Objective: Tuo-Min-Ding-Chuan decoction (TMDC), a traditional Chinese prescription, has demonstrated significant clinical efficacy in treating allergic asthma. This study aimed to investigate the mechanism of TMDC in treating asthma from the perspective of Treg cells and gut microbiota across distinct gut segments (jejunum, ileum, cecum, and colon). Methods: An ovalbumin (OVA)-induced asthma model was established in mice, followed by oral administration of TMDC at high, medium, and low dose. Immune cells and lung inflammation were examined to assess asthma severity. Microbial composition was determined by 16S rRNA sequencing. Antibiotic cocktail and Lactobacillus rhamnosus GG (LGG) were administrated to confirm the key role of specific bacteria. Results: TMDC attenuated lung inflammation (p < 0.01) and eosinophilic infiltration (p < 0.01) as well as IL-4 and IL-5 secretion (p < 0.01); it was also associated with an increase in Treg cells in the lung, small intestine (SI), and colon (p < 0.05). Meanwhile, TMDC restored the number of microbiota species and the Shannon index in the hindgut and reinstated beneficial bacteria, such as Allobaculum and Turicibacter, which were diminished in asthmatic mice. Notably, TMDC significantly enriched Bifidobacterium and Lactobacillus, particularly in the hindgut. Lactobacillus abundance was significantly correlated (p < 0.05) with Treg cells, IL-4, IL-5, and eosinophils. Furthermore, LGG supplementation restored elevated lung inflammation (p < 0.05) and decreased Treg cells (p < 0.01) due to antibiotic-induced microbiota depletion. Conclusion: TMDC alleviated asthma by promoting Treg cell expansion in a Lactobacillus-dependent manner across different gut segments, providing new insights into its therapeutic mechanisms.

ETHNOPHARMACOLOGICAL RELEVANCE:Several children with pneumonia (especially severe cases) have symptoms of cough and expectoration during the recovery stage after standard symptomatic treatment, which eventually results in chronic lung injury. Danggui yifei Decoction (DGYFD), a traditional Chinese formula, has shown clinical promise for the treatment of chronic lung injury during the recovery stage of pneumonia, however, its mechanism of action is yet to be deciphered.AIM OF THIS STUDY:To investigate the therapeutic mechanism of DGYFD for the treatment of chronic lung injury by integrating network pharmacology and transcriptomics.MATERIALS AND METHODS:BALB/c mice were used to establish the chronic lung injury mouse model by intratracheal instillation of lipopolysaccharide (LPS). Pathological analysis of lung tissue, lung injury histological score, lung index, protein levels in bronchoalveolar lavage fluid (BALF), immunohistochemical staining, blood rheology, inflammatory cytokines, and oxidative stress levels were used to evaluate the pharmacological effects of DGYFD. Chemical components of DGYFD were identified using ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). Integrated network pharmacology together with transcriptomics was used to predict potential biological targets. Western blot analysis was used to verify the results.RESULTS:In this study, we demonstrated that DGYFD could improve lung injury pathological changes, decreases lung index, down-regulate NO and IL-6 levels, and regulate blood rheology. In addition, DGYFD was able to reduce the protein levels in BALF, up-regulate the expression levels of occludin and ZO-1, improve the ultrastructure of lung tissues, and reverse the imbalance of AT I and AT II cells to repair the alveolar-capillary permeability barrier. Twenty-nine active ingredients of DGYFD and 389 potential targets were identified by UPLC-MS/MS and network pharmacology, and 64 differentially expressed genes (DEGs) were identified using transcriptomics. GO and KEGG analysis revealed that the MAPK pathway may be the molecular target. Further, we found that DGYFD inhibits phosphorylation levels of p38 MAPK and JNK in chronic lung injury mouse models.CONCLUSIONS:DGYFD could regulate the imbalance between the excessive release of inflammatory cytokines and oxidative stress, repair the alveolar-capillary permeability barrier and improve the pathological changes during chronic lung injury by regulating the MAPK signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE:Paeonol (PAE) and glycyrrhizic acid (GLY) are predominate components of 14 blood-entering ones of Piantongtang No. 1, which is a traditional Chinese medicine prescription for chronic migraine with minimal side effects. Both paeonol and glycyrrhizic acid exhibit analgesic, neuroprotective and anti-inflammatory properties individually. Our previous research has highlighted their combined effect (PAE + GLY) in ameliorating migraine symptoms. However, there are not yet any studies exploring the mechanism of action of PAE + GLY in the treatment of migraine. AIM OF THE STUDY:This research aimed to determine the mechanism of PAE + GLY in ameliorating the recurrent nitroglycerin-induced migraine-like phenotype in rats. MATERIALS AND METHODS:Using a nitroglycerin-induced migraine model via subcutaneous injection in the neck, we evaluated the effect of PAE + GLY on migraine-like symptoms. Behavioural tests and biomarkers analysis were employed, alongside transcriptome sequencing (RNA-seq). Mechanistic insights were further verified utilising reverse transcription quantitative PCR (RT-qPCR), Western blot (WB), ELISA and immunofluorescence (IF) techniques. RESULTS:Following treatment with PAE + GLY, hyperalgesia threshold and 5-hydroxytryptamine (5-HT) levels increased, and migraine-like head scratching, histamine and calcitonin gene-related peptide (CGRP) levels were reduced. RNA-Seq experiments revealed that PAE + GLY upregulated the expression of Glutamate decarboxylase 2 (GAD2) and γ-aminobutyric acid type B receptor subunit 2 (GABBR2) genes. This upregulation activated the GABAergic synapse pathway, effectively inhibiting migraine attacks. Further validation demonstrated an increase in γ-aminobutyric acid (GABA) content in cerebrospinal fluid post PAE + GLY treatment, coupled with increased expression of dural GAD2, GABBR2 and transient receptor potential channel M8 (TRPM8). Consequently, this inhibited the expression of dural cAMP-dependent protein kinase catalytic subunit alpha (PRKACA) and transient receptor potential channel type 1 (TRPV1), subsequently downregulating p-ERK1/2, p-AKT1, IL-1β and TNF-α. CONCLUSIONS:Our findings underscore that PAE + GLY ameliorates inflammatory hyperalgesia migraine by upregulating inhibitory neurotransmitters and modulating the GABBR2/TRPM8/PRKACA/TRPV1 pathway.

Background and aim: Dachengqi Decoction (DCQD) as a classic traditional Chinese medicine has been reported to be effective in treating asthma, but its mechanism remains unknown. This study aimed to reveal the mechanisms of DCQD on the intestinal complications of asthma mediated by group 2 innate lymphoid cells (ILC2) and intestinal microbiota.Experimental procedure: Ovalbumin (OVA) was used to construct asthmatic murine models. IgE, cyto-kines (e.g., IL-4, IL-5), fecal water content, colonic length, histopathologic appearance, and gut microbiota were evaluated in asthmatic mice treated with DCQD. Finally, we administered DCQD to antibiotic -treated asthmatic mice to measure the ILC2 in the small intestine and colon.Results and conclusion: DCQD decreased pulmonary IgE, IL-4, and IL-5 levels in asthmatic mice. The fecal water content, the colonic length weight loss, and the epithelial damage of jejunum, ileum, and colon of asthmatic mice were ameliorated by DCQD. Meanwhile, DCQD greatly improved intestinal dysbiosis by enriching Allobaculum, Romboutsia and Turicibacter in the whole intestine, and Lactobacillus gasseri only in the colon. However, DCQD caused less abundant Faecalibaculum and Lactobacillus vaginalis in the small intestine of asthmatic mice. A higher ILC2 proportion in different gut segments of asthmatic mice was reversed by DCQD. Finally, significant correlations appeared between DCQD-mediated specific bacteria and cytokines (e.g., IL-4, IL-5) or ILC2. These findings indicate that DCQD alleviated the concurrent in-testinal inflammation in OVA-induced asthma by decreasing the excessive accumulation of intestinal ILC2 in a microbiota-dependent manner across different gut locations.(c) 2023 Center for Food and Biomolecules, National Taiwan University. Production and hosting by Elsevier Taiwan LLC. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/ licenses/by-nc-nd/4.0/).

MicroRNAs (miRNAs) and transfer RNA-derived small RNAs (tsRNAs) play critical roles in the regulation of different biological processes, but their underlying mechanisms in diabetes mellitus (DM) are still largely unknown. This study aimed to gain a better understanding of the functions of miRNAs and tsRNAs in the pathogenesis of DM. A high-fat diet (HFD) and streptozocin (STZ)-induced DM rat model was established. Pancreatic tissues were obtained for subsequent studies. The miRNA and tsRNA expression profiles in the DM and control groups were obtained by RNA sequencing and validated with quantitative reverse transcription-PCR (qRT-PCR). Subsequently, bioinformatics methods were used to predict target genes and the biological functions of differentially expressed miRNAs and tsRNAs. We identified 17 miRNAs and 28 tsRNAs that were significantly differentiated between the DM and control group. Subsequently, target genes were predicted for these altered miRNAs and tsRNAs, including Nalcn, Lpin2 and E2f3. These target genes were significantly enriched in localization as well as intracellular and protein binding. In addition, the results of KEGG analysis showed that the target genes were significantly enriched in the Wnt signaling pathway, insulin pathway, MAPK signaling pathway and Hippo signaling pathway. This study revealed the expression profiles of miRNAs and tsRNAs in the pancreas of a DM rat model using small RNA-Seq and predicted the target genes and associated pathways using bioinformatics analysis. Our findings provide a novel aspect in understanding the mechanisms of DM and identify potential targets for the diagnosis and treatment of DM.

目的 基于"肺与大肠相表里"理论探讨鼠李糖乳杆菌对过敏性哮喘小鼠细胞外信号调节蛋白激酶1/2(ERK1/2)和p38丝裂原激活蛋白激酶(p38 MAPK)通路及相关免疫细胞的影响.方法 将18只雌性C57 BL/6小鼠随机分为对照组、模型组、鼠李糖乳杆菌组,每组6只.采用卵白蛋白(OVA)致敏加激发方法构建小鼠过敏性哮喘模型,鼠李糖乳杆菌组于雾化激发前给予鼠李糖乳杆菌灌胃,连续7 d,对照组和模型组给予等量生理盐水灌胃.末次灌胃后取各组小鼠肺组织和结肠组织,HE染色进行组织病理观察;ELISA法检测血清OVA特异性IgE含量,流式细胞术检测肺组织中2型固有淋巴样细胞(ILC2)比例,Western blot法检测肺组织中p-ERK1/2和p-p38 MAPK表达情况.结果 与对照组比较,模型组小鼠支气管及血管周围存在明显炎症细胞浸润,部分肺泡结构消失;结肠腺体排列紊乱,隐窝和杯状细胞明显减少,黏膜及黏膜下层炎性细胞浸润;血清OVA特异性IgE含量、肺组织中ILC2比例、肺组织中p-ERK1/2及p-p38 MAPK相对表达量均明显升高(P均<0.05).与模型组比较,鼠李糖乳杆菌组小鼠支气管炎性细胞浸润明显改善;结肠腺体、杯状细胞增多,排列较为整齐,炎细胞浸润明显减少;血清OVA特异性IgE含量、肺组织中ILC2比例、肺组织中p-ERK1/2及p-p38 MAPK相对表达量均明显降低(P均<0.05).结论 鼠李糖乳杆菌可能通过抑制ERK1/2及p38 MAPK磷酸化,减少肺部ILC2,从而减轻过敏性哮喘的肺部炎症.

目的 基于网络药理学和分子对接技术探讨四臣止咳颗粒治疗慢性支气管炎(CB)急性发作的作用机制,并对其进行实验验证.方法 通过TCMSP、TCMID数据库等筛选四臣止咳颗粒的潜在化学成分,采用SwissTargetPrediction数据库预测活性成分的潜在靶点;在OMIM、GeneCards数据库中检索CB急性发作的相关靶点,对四臣止咳颗粒与CB急性发作的靶点进行PPI网络构建,交互处理得到四臣止咳颗粒治疗CB急性发作的关键靶点,进行GO和KEGG分析;利用分子对接技术对成分与关键靶点蛋白进行分子对接,通过多次气道内雾化给药的方式建立脂多糖(LPS)致大鼠CB急性发作模型加以验证.结果 获得四臣止咳颗粒靶点823个,CB急性发作潜在靶点744个,共同靶点180个,关键靶点包括STAT3、AKT1、NFκB1等25个,涉及toll样受体信号通路、NF-κB信号通路等.分子对接结果显示,STAT3、AKT1、MAPK1、NFκB1等可能为四臣止咳颗粒治疗CB急性发作的关键靶点.体内实验验证结果表明,四臣止咳颗粒可明显减轻大鼠肺组织病理程度,肺泡结构紊乱明显好转,炎症明显减轻,WAm/Pbm、WAi/Pbm、WAt/Pbm显著降低(P＜0.01),肺泡灌洗液(BALF)中促炎因子IL-6和TNF-α含量显著降低(P＜0.01),肺组织NF-κBp-p65蛋白表达均降低.结论 四臣止咳颗粒可能通过下调炎症信号通路NF-κB的激活,减轻炎性细胞浸润和气道重塑程度,调节炎症反应进程,从而起到抗CB急性发作的作用,可为深入四臣止咳颗粒治疗CB急性发作分子机制提供理论依据.

目的:探讨炙甘草汤对特发性肺纤维化(Idiopathic pulmonary fibrosis,IPF)小鼠纤维化相关指标的影响,挖掘炙甘草汤治疗IPF的机制.方法:将60只SPF级ICR小鼠随机分为空白组、模型组、吡菲尼酮组和炙甘草汤组,除空白组外,其余组采用气管滴注博莱霉素(5 mg/kg)方法复制JPF小鼠模型,并给予相应的药物治疗.空白组和模型组小鼠灌胃生理盐水,吡菲尼酮组和炙甘草汤组小鼠分别灌胃吡菲尼酮(50mg/kg)和炙甘草汤(25.4 g/kg),各组均连续给药4周后取材,记录各组小鼠的死亡情况,计算各组肺系数;观察肺组织切片病理变化;碱水解法检测肺组织羟脯氨酸(HYP)含量;比色法检测肺组织丙二醛(MDA)含量、超氧化物歧化酶(SOD)和谷胱甘肽过氧化物酶(GSH-Px)的活性;免疫组化、荧光定量PCR检测α-SMA、COL1A蛋白和mRNA的表达水平.结果:炙甘草汤组小鼠死亡数减少,肺系数显著降低(P＜0.01),炎性细胞浸润和胶原沉积面积大量减少,肺泡结构逐渐修复,HYP、MDA含量降低(P＜0.01),SOD活性(P＜0.05)和GSH-Px活性(P＜0.01)显著增强;α-SMA、COL1A蛋白和mRNA表达均降低(P＜0.01).结论:炙甘草汤通过抑制氧化应激反应,从而抑制成纤维细胞活化,减少细胞质基质沉积,从而减缓IPF疾病进程.

目的 考察前列闭尔通栓对慢性非细菌性前列腺炎(chronic nonbacterial prostatitis,CNP)模型大鼠的治疗作用.方法 将60只大鼠随机均分为空白组、模型组、前列通栓组和前列闭尔通栓低、中、高剂量组,模型组和给药组均注射完全弗氏佐剂0.1 mL 1次.造模1天后直肠给药23 d(前列闭尔通栓低、中、高剂量组分别给药0.33、0.66、0.99 g/d,前列通栓组给药0.42 g/d).分别从前列腺质量,前列腺指数,HE染色病理情况,膀胱内压,卵磷脂小体,白细胞数量,肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白细胞介素-10(interleukin-10,IL-10)、超氧化物歧化酶(superoxide dismutase,SOD)、白细胞介素-8(interleukin-8,IL-8)、丙二醛(malondialdehyde,MDA)含量方面进行分析.结果 与空白组相比,模型组前列腺质量增加、前列腺指数升高、病理评分升高(P<0.05),膀胱内压降低、卵磷脂小体数量减少(P<0.05),白细胞数量增多、TNF-α含量升高(P<0.05),IL-10、SOD含量降低(P<0.05),IL-8、MDA含量升高(P<0.05).与模型组比较,前列闭尔通栓中、高剂量组前列腺质量降低(P<0.05);前列闭尔通栓中剂量组前列腺指数降低(P<0.05)、SOD含量升高(P<0.05);前列闭尔通栓各剂量组病理评分降低(P<0.05)、IL-10含量升高(P<0.05);前列通栓组及前列闭尔通栓各剂量组膀胱内压升高、白细胞数量降低(P<0.05);前列闭尔通栓中、高剂量组及前列通栓组卵磷脂小体数量升高(P<0.05);前列通栓组TNF-α含量降低(P<0.05);前列闭尔通栓高剂量组MDA含量降低(P<0.05).与前列闭尔通栓低剂量组比较,前列闭尔通栓高剂量组病理评分降低(P<0.05)、膀胱内压增加(P<0.05)、IL-8含量降低(P<0.05);前列闭尔通栓中剂量组SOD含量上升(P<0.05).与前列闭尔通栓中剂量组相比,前列闭尔通栓低、高剂量组及前列通栓组卵磷脂小体数量降低(P<0.05)、白细胞数量升高(P<0.05).结论 前列闭尔通栓可能通过降低CNP大鼠的前列腺指数和前列腺质量、升高膀胱内压、减轻病理损伤程度、增加CNP大鼠前列腺液中的卵磷脂小体数量、减少CNP大鼠前列腺液中的白细胞数量,对CNP模型大鼠发挥治疗作用.

With the concept of the gut-lung axis reinforced in recent years, emerging evidence has shown that intestinal homeostasis is vital for lung health. Nevertheless, the impacts of lung homeostasis on intestinal tracts and their mechanism are rarely studied. Our results showed that papain-induced asthmatic mice exhibited apparent colonic injuries compared with controls, including increased intestinal permeability, neutrophil and Th17 infiltration in the colonic lamina propria. Moreover, the intranasal administration of papain aggravated such colonic injuries in mice with dextran sulfate sodium-induced colitis, as evidenced by increased occult blood scores, shortened colon length, and accumulated neutrophils. The level of IL-17A was also higher in the serum of asthmatic mice than wild-type mice. Interestingly, the pathologic scores, the proportion of Th17 cells, and neutrophil infiltration in the colon were markedly reduced after IL-17A blocking. Similarly, longer length, lower pathologic scores, and fewer neutrophils were also observed in the colon of IL-17-deficient asthmatic mice. More importantly, we demonstrated that severe gastrointestinal symptoms could accompany clinical asthmatics. The frequencies of Th17 cells and the mRNA expression of IL-17A in the peripheral blood of these patients were significantly enhanced. Besides, the gastrointestinal symptom rating scale scores positively correlated with the frequencies of Th17 in asthmatics. These findings enlighten that IL-17A aggravates asthma-induced intestinal immune injury by promoting neutrophil trafficking, which facilitates the exploration of new potential biomarkers to treat asthma.

目的 观察中焦点穴对功能性消化不良(functional dyspepsia,FD)大鼠十二指肠肿瘤坏死因子 α(tumor necrosis factor,TNF-α)、干细胞因子(stem cell factor,SCF)/C-Kit原癌基因(C-Kit proto-oncogeneprotein,C-Kit)通路的调节,探讨其对FD大鼠十二指肠动力的影响.方法 将36只雄性SPF级SD大鼠随机分为空白组、模型组、模型阻断组、西药组、点穴组、点穴阻断组,每组各6只.除空白组外,其余各组用轻度夹尾刺激+间断饮食法造模,模型阻断组与点穴阻断组在此基础上注射C-Kit单克隆抗体(ACK2),21天造模成功后给予各组相应干预措施,干预持续14天.6组大鼠在最后1次灌胃后禁食,24小时后称体重,按照10 mL/kg灌服活性碳半固体营养糊,30分钟后麻醉断头,取出大鼠全部小肠,计算小肠推进率,苏木素-伊红染色法(hematoxylin-eosin staining,HE)观察十二指肠的形态及炎性细胞浸润情况.酶联免疫吸附法测定大鼠十二指肠中TNF-α、SCF、C-Kit的含量.蛋白免疫印迹法检测十二指肠中SCF、C-kit、磷脂酰肌醇-3激酶(phosphatidylinositol-3-kinase,PI3K)蛋白的表达.结果 (1)与空白组相比,模型组、模型阻断组、西药组小肠推进率明显降低(P<0.05);与模型组相比,点穴组、点穴阻断组小肠推进率明显上升(P<0.05);模型阻断组与点穴阻断组小肠推进率有显著差异(P<0.05).(2)与空白组相比,模型组、模型阻断组、点穴阻断组SCF、C-Kit表达显著降低、TNF-α表达显著升高(P<0.05);与模型组相比,西药组、点穴组SCF、C-Kit表达显著升高、TNF-α表达显著降低(P<0.05);与点穴组相比,点穴阻断组SCF、C-Kit表达显著降低、TNF-α表达显著升高(P<0.05).(3)与空白组相比,模型组、模型阻断组、点穴阻断组C-Kit、SCF、PI3K蛋白的表达均显著降低(P<0.05),西药组C-Kit、PI3K的表达显著降低(P<0.05);与模型组相比,西药组、点穴组上述蛋白的表达均显著升高(P<0.05);与点穴组相比,点穴阻断组SCF、PI3K的表达显著降低(P<0.05),C-Kit的表达有下降的趋势但无显著差异(P>0.05);西药组与点穴组相比PI3K的表达有显著差异(P<0.05).结论 中焦点穴可以抑制FD大鼠十二指肠TNF-α的生成,减轻十二指肠炎症,进而调节SCF/C-kit通路,激活Cajal间质细胞增殖的相关信号,从而恢复十二指肠动力.

Children are susceptible to pneumonia, which affects their growth and development. Immune disorders and unrepaired alveolar mucosal epithelium following pneumonia cause chronic lung injury. The mechanism of chronic lung injury is unknown and lacks animal models for reference. Therefore, we developed a chronic lung injury young mouse model to simulate the pathological process of children. 3-week-old mice were intratracheal instillation of lipopolysaccharide (LPS) every other day for six weeks. Consequently, the histopathology showed damaged integrity of lung tissue, fibrosis, and abnormally distributed alveolar epithelial cells. The total protein concentration in bronchoalveolar lavage fluid (BALF) was increased, alveolar epithelial type (AT) I cells were abnormal distribution, and AT II cells were reduced. The phosphorylation levels of IKBα and the expression levels of NF-κB p65 in lung tissue were up-regulated. In serum and BALF, the IL-6 was oversecretion, nitric oxide (NO) and superoxide dismutase (SOD) were perturbed secretion, oxidative stress imbalance. In addition, blood viscosity, plasma viscosity, and erythrocyte sedimentation rate (ESR) indexes in hemorheology were increased. In conclusion, it is feasible to construct the mouse model of chronic lung injury, and AT I and AT Ⅱ cells were imbalanced, which paves the way for further investigations on the pathogenesis of chronic lung injury and the efficacy of novel treatments.

The hepatic protective role of Sagittaria sagittifolia polysaccharide (SSP) and its possible mechanism were discussed in mice and L02 hepatocytes injured by heavy metals mixture of Cd + Cr (VI) + Pb + Mn + Zn + Cu. After 30-day intervention, blood and liver samples were collected for the relevant assessments. Methyl thiazolyl tetrazolium (MTT) assay showed 24 h was the best protecting point and the SSP protection at 1 mg/mL was strongest in L02 hepatocytes. SSP can alleviated hepatic injury, as evidenced by significantly decreased the activities of aspartate aminotransferase (AST), alanine aminotransferase (ALT), and the malondialdehyde (MDA) content, also increased the superoxide dismutase (SOD) activity and glutathione (GSH), total sulphydryl (T-SH) contents. SSP effectively reduced pathological damage of mice and accumulation of heavy metals in liver, as well as decreased the level of reactive oxygen species (ROS) in L02 hepatocytes. After SSP treatment, the protein expressions or gene transcription of nuclear factor erythroid 2-related factor 2 (Nrf2), NAD(P)H dehydrogenase, quinone 1 (NQO1) and heme oxygenase1 (HO-1) decreased in L02. The protein expression of Nrf2 and NQO1 were increased while HO-1 was decreased in liver. Besides, SSP can attenuates apoptosis through reducing the protein expression of Bcl-2-associated X protein (Bax) and caspase-3, and increasing B-cell lymphoma gene 2 (Bcl-2) and B-cell lymphoma-extra large (Bcl-xl). SSP protects against six-heavy-metal-induced hepatic injury in mice and L02 hepatocytes. Supported by Nrf2 gene silencing, the mechanisms may correlate with activating Nrf2 pathway to mitigate oxidative stress and apoptosis.

目的 研究芍药甘草汤对支气管哮喘小鼠肠道菌群结构的调节作用.方法 将50只SPF级BALB/c雌性小鼠随机分为正常组、模型组、醋酸地塞米松组、芍药甘草汤1:1组、芍药甘草汤2:1组.通过卵白蛋白致敏、雾化激发建立肠道菌群紊乱模型.从实验第15天开始,雾化前30 min灌胃,连续灌胃14 d.正常组、模型组给予等量蒸馏水,醋酸地塞米松组(0.676 mg/kg)、芍药甘草汤1:1组(6.01 g/kg)、芍药甘草汤2:1组(6.01 g/kg)给予相应的药物灌胃.实验第29天采集各组结肠组织及粪便样本,观察分析小鼠的一般状态、哮喘典型症状和结肠组织病理改变,各组随机选取6只小鼠进行16S rRNA测序分析肠道菌群多样性和物种丰富度的变化.结果 醋酸地塞米松组、芍药甘草汤1:1组、芍药甘草汤2:1组小鼠的一般状态和哮喘典型症状得到明显改善.病理组织检测显示:正常组结肠组织清晰,结构完整,模型组结肠黏膜及黏膜下层有大量炎性细胞和肿大淋巴结,醋酸地塞米松组充血水肿情况改善,芍药甘草汤1:1组仍可见炎性细胞浸润,芍药甘草汤2:1组结构较完整,黏膜及黏膜下层偶见散在炎性细胞.16S rRNA测序结果显示:与正常组比较,模型组Chao1指数、PD指数降低(P<0.01),observed species指数降低(P<0.05);与模型组比较,醋酸地塞米松组、芍药甘草汤2:1组PD指数升高(P<0.01),Chao1指数、observed species指数升高(P<0.05).肠道菌群菌属水平显示:与正常组相比,模型组毛螺菌属丰度增加(P<0.05),罗斯氏菌属丰度增加(P<0.01),拟普雷沃菌属丰度减少(P<0.05);与模型组相比,芍药甘草汤1:1组毛螺菌属丰度降低(P<0.05);与醋酸地塞米松组相比,芍药甘草汤1:1组拟普雷沃菌属、拟杆菌属、乳杆菌属、罗斯氏菌属丰度增加(P<0.01),芍药甘草汤2:1组罗斯氏菌属丰度降低(P<0.01);与芍药甘草汤1:1组相比,芍药甘草汤2:1组乳杆菌属、罗斯氏菌属丰度降低(P<0.01).结论 芍药甘草汤可改善哮喘小鼠的典型症状、结肠组织病理情况等,其作用机制可能与其恢复肠道菌群的多样性、丰富度及调节菌群结构有关.

目的:探讨麦门冬汤加减方对特发性肺纤维化小鼠转化生长因子-β1(Transforming growth factor-β1,TGF-β1)、平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)、胶原 Ⅰ(Collagen type Ⅰ,COL1A)表达的影响以及对 PI3K/AKT/mTOR 通路的调控作用.方法:将120只SPF级ICR小鼠随机分入空白组、模型组、吡菲尼酮组和麦门冬汤加减方组,用博来霉素(5 mg/kg)建立特发性肺纤维化模型,24 h后分别给予相应的药物治疗.吡菲尼酮组和麦门冬汤加减方组小鼠分别灌胃吡菲尼酮和中药麦门冬汤加减方,空白组和模型组小鼠灌胃生理盐水,各组均连续给药3周(21d)后取材.观察指标:各组小鼠的肺系数;肺组织病理变化;肺组织TGF-β1、α-SMA、COL1A的表达量(免疫组化);肺组织中α-SMA、COL1A、p-PI3K、p-AKT、mTOR的蛋白表达量(Western blot);肺组织中TGF-β1、α-SMA、COL1A的mRNA表达量(qPCR).结果:模型组小鼠的肺系数显著增加,麦门冬汤加减方组肺系数显著降低;模型组小鼠肺组织中有较多炎性细胞浸润,胶原沉积明显,肺泡结构破坏严重,麦门冬汤加减方组小鼠肺组织病理改变较模型组明显减轻,胶原沉积大量减少,肺泡结构逐渐修复;麦门冬汤加减方组较模型组α-SMA、COL1A、TGF-β1的蛋白表达量显著降低(P＜0.01);麦门冬汤加减方组较模型组α-SMA、COL1A、p-PI3K、p-AKT、mTOR的蛋白表达量显著下调(P＜0.01);麦门冬汤加减方组较模型组α-SMA、COL1A、TGF-β1的mRNA表达量显著降低(P＜0.01).结论:麦门冬汤加减方能有效改善博来霉素诱导的特发性肺纤维化,降低α-SMA、COL1A、TGF-β1的表达,可能是通过调控PI3K/AKT/mTOR信号通路,抑制上皮间充质转化,减少细胞外基质沉积而发挥作用.

目的:观察诺丽对氢化可的松致肾阳虚模型小鼠的影响,研究诺丽补肾改善记忆的功效,探讨其作用机制.方法:60只昆明小鼠随机分为对照组,模型组,人参组,诺丽高、中、低剂量组,每组10只,除对照组,其他组均灌胃氢化可的松建立肾阳虚模型,观察小鼠给药后记忆行为学变化,尼氏染色观察脑组织海马CA3区锥体细胞形态和尼氏体数量病理改变,ELISA检测小鼠脑组织神经递质、cAMP、cGMP及PKA含量变化.结果:与对照组相比,模型组跳台实验和避暗实验潜伏期明显缩短(P<0.01),错误次数明显增多(P<0.01),消退潜伏期明显缩短(P<0.01),消退错误次数明显增多(P<0.01),大脑海马CA3区尼氏体数量较少(P<0.01),脑组织神经递质5-HT、ACh、DA、NA表达降低(P<0.01),cAMP、PKA含量下降(P<0.01,P<0.05),cAMP/cGMP比值降低(P<0.05).与模型组相比,人参和诺丽高、中剂量能明显延长小鼠跳台实验和避暗实验的潜伏期时间(P<0.01,P<0.05),降低错误次数(P<0.01,P<0.05),延长消退潜伏期时间(P<0.01,P<0.05),降低消退错误次数(P<0.01,P<0.05),增加大脑海马CA3区尼氏体数量(P<0.01),增加脑组织神经递质ACh、5-HT、DA、NA表达(P<0.01,P<0.05),升高cAMP、PKA含量(P<0.01),升高cAMP/cGMP比值(P<0.01).结论:诺丽对氢化可的松致肾阳虚小鼠的记忆损伤有保护作用,其补肾改善记忆是通过调节cAMP/PKA信号通路蛋白,影响下游神经递质,促进神经元蛋白合成实现的.

目的 观察四神方对三硝基苯磺酸(trinitrobenzene sulfonic acid,TNBS)诱导的小鼠实验性炎症性肠病(inflammatory bowel disease,IBD)的免疫调节作用.方法 取56只BALB/c小鼠,随机分为乙醇对照组、模型组、柳氮磺胺吡啶(salicylazosulfapyridine,SASP)组和四神方组.小鼠麻醉后直肠内灌注50％乙醇溶液(含有1.5 mg的TNBS)制备IBD模型.造模2 d后,开始分别使用0.5 g/kgSASP或者2.3 g/kg四神方灌胃给药,连续8 d.观察小鼠的一般状态和疾病活动指数(disease activity index,DAI)评分、结肠组织学、结肠中髓过氧化物 酶(myeloperoxidase,MPO)变化.使用ELISA法检测人重组干扰素γ(IFN-γ)和白细胞介素(IL-12/23 p40)的水平;使用MTT法检测脾脏中以及肠系膜淋巴结(MLNs)中T淋巴细胞增殖;使用流式细胞术检测相关协同刺激因子的表达.结果 四神方有效改善小鼠IBD模型一般状况及DAI评分,缓解结肠局部炎性损伤,减轻炎细胞浸润,减少MPO的产生,降低结肠局部IL-12/23 p40和IFN-γ的水平,引起脾脏中T淋巴细胞数目增加和MLNs中T淋巴细胞数目减少并抑制MLNs中T淋巴细胞协同分子的表达.结论 四神方对TNBS所诱导的小鼠IBD具有显著的治疗作用,可能通过抑制MPO的产生、调节T细胞免疫作用、降低IL-12/23 p40和IFN-γ 的水平缓解结肠中过度的炎性反应.

Viral pneumonia is a common complication caused by Influenza A virus infection and is characterized by severe pulmonary inflammation. A previous study showed that berberine (BBR) significantly ameliorated the pulmonary inflammation in mice with influenza viral pneumonia but its underlying mechanism is not entirely understood. In this study, we reproduced the mouse model of influenza viral pneumonia through intranasal infection of A/Puerto Rico/8/34 (H1N1), to further investigate the anti-inflammatory mechanism of BBR based on nucleotide-binding oligomerization domain-like receptor protein (NLRP) 3 inflammasome activation and Gasdermin D (GSDMD)-mediated pyroptosis. Consistent with MCC950 (10 mg/kg, a specific NLRP3 inflammasome inhibitor), BBR (10 mg/kg) obviously improved the weight loss and survival rate of infected mice, alleviated their pulmonary inflammation, and suppressed the accumulation of tumor necrosis factor and interleukin (IL)-6 in lungs without obvious inhibition on viral multiplication (hemagglutinin titer and nucleoprotein messenger RNA). Moreover, BBR (10 mg/kg) reduced the expressions of NLRP3, apoptosis-associated speck-like protein containing a CARD (ASC), and cysteinyl aspartate-specific proteinase (Caspase)1 (Caspase1 precursor [Pro-caspase1] + Caspase1p20 subunit) and the ratio of Caspase1p20 subunit to Caspase1, thus inhibiting the NLRP3 inflammasome activation and resulting in the decreased contents of mature IL-1 beta and IL-18 in lungs. The GSDMD expression (GSDMD precursor [Pro-GSDMD] + GSDMD-N terminal [NT]) and the ratio of GSDMD-NT to GSDMD were also declined by BBR (10 mg/kg). These evidence indicate that BBR may ameliorate pulmonary inflammation in mice with influenza viral pneumonia through inhibiting NLRP3 inflammasome activation, as well as depressing GSDMD-mediated pyroptosis via declining GSDMD expression and restraining NLRP3 inflammasome-mediated GSDMD activation.

Objective Ulcerative colitis (UC) as one of the intractable diseases in gastroenterology seriously threatens human health. Respiratory pathology is a representative extraintestinal manifestation of UC affecting the quality of life of patients. Gegen Qinlian Decoction (GQD) is a classical traditional Chinese medicine prescription for UC or acute lung injury. This study was aimed to reveal the therapeutic effect of GQD on UC and its pulmonary complications and uncover its molecular mechanism mediated by myeloid cells and microbiota. Methods Mice with DSS-induced colitis were orally administrated with GQD. Overall vital signs were assessed by body weight loss and disease activity index (DAI). Pulmonary general signs were evaluated by pulmonary pathology and lung function. The mechanism of GQD relieving UC was characterized by detecting myeloid cells (neutrophils, macrophages, inflammatory monocytes, and resident monocytes) in colonic and lung tissues, related inflammatory cytokines, as well as the microbiota in bronchoalveolar lavage fluid (BALF) and feces. Results GQD significantly reduced weight loss, DAI scores, and lung injury but improved the lung function of colitis mice. The DSS-induced colonic and concurrent pulmonary inflammation were also alleviated by GQD, as indicated by the down-regulated expressions of inflammatory cytokines (TNF-α, IL-1β, IL-6, CCR2, and CCL2) and the suppressed recruitment of neutrophils and inflammatory monocytes. Meanwhile, GQD greatly improved intestinal microbiota imbalance by enriching Ruminococcaceae UCG-013 while decreasing Parabacteroides, [Eubacterium]_fissicatena_group, and Akkermansia in the feces of colitis mice. Expectantly, GQD also restored lung microbiota imbalance by clearing excessive Coprococcus 2 and Ochrobactrum in the BALF of colitis mice. Finally, significant correlations appeared between GQD-mediated specific bacteria and inflammatory cytokines or immune cells. Conclusion GQD could alleviate UC by decreasing excessive inflammatory myeloid cells and cytokines, and reshaping the microbiota between the colon and lung, which contributes to clarifying the mechanism by which GQD ameliorates colitis-associated pulmonary inflammation.

目的:基于网络药理学和动物实验探讨养心生脉颗粒治疗急性心肌梗死合并抑郁症的作用机制.方法:通过中药系统药理学数据库与分析平台(TCMSP)、中医药综合数据库(TCMID)、中国知网及SwissTargetPrediction收集养心生脉颗粒的化学成分和对应靶点,基于GeneCards和OMIM数据库检索急性心肌梗死和抑郁症靶点,韦恩图获取药物和疾病的交集靶点,采用STRING数据库构建蛋白质-蛋白质相互作用(PPI)网络,导入Cytoscape 3.8.2软件绘制中药-成分-靶点网络和成分-靶点-通路网络,并利用DAVID 6.8数据库进行基因本体(GO)生物学过程和京都基因与基因组百科全书(KEGG)通路富集分析.结合网络药理学预测的核心靶点和关键通路,以冠状动脉左前降支结扎结合慢性不可预见性轻度应激(CUMS)方法制备大鼠急性心肌梗死合并抑郁模型,经养心生脉颗粒干预后,观察大鼠抑郁样行为、血流动力学变化及心肌组织病理情况,采用蛋白质印迹法(Western blot)检测关键通路靶点,验证养心生脉颗粒治疗急性心肌梗死合并抑郁的疗效及作用机制.结果:通过网络药理学共筛选出养心生脉颗粒活性成分靶点1451个,急性心肌梗死靶点615个,抑郁症靶点590个,交集靶点54个.PPI网络及KEGG富集分析显示,多个关键靶蛋白如白细胞介素-6(IL-6)、蛋白激酶B1(Akt1)、肿瘤坏死因子-α(TNF-α)、血管内皮生长因子(VEGF)、IL-10、哺乳动物雷帕霉素靶蛋白(mTOR)等,以及5-羟色胺能突触(serotonergic synapse)、TNF信号通路、丝裂原活化蛋白激酶(MAPK)信号通路、磷脂酰肌醇3-激酶/蛋白激酶B(PI3K/Akt)信号通路、神经营养素(neurotrophin)信号通路等与炎症关系密切.选取关键蛋白Akt1、mTOR及PI3K/Akt信号通路进行动物实验验证.结果 显示,与假手术组相比,模型组大鼠的糖水偏嗜度下降,游泳不动时间增加,心肌损伤程度加重,PI3K、Akt、mTOR磷酸化水平明显下调(P<0.05,P<0.01);与模型组相比,养心生脉颗粒低、中剂量组糖水偏嗜度增加,心肌损伤程度明显减轻,PI3K、Akt、mTOR磷酸化水平上调(P<0.05,P<0.01).结论:基于网络药理学和动物实验验证,养心生脉颗粒可能通过PI3K/Akt/mTOR信号通路逆转急性心肌梗死合并抑郁大鼠的抑郁样行为,缓解心肌组织损伤.