痛痒消洗剂可促进大鼠肛瘘术后的创面愈合

doi:10.12122/j.issn.1673-4254.2024.07.05

南方医科大学学报 ›› 2024, Vol. 44 ›› Issue (7): 1256-1265.doi: 10.12122/j.issn.1673-4254.2024.07.05

痛痒消洗剂可促进大鼠肛瘘术后的创面愈合

王琳月¹(), 戚文月², 高记华³(), 田茂生³, 许建成³

^1.河北中医药大学研究生学院，河北石家庄 050091
^2.河北师范大学校医院，河北石家庄 050010
^3.河北中医药大学第一附属医院肛肠科，河北石家庄 050013

收稿日期:2024-04-11 出版日期:2024-07-20 发布日期:2024-07-25
通讯作者: 高记华 E-mail:tgwangly@163.com;gaojihua2005@163.com
作者简介:王琳月，在读博士研究生，医师，E-mail: tgwangly@163.com
基金资助:
国家自然科学基金(82174381);河北中医学院研究生创新能力培养资助项目(XCXZZBS2023021)

Tongyangxiao Lotion promotes postoperative wound healing in a rat model of anal fistula by downregulating inflammatory factors and suppressing inflammation

Linyue WANG¹(), Wenyue QI², Jihua GAO³(), Maosheng TIAN³, Jiancheng XU³

^1.Graduate School, Hebei University of Traditional Chinese Medicine, Shijiazhuang 050091, China
^2.School Hospital, Hebei Normal University, Shijiazhuang 050010, China
^3.Department of Anorectal Medicine, First Affiliated Hospital of Hebei University of Traditional Chinese Medicine, Shijiazhuang 050013, China

Received:2024-04-11 Online:2024-07-20 Published:2024-07-25
Contact: Jihua GAO E-mail:tgwangly@163.com;gaojihua2005@163.com
Supported by:
National Natural Science Foundation of China(82174381)

摘要/Abstract

摘要：

目的通过网络药理学及分子对接技术预测痛痒消洗剂促进创面愈合的潜在靶点，并通过动物实验进行验证。方法通过TCMSP、BATMAN数据库筛选痛痒消洗剂活性成分及药物靶点，通过GeneCards、OMIM数据库筛选创面愈合相关靶点，对药物及创面相关靶点取交集，进行PPI分析、GO和KEGG富集分析，25只SD大鼠随机分为模型组（Model），对照组（PP），痛痒消洗剂低、中、高剂量组（TYX-L、TYX-M、TYX-H），n=5。每组大鼠造模后，Model、PP及TYX各组分别予生理盐水、高锰酸钾溶液及低、中、高浓度痛痒消洗剂药液（药物浓度分别为171 mg·mL^-1·d^-1，342 mg·mL^-1·d^-1及684 mg·mL^-1·d^-1）塌渍创面，10 min/次，1次/d，治疗14 d。HE染色观察肉芽组织病理变化，免疫组化观察肉芽组织TNF-α蛋白表达，RT-PCR法检测肉芽组织白介素-1β（IL-1β）、肿瘤坏死因子-α（TNF-α）、白介素-6（IL-6） mRNA水平，Western blotting法检测肉芽组织中IL-1β、TNF-α蛋白表达，ELISA法检测肉芽组织IL-6蛋白表达。结果网络药理学获取痛痒消洗剂与创面的共同靶点156个，炎症因子IL-1β、TNF-α、IL-6是痛痒消洗剂促进创面愈合的潜在靶点。痛痒消洗剂6个核心成分与IL-1β、TNF-α、IL-6的结合能都小于-6.0 kcal/mol，较好地与关键蛋白结合。Model组比较，TYX各组、PP组炎细胞浸润较少、可见大量成纤维细胞且有较多胶原沉积，TYX各组大鼠肉芽组织IL-6、IL-1β、TNF-α mRNA及IL-6蛋白表达降低（P<0.05），TYX-M、TYX-H组TNF-α蛋白表达降低（P<0.05）；与PP组相比，TYX-M、TYX-H组TNF-α mRNA，TYX-L、TYX-H组IL-6 mRNA表达降低（P<0.05）。结论痛痒消洗剂可通过下调炎症因子表达、促进炎症消退而加快组织愈合。

关键词: 创面愈合, 肛瘘, 网络药理学, 白介素, 肿瘤坏死因子

Abstract:

Objective To explore the mechanism of Tongyangxiao Lotion (TYX) for promoting wound healing following surgery for anal fistula. Methods The active ingredients and drug targets of TYX were explored using TCMSP and BATMAN databases, and the targets associated with wound healing were screened using GeneCards and OMIM databases; the intersecting drug and wound-related targets were analyzed with protein-protein interaction (PPI) analysis and GO and KEGG enrichment analyses. In 25 SD rat models with simulated anal fistula surgery, the effect of wound dressing with TYX at low, medium and high doses (once daily for 14 days) on wound healing were assessed in comparison with potassium permanganate (PP) solution. The granulation tissues collected from the wounds were examined for pathological changes with HE staining and for TNF-α expression using immunohistochemistry. The expressions of 1β, TNF-α, IL-6 mRNA and proteins in the granulation tissue were detected using RT-qPCR, Western blotting or ELISA. Results Network pharmacology analysis yielded 156 common targets between TYX and wound healing, and among them IL-1β, TNF-α, and IL-6 were identified as potential targets of TYX for promoting wound healing. Six core components of TYX were capable of binding to IL-1β, TNF-α, and IL-6 with binding energies all below -6.0 Kcal/mol. In the rat models, the wounds with TYX and PP solution dressing showed significantly reduced inflammatory cell infiltration and increased fibroblasts and collagen deposition. TYX at the 3 doses and PP solution all significantly reduced the expressions of IL-6, IL-1β, TNF-α mRNA and IL-6 protein in the granulation tissues, but TYX at the medium and high doses produced significantly stronger effects than PP solution for lowering TNF-α protein expression and mRNA expressions of TNF-α and IL-6. Conclusion TYX accelerates wound healing by down-regulating the inflammatory factors and reducing inflammation in the wounds.

Key words: wound healing, anal fistula, network pharmacology, interleukin, tumor necrosis factor-α

王琳月, 戚文月, 高记华, 田茂生, 许建成. 痛痒消洗剂可促进大鼠肛瘘术后的创面愈合[J]. 南方医科大学学报, 2024, 44(7): 1256-1265.

Linyue WANG, Wenyue QI, Jihua GAO, Maosheng TIAN, Jiancheng XU. Tongyangxiao Lotion promotes postoperative wound healing in a rat model of anal fistula by downregulating inflammatory factors and suppressing inflammation[J]. Journal of Southern Medical University, 2024, 44(7): 1256-1265.

图/表 13

图 1 造模过程图

Fig.1 Establishment of rat models following simulated anal fistula surgery. A: Preparation of a depilation area on the back. B: Creating a 2 cm-diameter circular wound and excision of the whole skin. C: Spraying E. coli suspension and application of waterproof stickers. D: Purulent secretion visible on the wound surface.

表1 引物序列

Tab.1 Primer sequences for RT-qPCR

Gene	bp	Sequence of primer
GAPDH	74	F: AGGAAATGATGACCTCCTGAACT
GAPDH	74	R: TGTTTTTGTAAGTATCTTGGTGCCT
IL-6	85	F: AGAGACTTCCAGCCAGTTGC
IL-6	85	R: AGTCTCCTCTCCGGACTTGT
TNF-α	164	F: GGCTTTCGGAACTCACTGGA
TNF-α	164	R: GGGAACAGTCTGGGAAGCTC
IL-1β	121	F: TGGCAACTGTCCCTGAACTC
	121	R: CCCAAGTCAAGGGCTTGGAA

图2 痛痒消洗剂药物-化合物-靶点映射图

Fig.2 Drug-compound-target mapping for TYX Lotion.

图3 痛痒消洗剂促进创面愈合核心靶点PPI图

Fig.3 PPI map of the core targets of TYX Lotion for promoting wound healing.

图4 痛痒消洗剂作用于创面的交集靶点GO富集分析图

Fig.4 GO enrichment analysis of intersecting TYX Lotion and wound targets.

图5 痛痒消洗剂作用于创面的交集靶点KEGG富集图

Fig.5 KEGG enrichment of intersecting TYX Lotion and wound targets.

图6 核心成分与靶点对接热图

Fig.6 Heat map of molecular docking of the core components to the core targets.

图7 活性成分与核心靶点分子对接图

Fig.7 Molecular docking between the active ingredients and the core targets. A: β-sitosterol+IL-1β. B: Apigenin+IL-6. C: Wogonin+TNF-α. Hydrogen bonding and hydrophobic interactions are shown in yellow and blue, respectively.

图8 各组大鼠肉芽组织HE染色

Fig. 8 HE staining of granulation tissue in each group (Original magnification: ×40). A: Model. B: PP. C: TYX-L. D: TYX-M. E: TYX-H.

图9 各组大鼠肉芽组织免疫组化TNF-α的表达

Fig.9 Expression of TNF-α in the granulation tissue of rats in each group detected by immunohistochemical staining. A: Model. B: PP. C: TYX-L. D: TYX-M. E: TYX-H. F: AOD of TNF‑α. *P<0.05.

图10 各组肉芽组织TNF-α、IL-1β、IL-6 mRNA相对表达量

Fig. 10 Relative expression of TNF-α (A), IL-1β (B) and IL-6 (C) mRNA in granulation tissue in each group. *P<0.05 vs Model group. #P<0.05 vs PP group.

表2 肉芽组织IL-6表达量

Tab.2 Expression level of IL-6 in the granulation tissue in each group detected by ELISA

Group	IL-6 (pg/mL)
Model	18.77±1.63
TYX-L	12.70±3.06*
TYX-M	14.45±1.38*
TYX-H	14.05±2.25*
F	4.361
P	0.04

图11 各组肉芽组织TNF-α、IL-1β蛋白表达

Fig. 11 TNF-α and IL-1β protein expression in the granulation tissue in each group. A: Western blotting for detecting protein expression of TNF-α and IL-1β. B, C: Quantitative analysis of TNF-α and IL-1β protein levels. *P<0.05 vs Model group.

参考文献 32

1	盛惠英, 马天林. 肛周疾病与行为方式的分析[J]. 中国当代医药, 2009, 16(9): 44-6. DOI: 10.3969/j.issn.1674-4721.2009.09.024
2	朱煜璋, 郭修田. 中医外治法在良性肛肠疾病中的运用研究进展[J]. 上海中医药杂志, 2023, 57(11): 95-100.
3	杜美萱, 丁康, 谭妍妍, 等. 祛腐生肌中药外用治疗肛瘘术后的Meta分析[J]. 中医临床研究, 2021, 13(26): 123-7. DOI: 10.3969/j.issn.1674-7860.2021.26.040
4	张晓, 余先昊. 益气祛腐生肌法对2型糖尿病老年患者肛瘘术后创面的影响[J]. 中国老年学杂志, 2024, 44(2): 323-7.
5	田茂生, 高记华, 许建成, 等. 贻贝粘蛋白联合痛痒消洗剂治疗Ⅰ度和Ⅱ度混合痔的临床疗效研究[J]. 中国全科医学, 2023, 26(8): 955-62.
6	许建成, 师晨松, 张海深, 等. 痛痒消洗剂对肛周脓肿术后患者的临床疗效[J]. 中成药, 2023, 45(2): 439-43.
7	戚文月, 高记华, 田茂生, 等. 痛痒消洗剂对肛瘘术后创伤大鼠创面愈合的影响[J]. 中华中医药杂志, 2023, 38(3): 1246-9.
8	谢成志, 任建勋. 中药及复方功效的网络药理学研究与思考[J]. 中国实验方剂学杂志, 2024, 30(1): 198-207.
9	张雅明, 叶伟成. 痔科熏洗方对大鼠术后伤口愈合的实验研究[J]. 中国肛肠病杂志, 2000, 447(3): 3-5.
10	Tsai CF, Chen GW, Chen YC, et al. Regulatory effects of quercetin on M1/M2 macrophage polarization and oxidative/antioxidative balance[J]. Nutrients, 2021, 14(1): 67.
11	Beken B, Serttas R, Yazicioglu M, et al. Quercetin improves inflammation, oxidative stress, and impaired wound healing in atopic dermatitis model of human keratinocytes[J]. Pediatr Allergy Immunol Pulmonol, 2020, 33(2): 69-79.
12	Mi YH, Zhong L, Lu SJ, et al. Quercetin promotes cutaneous wound healing in mice through Wnt/β‑catenin signaling pathway[J]. J Ethnopharmacol, 2022, 290: 115066.
13	Liu Y, Li ZN, Li WD, et al. Discovery of β-sitosterol's effects on molecular changes in rat diabetic wounds and its impact on angiogenesis and macrophages[J]. Int Immunopharmacol, 2024, 126: 111283.
14	Cui SY, Jiang HL, Chen L, et al. Design, synthesis and evaluation of wound healing activity for β-sitosterols derivatives as potent Na+/K+-ATPase inhibitors[J]. Bioorg Chem, 2020, 98: 103150.
15	Bai FF, Chen Z, Xu SL, et al. Wogonin attenuates neutrophilic inflammation and airway smooth muscle proliferation through inducing caspase-dependent apoptosis and inhibiting MAPK/Akt signaling in allergic airways[J]. Int Immunopharmacol, 2022, 113(Pt B): 109410.
16	孙鹏. 辐照后皮肤伤口愈合障碍与P53相关凋亡通路的关系[D]. 上海: 中国人民解放军海军军医大学, 2019.
17	叶杰阳, 庞苏滨, 张圣江, 等. 消肿生肌汤对肛瘘术后患者肉芽组织Bax、P53、Caspase-3水平的影响[J]. 长春中医药大学学报, 2021, 37(3): 567-70. DOI: 10.13463/j.cnki.cczyy.2021.03.025
18	李剑庆. 基于IL-17A/JNK/c-Jun通路研究补肺汤对COPD凋亡的影响[D]. 长沙: 湖南中医药大学, 2023.
19	Zhang MY, Ma LJ, Jiang L, et al. Paeoniflorin protects against cisplatin-induced acute kidney injury through targeting Hsp90AA1-Akt protein-protein interaction[J]. J Ethnopharmacol, 2023, 310: 116422.
20	李媛媛, 赵富生, 张可心, 等. 硫化氢对糖尿病大鼠皮肤创面自噬和血管形成的影响[J]. 中国病理生理杂志, 2023, 39(12): 2223-33.
21	魏晓涛, 何志军, 刘涛, 等. 消肿止痛合剂对糖尿病溃疡模型大鼠创面愈合的影响[J]. 中国病理生理杂志, 2023, 39(5): 902-9.
22	Mei H, Yao PL, Wang SS, et al. Chronic low-dose cadmium exposure impairs cutaneous wound healing with defective early inflammatory responses after skin injury[J]. Toxicol Sci, 2017, 159(2): 327-38.
23	Wang YL, Rouabhia M, Zhang Z. Pulsed electrical stimulation benefits wound healing by activating skin fibroblasts through the TGFβ1/ERK/NF‑κB axis[J]. Biochim Biophys Acta, 2016, 1860(7): 1551-9.
24	杨影, 陈东升, 赵艾艾, 等. TNF-α和VEGF协同作用小鼠胚胎干细胞衍生的血管内皮祖细胞促伤口愈合[J]. 中国生物化学与分子生物学报, 2024, 40(2): 240-50. DOI: 10.13865/j.cnki.cjbmb.2023.12.1300
25	Basso FG, Pansani TN, Turrioni AP, et al. Tumor necrosis factor-α and interleukin (IL)-1β, IL-6, and IL-8 impair in vitro migration and induce apoptosis of gingival fibroblasts and epithelial cells, delaying wound healing[J]. J Periodontol, 2016, 87(8): 990-6.
26	何咏磬. IL-1β通过调节stathmin/FAK通路促进人真皮成纤维细胞迁移的作用及机制[D]. 重庆: 中国人民解放军陆军军医大学, 2023.
27	靳立强, 郭海涛, 韩书明. 复方黄柏液涂剂外敷联合泡沫硬化治疗下肢静脉曲张性溃疡疗效及其对TNF-α、IL-6、CRP水平的影响[J]. 中华中医药学刊, 2023, 41(11): 250-3.
28	李锐阳, 董秋霖, 胡金华, 等. 成纤维细胞与间充质干细胞来源外泌体修复急性皮肤创面的比较[J]. 中山大学学报: 医学科学版, 2023, 44(3): 369-78.
29	吴许雄, 汪昭楚, 黄娟, 等. 国医大师陈民藩教授“存体寡损-形神兼顾” 治瘘护肛理论的形成与应用[J]. 时珍国医国药, 2023, 34(11): 2779-81.
30	夏庭伟, 杨越, 李炜弘, 等. 中医学“腐” 的语义内涵探赜[J]. 中华中医药杂志, 2023, 38(5): 2201-3.
31	梅苹苹, 冯文哲, 石鹏, 等. 基于NF-κB/VEGF信号通路探究加味生肌玉红膏促进湿热型肛瘘术后创面愈合的机制[J]. 浙江中医药大学学报, 2024, 48(3): 255-63, 281.
32	杨露, 张永萍, 彭丽华. 中药及其外用制剂调控细胞因子促创面愈合的研究进展[J]. 中国中药杂志, 2021, 46(20): 5173-84. DOI: 10.19540/j.cnki.cjcmm.20210705.601

[1]	王瑾瑾, 崔文飞, 窦雪伟, 尹冰磊, 牛钰琪, 牛羚, 闫国立. 鬼箭羽通过调节EGFR酪氨酸激酶抑制剂耐药信号通路延缓糖尿病肾病的进展[J]. 南方医科大学学报, 2024, 44(7): 1243-1255.
[2]	李睿镈, 高歌, 谢曦, 罗海彬. 槟榔活性成分诱导口腔黏膜下纤维化的机制：基于网络药理学结合临床样本验证[J]. 南方医科大学学报, 2024, 44(5): 930-940.
[3]	李云飞, 杨婧怡, 张颖, 张财霞, 韦宇翔, 王怡颖, 吴宁, 孙见飞, 吴遵秋. 苗药四大血减轻大鼠的类风湿性关节炎：基于下调基质金属蛋白表达[J]. 南方医科大学学报, 2024, 44(4): 739-747.
[4]	陈君洁, 黄传兵, 李明. 健脾滋肾方抑制系统性红斑狼疮患者的足细胞自噬：基于网络药理学和临床研究[J]. 南方医科大学学报, 2024, 44(3): 465-473.
[5]	崔艺馨, 王德财, 谢东晴, 王海明, 徐睿鑫, 唐潇然, 张印. 健脾温阳凝胶剂脐疗治疗脾胃虚弱型慢性腹泻的疗效及机制：一项临床随机对照试验[J]. 南方医科大学学报, 2024, 44(2): 217-225.
[6]	张倩, 张梅奎, 刘颖璐, 王妍, 吕菲菲, 王毓国. 六味酸枣汤治疗围绝经期失眠的作用机制：基于网络药理学与动物实验[J]. 南方医科大学学报, 2023, 43(9): 1536-1547.
[7]	罗敏, 胡鸿彬, 孙烨, 赵新, 曾振华, 刘易林, 武钢. 西维来司他钠治疗COVID-19相关急性呼吸窘迫综合征重症患者的疗效：一项回顾性、单中心、队列研究[J]. 南方医科大学学报, 2023, 43(8): 1259-1267.
[8]	张雪芳, 陈延华, 李宗恒, 尚靖, 袁泽婷, 邓皖利, 骆莺, 韩娜, 殷佩浩, 殷军. 六神丸治疗小鼠结肠炎相关性结直肠癌的作用机制：基于网络药理学和体内验证方法[J]. 南方医科大学学报, 2023, 43(7): 1051-1062.
[9]	刘芳, 张远芳, 刘鹏, 刘佳敏, 刘思妤, 王俊杰. 东革阿里抗炎的物质基础及其作用机制：基于UPLC-Q-TOF-MS/MS和网络药理学方法[J]. 南方医科大学学报, 2023, 43(6): 879-888.
[10]	罗冠峰, 刘华熙, 谢钡, 邓伊健, 谢鹏辉, 赵晓山, 孙晓敏. 肾病III号方治疗慢性肾脏病大鼠肾脏纤维化的作用机制:基于网络药理学和分子对接技术[J]. 南方医科大学学报, 2023, 43(6): 924-934.
[11]	孙洋, 许轶博, 肖林雨, 朱国庆, 李静, 宋雪, 徐磊, 胡建国. 乙酰紫堇灵促进大鼠脊髓损伤后的功能恢复：基于调控EGFR/MAPK信号通路抑制小胶质细胞活化[J]. 南方医科大学学报, 2023, 43(6): 915-923.
[12]	赵宇曦, 赵旭, 朱清楠, 朱炳睿, 张震彬, 陈静. 桂枝甘草汤通过多靶点、多通路治疗心力衰竭：基于网络药理学方法[J]. 南方医科大学学报, 2023, 43(5): 772-782.
[13]	张钰, 涂星, 张燕, 文德鉴, 赵方毓, 袁林, 李文慧. 文王一支笔的抗炎作用机制：基于网络药理学、分子对接及实验验证[J]. 南方医科大学学报, 2023, 43(3): 383-392.
[14]	张铭, 于浩, 邵阳, 宋国栋, 陈铭锐, 刘顺利. 纳米脂肪联合富血小板血浆治疗大鼠压力性损伤创面的实验研究[J]. 南方医科大学学报, 2023, 43(12): 2061-2070.
[15]	徐朦, 张鹏, 张国梁. 益气解毒方治疗原发性肝癌的作用机制：基于网络药理学及分子对接方法[J]. 南方医科大学学报, 2022, 42(6): 805-814.

痛痒消洗剂可促进大鼠肛瘘术后的创面愈合

Tongyangxiao Lotion promotes postoperative wound healing in a rat model of anal fistula by downregulating inflammatory factors and suppressing inflammation

RichHTML

PDF

可视化

摘要/Abstract

引用本文

使用本文

图/表 13

参考文献 32

相关文章 15

编辑推荐

Metrics

本文评价