复方苦参汤对溃疡性结肠炎小鼠结肠组织中炎症因子及自噬水平的影响

目的 观察复方苦参汤对溃疡性结肠炎（UC）小鼠结肠组织中炎症因子及自噬水平的影响,并探讨其作用机制。方法 将20只SPF级C57BL/6雄性小鼠随机分为正常组、模型组、复方苦参汤组、美沙拉嗪组,每组5只。适应性喂养7 d后,除正常组外,其余3组予以3%葡聚糖硫酸钠（DSS）自由饮用7 d制备UC小鼠模型。同时在造模第1天开始,正常组和模型组每日均以质量分数为0.9%的氯化钠溶液灌胃,复方苦参汤组与美沙拉嗪组分别予复方苦参汤（7.28 g/kg）、美沙拉嗪（0.52 g/kg）灌胃,每日1次,连续灌胃7 d。每天记录小鼠体质量、大便性状及便血情况,进行疾病活动指数（DAI）评分。末次灌胃结束后,取各组小鼠结肠组织,测量结肠长度,肉眼观察肠黏膜炎症损伤程度进行结肠黏膜损伤指数（CMDI）评分。采用苏木精-伊红染色（HE）法观察结肠组织病理变化,酶联免疫吸附试验（ELISA）检测白介素-1β(IL-1β)、白介素-18(IL-18)含量;免疫印迹（Western blot）法及实时荧光定量逆转录聚合酶链式反应（RT-qPCR）法检测自噬相关分子蛋白及mRNA表达。结果 与正常组比较,模型组大...     

岭南山竹子醇提物对溃疡性结肠炎小鼠的作用＊

目的 研究岭南山竹子乙醇提取物（Garcinia oblongifolia ethanol extract， GOEE）对小鼠溃疡性结肠炎（Ulcerative colitis， UC）的作用。方法 UC以3%葡聚糖硫酸钠（Dextran sulphate sodium， DSS）自由饮水造模，将C57BL/C小鼠随机分正常组、模型组、美沙拉嗪组、GOEE低（200 mg?kg^-1）、中（400 mg?kg^-1）、高（800 mg?kg^-1）剂量组，观察小鼠的结肠长度、疾病活动指数（Disease activity index， DAI）、病理组织学评分；检测血清和结肠的白介素-6（IL-6）、肿瘤坏死因子-α（TNF-α）、白介素-1β（IL-1β）、白介素-4（IL-4）、白介素-10（IL-10）、髓过氧化物酶（MPO）、丙二醛（MDA）含量和超氧化物歧化酶（SOD）、谷胱甘肽过氧化物酶（GSH-px）活力。结果 GOEE显著改善溃疡性结肠炎小鼠的结肠长度、DAI指数和结肠病理评分（P < 0.05）；与模型组比，GOEE显著降低UC小鼠血清IL-6、TNF-α、IL-1β的水平和结肠中MPO、MDA的含量（P < 0.05）；提高IL-4、IL-10水平和SOD、GSH-px活力（P < 0.05）。结论 GOEE对小鼠UC具有治疗作用，其机制可能通过调节炎症相关因子的释放和恢复结肠组织的抗氧化能力起作用。     

自噬在溃疡性结肠炎中的作用及中药干预研究进展

溃疡性结肠炎（UC）是一种病因复杂的慢性肠道炎症性疾病。该病发病机制复杂，至今尚未阐明，由多种因素共同促成。其中肠黏膜屏障损伤是UC的基本病理改变。自噬作为细胞的非损伤性应答，通过降解及重吸收等调节肠黏膜免疫、炎症、氧化应激及菌群稳态等多种过程，从而修复受损的肠黏膜屏障，在UC的发生发展中起关键作用。该病在临床上主要使用氨基水杨酸制剂、糖皮质激素及免疫抑制剂等治疗，西医治疗本病起效快，短期疗效确切，但长期使用容易伴随较多不良反应，且部分药品价格昂贵，给患者带来了极大的身心痛苦与经济负担。因此，探索疗效稳定、不良反应小的新疗法刻不容缓。近年来大量研究表明，中药能够多靶点、多效应调节肠黏膜细胞自噬，修复肠黏膜屏障功能，从而遏制UC发展。众多实验显示，中药活性成分或单体、复方可通过调节细胞自噬水平改善肠黏膜免疫、炎症、氧化应激及菌群等以维持肠黏膜屏障正常功能，从而有效干预UC，为防治UC提供了新举措。但目前尚缺乏对中药调节肠黏膜细胞自噬水平防治UC的系统综述。因此，该文基于UC研究现状、自噬过程、中药治疗等，综述了自噬及其关键靶点蛋白与UC的关系，以阐明自噬在UC产生中的关键作用。同时，对近年来靶向调节细胞自噬以治疗UC的中药进行系统总结，以期为UC的治疗及药物研发提供新思路。     

基于数据挖掘对溃疡性结肠炎中医用药规律的分析＊

目的 基于数据挖掘分析溃疡性结肠炎（ulcerative colitis，UC）的中医用药规律。方法 搜集并筛选近十年发表与中医药治疗UC相关的文献，对证型和药物信息进行提取、规范，构建数据库并应用聚类分析、关联规则等方法进行数据挖掘。结果 本研究共提取方剂243首，药物188种，高频用药为甘草、白术、黄连、党参、白芍等28种，以补虚药及清热药为主，药性以温、寒、平为先，五味则偏苦、甘味，用药大多归脾胃经。运用Modeler 15.0软件在S≥14%，C≥90%，L≥1.0条件下挖掘出28个药物关联规则，聚类出6个组方并与各证型相对应。结论 UC证型分布以大肠湿热证、脾肾阳虚证、肝郁脾虚证、脾虚湿困证、脾气虚证、寒热错杂证为主。核心用药多味苦、甘，入脾胃经，功效上以补虚、清热、理气、收涩、渗湿、温里药为主。组方多为四君子汤、参苓白术散等经典方加减而成，药物配伍上体现了从脾论治的治疗思想，清热除湿、调气理血并重，后期注重补火助阳，多药相合，多法并用以标本兼治。     

溃结康激活自噬及调节肠道菌群防治溃疡性结肠炎的作用机制研究

目的:探讨溃结康调控自噬及肠道菌群防治溃疡性结肠炎的作用机制。方法:采用葡聚糖硫酸钠(Dextran sulphate sodium, DSS)制备急性溃疡性结肠炎(Ulcerative colitis, UC)小鼠模型,随机分为正常对照组、模型对照组、柳氮磺胺吡啶0.45 g/kg组、溃结康3.2、6.4、12.8 g/kg组。除正常对照组外,各组小鼠自由饮用3%DSS水溶液7 d,隔天更换新鲜DSS水溶液。每天观察各组小鼠体质量及疾病活动指数变化,并于第8 d试验结束时,脱颈椎处死各组小鼠收集结肠和粪便;HE染色观察结肠组织病理学改变;电镜检测结肠上皮损伤及自噬体形成数量;RT-qPCR及Western-blot法检测结肠微管相关蛋白轻链(Lc3)、苄氯素1(Beclin1)、自噬蛋白(P62)mRNA及蛋白表达;16S核糖体RNA(16S rRNA)测序技术检测小鼠粪便肠道菌群的变化。结果:与正常对照组比较,模型对照组小鼠体质量显著下降、DAI评分显著升高,结肠长度显著降低,结肠黏膜损伤严重,病理CMDI评分显著升高(P<0.01),视野内未见自噬小体,结肠组织中Lc3、B...     

人参败毒散调控自噬修复溃疡性结肠炎黏膜的机制

目的：从调控自噬清除过氧化物角度,探究人参败毒散修复溃疡性结肠炎(UC)肠黏膜的机制。方法：自由饮用3.0%葡聚糖硫酸钠(DSS)溶液诱导UC小鼠模型,雄性C57BL/6J小鼠共60只,随机分为正常组、模型组、阳性药美沙拉嗪组(0.3 g·kg^-1)、人参败毒散高、中、低剂量组(12.35、8.22、4.11 g·kg^-1),每组10只,连续给药灌胃7 d。选用苏木素-伊红(HE)染色观察结肠组织病变,阿利新蓝-过碘酸雪夫氏(PAS/AB)染色观察杯状细胞改变。免疫荧光法检测结肠组织活性氧自由基(ROS)荧光表达;生化法检测超氧化物歧化酶(SOD)活性、丙二醛(MDA)含量。蛋白免疫印迹法(Western blot)检测结肠组织细胞增殖核抗原(PCNA)、微管相关蛋白1轻链3(LC3)、富含亮氨酸重复序列G蛋白偶联受体5(LGR5)、p62的表达水平。结果：与正常组比较,UC模型组结肠HE染色见黏膜上皮结构大量缺失,肠腺破坏,炎性细胞大量浸润,结肠病理损伤评分(TDI)明显上调(P<0.05),杯状细胞大量丢失,SOD活性及LC3Ⅱ/Ⅰ...     

重楼皂苷Ⅰ通过JNK信号通路诱导乳腺癌细胞凋亡和自噬＊

目的 探究重楼皂苷Ⅰ对乳腺癌细胞凋亡、自噬的影响及其作用机制。方法 用不同浓度的重楼皂苷（20、50、80、110、140 μg·mL^-1）处理乳腺癌细胞，细胞计数试剂盒（Cell counting kit，CCK-8）检测细胞增殖抑制率，筛选最适浓度80 μg·mL^-1。JNK信号通路特异性抑制剂SP联合重楼皂苷Ⅰ处理乳腺癌细胞，用膜联蛋白V-碘化丙锭（Annexin V- PI）双染法检测不同处理方式下的细胞凋亡率，蛋白印迹（Western blot，WB）检测不同处理方式下细胞凋亡相关蛋白裂解的半胱氨酸天冬氨酸蛋白酶（cleaved-caspase 3，cle.cap.3）、B淋巴细胞瘤相关的2蛋白（B lymphocytoma-2-associated X protein，Bax），自噬相关蛋白微管相关蛋白轻链3 Ⅰ（Microtubule-associated protein light chain 3 Ⅰ，LC3Ⅰ）、LC3Ⅱ、自噬相关基因（Beclin），Jun氨基末端激酶（c-Jun N-terminal kinase，JNK）信号通路相关蛋白磷酸化JNK（phosphorylation JNK，p-JNK）、JNK。结果 从20、50、80、110、140 μg·mL^-1重楼皂苷中筛选最适浓度80 μg·mL^-1。与阴性对照组相比，实验组乳腺癌细胞的凋亡率明显升高（P<0.05），cle.cap.3、Bax、LC3Ⅱ、Beclin、蛋白表达均显著上调（P<0.05），p-JNK、LC3Ⅰ蛋白表达显著下调（P<0.05）。与实验+DMSO组相比，实验+SP组癌细胞的抑制率和凋亡率均显著降低（P<0.05），cle.cap.3、Bax、LC3Ⅱ、Beclin、下调（P<0.05），p-JNK、LC3Ⅰ上调（P<0.05）。结论 重楼皂苷Ⅰ促进乳腺癌细胞发生凋亡和自噬，JNK信号通路的活性参与药物的调控作用，为重楼皂苷Ⅰ在乳腺癌治疗中的应用提供支持。     

时间医学对溃疡性结肠炎患者排便时间规律的认识及展望＊

溃疡性结肠炎是结肠黏膜连续性的慢性炎症性疾病，目前病因及发病机制不清楚，也无确切有效的治疗方法。时间医学是研究生物体与时间关系以应用到医学领域的一门学科，其主要研究对象是生物的时间节律，这与溃疡性结肠炎患者的排便时间规律相契合。因此，本文分别从中医、西医两个角度，挖掘时间医学对溃疡性结肠炎排便时间规律的认识及启示，探索溃疡性结肠炎治疗的新思路。     

白术黄芪汤及其有效部位组方对小鼠溃疡性结肠炎的疗效观察

目的观察白术黄芪汤及其有效部位组方对小鼠溃疡性结肠炎模型的疗效。方法采用三硝基苯磺酸(TNBS)灌肠法对小鼠进行造模,以疾病活动指数、结肠黏膜组织损伤的大体和病理评分、髓过氧化物酶(MPO)活性为观察指标对其疗效进行综合评价。结果白术黄芪汤及其有效部位组方对造模小鼠的疾病活动指数、结肠黏膜损伤指数、病理组织学评分和MPO活性均有一定疗效;有效部位组方疗效略优,其与模型组比较差异均有显著性意义(P<0.01),与白术黄芪汤组比较差异也有显著性意义(P<0.05)。结论白术黄芪汤及其有效部位组方对小鼠溃疡性结肠炎均有一定疗效,以有效部位组方疗效较好。     

白术内酯Ⅲ介导的肌动蛋白相关蛋白2/3抑制肠上皮间质转化以缓解溃疡性结肠炎的研究

目的：探寻白术内酯Ⅲ（AT-Ⅲ）的关键靶点,探究肌动蛋白相关蛋白2/3（Arp2/3）抑制溃疡性结肠炎（UC）的作用机制。方法：采用有限酶切-质谱法（lip-SMap）筛选大鼠小肠隐窝上皮细胞（IEC-6）特征蛋白,进行基因本体（GO）和京都基因与基因组百科全书（KEGG）富集分析,确定AT-Ⅲ的关键靶点。脂多糖（LPS）与IEC-6共孵育模拟UC模型,酶联免疫吸附实验（ELISA）检测白细胞介素-6（IL-6）,肿瘤坏死因子-α（TNF-α）含量;划痕实验测定IEC-6迁移速率;蛋白质印迹法测定上皮钙黏素（E-cadherin）、波形蛋白（Vimentin）和Arp2/3含量;分子对接评估AT-Ⅲ与关键靶点的结合能力。结果：基于lip-SMap获得的特征蛋白参与调控磷脂酰肌醇3激酶-丝苏氨酸激酶（PI3K-AKT）信号通路和紧密连接（TJ）信号通路;与对照组比较,LPS组中IL-6和TNF-α释放量升高（均P<0.05）,迁移速率变快（P<0.05）,E-cadherin和Vimentin的表达都有不同程度的影响（均P<0.05）;与LPS组比较,LPS+CK666+AT-Ⅲ组显著降低IL-6和TNF-α含量（均P<0.01）,有效逆转LPS诱导的细胞迁移变化及E-cadherin和Vimentin表达（均P<0.05）;分子对接显示,Arp2/3与AT-Ⅲ有较好的结合活性。结论：Arp2/3作为AT-Ⅲ的关键靶点,可通过上皮细胞-间充质转化（EMT）缓解LPS诱导的UC。     

半夏泻心汤通过抑制炎症抑制小鼠溃疡性结肠炎

[目的]观察半夏泻心汤古方剂量及现代临床常用剂量对采用葡聚糖硫酸钠（DSS）自由饮用诱导建立的溃疡性结肠炎（UC）模型小鼠（DSS-UC小鼠）炎症因子的影响,探索其合理的临床用量并从免疫角度探讨其治疗溃疡性结肠炎的作用机制。[方法]将32只C57BL6/J小鼠随机分为空白组8只、模型组8只、半夏泻心汤古方组8只和半夏泻心汤药典组8只。采用2.5%的DSS自由饮用7 d诱导溃疡性结肠炎模型,造模同时对各组小鼠每日对应给予生理盐水、半夏泻心汤古方量方、半夏泻心汤药典量方灌胃14 d。随后苏木精-伊红（HE）染色观察结肠组织病理学变化,酶联免疫吸附测定法（ELISA）检测血清及结肠组织中炎症因子水平。[结果]半夏泻心汤可显著恢复DSS-UC模型小鼠的体质量、降低疾病指数（DAI）。升高DSS-UC小鼠血清及结肠组织中5-羟色胺（5-HT）、白介素（IL）-10水平,降低血清及结肠组织中IL-4、肿瘤坏死因子（TNF）-α水平,但对IFN-γ水平无显著改善。[结论]半夏泻心汤可以通过升高DSS-UC小鼠血清及结肠组织中IL-10、5-HT水平,降低血清及结肠组织中IL-4、TNF-α水平进而改...     

五味子甲素对小鼠溃疡性结肠炎的治疗作用

目的:考察五味子甲素对小鼠溃疡性结肠炎的治疗作用。方法:以三硝基苯磺酸(TNBS)灌肠制备溃疡性结肠炎(ulcerative colitis,UC)小鼠模型,溃疡性结肠炎小鼠随机分为模型组、阳性对照组及五味子甲素高、中、低剂量组(80、40、20 mg/kg),同时设立正常对照组,每组10只。灌胃给药,正常组、模型组分别给予等体积蒸馏水,阳性对照组给予500 mg/kg柳氮磺吡啶(SASP),连续给药14 d。观察五味子甲素对溃疡性结肠炎小鼠临床症状的影响;酶联免疫吸附法(ELISA)检测溃疡性结肠炎小鼠结肠中肿瘤坏死因子-α(TNF-α)及白介素-6(IL-6)水平;用试剂盒检测结肠中髓过氧化物酶(MPO)、超氧化物歧化酶(SOD)及一氧化氮合成酶(TNOS)的活性和丙二醛(MDA)、一氧化氮(NO)的含量。取结肠后HE染色观察组织形态学变化。结果:五味子甲素可改善三硝基苯磺酸诱导的溃疡性结肠炎小鼠临床症状,延长溃疡性结肠炎小鼠结肠转运时间,可显著降低MPO、TNOS活性,升高SOD活性,并降低MDA、NO、IL-6及TNF-α水平。组织病理学检查可见,给予五味子甲素后,溃疡性结肠炎小鼠病变严重程度减轻,动物结肠组织损伤范围明显减少,炎性细胞浸润程度减轻。结论:五味子甲素对三硝基苯磺酸诱导的小鼠溃疡性结肠炎有治疗作用,作用机制可能与其抑制炎症因子NO、IL-6及TNF-α的产生有关。     

反相高效液相色谱法同时测定参苓白术颗粒中白术内酯Ⅱ和白术内酯Ⅲ的含量

目的 建立一种同时测定白术内酯Ⅱ和白术内酯Ⅲ的高效液相色谱检测方法,并分析中成药参苓白术颗粒中白术内酯Ⅱ和白术内酯Ⅲ的含量.方法 样品采用液-液萃取,以乙腈-水(50:50)为流动相,流速1.0 ml· min-1,采用SHIMADZU ODS柱(150 mm×4.6 mm; 5 μm)色谱柱分离;检测波长220 nm.结果 标准曲线线性范围为0.5～50.0 μg·ml-1,线性关系良好(r>0.99),定量限为0.5 μg/ml,回收率98%以上,方法的稳定性、重复性和精密度均较好(RSD<3%).结论 该法操作简便、快速、灵敏、准确,已经成功地用于中成药参苓白术颗粒中白术内酯Ⅱ和白术内酯Ⅲ的含量测定.     

调脾胃升降温针法对溃疡性结肠炎患者Th17/Treg平衡及其细胞因子水平的影响

目的 观察调脾胃升降温针法治疗溃疡性结肠炎的临床疗效及对患者外周血辅助T细胞17(Th17)与调节性T细胞(Treg)比例及相关细胞因子水平的影响.方法 将2018年6月—2020年1月唐山市中医医院收治的溃疡性结肠炎脾虚证患者随机分为2组,对照组38例予美沙拉嗪肠溶片口服,观察组39例在对照组基础上加用调脾胃升降温针...     

Electro-acupuncture promotes gut motility and alleviates functional constipation by regulating gut microbiota and increasing butyric acid generation in mice

ObjectiveAbnormalities in the gut microbiota and intestinal short-chain fatty acid (SCFA) levels are implicated in the pathogenesis of functional constipation (FC). Electro-acupuncture (EA) has been shown to improve constipation-related symptoms and rebalance the gut microbiota. However, it is currently unknown whether the gut microbiota is a key mechanistic target for EA or how EA promotes gut motility by regulating the gut microbiota and SCFAs. Therefore, we assessed the effects of EA in FC mice and pseudo-germfree (PGF) mice to address these questions.MethodsForty female Kunming mice were randomly separated into a normal control group (n = 8), an FC group (n = 8), an FC + EA group (n = 8), a PGF group (n = 8) and a PGF + EA group (n = 8). The FC group and FC + EA group were treated with diphenoxylate to establish the FC model; the PGF group and PGF + EA group were given an antibiotic cocktail to initiate the PGF model. After maintaining the model for 14 d, mice in the FC + EA and PGF + EA groups received EA stimulation at the ST25 and ST37 acupoints, once a day, 5 times per week, for 2 weeks. Fecal parameters and intestinal transit rate were calculated to assess the efficacy of EA on constipation and gastrointestinal motility. Colonic contents were used to quantify gut microbial diversity using 16S rRNA sequencing, and measure SCFA concentrations using gas chromatography-mass spectrometry.ResultsEA significantly shortened the first black stool defecation time (P < 0.05) and increased the intestinal transit rate (P < 0.01), and fecal pellet number (P < 0.05), wet weight (P < 0.05) and water content (P < 0.01) over 8 h, compared with the FC group, showing that EA promoted gut motility and alleviated constipation. However, EA treatment did not reverse slow-transit colonic motility in PGF mice (P > 0.05), demonstrating that the gut microbiota may play a mechanistic role in the EA treatment of constipation. In addition, EA treatment restored the Firmicutes to Bacteroidetes ratio and significantly increased butyric acid generation in FC mice (P < 0.05), most likely due to the upregulation of Staphylococcaceae microorganisms (P < 0.01).ConclusionEA-mediated resolution of constipation occurs through rebalancing the gut microbiota and promoting butyric acid generation.Please cite this article as: Xu MM, Guo Y, Chen Y, Zhang W, Wang L, Li Y. Electro-acupuncture promotes gut motility and alleviates functional constipation by regulating gut microbiota and increasing butyric acid generation in mice. J Integr Med. 2023; Epub ahead of print.     

Salidroside alleviates ulcerative colitis via inhibiting macrophage pyroptosis and repairing the dysbacteriosis-associated Th17/Treg imbalance

Ulcerative colitis (UC) is a chronic inflammatory bowel disease characterized by flora disequilibrium and mucosal immunity disorder. Here, we report that salidroside effectively restricts experimental colitis from two aspects of intestinal macrophage pyroptosis and dysbacteriosis-derived colonic Th17/Treg imbalance. In innate immunity, the upregulated TREM1 and pyroptosis-related proteins in inflamed colons were inhibited by salidroside administration and further experiments in vitro showed that salidroside suppressed LPS/ATP-induced bone marrow-derived macrophages (BMDMs) pyroptosis evident by the decline of LDH and IL-1β release as well as the protein level of NLRP3, caspase-1, and GSDMD p30. Moreover, the TREM1 inhibitor weakened the effect of salidroside on BMDMs pyroptosis, whereas salidroside still could downregulate TREM1 when NLRP3 was inhibited. In adaptive immunity, salidroside improved the gut microflora diversity and Th17/Treg ratio in DSS-induced mice, especially promoting the abundance of Firmicutes. Clearance of the gut flora blocked the benefit of salidroside on colonic inflammation and Th17/Treg adaptive immunity, but transplanting salidroside-treated foecal bacterium into flora-depleted wild mice reproduced the resistance of salidroside to gut inflammation. Taken together, our data demonstrated that salidroside protected experimental colitis via skewing macrophage pyroptosis and Th17/Treg balance, indicating its potential effect on UC and other immune disorders.     

Nigakinone alleviates DSS-induced experimental colitis via regulating bile acid profile and FXR/NLRP3 signaling pathways

The correlation of bile acid (BA) metabolism disorder with the pathogenesis of ulcerative colitis (UC) is realized nowadays. Farnesoid X receptor (FXR), a controller for BA homeostasis and inflammation, is a promising target for UC therapy. Nigakinone has potential therapeutic effects on colitis. Herein, we investigated the anti-UC effects and mechanism of nigakinone in colitic animals induced by dextran sulfate sodium (DSS). The related targets involved in the nucleotide-binding oligomerization domain, leucine-rich repeat, and pyrin domain-containing 3 (NLRP3) signaling pathway were measured. BA-targeted metabolomics was employed to reveal the regulatory effects of nigakinone on BA profile in colitis, while expressions of FXR and its mediated targets referring to BA enterohepatic circulation were determined. The critical role of FXR in the treatment of nigakinone for colitis was studied via molecule-docking, dual-luciferase reporter® (DLR™) assays, FXR silencing cells, and FXR knockout mice. Results showed nigakinone attenuated DSS-induced colitis symptoms, including excessive inflammatory response by NLRP3 activation, and injury of the intestinal mucosal barrier. Nigakinone regulated BA disorders by controlling cholesterol hydroxylase and transporters mediated by FXR, then decreased BA accumulation in colon. Molecular-docking and DLR™ assays indicated FXR might be a target of nigakinone. In vitro, nigakinone restrained BA-induced inflammation and cell damage via FXR activation and inhibition of inflammatory cytokines. However, ameliorating effects of nigakinone on colitis were suppressed by FXR knockout or silencing in vivo or in vitro. Taken together, nigakinone ameliorated experimental colitis via regulating BA profile and FXR/NLRP3 signaling pathway.     

Anti-inflammatory effects of Pulvis Fellis Suis extract in mice with ulcerative colitis

Ethnopharmacological relevance

Pulvis Fellis Suis is used in folk medicines to treat intestinal diseases, acute pharyngitis, whooping cough and asthma in China. Although several reports indicate that Pulvis Fellis Suis display diverse biological activities, such as antibacterial, anti-inflammatory and anti-infusorian effects, its effects on ulcerative colitis have not been previously explored.

Aim of the study

The purpose of the present study is to assess the anti-inflammatory effect of Pulvis Fellis Suis (PFS) extract in acute ulcerative colitis model induced by trinitrobenzene sulfonic acid (TNBS) in mice.

Materials and methods

Different doses of Pulvis Fellis Suis extract (100, 200 and 400 mg/kg/day) and sulfasalazine (500 mg/kg/day) were administered by gavage for 7 days after the induction of colitis with TNBS. The efficacy of PFS was studied by macroscopical and histological scoring systems as well as myeloperoxidase (MPO) activity. Serum levels, including tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) were assayed by enzyme-linked immunoassay. The expression of cyclooxygenase (COX)-2 in the colons was assessed by immunohistochemical analysis.

Results

Treatment with PFS significantly attenuated macroscopic damage as compared with TNBS (P < 0.01). Histological analysis showed that PFS improved the microscopic structure and preserved some areas of the colonic mucosa structure. In addition, administration of PFS effectively inhibited COX-2 protein expression and MPO activity accumulation. TNF-α and IL-6 levels were also diminished dose-dependently (P < 0.05, P < 0.01), and IL-6 level obtained had no significant results by small dose of PFS. All the effects of these parameters were comparable to that of the standard sulfasalazine, especially at the highest dose level.

Conclusions

We have shown for the first time that PFS has an anti-inflammatory effect in TNBS-induced ulcerative colitis which might be related to the reduction of up-regulated TNF-α and IL-6 production, and that it may have therapeutic value in the setting of inflammatory bowel disease (IBD).     

黄芩素调控NLRP3/Caspase-1/GSDMD通路介导的焦亡减轻小鼠急性肺损伤作用

目的 探究黄芩素对急性肺损伤（acute lung injure,ALI）小鼠的治疗作用以及对NOD样受体热蛋白结构域3（NOD-like receptor family pyrin domain containing 3,NLRP3）/半胱氨酸天冬氨酸蛋白酶-1（cystein-asparate protease-1,Caspase-1）/消皮素D（gasdermin D,GSDMD）焦亡通路的调控作用。方法 60只雄性C57BL/6小鼠随机分为对照组、模型组、Caspase-1抑制剂VX-765（30 mg/kg）组和黄芩素低、中、高剂量（10、20、40 mg/kg）组,每组10只。除对照组外,其余各组ip脂多糖（lipopolysaccharide,LPS,15 mg/kg）,分别于造模前24 h和造模0.5 h后给予黄芩素或VX-765（30 mg/kg）干预。造模24 h后,检测小鼠肺湿干质量比以评价肺肿胀;苏木素-伊红（HE）染色法检测肺组织病理变化;透射电子显微镜观察肺组织中巨噬细胞损伤情况;ELISA法检测小鼠血清和肺泡灌洗液中白细胞介素-1β（interleukin-1β,IL-1β）、IL-6、IL-18、IL-1α和肿瘤坏死因子-α（tumor necrosis factor-α,TNF-α）水平;采用Western blotting检测肺组织NLRP3、凋亡相关的斑点样蛋白（apoptosis-associated speck-like protein containing CARD,ASC）、cleaved Caspase-1、pro Caspase-1、GSDMD-N以及GSDMD蛋白表达。人源单核细胞白血病THP-1细胞加入100 nmol/L佛波酯诱导24 h,贴壁后随机分为对照组、模型组、VX-765（500 nmol/L）组和黄芩素低、中、高剂量（3、10、30 μmol/L）组,除对照组外,采用1 μg/mL LPS联合5 mmol/L三磷酸腺苷（adenosine triphosphate,ATP）刺激THP-1细胞建立细胞炎症模型,给予黄芩素或VX-765干预,采用CCK-8法检测细胞活力;ELISA法检测上清液中IL-1β、IL-18和IL-1α水平;LDH试剂盒检测上清液中乳酸脱氢酶（lactate dehydrogenase,LDH）活力;免疫荧光评估NLRP3炎症小体组装情况;采用膜联蛋白-V-PE（Annexin-V-PE）/7-氨基放线菌素（7-aminoactinomycin,7-AAD）试剂盒评价细胞焦亡情况;Western blotting检测细胞NLRP3/Caspase-1/GSDMD通路相关蛋白表达。结果 在ALI小鼠模型中,与模型组比较,VX-765组和黄芩素组小鼠肺肿胀程度明显减轻（P<0.01）,肺组织损伤评分显著降低（P<0.01）,肺组织中巨噬细胞炎性病变得到缓解,血清及肺泡灌洗液中IL-6、IL-1β、TNF-α、IL-18、IL-1α水平明显降低（P<0.05、0.01）,肺组织NLRP3、ASC、cleaved Caspase-1、pro Caspase-1、GSDMD-N以及GSDMD蛋白表达水平均显著降低（P<0.05、0.01）。在细胞炎症模型中,与模型组比较,VX-765组和黄芩素组THP-1细胞IL-1β、IL-18、IL-1α分泌量显著降低（P<0.01）,NLRP3/Caspase-1/GSDMD通路相关蛋白表达水平明显降低（P<0.05、0.01）,NLRP3炎症小体组装明显被抑制（P<0.05、0.01）,LDH释放量及细胞焦亡率显著降低（P<0.01）。结论 黄芩素通过抑制NLRP3/Caspase-1/GSDMD通路介导的细胞焦亡,从而改善LPS诱导的小鼠ALI。     

Atractylenolide I and atractylenolide III inhibit Lipopolysaccharide-induced TNF-alpha and NO production in macrophages

In order to clarify the mechanism involved in the antiinflammatory activity of atractylenolide I and atractylenolide III from the rhizomes of Atractylodes macrocephala Koidz, their effects on tumor necrosis factor-alpha (TNF-alpha) and nitric oxide (NO) production in peritoneal macrophages were examined. Atractylenolide I and atractylenolide III decreased the TNF-alpha level in LPS-stimulated peritoneal macrophages in a dose-dependent manner, their IC(50) values were 23.1 microm and 56.3 microm, respectively. RT-PCR analysis indicated that they inhibited TNF-alpha mRNA expression. Furthermore, they inhibited NO production in LPS-activated peritoneal macrophages, the IC(50) value of atractylenolide I was 41.0 microm, and the inhibition ratio of 100 microm of atractylenolide III was 45.1% +/- 6.2%. The activity analysis of inducible nitric oxide synthase (iNOS) indicated that they could inhibit the activity of iNOS, their IC(50) values were 67.3 microm and 76.1 microm, respectively. Western blot analysis showed that atractylenolide I and atractylenolide III attenuated LPS-induced synthesis of iNOS protein in the macrophages, in parallel. These results imply that the antiinflammatory mechanism of atractylenolide I and atractylenolide III may be explained at least in part, by the inhibition of TNF-alpha and NO production. Atractylenolide I showed more potent inhibition than atractylenolide III in the production of TNF-alpha and NO in LPS-activated peritoneal macrophages. So, atractylenolide I could be a candidate for the development of new drugs to treat inflammatory diseases accompanied by the overproduction of TNF-alpha and NO.