Inhibition of glutaminolysis alleviates myocardial fibrosis induced by angiotensin II / 生理学报

The present study was aimed to investigate the role and mechanism of glutaminolysis of cardiac fibroblasts (CFs) in hypertension-induced myocardial fibrosis. C57BL/6J mice were administered with a chronic infusion of angiotensin II (Ang II, 1.6 mg/kg per d) with a micro-osmotic pump to induce myocardial fibrosis. Masson staining was used to evaluate myocardial fibrosis. The mice were intraperitoneally injected with BPTES (12.5 mg/kg), a glutaminase 1 (GLS1)-specific inhibitor, to inhibit glutaminolysis simultaneously. Immunohistochemistry and Western blot were used to detect protein expression levels of GLS1, Collagen I and Collagen III in cardiac tissue. Neonatal Sprague-Dawley (SD) rat CFs were treated with 4 mmol/L glutamine (Gln) or BPTES (5 μmol/L) with or without Ang II (0.4 μmol/L) stimulation. The CFs were also treated with 2 mmol/L α-ketoglutarate (α-KG) under the stimulation of Ang II and BPTES. Wound healing test and CCK-8 were used to detect CFs migration and proliferation respectively. RT-qPCR and Western blot were used to detect mRNA and protein expression levels of GLS1, Collagen I and Collagen III. The results showed that blood pressure, heart weight and myocardial fibrosis were increased in Ang II-treated mice, and GLS1 expression in cardiac tissue was also significantly up-regulated. Gln significantly promoted the proliferation, migration, mRNA and protein expression of GLS1, Collagen I and Collagen III in the CFs with or without Ang II stimulation, whereas BPTES significantly decreased the above indices in the CFs. α-KG supplementation reversed the inhibitory effect of BPTES on the CFs under Ang II stimulation. Furthermore, in vivo intraperitoneal injection of BPTES alleviated cardiac fibrosis of Ang II-treated mice. In conclusion, glutaminolysis plays an important role in the process of cardiac fibrosis induced by Ang II. Targeted inhibition of glutaminolysis may be a new strategy for the treatment of myocardial fibrosis.

New eudesmane sesquiterpenoids from Atractylodis Macrocephalae Rhizoma and their inhibitory activities against SREBPs / 中国中药杂志

Three sesquiterpenoids were isolated and purified from the 95% ethanol extract of Atractylodis Macrocephalae Rhizoma by column chromatography on silica gel, Sephadex LH-20, ODS, and high-performance liquid chromatography(HPLC). Their chemical structures were identified on the basis of spectroscopic analysis and physiochemical properties as(7Z)-8β,13-diacetoxy-eudesma-4(15),7(11)-diene(1), 7-oxo-7,8-secoeudesma-4(15),11-dien-8-oic acid(2), and guai-10(14)-en-11-ol(3). Compounds 1 and 2 are new compounds and compound 3 was obtained from Compositae family for the first time. Compounds 1, 2, and 3 showed weak inhibitory activities against sterol regulatory element-binding proteins(SREBPs).

Effects of compound deer bone extract on osteoporosis model mice and intestinal microflora.

The preventive and therapeutic mechanisms of CDBE on osteoporosis were studied by observing the serum bone-related biochemical indicators, bone trabecular micro-structure and intestinal flora in ovariectomized osteoporosis model mice, in order to provide a scientific theoretical basis for the further study on the effect of CDBE on osteoporosis, and the prevention and treatment of osteoporosis with clinical traditional Chinese medicines. The components in CDBE were detected by UHPLC-MS. A mouse osteoporosis model was established by the bilateral ovariectomy in female ICR mice. The biochemical indicators related to osteoporosis were detected, the right proximal tibia was scanned by Micro-CT, the intestinal microflora in the colon contents were examined, and the changes of microflora were taken as the main target to evaluate the effect of CDBE on the intestinal microflora in the model mice. A total of 16 compounds were obtained by the combined application of UHPLC-MS. CDBE could significantly increase the contents of E2, Ca2+ , CT, HyP, OCN, FOXP3, P1NP and CTX-II, in the model mice. CDBE could significantly improve the trabecular micro-structure, Tb.N, Tb.Sp, SMI and Conn.D. CDBE could make the intestinal flora of osteoporosis model mice tend to healthy mice in species and quantity. CDBE can improve the symptoms of postmenopausal osteoporosis in mice, with a positive effect on the intestinal flora of postmenopausal mice. Its mechanism of regulating the symptoms of osteoporosis may be related to the regulation of bone-related biochemical indicators in the serum of mice. PRACTICAL APPLICATIONS: This research has a positive impact on the development of functional food with anti-osteoporosis in the future.

Combination of flavonoids in Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle and Angelicae Sinensis Radix in treatment of scalds: a preliminary study of dosage form / 中国中药杂志

The anti-inflammatory activity and compatibility ratio of flavonoids in Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle(GR) and Angelicae Sinensis Radix(AS) were evaluated by the superoxide anion scavenging test. The matrix formula of gel was optimized by orthogonal test design and the model of deep partial-thickness scald in mice was induced. The gel was applied to the wound. The tissue water content, wound healing rate, serum TNF-α and IL-1, and EGF and VEGF in tissues were measured at diffe-rent periods. The results revealed that when the compatibility ratio of GR and AS was 1∶2, the maximal scavenging efficacy on supe-roxide anion was observed. The gel displayed the optimal properties when carbomer(1%), glycerol(5%), propylene glycol(10%) were added into the matrix. Gel external application can significantly improve the wound healing rate, relieve tissue edema, diminish tissue water content, alleviate inflammatory reaction, and increase the content of EGF and VEGF in tissues(P<0.05). The gel prepared in the present study is effective in promoting granulation, relieving pain, resisting inflammation, and alleviating edema, and is potent in healing scalds.

Molecular Mechanism Analysis of Jiangtang Xiaozhi Tablets in Treatment of NAFLD / 中国实验方剂学杂志

Objective:To explore the molecular mechanism of Jiangtang Xiaozhi tablets （JTXZT） in the treatment of non-alcoholic fatty liver disease （NAFLD） by means of network pharmacology and molecular docking. Method:With the help of traditional Chinese medicine （TCM） Systems Pharmacology Database and Analysis Platform （TCMSP）， TCMs Integrated Database （TCMID）， Encyclopedia of TCM （ETCM） and Bioinformatics Analysis Tool for Molecular Mechanism of TCM （BATMAN-TCM）， the chemical compositions of medicinal materials in JTXZT were obtained， the compound targets were predicted in SwissTargetPrediction database and STITCH database. The targets of NAFLD were searched by The Human Gene Database （GeneCards）， Online Mendelian Inheritance in Man （OMIM）， Therapeutic Target Database （TTD） and DisGeNET， and intersection analysis was performed with the targets of the active ingredients to obtain the targets of JTXZT for treatment of NAFLD. Based on STRING 11.0 database， the protein-protein interaction （PPI） network of therapeutic targets was constructed， and the enrichment analysis of therapeutic targets was carried out by DAVID 6.8. Finally， the interaction characteristics of key components and core therapeutic targets of JTXZT for treatment of NAFLD were verified based on molecular docking. Result:The key components of JTXZT for treatment of NAFLD were quercetin， luteolin， kaempferol， berberine， isorhamnetin， betulinic acid， oleanolic acid， ursolic acid. formononetin and hexitol， and the core targets of JTXZT for treatment of NAFLD were mitogen-activated protein kinase 1 （MAPK1）， Jun proto-oncogene， activator protein-1 （AP-1） transcription factor subunit （JUN）， MAPK3， protein kinase B1 （AKT1 or Akt1）， tumor protein p53 （TP53）， E1A binding protein p300 （EP300）， Fos proto-oncogene， AP-1 transcription factor subunit （FOS）， tumor necrosis factor （TNF），amyloid beta precursor protein （APP） and cytochrome P450 family 2 subfamily E member 1 （CYP2E1）. Biological function and pathway enrichment analysis showed that JTXZT mainly through xenobiotic metabolic process， oxidation-reduction process， cholesterol metabolic process and other biological processes， regulating phosphatidylinositol 3-kinase/protein kinase B （PI3K/Akt） signaling pathway， MAPK signaling pathway， NAFLD and insulin signaling pathway to play a role in the treatment of NAFLD. The results of molecular docking showed that the active components of JTXZT had a good affinity with the core targets of JTXZT for the treatment of NAFLD. Conclusion:JTXZT treats NAFLD through multiple active components， multiple key targets and multiple action pathways.

Network Pharmacology Approach to Explore Mechanisms of Qizhu Granules for Treatment of Diabetic Nephropathy / 中国实验方剂学杂志

Objective:To explore the mechanism of Qizhu granules in the treatment of diabetic nephropathy by using network pharmacology. Method:The Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform database （TCMSP） and The Encyclopedia of Traditional Chinese Medicine（ETCM） database were used to screen out the chemical constituents and protein targets of each drug in the Qizhu granules based on oral bioavailability and drug-like properties. The protein target was standardized into the corresponding gene name through the UniProt database. Online Mendelian Inheritance in Man（OMIM）， DisGeNET， Therapeutic Target Database （TTD）， ETCM database were used to search for related targets of diabetic nephropathy， after the intersection of the two， construct a protein interaction network through protein interaction database （STRING）， use Cytoscape to analyze the core target of the network， and the relevant targets were analyzed by KOBAS 3.0 database for Gene Ontology （GO） pathway enrichment analysis and Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment analysis. Result:A total of 93 chemical components were obtained from Qizhu granules， involving 254 targets， and 607 targets related to diabetic nephropathy. After the intersection， 76 sputum granules were determined to treat diabetic nephropathy， including protein kinase B1 （Akt1）， vascular endothelial growth factor （VEGFA）， interleukin （IL）-6， tumor necrosis factor （TNF）， mitogen-activated protein kinase 1 （MAPK1）， matrix metalloproteinase （MMP）-9 and other core targets， after GO analysis and KEGG analysis， Qizhu granules can affect cellular response to nitrogen compound， regulation of reactive oxygen species metabolic process and other biological processes， regulate advanced glycation end product （AGE）/advanced glycation end product receptor （RAGE） signaling pathway in diabetic complications， fluid shear stress and atherosclerosis， IL-17 signaling pathways， HIF-1 signaling pathways TNF signaling pathways and other pathways. Conclusion:The therapeutic effect of Qizhu granules on diabetic nephropathy may affect Akt1，VEGFA， IL-6， TNF， MAPK1， MMP-9 and other targets， and regulate AGE/RAGE signaling pathway in diabetic complications， fluid shear stress and atherosclerosis， IL-17 signaling pathways， hypoxia-inducing factor-1（HIF-1）signaling pathways TNF signaling pathways and other pathways， which can provide a theoretical reference for further basic experimental research.

Intervention of phlegm and blood stasis inhibits TGF-β1/Smad3 signaling pathway in the kidney of diabetic rats / 南方医科大学学报

OBJECTIVE@#To observe the effect of traditional Chinese medicine for intervention of phlegm and blood stasis in regulating TGF-β1/Smad3 signaling and relieving nephropathy in diabetic rats.@*METHODS@#SD rats were divided into blank group (NC), diabetic model group (MC group), intervention of phlegm and blood stasis (RPDBS) group, phlegm-removing (RP) group and blood-removing (DBS) group. Diabetic models were established in all the rats except for those in the blank group. After 4 weeks of feeding, the rats in RPDBS group, RP group and DBS group were given corresponding drug intervention for 8 weeks. HE staining was used to observe the changes in renal histopathology. Western blotting and real-time fluorescence quantitative PCR were used to detect the expression levels of transforming growth factor-β1 (TGF-β1) and Smad3.@*RESULTS@#The structure and arrangement of the glomeruli and renal tubules improved significantly in the treatment groups in comparison with those in the MC group. The expression levels of TGF-β1, Smad3 and p-Smad3 were significantly downregulated at both the protein and mRNA levels in the treatment groups ( < 0.05), and the down-regulation was more obvious in RPDBS group than in RP group and DBS group ( < 0.05).@*CONCLUSIONS@#Intervention of phlegm and blood stasis may inhibit the activation of TGF-β1/Smad3 signaling pathway and delay diabetic nephropathy and fibrosis to protect the renal function in diabetic rats.

Effect of Selenium Deficiency on Phosphorylation of the AMPK Pathway in Rats.

Selenium is an important trace element for human health. Previous studies have raised concern that dietary selenium intake may change energy metabolism. AMP-activated protein kinase (AMPK) is a sensor of energy status that controls cellular energy homeostasis. We aimed to determine the effect of selenium on the phosphorylation of AMPK pathway between Se-deficient and normal Sprague-Dawley rats. Twenty-four weaning rats were fed either a Se-deficient diet (0.02 mg Se/kg) or a standard diet (0.18 mg Se/kg). After 109 days, total serum levels of non-esterified fatty acid and total amino acids were significantly higher and the serum insulin concentration was significantly lower in Se-deficient rats than in healthy controls. Selenium concentration and the activity of glutathione peroxidase (GPx) in myocardial tissue were significantly lower in Se-deficient rats. Importantly, mRNA levels of acetyl-CoA carboxylase beta (ACACB), peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), and protein levels of p-AMPKα were increased in the Se-deficient group compared to normal controls (p < 0.05). In conclusion, our results suggest that selenium deficiency induces changes in metabolic and molecular parameters involved in energy metabolism in the AMPK pathway.

Research on Acute Toxicity and Pharmacodynamics of Jinyulian Oral Solution.

A research on Jinyulian Oral Solution was conducted and the objectives were to discover its possible acute toxicity and antibacterial effects when used in vitro and in vivo. Regarding the acute toxicity test, Kunming mice were fed a maximum amount of the solution as their stomachs could hold, i.e., 40 mL kg(-1). To ascertain the minimum inhibitory concentration (MIC) of the solution, two types of germs, i.e., Staphylococcus aureus and Escherichia coli, were selected and tube dilution method was adopted. An antibacterial experimental model relying on animals' body was developed for the researchers to observe the solution's antibacterial effects. Test results showed that no abnormalities were discovered within 14 days after the initial date of testing and the mice grew as normal when fed with an amount of the solution 250 times of a normal clinical doze (In this case a man was assumed to weigh 60 kg.) and that the solution demonstrated obvious antibacterial effects on the two types of selected germs. The respective measured MIC50 and MIC90 values of the two germs were 3.2, 12.8, 6.4, and 25.6 mg L(-1). Therefore, it is reasonable to conclude that Jinyulian Oral Solution possesses no acute toxicity but obvious antibacterial effects on the two before-mentioned germs.

HPLC simultaneous determination of contents of 5 saponin constituents in Ophiopogonis Radix / 中国中药杂志

This research is to establish an HPLC method for the simultaneous determination of ophiopogonin D, ophiopogonin D', ophiopogonin C, deacetylophiopojaponin A and ophiogenin-3-O-α-L-rhamnosyl-(1-->2)-β-D-glucoside in Ophiopogonis Radix. HPLC-ELSD analysis was performed on a Kromasil 100-5 C₁₈ column (4.6 mm x 250 mm, 5 µm), with the mobile phase of acetonitrile (A) -water (B) in gradient elution mode (0-45 min, 35%-55% A), at a flow rate of 1 mL · min⁻¹. The column temperature was 35 °C and the drift tube temperature was 100 °C in a gas flow rate of 3.0 L · min⁻¹. The result showed that baseline of all the 5 constituents was well separated, and every constituent had wide linearity range and good linear relation (r > 0.999). The recovery rate was between 95.75% and 103.1%. The new established method for simultaneous determination of saponin constituents in Ophiopogonis Radix was sensitive and has good, repeatability. It could be applied to quality evaluation of Ophiopogonis Radix.

Intervention of berberine on lipid deposition in liver cells of non-alcoholic fatty liver disease rats induced by high fat diet / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To explore the effect of berberine on lipid metabolism disorder and lipid deposition in liver cells of non-alcoholic fatty liver disease (NAFLD) rats induced by high fat diet.</p><p><b>METHODS</b>After one week adaptable feeding, 45 SPF level male SD rats were randomly divided into 3 groups, the normal control group, the model group, and the berberine group, 15 in each group. Except those in the normal control group, all rats were fed with high fat diet to prepare NAFLD model. As for rats in the berberine group, Berberine Hydrochloride was administered by gastrogavage. HE staining and oil red O staining were performed to identify the model after 8 weeks. Hepatocytes were isolated, and their activities and purities were tested by Typan blue staining and flow cytometry (FCM). Serum levels of TC, TG, HDL-C, and LDL-C were detected using automatic biochemical analyzer. mRNA expression levels of LXRα and FAS in liver cells were analyzed by Real-time quantitative polymerase chain reaction (PCR). Protein levels of LXRα and FAS in liver cells were examined by Western blot.</p><p><b>RESULTS</b>The NAFLD rat model was successfully established by high fat diet. The yields of purified liver cells in each rat were (6.0-7.5) x 10(8). The viability of isolated liver cells with purity over 90% (tested by FCM analysis) was higher than 95%. Compared with the normal control group,the expression of LXRα and FAS at mRNA and protein levels was higher in the model group (P < 0.01). Compared with the model group, the expression of LXRα and FAS at mRNA and protein levels was obviously down-regulated in the berberine group (P < 0.01).</p><p><b>CONCLUSIONS</b>LXRα/FAS signaling pathway was one of important signaling pathways of NAFLD lipid metabolism disorders. Berberine could recover hepatocyte fatty deposits in NAFLD rats by adjusting the LXR/FAS signaling pathway of hepatocytes, which might be one of important mechanisms for fighting against NAFLD.</p>