Inosine:A broad-spectrum anti-inflammatory against SARS-CoV-2 infection-induced acute lung injury via suppressing TBK1 phosphorylation / 药物分析学报

Severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)-induced cytokine storms constitute the primary cause of coronavirus disease 19(COVID-19)progression,severity,criticality,and death.Gluco-corticoid and anti-cytokine therapies are frequently administered to treat COVID-19,but have limited clinical efficacy in severe and critical cases.Nevertheless,the weaknesses of these treatment modalities have prompted the development of anti-inflammatory therapy against this infection.We found that the broad-spectrum anti-inflammatory agent inosine downregulated proinflammatory interleukin(IL)-6,upregulated anti-inflammatory IL-10,and ameliorated acute inflammatory lung injury caused by mul-tiple infectious agents.Inosine significantly improved survival in mice infected with SARS-CoV-2.It indirectly impeded TANK-binding kinase 1(TBK1)phosphorylation by binding stimulator of interferon genes(STING)and glycogen synthase kinase-3β(GSK3β),inhibited the activation and nuclear trans-location of the downstream transcription factors interferon regulatory factor(IRF3)and nuclear factor kappa B(NF-κB),and downregulated IL-6 in the sera and lung tissues of mice infected with lipopoly-saccharide(LPS),H1N1,or SARS-CoV-2.Thus,inosine administration is feasible for clinical anti-inflammatory therapy against severe and critical COVID-19.Moreover,targeting TBK1 is a promising strategy for inhibiting cytokine storms and mitigating acute inflammatory lung injury induced by SARS-CoV-2 and other infectious agents.

Shaoyaotang Alleviates Ulcerative Colitis by Regulating Th17/Treg Balance Through IL-6/STAT3 Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo explore the role of interleukin-6 （IL-6）/signal transducer and activator of transcription 3 （STAT3） pathway in the balance of T helper 17 （Th17）/regulatory T （Treg） cells in ulcerative colitis （UC） with internal dampness-heat accumulation syndrome and the intervention mechanism of Shaoyaotang. MethodA total of 60 SD rats were randomized into blank group （equivalent volume of normal saline）， model group （equivalent volume of normal saline）， western medicine control group （0.42 g·kg-1 mesalazine）， and low-dose （11.1 g·kg-1）， medium-dose （22.2 g·kg-1）， and high-dose （44.4 g·kg-1） Shaoyaotang groups. UC with internal dampness-heat accumulation syndrome was induced in rats with the compound method except for the blank group. The administration lasted 14 days for each group. At 24 h after the last administration， rats were killed and the spleen and colon tissues were separated. The histopathological changes of colon were observed based on hematoxylin and eosin （HE） staining and the levels of interleukin-17 （IL-17） and transforming growth factor-β1 （TGF-β1） in colon tissue were detected by immunohistochemistry （IHC）. Flow cytometry was employed to determine the levels of Th17/Treg cells in the spleen， and Western blot to measure the levels of IL-6 and STAT3 proteins in colon tissue. ResultCompared with the blank group， the model group had lesions such as congestion and erosion， low percentage of spleen Treg cells （P<0.01）， high percentage of Th17 cells （P<0.01）， and high levels of IL-6 and STAT3 proteins in colon tissue （P<0.01）. Compared with the model group， the administration groups showed alleviation of colon injury， high percentage of spleen Treg cells （P<0.05， P<0.01）， low percentage of Th17 cells （P<0.01）， and low levels of IL-6 and STAT3 proteins in colon tissue （P<0.01）. ConclusionShaoyaotang regulates the balance of Th17/Treg by inhibiting the IL-6/STAT3 pathway， thereby relieving the pathological damage of UC rats with internal dampness-heat accumulation syndrome and affecting their immune function.

Role of interleukin-6 in ocular inflammatory diseases / 国际眼科杂志(Guoji Yanke Zazhi)

Interleukin-6(IL-6)is a pleiotropic cytokine produced by monocytes, macrophages, T lymphocytes and other cell types. It is significantly up-regulated in the process of infection and inflammation, and is the core cytokine of the host's defense against environmental stresses(such as injury and infection). Abnormal and persistent IL-6 production is closely associated with the development of various autoimmune and inflammatory diseases. A growing number of studies have shown that IL-6 plays a critical role in ocular inflammation and angiogenesis in the conjunctiva, cornea, uvea and retina. Blockade of IL-6 ameliorates chronic and refractory intraocular inflammation. This article aims to review the role as well as the mechanism of IL-6 in ocular inflammatory diseases, attempting to have a deep and systematic understanding of the role of IL-6 in ocular inflammatory diseases.

Molecular Mechanism of Bailing Capsules in Treatment of Chronic Obstructive Pulmonary Disease Based on Network Pharmacology / 中国实验方剂学杂志

ObjectiveThis study aimed to predict the pharmacodynamic material basis and core targets of Bailing capsules in the treatment of chronic obstructive pulmonary disease （COPD） based on network pharmacology and molecular docking， which were further verified by cell experiments to explore the mechanism. MethodThe main active ingredients and related targets of Bailing capsules were screened in Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform （TCMSP） and SwissTargetPrediction. The main COPD targets were searched from GeneCards， DrugBank， Online Mendelian Inheritance in Man （OMIM） and Therapeutic Target Database （TTD）. The protein-protein interaction （PPI） network was constructed by STRING and Cytoscape 3.6.1. Gene Ontology （GO） function annotation and Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment analysis were performed by the Database for Annotation， Visualization and Integrated Discovery （DAVID）. Molecular docking verification was carried out using AutoDock Vina. The cell viability was detected by 3-（4，5-dimethylthiazol-2-yl）-5-（3-carboxymethoxyphenyl）-2-（4-sulfophenyl）-2H-tetrazolium （MTS） assay， and the mRNA level of the targets was detected by real-time polymerase chain reaction （Real-time PCR）. ResultA total of 11 active ingredients of Bailing capsules such as cerevisterol， 270 related drug targets， and 1 020 COPD target proteins were obtained， with 74 intersection targets. The visualization analysis of the PPI network showed that the core targets of Bailing capsules in the treatment of COPD were tumor protein P53 （TP53）， catenin beta 1 （CTNNB1）， tumor necrosis factor （TNF）， interleukin-6 （IL-6） and insulin （INS）. Further， 20 signaling pathways were screened by KEGG enrichment analysis as the main pathways for Bailing capsules to treat COPD， involving phosphatidylinositol 3-kinase/protein kinase B （PI3K/Akt）， cyclic adenosine monophosphate （cAMP）， forkhead box O （FoxO）， TNF， and hypoxia inducible factor-1 （HIF-1） signaling pathways. Molecular docking validation demonstrated that four active ingredients had stable binding to IL-6， with the lowest energy. Bailing capsules could reduce the mRNA level of IL-6 in RAW264.7 cells induced by lipopolysaccharide （LPS） （P<0.01） compared with the control group. ConclusionThe pharmacological mechanism of Bailing capsules in the treatment of COPD might be that its main active ingredients improved the inflammatory response by acting on TP53， CTNNB1， TNF， IL-6 and other targets and regulating PI3K/Akt， cAMP and other signaling pathways， thereby ameliorating COPD symptoms. This study provided experimental basis for subsequent in-depth research， and provided a diagnosis and treatment direction for disease-related clinical treatment.

Da Chaihutang Inhibits Hepatocellular Carcinoma by Regulating p38 MAPK/IL-6/STAT3 Signaling Pathway / 中国实验方剂学杂志

ObjectiveThis study aims to investigate the efficacy and underlying mechanism of Da Chaihutang （DCHT） in treating hepatocellular carcinoma （HCC） in vitro and in vivo. MethodWe employed methyl thiazolyl tetrazolium （MTT） assay and crystal violet staining to observe the proliferation of Hepa1-6 liver cancer cells treated with DCHT at different doses （0， 125， 250， 500， 1 000 mg·L-1） for different time periods （1， 2， 4， 8 days）. The orthotopic liver cancer model was established by injection of 1×106 Hepa1-6 cells into mouse， and then the model mice were randomly assigned into six groups： blank， model， DCHT （0.21， 0.625， 1.875 g·kg-1， ig， qd）， and positive control （5-fluorouracil， 25 mg·kg-1， ip， qod）. After 14 days of administration， the mice were sacrificed， and the liver samples were collected and fixed in 4% paraformaldehyde for hematoxylin-eosin （HE） staining. The Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform （TCMSP）， Cytoscape 3.7.2， STRING， and DAVID were used for the searching of the key targets of DCHT in treating HCC， the construction of protein-protein interaction （PPI） network， and the Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment. Quantitative real-time PCR was performed to determine the mRNA level of interleukin-6 （IL-6） in Hepa1-6 cells and liver tissue. Western blotting was employed to measure the protein levels of the proteins involved in the mitogen-activated protein kinase （MAPK） and signal transducers and activators of transcription 3 （STAT3） signaling pathways. ResultDCHT （500， 1 000 mg·L-1） treatment for 4 and 8 days inhibited the proliferation of Hepa1-6 cells in a dose- and time-dependent manner （P<0.05）. The in vivo assay showed that DCHT （high dose， 1.875 g·kg-1） treatment for 14 days led to high differentiation and unobvious heterogeneity of HCC cells and small necrotic area compared with the model group. Network pharmacology analysis predicted that the potential targets of DCHT in the treatment of HCC were mainly the inflammation cytokines such as IL-6， interleukin-1β （IL-1β）， and tumor necrosis factor-alpha （TNF-α） in HCC microenvironment. The potential signaling pathways involved in the treatment were mainly associated with HCC growth and differentiation， including MAPK and STAT3 signaling pathways. Compared with the blank group， DCHT （1 000 mg·L-1） treatment for 1， 2， 4， and 8 days down-regulated the mRNA level of IL-6 in Hepa1-6 cells （P<0.05）. Similar results were observed in the livers of mice treated with DCHT （0.625， 1.875 g·kg-1）. The in vitro assay demonstrated that DCHT （1 000 mg·L-1） treatment for 4 and 8 days and DCHT （500， 1 000 mg·L-1） treatment inhibited the phosphorylation of extracellular signal-regulated kinases 1/2 （ERK1/2）， c-Jun NH2-terminal kinase/stress-activated protein kinase （JNK）， p38 MAPK， and STAT3 in a dose- and time-dependent manner （P<0.05）. The in vivo assay showed that DCHT （0.625 and 1.875 g·kg-1） treatment only inhibited the phosphorylation of p38 MAPK and STAT3 （P<0.05）. ConclusionThe present study indicates that DCHT can inhibit liver cancer cell proliferation by regulating p38 MAPK/IL-6/STAT3 signaling pathway.

Regulatory Effect of Herbal Pair Astragali Radix-Puerariae Lobatae Radix on PI3K/Akt/FoxO1 Signaling Pathway in Rats with T2DM Based on Network Pharmacology / 中国实验方剂学杂志

ObjectiveTo explore the mechanism of herbal pair Astragali Radix-Puerariae Lobatae Radix （AR-PLR） against type 2 diabetes mellitus （T2DM） based on network pharmacology and experimental verification. MethodThe active ingredients and targets of AR and PLR were retrieved from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform （TCMSP）. The related targets of T2DM were retrieved from disease databases and the common targets of drugs and diseases were extracted. The protein-protein interaction （PPI） network was analyzed and constructed by STRING and the network topology of key targets was analyzed by Cytoscape 3.7.1. Then gene ontology（GO） and Kyoto encyclopedia of genes and genomes（KEGG） enrichment analyses of core targets were carried out by DAVID to explore its possible molecular mechanism. The T2DM model was induced in rats by the high-fat diet combined with tail intravenous injection of streptozocin. The rats were divided into a normal group，a model group，a metformin group，and high-，medium- and low-dose AR-PLR groups. After four weeks of intragastric administration，the serum levels of fasting blood sugar （FBS），fasting insulin（FINS），aspartate aminotransferase（AST），alanine aminotransferase（ALT），triglyceride（TG），total cholesterol（TC），low-density lipoprotein cholesterin（LDL-C），high-density lipoprotein cholesterin （HDL-C），interleukin-6（IL-6）， and tumor necrosis factor-α（TNF-α） of rats in each group were measured. The protein expression of insulin receptor substrate-2（IRS-2），phosphatidylinositol 3-kinase（PI3K），protein kinase B（Akt）， and forkhead box transcription factor O1（FoxO1） in rat liver was detected by Western blot. ResultA total of 131 core targets of AR-PLR in the treatment of T2DM were screened out by network pharmacology， where Akt1，mitogen-activated protein kinase 1 （MAPK1），TNF-α，and IL-6 were critical. As revealed by KEGG enrichment analysis， AR-PLR exerted the hypoglycemic effect mainly through the PI3K/Akt，TNF， and FoxO signaling pathways. Compared with the model group，the high- and medium-dose AR-PLR groups showed reduced FBS and FINS levels and increased glycogen level （P<0.05，P<0.01），all the AR-PLR groups showed decreased levels of AST，ALT，TG， and LDL-C （P<0.05，P<0.01）， the high- and low-dose AR-PLR groups showed decreased TC levels （P<0.05）. Compared with the model group， the high- and medium-dose AR-PLR groups showed reduced levels of IL-6 and TNF-α（P<0.05，P<0.01）， and the high-dose AR-PLR group showed increased expression of IRS-2， Akt， p-Akt， PI3K， and p-PI3K， and decreased expression of FoxO1 protein（P<0.05）. ConclusionAR-PLR has the characteristics of multi-component，Multi-target and multi-pathway in the treatment of T2DM. This herbal pair may regulate the PI3K/Akt/FoxO1 signaling pathway through IL-6， TNF-α， and other targets to affect insulin resistance， glycogen synthesis， gluconeogenesis， glucose transport， inflammation， immune response， and other processes， thereby treating T2DM.

Molecular Mechanism Analysis of Dengzhan Shengmai Capsules Against Chronic Cerebral Hypoperfusion / 中国实验方剂学杂志

ObjectiveBased on the protective effect of Dengzhan Shengmai capsules （DZSM） on chronic cerebral hypoperfusion （CCH）， network pharmacology was employed to investigate the molecular mechanism. MethodCCH model was established by right common carotid artery ligation. The mice were divided into sham operation group， model group， ginaton group （48 mg·kg-1）， DZSM low- and high-dose groups （0.040 5， 0.162 g·kg-1）. The efficacy was evaluated by the Morris water maze test and open-field test. The underlying mechanism of DZSM for CCH was analyzed by network pharmacology and verified by molecular biology experiments. PubChem， GeneCards， Metascape and other databases were used for targets collection and enrichment analysis. Besides， the association of ingredients targets of DZSM with disease targets of CCH， core target network and chemical components-core targets-pathways network were constructed by STRING 11.0 and Cytoscape 3.7.1. ResultThe escape latency of CCH mice significantly shortened on the 3rd to 5th day after DZSM low-dose treatment， the crossing times， time spent in the target quadrant， movement distance and distance in the central region of CCH mice significantly increased after DZSM low-dose and high-dose treatment. The results of network pharmacology indicated that DZSM might play a key role by regulating inflammatory response， phosphatidylinositol 3-kinase/protein kinase B （PI3K/Akt） signaling pathway， cytokine-cytokine receptor interaction， tumor necrosis factor （TNF） signaling pathway， blood circulation， angiogenesis， extracellular matrix and other related biological processes and pathways， and acting as targets such as interleukin-6 （IL-6）， TNF， insulin-like growth factor 1 （IGF1）， vascular endothelial growth factor A （VEGFA）， epidermal growth factor （EGF）. The results of biological experiments showed that DZSM could reduce the expression of IL-6 in brain tissue of CCH mice. ConclusionDZSM provides a protective effect during CCH， and its multi-component， multi-pathway， multi-target mechanism is also revealed， which provides a basis for further study of the mechanism.

Citoquinas pro y anti-inflamatorias en la infección por SARS-CoV-2, en población de Colombia / Pro- and anti-inflammatory cytokines associated to SARS-COV-2 infection in the Colombian population

Resumen La infección por SARS-CoV-2 es hoy el principal problema de salud pública en el mundo. No es claro el papel de las citoquinas en la fisiopatología del COVID-19,que en algunos individuos presenta una progresión rápida, severa y mortal asociada con proinflamación sistémicos relacionada con coagulopatías y fallas multiorgánicas. En este estudio, evaluamos los niveles séricos de citoquinas y su correlación con IgM, IgG e IgA, en 24 muestras de individuos positivos y 8 muestras de individuos negativos, para SARS-CoV-2. Hallamos concentraciones significativamente menores de IFN-g, TNF, IL-2 e IL-4 y un aumento significativo de IL-6 en el grupo de infectados hospitalizados respecto a los no infectados, así como una tendencia significativa al aumento, para IgG e IgA en el mismo grupo de individuos, respecto a infectados asintomáticos. Nuestros datos soportan el papel de la IL-6 en la severidad de la enfermedad destacando su potencial papel como biomarcador en la prognosis de esta patología. También, soportan la hipótesis sobre la función de los anticuerpos en el control efectivo del patógeno; se observa una respuesta inmune humoral más débil, frente a la proteína de la nucleocápside viral, en individuos con un mejor curso de la enfermedad.

Abstract The emergency caused by the infection in humans of SARS-COV-2 and the clinical syndrome resulting from the infection (COVID-19) is a major public health crisis with global repercussions. Currently, the role of different cytokine profiles in the infection pathophysiology and its outcome remains unclear despite the coordina ted efforts of the scientific community. COVID-19 shows a rapid progression where the disease severity and mortality are linked to systemic pro-inflammatory pro cesses associated to a dysregulation in the cytokine production balance, resulting in blood clothing disorders and multiorgan failure. Here we evaluate the serum concentration for a cytokine panel as well as the antibody titers of IgM, IgG and IgA from 24 individuals who tested positive for SARS-CoV-2 by RT-PCR (divided into three separate groups according to disease severity) and eight RT-PCR-negative controls. Significantly lower concentrations of IFN-g, TNF, IL-2 and IL-4, and a higher production of IL-6 were observed in hospitalized COVID-19 patients when compared to SARS-CoV-2-negative individuals. Furthermore, a significant and sustained increase in the levels of IgG and IgA was found for the group of hospitalized patients compared to asymptomatic SARS-CoV-2-positive individuals. Our data support previous findings on the role of cytokines like IL-6 in the severity of the disease and highlight their potential use as biomarkers for the prognosis of COVID-19. Finally, we provide evidence supporting the potential function of the antibody response in the effective control of the virus, showing that a somehow weaker humoral immune response can be associated to milder forms of COVID-19.

Effect of Tongxie Yaofang on Expressions of Colon SERT and Liver 5-HT2AR Proteins in Rats with Ulcerative Colitis Model of Liver Stagnation and Spleen Deficiency / 中国实验方剂学杂志

Objective:To observe the effect of Tongxie Yaofang on the expressions of colon serotonin transporter （SERT）， liver 5-hydroxytryptamine_2A receptor （5-HT_2AR） protein， serum 5-HT and inflammatory factors in ulcerative colitis （UC） model rats of liver stagnation and spleen deficiency， in order to explore the basis of syndrome of liver stagnation and spleen deficiency and the intervention mechanism of Tongxie Yaofang. Method:Fifty male SD rats were randomly divided into blank control group， model group， high， medium and low-dose Tongxie Yaofang group （10，5，2.5 g·kg^-1）， and salazosulacil group （0.3 g·kg^-1）. The ulcerative colitis model of liver depression and spleen deficiency was established by 2，4，6-trinitrobenzene sulfonic acid （TNBS）/ethanol solution enema + restraint stress + diet loss. After successful modeling， the samples were collected after 21 days of drug intervention. Htoxylin eosin （HE） staining and oil red staining were used to observe the pathological changes of colon and liver in each group. Serum interleukin-6 （IL-6）， IL-9， 5-HT and superoxide dismutase （SOD） were detected by enzyme linked immunosorbent assay （ELISA）. Protein expressions of SERT in the colons and 5-HT_2AR in liver of rats were detected by Western blot. Result:Compared with the normal group， obvious ulcers were formed in the colon and lipid droplets in the liver increased in the model group， serum levels of IL-6， IL-9 and 5-HT in the model group increased， while the level of SOD decreased （<italic>P</italic><0.05）. The protein expression of SERT in colon decreased， whereas the protein expression of 5-HT_2AR in liver increased （<italic>P</italic><0.05）. Compare with model group， the pathological damage of colon was improved， and the formation of lipid droplets in liver was reduced in high， medium-dose Tongxie Yaofang groups and sulfasalazine group. The serum levels of IL-6， IL-9 and 5-HT decreased， while the level of SOD increased in Tongxie Yaofang group and sulfasalazine group （<italic>P</italic><0.05）. The protein expression of SERT in colon increased in high，low-dose Tongxie Yaofang groups and sulfasalazine group， and the protein expression of 5-HT_2AR in liver decreased in medium， low dose Tongxie Yaofang groups and sulfasalazine group （<italic>P</italic><0.05）. Conclusion:Tongxie Yaofang may reduce the content of 5-HT， and regulate the intestinal motility and sensory system by up-regulating the expression of SERT in the colon， inhibit the expressions of IL-6，IL-9 and other inflammatory factors， and play an anti-inflammatory role， reduce the content of 5-HT and the expression of 5-HT_2AR in the liver， increase the level of SOD， regulate emotion and lipid metabolism in the liver， and then exert the intervention effect on ulcerative colitis with liver depression and spleen deficiency on the whole.

Mechanism of Scutellariae Radix-Hedyotidis Herba Against Proliferation of Lung Adenocarcinoma A549 Cells / 中国实验方剂学杂志

Objective:To observe the effects of Scutellariae Radix-Hedyotidis Herba on the proliferation of human lung adenocarcinoma A549 cells and the expression of interleukin-6（IL-6）， phosphatidylinositol 3-kinase （PI3K），protein kinase B （Akt）， p-protein kinase B （p-Akt）， mechanistic target of rapamycin （mTOR）， hypoxia-inducible factor-1<italic>α </italic>（HIF-1<italic>α</italic>）， and Cyclin D₁ at the cellular level， and to explore their molecular mechanism. Method:Following the set-up of the blank group （complete medium）， low-， moderate-，and high-dose （20， 40， and 60 mg·L^-1） Scutellariae Radix-Hedyotidis Herba groups， and low-， moderate-， and high-dose （5， 10， and 20 mg·L^-1） cisplatin groups， the cell were treated with the corresponding drugs for 24， 48， and 72 h for detecting their viability by tetrazolium bromide （MTT） colorimetry. A549 cells were then divided into the blank group， Scutellariae Radix-Hedyotidis Herba group， cisplatin group， and combined medication group and intervened with thecomplete medium， 40 mg·L^-1Scutellariae Radix-Hedyotidis Herba， 10 mg·L^-1 cisplatin， and 40 mg·L^-1Scutellariae Radix-Hedyotidis Herba + 10 mg·L^-1cisplatin， respectively， for 24， 48 and 72 h， followed by the measurement of inhibitory effects against the proliferation of A549 cells in each experimental group. The level of IL-6 in cell culture supernatant was determined by enzyme-linked immunosorbent assay （ELISA） after 72 h. The mRNA expression levels of HIF-1<italic>α</italic> and Cyclin D₁ in each group were assayed by real-time polymerase chain reaction （Real-time PCR）， and the protein expression levels of PI3K， Akt， p-Akt， mTOR， HIF-1<italic>α</italic>， and Cyclin D₁ by Western blot. Result:After 24 h intervention， Scutellariae Radix-Hedyotidis Herba did not significantly inhibit the proliferation of A549 cells. However， 48 h later， the inhibitory effect in Scutellariae Radix-Hedyotidis Herba groups were significantly enhanced in comparison with that in the blank group （<italic>P</italic><0.05）， exhibiting a time-dependent response. After 72 h of action， no significant change was present in the inhibitory effect of each Scutellariae Radix-Hedyotidis Herba group， so the optimal concentration of Scutellariae Radix-Hedyotidis Herba was set at 40 mg·L^-1 for follow-up experiments. As demonstrated by the comparison with the blank group， cisplatin at each concentration inhibited the cell proliferation in a time-dependent manner （<italic>P<</italic>0.05）. Considering the cell survival rate， the best concentration of cisplatin was set at 10 mg·L^-1. Compared with the blank group， Scutellariae Radix-Hedyotidis Herba combined with cisplatin remarkably inhibited the proliferation of A549 cells in a time-dependent manner （<italic>P<</italic>0.05）， and the differences between the combined medication group and the other two groups became more significant after 72 h of medication （<italic>P<</italic>0.01）. The IL-6 level in each experimental group， especially in the combined medication group， significantly declined in contrast to that in the blank group （<italic>P<</italic>0.01）. The mRNA expression levels of HIF-1<italic>α</italic> and Cyclin D₁ in all experimental groups were obviously lower than those in the blank group， with the most significant changes observed in the combined medication group （<italic>P</italic><0.05，<italic>P</italic><0.01）. The protein expression levels of PI3K， p-Akt， mTOR， HIF-1<italic>α</italic>， and Cyclin D₁ in each experimental group was significantly down-regulated（<italic>P</italic><0.05，<italic>P</italic><0.01）， and the levels in the combined medication group were even lower than those in the cisplatin group （<italic>P<</italic>0.01）. Conclusion:Scutellariae Radix-Hedyotidis Herba has an inhibitory effect on the proliferation of A549 cells， which may be related to its inhibition against the expression and secretion of IL-6/PI3K/Akt/mTOR-HIF-1<italic>α</italic> axis.

Therapeutic Effect Of Silver Nanoparticles Against Diethyl Nitrosamin And Carbon Tettrachloride-Induced Hepatocellular Carcinoma In Rats

Objective: Hepatic cancer is known as primary liver cancer and hepatocellular carcinoma (HCC). Newly silver nanoparticles gained importance due to its advantages and multiple potential such as molecular imaging agent, antimicrobial, wound healing, anti-inflammatory and anticancer activity. The current study deals to assess therapeutic property silver nanoparticles (AgNPs) against diethylnitrosamine (DENA), and carbon tetrachloride (CCL4) induced hepatic cancer. Methods: Thirty male albino rats (200-250g) were distributed into four groups and hepatic cancer was induced with a single intraperitoneal dose of 200 mg/kg body weight of DENA. Two weeks later, animals received subcutaneous injections of CCl4 once a week in a dose of 3 ml/kg body weight for 6weeks. Serum biomarkers, antioxidants enzymes, inflammatory markers were evaluated to find the anti-proliferative potential of silver nanoparticles. Histological evaluation and microscopic reports were also done to document the results of the current work. Results: AgNPs significantly recover the serum marker enzymes of hepatic parameter AST, ALT, ALP, and total bilirubin and also decreased the levels of NO, IL-6 and TNF-α. Histopathological features also exhibited recovery of a hepatic architecture in cancer-induced rats. Moreover, the immunohistochemical investigation demonstrated that the levels of PCNA, and Caspase-3, which are hepatocarcinogenic markers, were significantly improved by AgNPs. Conclusion: These results concluded that AgNPs showed promising curing effects on hepatocellular ailments.

Assessment of Cardiovascular Disease Risk in Prediabetes

This study was aimed at investigating the nutrient and bioactive components of Annona muricataand Fagara zanthxoyloidefrom south-southern Nigeria. The roots and leaves of these plants were collected from communities within this region and an analysis of the phytochemical, mineral and vitamin components of these plant parts were carried out using standard methods. The results of the investigation revealed the a high presence of alkaloids (27.34 ± 0.15 and 12.98 ± 0.98), flavonoids (19.66 ± 0.04 and 3.71 ± 0.46) and phenols (15.10 ± 0.11 and 0.07 ± 0.42) in the leaves and roots of Annona muricatawhile alkaloids (35.55 ± 0.95 and 50.90 ± 0.83), tannins (28.70 ± 0.19 and 55.37 ± 0.47) and terpenoids (18.23 ± 0.08 and 41.21 ± 0.16) were observed in leaves and roots of Fagara zanthoxyloide. Mineral analysis revealed the presence of iron (20.23 ± 0.01 and 5.21 ± 0.02), calcium (3.67 ± 0.06 and 1.59 ± 0.01), copper (2.17 ± 0.011 and 0.16 ± 0.01) and magnesium (3.04 ± 0.01 and 2.18 ± 0.005) in leaves and roots of Annona muricataand iron, copper (2.53 ± 0.011and 7.38 ± 0.017) and zinc (5.16 ± 0.02 and 5.32 ± 0.011) in leaves and roots of Fagara zanthoxyloide.The leaves and roots of both plants also showed the presence of folate (26.82±0.48 and 23.47±0.03 for A. muricata and 15.82±0.18 and 20.63±0.91 for F. zanthoxyloide) and ascorbate (31.97±0.03and 26.89±0.19 for A. muricataand13.86±0.13 and 30.21±0.01for F. zanthoxyloide) in appreciable quantities while vitamins D, E and K were also observed in minute concentrations in both plant samples. These results may thus suggest that these plants from this region as a result of their rich nutrients and bioactive compositions may play a large role in alleviating the salient nutritional, physiological and medical challenges observed among people within this region.

Regulatory Mechanism of Youguiwan on Articular Cartilage Degeneration via IL-6/STAT3 Signaling Pathway in Rats with Knee Osteoarthritis / 中国实验方剂学杂志

<b>Objective::To observe the effect of Youguiwan on the levels of cartilage transducers and activators of transcription 3 (STAT3) and interleukin-6 (IL-6) in rats with knee osteoarthritis (KOA). <b>Method::Sixty SD rats were randomly divided into six groups: sham control group, model group, glucosamine sulfate group and Youguiwan (high, middle and low-dose) groups. The modified Hulth method was used to prepare KOA models for 6 weeks. The shame control group and the model group were treated with normal saline, Youguiwan high, middle, low-dose groups received Youguiwan 4.8, 2.4, 1.2 g·kg^-1 by gavage respectively, and the glucosamine sulfate group was treated with glucosamine sulfate 0.17 g·kg^-1. The rats were administrated for 8 weeks according to the dose. After intervention, each group was put to death by femoral artery blood collection, and the keen cartilages of the rats were collected. The pathological changes were observed by htoxylin eosin (HE) staining method, and Mankin score was evaluated. The expressions of STAT3, superoxide dismutase3(SOD3) and Wnt inhibitory factor 1(WIF1) in articular cartilage were detected by immunohistochemistry. The expressions of IL-6 mRNA in articular cartilage were detected by quantitative real-time fluorescence polymerase chain reaction (Real-time PCR). The expression of WIF1 in articular cartilage was detected by Western blot. <b>Result::Compared with the sham control group, the Makin score was obviously increased in the model group, the protein expression of STAT3 was increased significantly, the mRNA of IL-6 was raised significantly, but the protein expression of WIF1 was decreased significantly (<italic>P</italic><0.01), articular cartilage was seriously damaged, and chondrocytes were arranged in disorder. Compared with the model group, the Makin score was declined obviously in the high-dose Youguiwan group, the protein expression of STAT3 was significantly reduced, the mRNA expression of IL-6 was significantly reduced in Youguiwan treatment group, while the protein expression of WIF1 was significantly increased (<italic>P</italic><0.05, <italic>P</italic><0.01), the cartilage structure returned to be normal, the chondrocytes distribution was uneven, and articular cartilage surface was not smooth. <b>Conclusion::Youguiwan could significantly improve the articular cartilage degeneration of KOA rats, and inhibit the inflammation of chondrocytes, which may be related to the suppression of STAT3 and IL-6 expression.

Effect of acupuncture on patients with cancer-related fatigue and serum levels of CRP, IL-6, TNF-α and sTNF-R1 / 中国针灸

OBJECTIVE@#To observe the therapeutic effect of acupuncture on cancer-related fatigue (CRF) and to explore its possible mechanism.@*METHODS@#A total of 80 patients with CRF were randomized into an observation group and a control group, and finally 67 patients completed the trial (36 patients in the observation group, 31 patients in the control group). Patients in the control group were treated with conventional chemoradiotherapy and symptomatic treatment, while no particular anti-fatigue intervention was adopted. On the basis of treatment in the control group, acupuncture was applied at Baihui (GV 20), Guanyuan (CV 4), Qihai (CV 6), Fengchi (GB 20), Zusanli (ST 36), Sanyinjiao (SP 6) in the observation group, once a day, 5 times as one course, with 2 days interval between each course, totally 4 courses were required. Before and after treatment, scores of functional assessment of cancer therapy-fatigue (FACT-F) in Chinese and McGill quality of life questionnaire (MQOL), serum levels of C-reactive protein (CRP), interleukin-6 (IL-6), tumor necrosis factor-α(TNF-α) and soluble TNF receptor-1 (sTNF-R1) were observed in the two groups.@*RESULTS@#①Compared before treatment, the FACT-F score was decreased after treatment in the observation group (<0.05), while there was no significant difference in the control group (<0.05). The change of the FACT-F score in the observation group was larger than that in the control group (<0.05). ②In the observation group, scores of physiological and psychological dimension were decreased (<0.05), score of social support dimension was increased after the treatment (<0.05). The score changes of physiological, psychological and social support dimension in the observation group were larger than those in the control group (all <0.05). ③After treatment, the serum levels of IL-6, TNF-α and sTNF-R1 were decreased in the observation group (<0.05), while the serum levels of CPR and IL-6 were increased in the control group (<0.05). The serum levels of CPR, IL-6 and TNF-α in the observation were lower than those in the control group (<0.05).@*CONCLUSION@#①Acupuncture can improve the related symptoms of depression, weakness and headache in patients with CRF, strengthen their cognition of the support from society and family, and boost the confidence in curing the disease. ②Acupuncture can effectively down-regulate serum levels of the relative inflammatory factors, which may be its possible mechanism on treating CRF.

Effect of Shaoyaotang on Expressions of TLR4, NF-κB p65 and IL-6 in Rats with Damp-heat Ulcerative Colitis / 中国实验方剂学杂志

Objective:To explore the mechanism of Shaoyaotang in the treatment of ulcerative colitis (UC) based on toll-like receptor 4 (TLR4)/nuclear factor kappaB (NF-κB) signaling pathway. Method:A total of 50 Wistar rats were selected, including half male and half female. The damp-heat UC rat model was replicated by the methods of the combination of diseases and syndromes and the combination of 2, 4, 6-nitrobenzene sulfonic acid (TNBS) and ethanol. After the successful modeling, the model rats were randomly divided into model group, salazulesulfonate group, and low, medium and high-dose Shaoyaotang groups, and 10 rats (half male and half female) were selected as the blank control group. Low, medium and high-dose Shaoyaotang groups were given 6, 12, 24 g·kg-1 by gavage, and salazonyl arsenic group was given 1 g·kg-1 by gavage. Blank control group was given the equal volume of normal saline for 21 consecutive days. Colon samples were collected after the last administration, and the expressions of TLR4, NF-κB p65 and IL-6 mRNA in colon tissues were detected by fluorescent quantitative polymerase chain reaction (Real-time PCR）, and the expressions of TLR4, NF-κB p65 and IL-6 protein in colon tissues were detected by Western blot. Result:Compared with the blank control group, the relative expressions of TLR4, NF-κB p65, IL-6 mRNA and protein in the model group were significantly increased (P<0.05). Compared with the model group, the expression levels of TLR4, NF-κB p65 and IL-6 mRNA and protein in the salazopyridine group and Shaoyaotang groups were significantly decreased (P<0.05). Conclusion:Shaoyaotang can inhibit the development of UC by regulating the expressions of TLR4, NF-κB p65 and IL-6 mRNA and proteins in the TLR4/NF-κB pathway.

Effect of Modified Taohe Chengqitang Combined with Colon Hydrotherapy on Inflammatory Factors and Insulin Resistance in Patients with Severe Nonalcoholic Fatty Liver Disease / 中国实验方剂学杂志

Objective::To define the clinical efficacy of modified Taohe Chengqitang combined with colon hydrotherapy in patients with severe nonalcoholic fatty liver disease (NAFLD) accompanied by phlegm-heat stagnation syndrome and its mechanism. Method::Totally 100 patients with severe NAFLD by phlegm-heat stagnation syndrome were enrolled in the study.They were all given Shanzha Xiaozhi capsule.According to the random number table, the patients were randomly divided into the observation group (50 patients, colon hydrotherapy combined with traditional Chinese medicine) and the control group (50 patients, Shanzha Xiaozhi capsule alone). The observation period was 4 weeks.The therapeutic effect of colon hydrotherapy was verified through determinations of the liver function, blood lipid, insulin resistance index (IRI), controlled attenuation parameter (CAP), tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) before and after treatment.The mechanism of colon hydrotherapy combined with modified Taohe Chengqitang was preliminarily analyzed based on changes of IR, TNF-α and IL-6. Result::Alanine aminotransferase (ALT), aspartate aminotransferase (AST), γ-glutamyl transpeptadase (γ-GT), total cholesterol (TCH), triglyceride (TG), fasting plasma glucose (FPG), fasting insulins (FINS), IRI, CAP, TNF-αand IL-6 of NAFLD patients in both of two groups were significantly lower than those before treatment (P<0.01). ALT, AST, γ-GT, TCH, TG, FPG, IRI, CAP, TNF-α and IL-6 in observation group were significantly lower than those in the control group after treatment (P<0.01). FINS in observation group was significantly lower than that in the control group (P<0.05). Conclusion::Colon hydrotherapy combined with modified Taohe Chengqitang is an effective method for treating NAFLD accompanied by phlegm-heat stagnation syndrome.Its mechanism may be mainly correlated with the reduction of IRI, serum TNF-α and IL-6.The course of colon hydrotherapy, the therapeutic mechanism and the long-term efficacy need to be further studied in the future.

Therapeutic Effect of Anemoside B4 on Chronic Endometritis in Rats and Its Related Factors / 中国实验方剂学杂志

Objective:The effects of anemoside B4 on endometritis rats were studied through in vivo and in vitro experiments. Method:Animal experiments used 25% phenol glue to prepare endometritis models. 60 female SD rats were randomly divided into blank group， model group， Kushen gel group（0.005 g·kg-1），anemoside B4 gel low，medium and high dose groups（0.005，0.01，0.02 g·kg-1），10 rats in each group，except for the blank group，rats in each group were injected with 25% phenol glue into their vagina every 2 days，and the modeling continued for 30 days. Administration started on the day after modeling. Anemoside B4 gel low， medium and high dose groups were administered rectal daily，Kushen gel group was given daily vaginal administration. The blank group and model group were given the same amount of normal saline in the same way for 30 consecutive days. After the last administration，the uterus and its attachments of each group of rats were taken to analyze the uterine morphology and index. Hematoxylin-eosin （HE） staining was used to observe the pathological changes of rat uterus. Real-time PCR was used to detect tumor necrosis factor-α （TNF-α），interleukin-1β （IL-1β），and interleukin-6 （IL-6），signal transduction protein 130 （gp130），signal transduction and transcription activator 3 （STAT3）mRNA expression. Detection of IL-6 and STAT3 protein expression in rat uterus by Western blot. In cell experiments，lipopolysaccharide （LPS）was used to induce rat endometrial epithelial cells to prepare an in vitro inflammation model， and Real-time PCR was used to detect the expression of IL-6，gp130 and STAT3 mRNA in each group of rat endometrial epithelial cells. Result:The results of animal experiments showed that compared with the blank group， the model group had inadequate uterine cavity adhesions， endometrial edema and hyperemia. Compared with model group， there was no adhesion in the uterine cavity of the rats in the high dose anemoside B4 gel group and the Kushen gel group. The uterine tissue was relatively complete， and the uterine pathological structure was significantly improved. Compared with the blank group， the uterine index of the model group was significantly increased（P<0.05）， the expression of IL-1β mRNA in the uterine tissue was significantly increased （P<0.05）， the expression of mRNA and protein of IL-6 and STAT3 in the uterine tissue significantly increased （P<0.05）. Compared with model group， the uterine index in anemoside B4 gel high dose group was significantly reduced （P<0.05）， and the mRNA and protein expression levels of IL-6 and STAT3 in the uterine tissue were significantly reduced （P<0.05）. There was no statistically significant difference between the TNF-α and IL-1β mRNA expression compared with the model group. Cell experiment results showed that compared with the blank group， the mRNA expression of IL-6 and gp130 in model group endometrial epithelial cells was significantly increased （P<0.01）， STAT3 mRNA expression was significantly increased （P<0.05）. Compared with model group， the mRNA expression levels of IL-6， gp130 and STAT3 in anemoside B4 high dose group decreased significantly （P<0.05）. Conclusion:Anemoside B4 can improve the inflammatory response of chronic endometritis in rats and reduce the release of inflammatory factor IL-6. The mechanism may be related to the down-regulation of IL-6/STAT3 pathway.

Effect of Plantaginis Semen on Inflammatory Cytokines and mRNA and Protein Expressions of AQP4 in Colon Tissue of Diarrhea Rats / 中国实验方剂学杂志

Objective:To observe Plantaginis Semen's mechanism in treating diarrhea by observing the effect on inflammatory factors in serum and mRNA and protein expressions of aquaporin4 (AQP4) in colon tissue of diarrhea rats. Method:Senne Folium was orally administered to duplicate diarrhea rats. Sixty male SD rats were randomly divided into normal group, model group, hydrochlorothiazide group (9 mg·kg-1), and low, middle, and high-dose Plantaginis Semen groups (0.95, 1.9, 3.8 g·kg-1). Senne Folium (20 mL·kg-1) was intragastrically administered in 5 groups in the morning, except for normal group that was orally given the same dose of distilled water. In the afternoon, each treatment group was orally given the corresponding drugs, while normal group and model group were orally given the same dose of distilled water. The loose stool rate, average degree of loose stool, and diarrhea index were compared according to fecal traits and stool times after 14 days of treatment. The serum and colon tissue were collected to detect the contents of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and C-reactive protein (CRP) in serum. Hematoxylin-eosin (HE) staining was used to observe the pathological morphological changes of colon tissue, and quantiative Real-time fluorescent quantitative polymerase chain reaction (Real-time PCR) and Western blot were used to detect the mRNA and protein expressions of AQP4 in colon tissue. Result:In the model group, the loose stool rate, average degree of loose stool, and diarrhea index were significantly increased (P<0.01), apoptosis and necrosis were observed in the epidermal cells of colonic mucosa, telangiectasia and congestion in lamina propria were obvious, and a few neutrophils were infiltrated, and the contents of TNF-α, IL-6 and CRP in serum increased (P<0.05, P<0.01), the mRNA and protein expressions of AQP4 significantly decreased (P<0.01). Compared with the model group, the loose stool rate, average degree of loose stool, and diarrhea index were significantly decreased in low, middle, and high-dose Plantaginis Semen groups (P<0.01), the apoptosis and necrosis of epidermal cells, telangiectasia and hyperemia and neutrophil infiltration in colonic mucosa were obviously improved, and the contents of TNF-α and CRP in serum significantly decreased (P<0.05, P<0.01), the mRNA and protein expressions of AQP4 increased （P<0.05, P<0.01）. Conclusion:Plantaginis Semen has a better antidiarrheal effect, and its mechanism may be related to inhibition of inflammatory reaction, repair of pathological damage of colonic mucosa, up-regulation of AQP4 expression and promotion of water and fluid metabolism.

Changes in peripheral blood inflammatory factors (TNF-a and IL-6) and intestinal flora in AIDS and HIV-positive individuals / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Objective: In this study, we investigated the changes in peripheral blood inflammatory factors and intestinal flora in acquired immune deficiency syndrome (AIDS) and human immunodeficiency virus (HIV)-positive individuals (AIDS/HIV patients), and explored the relationships among intestinal flora, peripheral blood inflammatory factors, and CD4+ T lymphocytes. Methods: Thirty blood and stool samples from an AIDS group and a control group were collected. The levels of tumor necrosis factor-a (TNF-a) and interleukin-6 (IL-6) were determined by enzyme-linked immunosorbent assay (ELISA), and the number of CD4+ T lymphocytes by a FACSCount automated instrument. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to determine the messenger RNA (mRNA) levels of Bifidobacterium, Lactobacillus, Escherichia coli, Enterococcus faecalis, and Enterococcus faecium. Correlations among intestinal flora, inflammatory factor levels, and CD4+ T lymphocyte values were evaluated using the Spearman correlation coefficient. Results: The levels of TNF-a and IL-6 in the AIDS group were higher than those in the control group, while the number of CD4+ T lymphocytes was lower. The amounts of Bifidobacterium and Lactobacillus in the AIDS group were significantly lower than those in control group, while the amounts of E. coli, E. faecalis, and E. faecium were much higher. The amounts of Bifidobacterium and Lactobacillus were negatively correlated with the content of TNF-a and IL-6 and the CD4+ T lymphocyte count, while those correlations were reversed for E. coli, E. faecalis, and E. faecium. Conclusions: The intestinal microbiota of AIDS/HIV patients were disordered, and there was a correlation between the amount of intestinal flora and the number of CD4+ T lymphocytes and the levels of TNF-a and IL-6.

Clinical Efficacy of Modified Xiaofengsan and Its Effect on Cytokines in Persistent Asthma Patients with Wind-asthma Pattern / 中国实验方剂学杂志

Objective: To observe the clinical efficacy of modified Xiaofengsan and its effect on cytokines in persistent asthma patients with wind-asthma pattern. Method: The 120 eligible patients with chronic and persistent asthma were randomly divided into control group (60 cases) and treatment group (60 cases). Patients in control group received the budesonide/formoterol (160 μg/4.5 μg, 2 inhales/time, bid). Patients in treatment group additionally received the modified Xiaofengsan combined with budesonide/formoterol (160 μg/4.5 μg). The treatment course was 1 month in both groups. The clinical efficacy of the two groups was observed. Before and 3 months after treatment, asthma control test (ACT) and the average times of acute exacerbation in 6 months before and after treatment between two groups were observed; percentage of forced expiratory volume in first second and its predicted value (FEV1%), variety ratio of Peak expiratory flow (PEF), eosinophilic granulocyte (EOS) count in peripheral blood and the levels of interleukin-4(IL-4), interleukin-6(IL-6) and interleukin-13(IL-13) in serum were measured before and after treatment respectively. In addition, the safety of the two groups was evaluated. Result: The total effective rate of the treatment group was higher than that of the control group (P1% in treatment group were higher(PPConclusion: Modified Xiaofengsan combined with budesonide/formoterol could relieve symptoms of asthma, improve pulmonary function and lower the levels of IL-4, IL-6 and IL-13 in serum.