Ethnopharmacological relevanceThe traditional Chinese medicine (TCM) formula Banxia Xiexin decoction (BXD) has definite therapeutic effect in treating stress-induced gastric ulceration (SIGU) and many other gastrointestinal diseases, but its effect on gastric lymphatic pumping (GLP) remains unclear.Aim of the studyElucidating the role of GLP in SIGU and BXD treatment, and exploring the molecular mechanisms of GLP regulation.Materials and methodsIn vivo GLP imaging were performed on SIGU rat model, and the lymphatic dynamic parameters were evaluated. Gastric antrum tissues and serum were collected for macroscopic, histopathological and ulcerative parameters analysis. Gastric lymphatic vessel (GLV) tissues were collected for RNA-Seq assays. Differentially expressed genes (DEGs) were screened from RNA-Seq result and submitted for transcriptomic analysis. Key DEGs and their derivative proteins were measured by qRT-PCR and WB.ResultsGLP was significantly suppressed in SIGU rats. BXD could recover GLP, ameliorate stomach lymphostasis, and alleviate the ulcerative damage. Transcriptome analysis of GLV showed the top up-DEGs were concentrated in smooth muscle contraction signaling pathway, while the top the down-DEGs were concentrated in energy metabolism pathways especially fatty acid degradation pathway, which indicated BXD can promote lymphatic smooth muscle contraction, regulate energy metabolism, and reduce fatty acid degradation. The most possible target of these mechanisms was the lymphatic smooth muscle cells (LSMCs) which drove the GLP. This speculation was further validated by the qRT-PCR and WB assessments for the level of key genes and proteins.ConclusionsBy activating the smooth muscle contraction signaling pathway, restoring energy supply, modulating energy metabolism program and reducing fatty acid degradation, BXD effectively recovered GLP, mitigated the accumulation of inflammatory cytokines and metabolic wastes in the stomach, which importantly contributes to its efficacy in treating SIGU.

ETHNOPHARMACOLOGICAL RELEVANCE:Haizao Yuhu decoction (HYD) is a classic Chinese herbal formula described in the surgical monographs of the Ming Dynasty "Waikezhengzong." It has been widely used to treat goiter for approximately 500 years and found to be particularly effective. HYD contains glycyrrhiza and sargassum. This pair of herbs belongs to "18 incompatible medicaments" of traditional Chinese medicine theory. Although these two herbs are opposite, our preliminary study proved that they have superior effect when added into HYD at 2 times the dose of Chinese Pharmacopoeia. However, the species of glycyrrhiza in HYD that are the most effective have not been recorded in ancient Chinese medical texts. According to the Chinese Pharmacopoeia, glycyrrhiza is divided into the following three species: Glycyrrhiza uralensis Fish., G. glabra L., and G. inflata Bat. The effect of HYD containing different species of glycyrrhiza and their mechanisms remain to be further explored.AIM OF THE STUDY:To investigate the effect of HYD containing three species of glycyrrhiza on goiter, and to elucidate the molecular mechanism using network pharmacology combined with RNA sequencing (RNA-seq).MATERIALS AND METHODS:A rat model of goiter was established by 14 days of intragastric gavage of propylthiouracil (PTU), and the rats were treated for 4 weeks with HYD containing three different species of glycyrrhiza. The body weight and rectal temperature of rats were tested weekly. At the end of the experiment, the serum and thyroid tissues of rats were collected. The effect of the three HYDs was assessed based on general observations (including body weight, rectal temperature, and living status of rats), absolute/relative thyroid weight, thyroid function (including triiodothyronine, thyroxine, free triiodothyronine, free thyroxine, and thyroid-stimulating hormone levels), and thyroid tissue pathology. Next, we explored their pharmacological mechanisms using network pharmacology combined with RNA-seq and validated key targets using real-time quantitative reverse-transcription polymerase chain reaction (RT-qPCR), western blotting (WB), and immunofluorescence (IF) assays.RESULTS:The three HYDs reduced the absolute/relative weights of thyroid tissues and improved the pathological structure, thyroid function, and general findings of rats with goiter. Overall, the effect of HYD-G. uralensis Fish. (HYD-U) was better. Results from network pharmacology and RNA-seq jointly suggested that both the pathogenesis of goiter and the mechanism of action of HYD for goiter were related to the phosphatidylinositol 3-kinase-protein kinase B (PI3K-Akt) pathway. We validated the key targets in the pathway, namely, vascular endothelial growth factor (VEGF) A, VEGF receptor 2, phosphoinositide-3-kinase regulatory subunit 1 (PIK3R1) and its encoded protein PI3K (p85), AKT serine/threonine kinase 1 (AKT1), phospho-AKT and cyclin D1 using RT-qPCR, WB, and IF assays. The PI3K-Akt pathway was hyperactivated in rats with PTU-induced goiter, whereas the three HYDs could inhibit the pathway.CONCLUSION:This study confirmed the definite effect of the three HYDs in the treatment of goiter, and HYD-U was found to be more effective. The three HYDs inhibited angiogenesis and cell proliferation in goiter tissue by inhibiting the PI3K-Akt signaling pathway.

目的 明确清开灵口服液对CCl4 大鼠的抗肝纤维化作用,并基于NOD 样受体热蛋白结构域3(NOD-like receptor protein domain 3,NLRP3)炎症小体通路揭示其抗肝纤维化的机制.方法 44只SD大鼠随机分为空白组、模型组、清开灵口服液组(QKL组),应用CCl4 灌胃的方法建立中毒性肝纤维化大鼠模型,设置4周与8周两个时间点.实验结束后,检测各组大鼠肝、脾指数;HE染色和Masson染色观察肝组织结构变化与胶原纤维增生情况;检测血清天冬氨酸氨基转移酶(aspartate aminotransferase,AST)、丙氨酸氨基转移酶(alanine aminotransferase,ALT)、总胆红素(total bilirubin,T-BIL)、羟脯氨酸(hydroxyproline,HYP)、谷胱甘肽过氧化物酶(glutathione peroxidation,GSH-Px)含量;ELISA法检测肝组织肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白细胞介素-1β(interleukin-1β,IL-1β)水平;免疫组化检测肝组织α-平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)的表达;采用West-ern blot、RT-PCR法检测肝组织NLRP3、衔接蛋白凋亡相关斑点样蛋白(adaptor protein apoptosis-related dot-like protein,ASC)、半胱氨酸天冬氨酸蛋白水解酶-1(Caspase-1)、α-SMA的蛋白和基因的表达水平.结果 与空白组相比,4周与8周模型组肝脾指数及AST、ALT、T-BIL、HYP含量均明显升高(P＜0.01)、GSH-Px活力显著下降(P＜0.01),IL-1β和TNF-α含量明显升高(P＜0.01),α-SMA、NL-RP3、ASC、Caspase-1的蛋白及基因表达水平均明显增加(P＜0.05).HE和Masson染色显示模型组肝组织形态结构紊乱,炎症细胞大量浸润,肝脏纤维组织增生更加明显.与模型组相比,同期清开灵组均可明显降低肝脾指数及血清AST、ALT、T-BIL含量和HYP含量(P＜0.05),提高GSH-Px活力(P＜0.05,P＜0.01),抑制TNF-α、IL-1β合成(P＜0.01,P＜0.05),减轻肝组织结构紊乱和炎细胞浸润,抑制胶原纤维增生,抑制α-SMA蛋白表达(P＜0.05),NLRP3、ASC和Caspase-1在蛋白和基因水平表达呈不同程度降低(P＜0.05,P＜0.01).结论 清开灵口服液可显著抑制CCl4 导致的大鼠肝纤维化,提高肝脏抗氧化能力,改善肝功能,缓解炎症,其机制可能与抑制星状细胞活化、抑制NLRP3炎症小体通路活化相关.

Objective: To explore the effect of modified Haizao-Gancao anti-drug combination of Haizao Yuhu Decoction（HYD） on the expression of genes related to PI3K/Akt pathway in goiter rats under the condition that the dosage of Haizao and Gancao is twice the high limit dosage prescribed in the Pharmacopoeia of the Pharmacopoeia of the People’s Republic of China 2015 Edition. Methods: A total of 84 rats were randomly divided into: blank group, model group, positive drug euthyrox group,HYD group, HYD without Haizao group, HYD without Gancao group and HYD without Haizao and Gancao group, 12 rats in each group. The propylthiouracil（PTU） was used to establish the model of goitre, then, each group were given the corresponding oral liquid. Apoptosis was detected by TUNEL method, apoptosis index was calculated, and expression of PI3K, Akt, Cyclin D1and Bcl-2 mRNA were detected by real-time quantitative PCR, the protein expression of Akt, p-Akt were detected by Western Blot. Results: Compared with the model group, the apoptosis index of the HYD group was distinctly rised（P＜0.05）, the mRNA expression level of PI3K, Cyclin D1 and Bcl-2 showed varying degrees of decline in the groups which were given interventional medicine（P＜0.01, P＜0.05）, the expression of AKT mRNA was significantly decreased in the HYD group and HYD without Haizao and Gancao group（P＜0.05）, the expression of p-Akt, p-Akt/Akt protein was significantly decreased in the groups which were given interventional medicine（P＜0.01, P＜0.05）. Conclusion: Anti-drug combination of Haizao Yuhu Decoction modified Haizao or Gancao on propylthiouracil induced goiter rats have a significant therapeutic effect, decreasing the expression of PI3K and Akt genes, thus affecting the expression of Cyclin D1 and Bcl-2, inhibiting cell proliferation and promoting cell apoptosis, so it acts as an anti-goiter.

Objective: To explore the improvement effect of Flos Puerariae, Hoveniae Semen, and their compatibility on acute alcoholic gastric mucosal injury, and lay a foundation for further development of Flos Puerariae, Hoveniae Semen, and their compatibility in the prevention and treatment of alcohol-induced multiple organ injury. Method: The acute alcohol-induced gastric mucosal injury model of mice was established by multiple intragastric administration of 56% Hongxing Erguotou liquor(15 mL·kg -1 ). A total of 120 male ICR mice were randomly divided into 8 groups, namely, the blank group, model group, omeprazole group(0.026 g·kg -1 ), Flos Puerariae-Hoveniae Semen（compatibility） high, medium, and low-dose groups(29.2,14.6, 7.3 g·kg -1 ), Flos Puerariae group(19.5 g·kg -1 ), and Hoveniae Semen group(19.5 g·kg -1 ), with 15 mice in each group. After one week of adaptive feeding, the animals were pre-administrated with the corresponding drug at the rate of 10 mL·kg -1 for 3 d. From the 4th day, after 1 h of administration, Erguotou liquid was administrated at the rate of 15 mL·kg -1 and the blank group was administrated with the same volume of deionized water to record the drunkenness and sober up time. The administration was lasted for 3 d. One hour after the last administration, the eyeballs were removed and the mice were sacrificed. The concentration of ethanol in serum was determined by gas chromatograph, and the activity of ethanol dehydrogenase（ADH） in gastric mucosa was determined by ultraviolet-vis spectrophotometer. Hematoxylin-eosin（HE） staining was used to observe the pathological changes in gastric mucosa. Serum inflammatory factors were determined by enzyme-linked immunosorbent assay（ELISA）. The mRNA expression of nuclear transcription factor-κB（NF-κB） p65 and NF-κB inhibitory protein α（IκBα） were detected by real-time polymerase chain reaction（Real-time PCR）.Result: As compared with the normal group, the content of interleukin-6（IL-6）, interleukin-1β（IL-1β）, and tumor necrosis factor-α（TNF-α） in serum of mice in the model group was increased（P＜0.05）, the mRNA expression of NF-κB p65 in gastric mucosa tissues was increased（P＜0.01）, and the mRNA expression of IκBαwas decreased（P＜0.01）. As compared with the model group, the drunkenness time of the omeprazole group, high and medium-dose compatibility groups, and Flos Puerariae group was prolonged（P＜0.05）, the sober up time of the high and medium-dose compatibility groups was shortened（P＜0.05）, the ethanol concentration in the serum of the high-dose compatibility group was decreased（P＜0.05）, the ADH activity in the gastric mucosa of the omeprazole group and high and medium-dose compatibility groups was increased（P＜0.05）, the macroscopic injury score of the high, medium, and low-dose compatibility groups and Flos Puerariae group was decreased（P＜0.05）, the score of pathological injury in the omeprazole group, high, medium, and low-dose compatibility groups, and Flos Puerariae group was decreased（P＜0.01）, the expression of IL-6 in serum of all drug groups was decreased（P＜0.05）, the expression of IL-1β in serum of the omeprazole group, high, medium, and low-dose Flos Puerariae groups, and Hoveniae Semen group was decreased（P＜0.05）, the expression of TNF-α in serum of high and medium-dose groups was decreased（P＜0.05）, the mRNA expression of NF-κB p65 in gastric mucosa tissues of all drug groups was decreased（P＜0.05）, and the mRNA expression of IκBα in gastric mucosa tissues of the omeprazole group and high, medium, and low-dose compatibility groups was increased（P＜0.05）. As compared with the high-dose compatibility group, the drunkenness time in the low-dose compatibility group and Hoveniae Semen group was shortened（P＜0.01）, the sober up time in the Flos Puerariae and Hoveniae Semen groups was prolonged（P＜0.01）, the concentration of ethanol in the serum of the medium and low-dose compatibility groups, Flos Puerariae group, and Hoveniae Semen group increased（P＜0.05）, the macroscopic injury score of the medium and low-dose compatibility groups and Hoveniae Semen group was increased（P＜0.05）, the pathological injury score of the medium and low-dose compatibility groups, Flos Puerariae group, and Hoveniae Semen group was increased（P＜0.01）, the content of IL-1β in serum of lowdose compatibility group, Flos Puerariae group, and Hoveniae Semen group was increased（P＜0.01）, and the mRNA expression of IκBα in gastric mucosa of the Flos Puerariae group and Hoveniae Semen group was decreased（P＜0.05）. As compared with the medium-dose compatibility group, the drunkenness time in the Hoveniae Semen group was shortened（P＜0.05）, the sober up time in the Flos Puerariae group was prolonged（P＜0.05）, the pathological injury score in the Flos Puerariae group and Hoveniae Semen group was increased（P＜0.01）, and the content of IL-1β in serum of the low-dose compatibility group, the Flos Puerariae group, and Hoveniae Semen group was increased（P＜0.05）. As compared with the low-dose compatibility group, the pathological injury score of the Hoveniae Semen group was increased（P＜0.05）. Conclusion: Flos Puerariae, Hoveniae Semen, and their compatibility play a role in preventing and treating acute alcoholic gastric mucosal injury in mice, which may be related to the inhibition of the expression of NF-κB signal pathway in gastric mucosa, and the high-dose compatibility group has the optimal effect.

Background:There is growing evidence to suggest that ginsenoside Rd (GRd) has a therapeutic effect on depression, but the specific mechanisms behind its activity require further study.Objective:This study is designed to investigate the antidepressant-like effect and underlying mechanisms of GRd.Methods:In this study, the behavioral despair mouse model of depression and chronic unpredictable mild stress (CUMS) rat model of depression were established to explore the effects of GRd on depression-like behavior and its underlying mechanisms. Behavioral tests were used to evaluate the replication of animal models and depression-like behaviors. The hypoxia-inducible factor-1α (HIF-1α) blocker 2-methoxyestradiol (2-ME) was injected to determine the role of HIF-1α in the antidepressant-like effect of GRd. In addition, molecular biology techniques were used to determine the mRNA and protein expression of HIF-1ɑ signaling pathway and synaptic plasticity-related regulators, that is synapsin 1 (SYN 1) and postsynaptic density protein 95 (PSD 95). In silico binding interaction studies of GRd with focused target proteins were performed using molecular docking to predict the affinity and optimal binding mode between ligands and receptors.Results:Our data show that GRd significantly reversed depression-like behavior and promoted mRNA and protein expression of HIF-1ɑ signaling pathway and synaptic plasticity-related regulators. However, the antidepressant-like effect of GRd disappeared upon inhibition of HIF-1α expression following administration of 2-ME. Furthermore, molecular docking results showed that GRd possessed significant binding affinity for HIF-1α, VEGF, and VEGFR-2.Conclusion:Our results show that GRd exhibits significant antidepressant-like effect and that HIF-1α signaling pathway is a promising target for the treatment of depression.

目的:探究瞬时受体电位香草酸亚型1(TRPV1)通道在诃子制草乌减轻心肌细胞毒性中的作用机制.方法:以体外培养的H9c2心肌细胞为模型,采用噻唑蓝(MTT)法检测细胞活力;实时荧光定量聚合酶链式反应(Real-time PCR)检测心肌细胞中TRPV1 mnRNA的表达水平;酶联免疫吸附测定试验(ELISA)检测细胞中乳酸脱氢酶(LDH)漏出率,细胞核,活性氧(ROS),线粒体膜电位及钙离子(Ca2+)含量的变化.结果:与空白组比较,当质量浓度≥0.5 g·L-1时,生草乌、诃子制草乌均可显著降低细胞活力(P＜0.01),细胞中LDH漏出率,ROS及Ca2+释放量增加,线粒体膜电位降低,细胞核固缩甚至是破碎.当质量浓度≥0.5g·L-1时,与同质量浓度生草乌组相比,诃子制草乌组细胞活力显著上升(P＜0.01),细胞中LDH漏出率,ROS及Ca2+释放量减少,线粒体膜电位升高,细胞核形态有所改善.与作用于未经处理的H9c2心肌细胞比较,同质量浓度的生草乌作用于TRPV1抑制剂BCTC预处理后的H9c2 心肌细胞后,其细胞活力明显上升,细胞中LDH漏出率,ROS及Ca2+释放量减少,线粒体膜电位升高,细胞核固缩程度得到改善;而同质量浓度的诃子制草乌作用于BCTC预处理后的H9c2心肌细胞,其细胞活力下降,细胞中LDH漏出率,ROS及Ca2+释放量增加,线粒体膜电位降低.Real-time PCR结果表明,草乌和辅料诃子汤均能使心肌细胞中TRPVlmRNA表达量增多,且呈现浓度依赖性.结论:生草乌可通过激活TRPV1通道诱导心肌细胞凋亡,引发心脏毒性;而诃子制草乌可通过TRPV1通道介导起到减毒作用,其减毒作用的发挥可能与诃子中的酸性成分和草乌中的生物碱协同作用于TRPV1通道有关.

ETHNOPHARMACOLOGICAL RELEVANCE:Glycyrrhiza and sargassum are among the 18 incompatible medicaments according to traditional Chinese medicine (TCM) theory. Although it contains glycyrrhiza and sargassum, Haizao Yuhu decoction (HYD) is a classic prescription widely used as TCM to treat goiter. According to the Chinese Pharmacopoeia, glycyrrhiza is divided into three varieties: Glycyrrhiza uralensis Fish., Glycyrrhiza glabra L., and Glycyrrhiza inflata Bat. Whether the three varieties of glycyrrhiza have different efficacy or toxicity when applied in the HYD is unknown.AIM OF THE STUDY:To explore whether the HYDs comprising three varieties of glycyrrhiza have different efficacy or toxicity when used to treat goiter in rats and the underlying mechanisms of these HYDs.MATERIALS AND METHODS:For two weeks, the goiter model was replicated by intragastric propylthiouracil (PTU) administration. Samples were divided into the control group, model group, euthyrox group, HYD with glycyrrhiza uralensis (HYD-U) group, HYD with glycyrrhiza glabra (HYD-G) group, and HYD with glycyrrhiza inflata (HYD-I) group. After four weeks of treatment, body weight, rectal temperature, thyroid/liver/kidney coefficient, thyroid/liver/kidney function, thyroid/liver/kidney histomorphology, and thyroid ultrastructure were evaluated. Then, real-time quantitative reverse-transcription polymerase chain reaction (RTqPCR), Western blot, and immunofluorescence analyses were performed to detect genes and proteins affecting autophagy and apoptosis in thyroid cells in the AMP-activated Protein Kinases (AMPK)/Mammalian target of rapamycin (mTOR) pathway.RESULTS:All three HYDs increased thyroid hormones (THs) levels, relieved thyroid pathological tissue and ultrastructure, and activated vital proteins and genes in the AMPK/mTOR pathway. Comparisons among the efficacy of the three HYDs indicated that HYD-U restored the THs most effectively; however, no difference in the anti-goiter effect was observed. Moreover, the three HYDs resulted in no toxicity and promoted the recovery of impaired liver and kidney function caused by PTU. Comparisons among the recovery effects of the three HYDs on the liver and kidney were the same.CONCLUSION:Our experiments demonstrated that the three HYDs had outstanding anti-goiter effects and protected liver and kidney function. Their anti-goiter effects were attributed to AMPK/mTOR pathway-induced autophagy and apoptosis. HYD-U resulted in the best THs recovery. It was further indicated that in our present study, glycyrrhiza and sargassum were compatible in the three HYDs, thereby suggesting their safety of compounding in HYD and providing a basis for the research of the 18 incompatible medicaments.

目的:探究甘遂半夏汤中甘遂-甘草反药组合加减治疗腹水大鼠的利水药效及其机制.方法:雄性Wistar大鼠随机分为空白组、模型组、阳性药组、甘遂半夏汤全方组(全方组)、甘遂半夏汤全方去生甘遂组(去遂组)、甘遂半夏汤全方去炙甘草组(去草组)、甘遂半夏汤全方去生甘遂炙甘草组(双去组).腹腔注射Walker-256细胞复制大鼠腹水模型.造模后第2天给药,第3天收集24 h尿液,记录尿量,检测尿液上清液中Na+、K+、Cl-含量;第9天选取造模成功大鼠,腹主动脉取血后摘取双侧肾脏,检测血清Na+、K+、Cl-含量,血浆肾素活性(PRA)、血管紧张素Ⅱ(AngⅡ)、醛固酮(ALD)、抗利尿激素(ADH)含量及肾脏水通道蛋白2(AQP2)mRNA和蛋白表达.结果:与空白组比较,模型组尿量显著减少(P＜0.01),血浆PRA、AngⅡ、ALD、ADH含量显著升高(P＜0.01),肾脏AQP2 mRNA和蛋白表达显著升高(P＜0.01,P＜0.05),尿Na+量显著降低(P＜0.01),血清K+、Cl-量显著升高(P＜0.01).与模型组比较,阳性药组、全方组尿量显著增多(P＜0.01,P＜0.05),血浆PRA、AngⅡ、ALD、ADH含量显著降低(P＜0.01);全方组肾脏AQP2 mRNA和蛋白表达显著降低(P＜0.05);全方组尿Na+量显著升高(P＜0.01),阳性药组、全方组血清K+、Cl-量显著降低(P＜0.01,P＜0.05).与全方组比较,去草组和双去组尿量显著减少(P＜0.01,P＜0.05),去遂组、去草组和双去组血浆PRA、AngⅡ、ALD、ADH显著升高(P＜0.01),去遂组和去草组肾脏AQP2 mRNA表达显著升高(P＜0.01,P＜0.05),去草组和双去组尿Na+量显著降低(P＜0.01),双去组血清K+、Cl-量显著升高(P＜0.01).结论:甘遂半夏汤全方及全方加减一味或两味反药组合可增加腹水大鼠尿量,其中全方利水药效佳,属于"增效"配伍关系,其"增效"机制一方面可能通过增强全方对RAAS系统的抑制;另一方面可能通过增加全方对肾脏AQP2 mRNA和蛋白表达的抑制.通过以上两方面甘遂半夏汤全方对水液的重吸收减少,体液排出增多,利水药效增加,并且改善了腹水引起的血尿电解质紊乱.

Increasing evidence indicates that the pathogenesis of depression is closely linked to impairments in neuronal synaptic plasticity. Honokiol, a biologically active substance extracted from Magnolia Officinalis, has been proven to exert significant antidepressant effects. However, the specific mechanism of action remains unclear. In this study, PC12 cells and chronic unpredictable mild stress (CUMS) model rats were used to explore the antidepressant effects and potential mechanisms of honokiol in vitro and in rats. In vitro experiment, a cell viability detection kit was used to screen the concentration and time of honokiol administration. PC12 cells were administered with hypoxia-inducible factor-1α (HIF-1α) blocker, 2-methoxyestradiol (2-ME), and vascular endothelial growth factor receptor 2 (VEGFR-2) blocker, SU5416, to detect the expression of HIF-1α, VEGF, synaptic protein 1 (SYN 1), and postsynaptic density protein 95 (PSD 95) by western blotting. In effect, we investigated whether the synaptic plasticity action of honokiol was dependent on the HIF-1α-VEGF pathway. In vivo, behavioral tests were used to evaluate the reproducibility of the CUMS depression model and depression-like behaviors. Molecular biology techniques were used to examine mRNA and protein expression of the HIF-1α-VEGF signaling pathway and synaptic plasticity-related regulators. Additionally, molecular docking techniques were used to study the interaction between honokiol and target proteins, and predict their binding patterns and affinities. Experimental results showed that honokiol significantly reversed CUMS-induced depression-like behaviors. Mechanically, honokiol exerted a significant antidepressant effect by enhancing synaptic plasticity. At the molecular level, honokiol can activate the HIF-1α-VEGF signaling pathway in vitro and in vivo, as well as promote the protein expression levels of SYN 1 and PSD 95. Taken together, the results do not only provide an experimental basis for honokiol in the clinical treatment of depression but also suggest that the HIF-1α-VEGF pathway may be a potential target for the treatment of depression.

Background: There is growing evidence to suggest that ginsenoside Rd (GRd) has a therapeutic effect on depression, but the specific mechanisms behind its activity require further study. Objective: This study is designed to investigate the antidepressant-like effect and underlying mechanisms of GRd. Methods: In this study, the behavioral despair mouse model of depression and chronic unpredictable mild stress (CUMS) rat model of depression were established to explore the effects of GRd on depression-like behavior and its underlying mechanisms. Behavioral tests were used to evaluate the replication of animal models and depression-like behaviors. The hypoxia-inducible factor-1α (HIF-1α) blocker 2-methoxyestradiol (2-ME) was injected to determine the role of HIF-1α in the antidepressant-like effect of GRd. In addition, molecular biology techniques were used to determine the mRNA and protein expression of HIF-1ɑ signaling pathway and synaptic plasticity-related regulators, that is synapsin 1 (SYN 1) and postsynaptic density protein 95 (PSD 95). In silico binding interaction studies of GRd with focused target proteins were performed using molecular docking to predict the affinity and optimal binding mode between ligands and receptors. Results: Our data show that GRd significantly reversed depression-like behavior and promoted mRNA and protein expression of HIF1ɑ signaling pathway and synaptic plasticity-related regulators. However, the antidepressant-like effect of GRd disappeared upon inhibition of HIF-1α expression following administration of 2-ME. Furthermore, molecular docking results showed that GRd possessed significant binding affinity for HIF-1α, VEGF, and VEGFR-2. Conclusion: Our results show that GRd exhibits significant antidepressant-like effect and that HIF-1α signaling pathway is a promising target for the treatment of depression.

细胞膜局部区域可形成富含饱和脂质、胆固醇、鞘脂的脂筏域作为其信号转导调控平台.传统实验手段在研究脂筏及其功能时受到系统复杂度高及区域结构瞬时性强等制约.近年来,分子动力学模拟技术为细胞膜的组织原则提供了重要的理论支撑,从简单的单一组分模型到多组分系统转变,最终形成了越来越多的细胞膜仿真模型.其中,粗粒化模拟由于其简化模型,可大副拓展模拟体系的复杂程度与模拟时间,在细胞膜以及蛋白质-脂质相互作用相关研究中得到了广泛应用.本文采用MARTINI粗粒化力场模拟,构建了一种含有阴离子脂质磷脂酰肌醇二磷酸(phosphatidylinositol diphosphate,PIP2)的混合膜体系.模拟结果表明,该体系在适当温度及饱和度条件下,能自发分层形成脂筏域;膜厚度、膜组分分布、膜组分流动性等多种参数均表明,脂筏结构形成且符合其结构特征;少量PIP2添加不影响分层特性且PIP2对脂筏具有显著亲和性.此外,利用该模型以跨膜信号蛋白CD3ε为例研究了脂筏域体系中蛋白质-脂质相互作用.结果 表明,PIP2-CD3ε胞内区相互作用可能是脂筏招募CD3ε的驱动力,且该过程可受钙离子调控.本工作体现了粗粒化模拟在仿真膜相关研究中的巨大优势及良好应用前景.

In traditional Chinese medicine, Glycyrrhiza and Sargassum are one pair of the “18 incompatible medicaments,” which in theory cannot be used together. However, since ancient times, many reports have described using compounds containing both Glycyrrhiza and Sargassum to treat diseases. Haizao Yuhu Decoction (HYD), which contains both ingredients, is mainly used to treat goiter. Chinese Pharmacopoeia officially recorded three varieties of Glycyrrhiza: Glycyrrhiza uralensis, Glycyrrhiza inflata, and Glycyrrhiza glabra. These three varieties have certain differences in chemical composition and pharmacological effects. The purpose of the present study was to investigate whether the HYD containing different varieties of Glycyrrhiza and Sargassum had different therapeutic effects in rats with goiter and to elucidate the underlying mechanism of any difference. In this study, propylthiouracil (PTU) was used to replicate the goiter model, then HYDs containing different varieties of Glycyrrhiza were used for treatment for four weeks, and then the relevant indicators were tested. The results demonstrated that HYD had antigoiter effects, alleviated the pathological changes in the thyroid tissue, and restored the abnormal serum levels of hormones related to thyroid function induced by PTU. HYD containing Glycyrrhiza uralensis had the best therapeutic effect in rats with PTU-induced goiter. The antigoiter effect of HYD may function through the hypothalamic-pituitary-thyroid (HPT) axis, inhibit the expression of the Tg and NIS genes, and regulate the synthesis of thyroid hormones, thereby reducing the excessive stimulation of TSH in thyroid cells. In addition, HYD also prevented goiter by promoting thyroid cell apoptosis and inhibiting the ERK/RSK1 pathway of cell proliferation. In conclusion, three types of HYD had different therapeutic effects in rats with goiter, which might be caused by the compatibility of different varieties of Glycyrrhiza and Sargassum.

目的 探讨海藻玉壶汤中海藻与甘草加减应用对甲状腺肿大模型大鼠哺乳动物雷帕霉素靶蛋白(mTOR)-核糖体S6蛋白激酶(p70S6K)/真核起始因子4E结合蛋白(4E-BP1)信号通路的影响.方法 选取Wistar大鼠84只,雄性,随机分为空白组、模型组、优甲乐组(0.02μg/g)、海藻玉壶汤组(10.08 g/kg)、海藻玉壶汤减海藻组(9.00 g/kg)、海藻玉壶汤减甘草组(9.18 g/kg)和海藻玉壶汤减海藻甘草组(8.10 g/kg),除空白组其余各组给予丙硫氧嘧啶复制甲状腺肿大病理模型,以优甲乐作为阳性对照药,其余各给药组给予相应药液.计算各组大鼠甲状腺系数,观察甲状腺病理形态变化,用RT-PCR法检测甲状腺组织中mTOR、p70 S6 K、4 E-BP1 mRNA的表达情况.结果 与空白组比较,模型组大鼠甲状腺系数、甲状腺细胞核个数升高(P<0.01),滤泡腔面积降低(P<0.01);与模型组比较,海藻玉壶汤组及各拆方组大鼠甲状腺系数、甲状腺细胞核个数降低(P<0.01),滤泡腔面积升高(P<0.01),其中海藻玉壶汤组的恢复作用最明显.与空白组比较,模型组mTOR、p70 S6 K、4 E-BP1 mRNA表达均升高(P<0.01);与模型组比较,海藻玉壶汤组及拆方各组均可使mTOR、p70 S6 K、4 E-BP1 mRNA表达水平降低(P<0.01).结论 海藻玉壶汤及拆方对甲状腺肿大模型大鼠的甲状腺系数及甲状腺组织病理形态有纠正作用,其作用机制可能与其抑制mTOR-p70 S6 K/4 E-BP1信号通路进而抑制细胞增殖有关.

目的 探讨山奈酚和芒柄花黄素对缺氧损伤的H9c2心肌细胞超氧化物歧化酶(SOD)活性,丙二醛(MDA)含量和肌酸激西酶(CK)、乳酸脱氢酶(LDH)活性的影响.方法 将H9c2细胞分为对照组、缺氧模型组、阳性药夹竹桃组、山奈酚组、芒柄花黄素组,采用CCK8法检测细胞活力,采用试剂盒检测上清液中CK、LDH活性和细胞内SOD活性,MDA含量.结果 与正常组相比,模型组细胞活力显著降低(P＜0.01);山奈酚、夹竹桃和芒柄花黄素能升高模型组细胞活力(P＜0.01,P＜0.01,P＜0.05).与正常组相比,模型组CK、LDH释放量和MDA活性显著升高(P＜0.01),SOD活性下降(P＜0.05).山奈酚组CK、LDH释放量均明显低于模型组(P＜0.01),芒柄花黄素组CK、LDH释放量低于模型组(P＜0.01,P＜0.05).山奈酚与芒柄花黄素均能显著降低模型组MDA活性(P＜0.01),升高模型组SOD活性(P＜0.05).结论 山奈酚和芒柄花黄素能通过降低CK、LDH活性和MDA含量,升高SOD活性对缺氧损伤心肌细胞发挥保护作用.

目的:观察三七总皂苷(PNS)对小鼠巨噬细胞炎性反应的治疗作用,同时探讨其可能机制.方法:通过蛋白质免疫印迹实验、实时PCR法、酶联免疫吸附试验,对照研究PNS对小鼠巨噬细胞系RAW 246.7细胞中炎性反应调控因子过氧化物酶体增殖物激活受体-α(PPAR-α)以及其下游调控的炎性反应递质肿瘤坏死因子-α(TNF-α)、C反应蛋白(CRP)和重组人巨噬细胞集落刺激因子(M-CSF),白细胞介素6(IL-6),炎性反应相关氧化因子诱导型一氧化氮合酶(iNOS)和环氧化酶-2(COX-2)表达的影响.结果:小鼠巨噬细胞炎性反应模型制备后TNF-α、CRP、M-CSF、IL-6、iNOS及COX-2水平明显上调,三七总皂苷给药后,细胞炎性反应递质与造模组比较有显著降低,差异有统计学意义(P＜0.05).结论:三七总皂苷可明显抑制小鼠巨噬细胞炎性反应效应,其作用机制可能通过激活调控因子PPAR-α实现.

Hepatocellular carcinoma (HCC) is a serious threat to human health. Chemotherapy drugs such as cisplatin are widely used in cancer treatment, but can cause severe side effects. Hydroxyl safflower yellow A (HSYA) is a water-soluble chalcone glycoside substance extracted from safflowers (Carthamus tinctorius L.) that has been reported to inhibit tumor growth with few side effects. The tumor immune microenvironment is crucial for the proliferation and invasiveness of tumor cells, and it is mediated by forkhead box P3-positive (FOXP3+) regulatory T cells (Tregs) and retinoic acid receptor-related orphan receptor-γ (RORγ)-expressing Th17 cells. FOXP3+ Tregs inhibit immunoreaction and FOXP3 is a key indicator of Tregs. RORγ isoform 2, also known as RORγt, is an important transcription factor in Th17 cells that may promote cancer progression. In the present study, the antitumor effect of HSYA on HCC was investigated, as well as its impact on the tumor immune microenvironment. Following the establishment of a mouse model for HCC, hematoxylin and eosin staining were performed to observe histological changes in liver tumors, and the spleen and thymus were weighed to calculate the spleen and thymus indexes. The proportion of FOXP3+ Tregs in the spleen was determined by flow cytometry, and expression levels of Foxp3 and Rorγt were examined by reverse transcription-quantitative polymerase chain reaction and western blot analysis. The results of the present study showed that cisplatin inhibited tumor growth, caused weight loss and reduced the immunoreactivity of the mice. HSYA inhibited tumor growth without causing significant weight loss. The proportion of FOXP3-expressing Tregs in the spleen and the expression of Foxp3 and Rorγt mRNA decreased following treatment with certain doses of HSYA. In conclusion, HSYA inhibited tumor growth without detrimental effects on the weight of the mice, indicating that HSYA may be suitable as a novel therapy for HCC patients.

心力衰竭具有高发病率,高死亡率,预后不利等特点,已成为严重危害人类健康,影响人类生活质量的公共卫生问题.因心力衰竭发病机制复杂,累及全身重要的器官,以往研究其分子机制相对独立,难以全面揭示发病机理,并且西药在治疗心衰的过程中带来的不良反应难以避免,不利于心衰患者的预后,因此文章围绕中药在治疗心衰,心室重构,减轻心肌纤维化,改善心功能,降低心衰复发率等方面的研究进展进行系统的综述.

Lipid raft microdomain of the plasma membrane is implicated in various biological and pathological processes. The involvement of lipid raft in T cell receptor (TCR) signal transduction has been widely studied, whereas the role of these structures in immunoreceptor signaling by DAP12 in natural killing (NK) cells remains largely unknown. Here, we demonstrate that phosphatidylinositol 4,5-bisphosphate (PIP2) lipid localized to lipid raft boundary in our coarse-grained (CG) model raft-forming membrane, and this negatively charged lipid recruits DAP12 homodimer into lipid raft boundary through protein-lipid interaction between the basic-rich regions and signaling immunoreceptor tyrosine-based activation motifs (ITAMs) of DAP12 and PIP2. Furthermore, our results reveal that the protein-lipid interaction can be disrupted by Ca2+, which competitively binds to PIP2 instead of DAP12. As a result, the cytoplasmic region of DAP12 homodimer is dissociated from the membrane back to the nonraft domain, and the ITAMs are exposed to allow further downstream signaling. These findings provide fundamental insights to understand the mechanism of signal transduction in NK cells regulated by membrane microenvironment.

目的:当代研究表明胸腺基质淋巴细胞生成素(TSLP)可以活化树突细胞(以下简称DCs),通过改变DCs分泌的细胞因子,形成有利于Th0向Th2优势分化的微环境,从而诱发过敏性鼻炎.临床发现小青龙汤治疗过敏性鼻炎疗效确切,关于其抗过敏的作用机制目前尚不明确.实验在体外用小青龙汤对TSLP-DCs体系进行干预,观察小青龙汤能否改变Th2优势分化的微环境.方法:从C57小鼠获取骨髓前体细胞,体外诱导培养生成DCs,设置空白组、TSLP刺激组、小青龙汤大中小3个剂量干预组,结合酶联免疫吸附实验(以下简称ELISA)在12、24、48 h3个时间点分别检测IL-4、IL-12、IFN-γ、IL-10的表达.结果:与空白组相比,各TSLP刺激组IL-4的表达均明显升高(P＜0.01),而IL-10、IFN-γ的表达均明显降低(P＜0.01).与TSLP刺激组相比,大中小三个剂量的小青龙汤干预组IL-4的表达量均明显降低(P＜0.01),降低最为明显的是4μg/mL小青龙汤组;与TSLP刺激组相比,大中小三个剂量的小青龙汤干预组IFN-γ、IL-10的表达量均升高(P＜0.01),升高最为明显的是4 μg/mL小青龙汤组;IL-10的表达量随时间增加呈现降低趋势并在48 h时表达量达到最低;IFN-γ、IL-10的表达量随时间增加呈现先升高后降低的趋势并在24h时表达量达到最高.结论:小青龙汤可以降低TSLP-DCs体系IL-4的表达,升高IL-10、IFN-γ的表达,改变Th2优势分化的微环境起到抗过敏作用,在剂量为4 μg/mL时作用最为明显.