUI X L, et al. Uric acid induces inflammatory injury in HK-2 cells via PI3K/AKT/NF-κB signaling pathway[J]. Chin J Nephrol, 2021, 37(1):36-42. doi:10.3760/cma.j.cn441217-20200618-00006.
[12]	朱雨晴, 俞彩霞, 朱杰, 等. 柏蛇湿疹膏对急性,慢性湿疹大鼠皮肤及炎症因子的影响[J]. 中国地方病防治杂志, 2021, 36(1):16-18.
	ZHU Y Q, YU C X, ZHU J, et al. Effect of cypress eczema cream on skin and inflammatory factors in rats with acute and chronic eczema[J]. Chin J Ctrl Endem Dis, 2021, 36(1):16-18.
[13]	郑琳琳, 王星星, 朱新景, 等. 止痛消炎膏对慢性湿疹模型小鼠皮损及TEWL、SCH、湿度的影响[J]. 辽宁中医杂志, 2022, 49(5):192-195.
	ZHENG L L, WANG X X, ZHU X J, et al. Effect of Zhitong Xiaoyan Ointment on skin lesions,TEWL,SCH and humidity of chronic atopic dermatitis model mice[J]. Liaoning J Tradit Chin Med, 2022, 49(5):192-195.
[14]	汪园园, 金明华, 黄颖娟, 等. 苓桂术甘汤联合热量限摄对湿疹模型大鼠皮损的影响及其机制分析[J]. 中国中西医结合杂志, 2020, 40(4):465-469.
	WANG Y Y, JIN M H, HAUNG Y J, et al. Effect of Calorie-restriction Therapy combined Linggui Zhugan Decoction on eczema model rats[J]. Chin J Integ Trad Western Med, 2020, 40(4):465-469. doi:10.7661/j.cjim.20190928.286.
[15]	赵辨. 湿疹面积及严重度指数评分法[J]. 中华皮肤科杂志, 2004, 37(1):3-4.
	ZHAO B. Eczema area and severity index scoring method[J]. Chin J Dermatol, 2004, 37(1):3-4. doi:10.3760/j.issn:0412-4030.2004.01.002.
[16]	刘影, 董燕, 孙道红, 等. 参苓白术散联合多磺酸黏多糖封包对老年慢性湿疹血虚风燥证患者炎症因子和外周血T淋巴细胞亚群的影响[J]. 中华中医药学刊, 2020, 38(5):166-169.
	LIU Y, DONG Y, SUN D H, et al. Effect of Shenling Baizhu Powder combined with Polysulfonic Acid Mucopolysaccharide Encapsulation on inflammatory cytokines and peripheral blood T lymphocyte subsets in elderly patients with chronic eczema and blood deficiency[J]. Chin Arch Trad Chin Med, 2020, 38(5):166-169. doi:10.13193/j.issn.1673-7717.2020.05.039.
[17]	JIANG Y, ZHAO L, CHEN Q, et al. Exploring the mechanism of berberine intervention in ulcerative colitis from the perspective of inflammation and immunity based on systemic pharmacology[J]. Evid Based Complement Alternat Med, 2021, 2021:9970240. doi:10.1155/2021/9970240.
[18]	TEW X N, XIN LAU N J, CHELLAPPAN D K, et al. Immunological axis of berberine in managing inflammation underlying chronic respiratory inflammatory diseases[J]. Chem Biol Interact, 2020, 317:108947. doi:10.1016/j.cbi.2020.108947.
[19]	WOLLENBERG A, BARBAROT S, BIEBER T, et al. Consensus-based European guidelines for treatment of atopic eczema (atopic dermatitis) in adults and children:part Ⅱ[J]. J Eur Acad Dermatol Venereol, 2018, 32(6):850-878. doi:10.1111/jdv.14891.
[20]	ZHOU X L, CHEN L L, WANG J F. Study on the antipruritic mechanism of Zanthoxylum bungeanum and Zanthoxylum schinifolium volatile oil on chronic eczema based on H1R and PAR-2 mediated GRPR pathway[J]. Allergol Immunopathol (Madr), 2022, 50(4):83-96. doi:10.15586/aei.v50i4.486.
[21]	唐学武, 王文革, 张俊红, 等. 脐血TSLP、IgE水平对早期婴儿湿疹的预测价值[J]. 中国皮肤性病学杂志, 2020, 34(8):879-883.
	TANG X W, WANG W G, ZHANG J H, et al. Predictive value of cord blood TSLP,IgE level for early infantile eczema[J]. Chin J Dermatovenereol, 2020, 34(8):879-883. doi:10.13735/j.cjdv.1001-7089.201911057.
[22]	陶文婷, 陈姣姣, 高清华, 等. 小檗碱通过抑制Th1恢复IFN-γ/IL-4平衡减轻迟发型超敏反应[J]. 中国中医基础医学杂志, 2021, 27(3):423-426.
	TAO W T, CHEN J J, GAO Q H, et al. Berberine attenuates delayed type hypersensitivity by inhibiting Th1 and restoring IFN-γ/IL-4 balance[J]. J Basic Chin Med, 2021, 27(3):423-426.
[23]	LIU B, PIAO X, NIU W, et al. Kuijieyuan Decoction improved intestinal barrier injury of ulcerative colitis by affecting TLR4-dependent PI3K/AKT/NF-κB oxidative and inflammatory signaling and gut microbiota[J]. Front Pharmacol, 2020, 11:1036. doi:10.3389/fphar.2020.01036.
[24]	ZHU Q, ENKHJARGAL B, HUANG L, et al. Aggf1 attenuates neuroinflammation and BBB disruption via PI3K/Akt/NF-κB pathway after subarachnoid hemorrhage in rats[J]. J Neuroinflammation, 2018, 15(1):178. doi:10.1186/s12974-018-1211-8.
[25]	WANG L, MA H, XUE Y, et al. Berberine inhibits the ischemia-reperfusion injury induced inflammatory response and apoptosis of myocardial cells through the phosphoinositide 3-kinase/RAC-α serine/threonine-protein kinase and nuclear factor-κB signaling pathways[J]. Exp Ther Med, 2018, 15(2):1225-1232. doi:10.3892/etm.2017.5575.
[26]	WU Y S, LI Z M, CHEN Y T, et al. Berberine improves inflammatory responses of diabetes mellitus in zucker diabetic fatty rats and insulin-resistant HepG2 cells through the PPM1B pathway[J]. J Immunol Res, 2020, 2020:2141508. doi:10.1155/2020/2141508.

组别	给药0周	给药1周	给药2周
慢性湿疹组	7.95±0.90	8.15±0.86	7.65±0.84
黄连素低剂量组	8.05±0.84	6.35±0.68^a	5.20±0.75^a
黄连素中剂量组	7.80±0.96	5.60±0.74^a	4.05±0.68^ab
黄连素高剂量组	8.00±0.85	4.05±0.62^abc	1.85±0.59^abc
泼尼松组	7.90±0.92	4.50±0.75^abc	2.60±0.72^abc
F	0.115	49.105^＊＊	100.996^＊＊

组别	给药0周	给药1周	给药2周
慢性湿疹组	7.95±0.90	8.15±0.86	7.65±0.84
黄连素低剂量组	8.05±0.84	6.35±0.68^a	5.20±0.75^a
黄连素中剂量组	7.80±0.96	5.60±0.74^a	4.05±0.68^ab
黄连素高剂量组	8.00±0.85	4.05±0.62^abc	1.85±0.59^abc
泼尼松组	7.90±0.92	4.50±0.75^abc	2.60±0.72^abc
F	0.115	49.105^＊＊	100.996^＊＊

组别	组胺/（μg/L）	GRP/（ng/L）	IgE/（μg/L）
对照组	18.96±3.21	8.29±1.45	12.51±2.54
慢性湿疹组	85.27±6.52^a	25.64±3.36^a	32.84±4.50^a
黄连素低剂量组	73.16±5.48^b	19.52±3.01^b	28.16±3.38
黄连素中剂量组	58.26±5.34^bc	14.35±2.87^b	22.35±4.06^bc
黄连素高剂量组	34.58±4.65^bcd	10.16±2.05^bcd	17.17±3.65^bcd
泼尼松组	39.01±5.18^bcd	11.08±1.94^bc	18.02±3.27^bc
F	237.003^＊＊	67.590^＊＊	43.445^＊＊

组别	组胺/（μg/L）	GRP/（ng/L）	IgE/（μg/L）
对照组	18.96±3.21	8.29±1.45	12.51±2.54
慢性湿疹组	85.27±6.52^a	25.64±3.36^a	32.84±4.50^a
黄连素低剂量组	73.16±5.48^b	19.52±3.01^b	28.16±3.38
黄连素中剂量组	58.26±5.34^bc	14.35±2.87^b	22.35±4.06^bc
黄连素高剂量组	34.58±4.65^bcd	10.16±2.05^bcd	17.17±3.65^bcd
泼尼松组	39.01±5.18^bcd	11.08±1.94^bc	18.02±3.27^bc
F	237.003^＊＊	67.590^＊＊	43.445^＊＊

组别	IL-4	IL-6
对照组	34.28±5.01	46.58±5.86
慢性湿疹组	75.32±6.85^a	136.19±8.42^a
黄连素低剂量组	66.15±6.42^b	112.34±8.06^b
黄连素中剂量组	55.26±5.86^bc	98.62±7.68^bc
黄连素高剂量组	42.89±5.62^bcd	65.24±6.32^bcd
泼尼松组	40.51±5.13^bcd	68.41±6.84^bcd
F	74.910^＊＊	214.422^＊＊
组别	TNF-α	IFN-γ
对照组	56.48±6.35	39.52±3.15
慢性湿疹组	121.52±8.51^a	21.34±2.37^a
黄连素低剂量组	105.37±8.62^b	26.21±2.85^b
黄连素中剂量组	86.42±7.96^bc	31.16±2.64^bc
黄连素高剂量组	70.36±6.85^bcd	35.08±2.35^bcd
泼尼松组	71.58±7.03^bcd	34.65±2.58^bc
F	102.366^＊＊	61.164^＊＊