Mechanism of Yiqi Jiedu Prescriptions in Protection of Mitochondria in PC12 Cells Against Hypoxia Injury Based on Energy Metabolism / 中国实验方剂学杂志

ObjectiveTo establish an evaluation method for mitochondrial energy metabolism with Seahorse analyzer and investigate the protective effect of Yiqi Jiedu prescriptions （YQ） on mitochondria in rat adrenal pheochromocytoma （PC12） cells against hypoxia injury. MethodThe PC12 cell injury model was induced in vitro using hypoxic chambers. Five groups were set up， ie， a control group， a model group （model）， high- （25 µmol·L-1）， medium- （5 µmol·L-1） and low-dose （1 µmol·L-1） YQ groups， and a positive drug trimetazidine （TMZ） group， with three replicate wells in each group. The experiment was repeated three times. The established method for energy metabolism analysis was used to assay the activity of mitochondrial complex in cells and screen the optimal dosing concentration. Subsequently， the YQ group and modified YQ groups were set up， and the aerobic respiration and glycolysis function were assayed by the Seahorse analyzer. According to the non-mitochondrial oxygen consumption， proton leakage， basal respiration， maximum respiration， ATP production， and potentially improved respiration， the effects of modified YQ groups on the aerobic respiration of mitochondria damaged by hypoxia were evaluated by principal component analysis （PCA） and variable importance in projection （VIP）. The expression of cytochrome C， B-cell lymphoma-2 （Bcl-2）， and Bcl-2-associated X protein （Bax） was detected by Western blot. ResultCompared with the groups of other concentrations， the optimal dosing concentration of carbonyl cyanide-4 （trifluoromethoxy）phenylhydrazone （FCCP） was 2 µmol·L-1. Compared with the model group， the medium-dose YQ group showed enhanced mitochondrial complex activity （P<0.05）. The YQ groups were superior to the model group in improvement （P<0.01）. The combination of ginsenoside and geniposide showed the optimal effect among the modified YQ groups （P<0.01）. VIP analysis revealed that for the improvement of mitochondrial respiratory function， the contribution of geniposide in YQ was the greatest. Compared with the model group， the high-dose YQ group displayed reduced leakage of mitochondrial cytochrome C （P<0.01）， decreased expression of Bax protein （P<0.01）， and increased expression of Bcl-2 protein （P<0.05， P<0.01）. ConclusionA cellular， high-throughput quantitative evaluation method for mitochondrial energy metabolism was established， which demonstrated that YQ could significantly improve the impaired mitochondrial energy metabolism in PC12 cells damaged by hypoxia， and the underlying mechanism might be related to the protection against mitochondrial apoptosis.

Correlation of gut microbiota and ischemic stroke: a review / 中国中药杂志

With the widespread application of next-generation sequencing(NGS), especially 16 S rRNA and shotgun sequencing, researchers are no longer troubled with massive data on the gut microbiota, and the correlation between the gut microbiota and the brain(central nervous system) has been gradually revealed. Research on the microbiota-gut-brain axis(MGBA) based on the gut microbiota have provided insights into the exploration of the pathogenesis and risk factors of ischemic stroke(IS), a cerebrovascular disease with high disability and mortality rates, and also facilitate the selection of therapeutic targets of this class of drugs. This study reviewed the application of NGS in the study of gut microbiota and the research progress of MGBA in recent years and systematically collated the research papers on the correlation between IS and gut microbiota. Furthermore, from the bi-directional regulation of MGBA, this study also discussed the high-risk factors of IS under the dysregulation of gut microbiota and the pathophysiological changes of gut microbiota after the occurrence of IS and summarized the related targets to provide a reliable reference for the therapeutic research of IS from the gut microbiota.

Synergistic antibacterial effect and mechanisms of dihydroartemisinin and cefuroxime incombination / 中国中药杂志

In this study, we explored the antibacterial effect and mechanism of dihydroartemisinin(DHA) combined with cefuroxime(CFX) or ampicillin against Escherichia coli. The minimum inhibitory concentration(MIC) of DHA, cefuroxime, and ampicillin against E. coli was 300,25,25 μmol·L~(-1), respectively, determined by broth microdilution method and 2,3,5-triphenyltetrazolium chloride(TTC) method. The minimum bactericidal concentration(MBC) was 25 μmol·L~(-1) for cefuroxime, above 600 μmol·L~(-1) for DHA. The fractional inhibitory concentration index(FICI) of DHA combined with cefuroxime or ampicillin was 0.375 and 0.75, respectively, determined by checkerboard microdilution assay, suggesting the synergistic effect or additive effect of the drug combination. Moreover, the time-effect curve showed that the antibacterial activity of DHA and CFX combination was much stronger than that of either of the drugs, suggesting that combination with DHA can decrease the CFX dosage. Then we studied the synergistic mechanism of DHA combined with cefuroxime and found that the combination of the two drugs had a significant inhibitory effect on the total protein bands, as shown by the results of polypropylene gel electrophoresis. The results of conductivity method and alkaline phosphatase test respectively showed that its conductivity value and alkaline phosphatase(AKP) leak were significantly higher than either of the drugs, suggesting that the integrity of bacteria may be damaged. The scanning electron microscope(SEM) results showed that the morphology of E. coli was destroyed most in the combination group. The quantitative fluorescence PCR technology showed that the combination of two drugs can inhibit the expression of superoxide stress gene soxS. In summary, the combination of dihydroartemisinin and cefuroxime has a synergistic antibacterial effect on E. coli.

Effect and Mechanism of Dihydroartemisinin on Rheumatoid Arthritis Animal Models / 中国实验方剂学杂志

Objective:To study the protective effect of dihydroartemisinin (DHA) on adjuvant-induced arthritis (AIA) and collagen-induced arthritis (CIA) in rats, in order to explore its possible mechanism. Method:Wistar rats were randomly divided into eight groups, namely AIA control group, AIA model group, AIA DHA group and AIA methotrexate group, CIA control group, CIA model group, CIA DHA group and CIA methotrexate group. To establish adjuvant-induced arthritis (AIA) model, rheumatoid arthritis rats were induced through intradermal injection with 0.1 mL Freund's complete adjuvant (FCA) into right postpedes except for the control group. To establish the model of collagen-induced arthritis (CIA), except for control group, the caudal root of rats was immunized subcutaneously with 0.2 mL of emulsion containing 1 g·L-1 of Collagen type Ⅱ (CⅡ). One week later, CⅡ emulsion was injected for the second time. After the rheumatoid arthritis model was successfully established and the administration with DHA (30 mg·kg-1·d-1), the anti-inflammatory effect of DHA on AIA/CIA rats was observed, including the arthritis index (AI), paw swelling degree and effect of DHA on immune organ index of AIA/CIA rats. Interleukin (IL)-6 levels in serum was detected by enzyme-linked immunosorbent assay (ELISA) and pathological sections of ankle joints of AIA/CIA rats. RAW264.7 macrophage cells were cultured in vitro and treated with DHA at various doses (0.5, 1, 2, 4, 8, 16 μmol·L-1) for 24 h, and the cell viability was detected by methyl thiazolyl tetrazolium(MTT) assay. Lipopolysaccharides (LPS) group, LPS+DHA groups (0.5, 1, 2 μmol·L-1) and control group were established. The level of IL-6 was detected by enzyme-linked immunosorbent assay(ELISA). The protein expression levels of nuclear transcription factor-κB p65 (NF-κB p65) was tested by Western blot. Result:Compared with control group, the paw swelling, AI, spleen index and IL-6 levels of model group were significantly increased (PPPPPPPP-1) groups had a remarkable effect on the cell viability (PP-1. The level of IL-6 and the protein expression of NF-κB p65 in LPS group were higher than those of control group. Compared with LPS group, DHA (0.5 μmol·L-1) groups could significantly reduce the secretion of IL-6 (PκB p65. Conclusion:DHA can alleviate the ankle joint lesion on rheumatoid arthritis rats. Its mechanism may be related to NF-κB signal pathway.

Preliminary study on antibacterial activity of artemisinin and its derivatives / 中国中药杂志

In this study,in order to detect the antimicrobial activity of artemisinin and its derivatives artesunate and dihydroartemisinin,two methods including broth dilution and plate punching method were used to detect the antibacterial activity against gram-negative bacteria(Escherichia coli)and gram-positive bacteria(Staphylococcus aureus)of artemisinin,dihydroartemisinin and artesunate at various concentrations within 5 mmol·L~(-1)and at four time points(8,16,24,32 h).Two antibacterial positive drugs,streptomycin against E.coli and penicillin against S.aureus,were used as positive controls.Plate punching method showed that,unlike the results of 5 mmol·L~(-1)dihydroartemisinin or artesunate,no inhibition zone was detected at the same concentration of artemisinin after 24 h-treatment against E.coli.Broth dilution method showed that,the antibacterial activity of dihydroartemisinin against E.coli.was stronger than those of both artesunate and artemisinin;IC_(50)at24 h-treatment was 155.9μmol·L~(-1)for dihydroartemisinin,370.0μmol·L~(-1)for artesunate and none for artemisinin.Interestingly,dihydroartemisinin and artesunate showed the strongest antibacterial activity between 16-24 h,while artemisinin showed relatively stronger antibacterial activity between 8-16 h.Dihydroartermisinin showed no antibacterial activity against S.aureus.Above all,the antibacterial activity of artemisinins against E.coli is dihydroartemisinin>artesunate>artemisinin.Artemisinin and its derivatives have showed different antibacterial kinetics,and no antibacterial activity against S.aureus.has been detected with dihydroartemisinin.

Mechanism of anti-endotoxemia based on LPS-TLR4 signaling pathway:research advances / 国际药学研究杂志

Toll-like receptors 4(TLR4),an important member of the TLR family,is considered as gene encoding LPS recep?tors,and major receptors for identifying lipopolysaccharide(LPS).LPS-induced TLR4 signaling pathway plays a key role in endotox?emia.After TLR4 activated by LPS,the mitogen-activated protein kinase(MAPK)signaling pathway and nuclear transcription factor κB(NF-κB)are activated and large amounts of inflammatory factors are released,triggering inflammatory cascade reaction.This arti?cle reviews the mechanisms of endotoxemia in treatment based on TLR4 signal pathway in three perspectives:the effects of LPS-TLR4 signaling pathway on the upstream target proteins,LPS-induced TLR4 signal transduction,and the downstream target proteins regulat?ed by LPS-TLR4 signaling pathway.

Glucocorticoids Significantly Influence the Transcriptome of Bone Microvascular Endothelial Cells of Human Femoral Head / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Appropriate expression and regulation of the transcriptome, which mainly comprise of mRNAs and lncRNAs, are important for all biological and cellular processes including the physiological activities of bone microvascular endothelial cells (BMECs). Through an intricate intracellular signaling systems, the transcriptome regulates the pharmacological response of the cells. Although studies have elucidated the impact of glucocorticoids (GCs) cell-specific gene expression signatures, it remains necessary to comprehensively characterize the impact of lncRNAs to transcriptional changes.</p><p><b>METHODS</b>BMECs were divided into two groups. One was treated with GCs and the other left untreated as a paired control. Differential expression was analyzed with GeneSpring software V12.0 (Agilent, Santa Clara, CA, USA) and hierarchical clustering was conducted using Cluster 3.0 software. The Gene Ontology (GO) analysis was performed with Molecular Annotation System provided by CapitalBio Corporation.</p><p><b>RESULTS</b>Our results highlight the involvement of genes implicated in development, differentiation and apoptosis following GC stimulation. Elucidation of differential gene expression emphasizes the importance of regulatory gene networks induced by GCs. We identified 73 up-regulated and 166 down-regulated long noncoding RNAs, the expression of 107 of which significantly correlated with 172 mRNAs induced by hydrocortisone.</p><p><b>CONCLUSIONS</b>Transcriptome analysis of BMECs from human samples was performed to identify specific gene networks induced by GCs. Our results identified complex RNA crosstalk underlying the pathogenesis of steroid-induced necrosis of femoral head.</p>

Effects of Ginkgo biloba extract 50 on hypoxia induced endothelial dysfunction / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To preliminarily investigate the influence of hypoxia on human umbilical vein endothelial cells (HUVECs), and the effect of Ginkgo biloba extract 50 (GBE50) on it.</p><p><b>METHODS</b>Flow cytometry, TUNEL, RT-PCR, Western blot, etc. were applied, to study the effect of hypoxia and GBE50 on endothelial cells.</p><p><b>RESULTS</b>After being interfered by hypoxia for 24 h, the levels of reactive oxygen species (ROS) in HUVECs and the apoptotic rate either in the early or in the late stage significantly increased, and accompanied with the increased levels of endothelin-1 mRNA (ET-1) and endothelial oxide synthase (eNOS) protein. However, when HUVECs were pretreated with GBE50 (25 [microg/ml) 4 h before hypoxia, the apoptotic rate in the early or late stage and expression of ET-1 mRNA significantly decreased (P < 0.05), and the heightened ROS level and eNOS expression partially decreased (P > 0.05).</p><p><b>CONCLUSION</b>Hypoxia can induce endothelial dysfunction, which could be partially or significantly reversed by GBE50, it shows a certain protective effect on hypoxia induced endothelial dysfunction.</p>