UPLC-Q-TOF-MS metabolomic study on improvement of acute myocardial ischemia in rats by Dalbergia cochinchinensis heartwood / 中国中药杂志

This paper aimed to study the effect of Dalbergia cochinchinensis heartwood on plasma endogenous metabolites in rats with ligation of the left anterior descending coronary artery, and to analyze the mechanism of D. cochinchinensis heartwood in improving acute myocardial ischemic injury. The stability and consistency of the components in the D. cochinchinensis heartwood were verified by the establishment of fingerprint, and 30 male SD rats were randomly divided into a sham group, a model group, and a D. cochinchinensis heartwood(6 g·kg~(-1)) group, with 10 rats in each group. The sham group only opened the chest without ligation, while the other groups established the model of ligation. Ten days after administration, the hearts were taken for hematoxylin-eosin(HE) staining, and the content of heart injury indexes in the plasma creatine kinase isoenzyme(CK-MB) and lactate dehydrogenase(LDH), energy metabolism-related index glucose(Glu) content, and vascular endothelial function index nitric oxide(NO) was determined. The endogenous metabolites were detected by ultra-high-performance liquid chromatography-time-of-flight-mass spectrometry(UPLC-Q-TOF-MS). The results showed that the D. cochinchinensis heartwood reduced the content of CK-MB and LDH in the plasma of rats to relieve myocardial injury, reduced the content of Glu in the plasma, improved myocardial energy metabolism, increased the content of NO, cured the vascular endothelial injury, and promoted vasodilation. D. cochinchinensis heartwood improved the increase of intercellular space, myocardial inflammatory cell infiltration, and myofilament rupture caused by ligation of the left anterior descending coronary artery. The metabolomic study showed that the content of 26 metabolites in the plasma of rats in the model group increased significantly, while the content of 27 metabolites decreased significantly. Twenty metabolites were significantly adjusted after the administration of D. cochinchinensis heartwood. D. cochinchinensis heartwood can significantly adjust the metabolic abnormality in rats with ligation of the left anterior descending coronary artery, and its mechanism may be related to the regulation of cardiac energy metabolism, NO production, and inflammation. The results provide a corresponding basis for further explaining the effect of D. cochinchinensis on the acute myocardial injury.

Bletilla striata polysaccharide improves toxic and side effects induced by 5-FU: an untargeted metabolomics study / 中国中药杂志

This study aimed to analyze the effect of Bletilla striata polysaccharide(BSP) on endogenous metabolites in serum of tumor-bearing mice treated with 5-fluorouracil(5-FU) by untargeted metabolomics techniques and explore the mechanism of BSP in alleviating the toxic and side effects induced by 5-FU. Male BALB/C mice were randomly divided into a normal group, a model group, a 5-FU group, and a 5-FU + BSP group, with eight mice in each group. Mouse colon cancer cells(CT26) were transplanted into the mice except for those in the normal group to construct the tumor-bearing mouse model by subcutaneous injection, and 5-FU chemotherapy and BSP treatment were carried out from the second day of modeling. The changes in body weight, diarrhea, and white blood cell count in the peripheral blood were recorded. The mice were sacrificed and sampled when the tumor weight of mice in the model group reached approximately 1 g. TUNEL staining was used to detect the cell apoptosis in the small intestine of each group. The proportions of hematopoietic stem cells and myeloid progenitor cells in bone marrow were measured by flow cytometry. Five serum samples were selected randomly from each group for untargeted metabolomics analysis. The results showed that BSP was not effective in inhibiting colon cancer in mice, but diarrhea, leukopenia, and weight loss caused by 5-FU chemotherapy were significantly improved after BSP intervention. In addition, apoptotic cells decreased in the small intestinal tissues and the percentages of hematopoietic stem cells and myeloid progenitor cells in bone marrow were significantly higher after BSP treatment. Metabolomics results showed that the toxic and side effects of 5-FU resulted in significant decrease in 29 metabolites and significant increase in 22 metabolites in mouse serum. Among them, 19 disordered metabolites showed a return to normal levels in the 5-FU+BSP group. The results of pathway enrichment indicated that metabolic pathways mainly involved pyrimidine metabolism, arachidonic acid metabolism, and steroid hormone biosynthesis. Therefore, BSP may ameliorate the toxic and side effects of 5-FU in the intestinal tract and bone marrow presumably by regulating nucleotide synthesis, inflammatory damage, and hormone production.

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 Saponins in Pulsatillae Radix on Proliferation and Apoptosis of NCI-H460 Lung Cancer Cells and Screening of Its Differentially Expressed Proteins / 中国实验方剂学杂志

Objective: To screen the differentially expressed proteins of saponins in Pulsatillae Radix inhibiting the proliferation and induce apoptosis on NCI-H460 tumor cells based on proteome technology using nano LC-LTQ-Orbitrap-MS/MS, and preliminarily speculate the potential mechanism. Method: NCI-H460, SK-OV-3 and SGC-7901 tumor cells were cultured in vitro. Methylthiazoletetrazolium (MTT) assay was used to detect the inhibitory rate of saponins in Pulsatillae Radix on three tumor cell lines. Effect of saponins in Pulsatillae Radix on apoptosis was analyzed by Annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) staining flow cytometry and 4',6-diamidino-2-phenylindole (DAPI) staining. Apoptosis was analyzed using flow cytometry and DAPI stain. Nano LC-LTQ-Orbitrap-MS/MS was used to investigate the changes in the protein profiles on NCI-H460 cells treated with saponins in Pulsatillae Radix. Proteins exhibiting differential expression were analyzed by DAVID Bioinformatics Resources 6.8 and Kyoto encyclopedia of genes and genomes (KEGG) database. The differentially expressed proteins were verified by Western blot. Result: Saponins in Pulsatillae Radix could inhibit the proliferation of NCI-H460, SK-OV-3 and SGC-7901 tumor cells and induce apoptosis of NCI-H460 tumor cells. Effect of Saponins in Pulsatillae Radix on the proliferation and apoptosis of NCI-H460 tumor cells was mainly related to the regulation of biological function of ribosome, glycolysis/gluconeogenesis and other biological processes. It was possible to induce apoptosis of NCI-H460 tumor cells by interfering mitogen-activated protein kinase (MAPK) signaling pathway and regulating the Caspase pathway. Conclusion: Saponins in Pulsatillae Radix can inhibit the proliferation and induce the apoptosis of NCI-H460 tumor cells, the mechanism may be related to the intervention of MAPK signaling pathway and the regulation of Caspase pathway. These findings are helpful to elucidate the molecular mechanism of the anti-tumor effect of saponins in Pulsatillae Radix.

Effect of neoflavonoid latifolin isolated from Dalbergia odorifera on acute myocardial ischemia in rats and its mechanism of Nrf2 signaling pathway / 中国中药杂志

The present study was designed to evaluate the cardioprotective effect of latifolin on pituitrin(Pit) or isoproterenol(ISO)-induced myocardial injury in rats, and further investigate its underlying mechanisms. Rats were administrated sublingually with pituitrin or subcutaneously with isoproterenol to induce acute myocardial ischemia in rats, and lead II electrocardiograph was recorded. In rats with isoproterenol, ELISA assay or colorimetric method was used to detect the content or activity of myocardial injury markers in serum, and the SOD activity and MDA content in myocardium were detected by colorimetric assay; histopathological examination was conducted by HE staining; the frozen section of myocardial tissues was used for DCFH-DA fluorescent staining to detect the content of ROS in myocardium; Western blot was used to detect the protein expression levels of Nrf2, Keap1, HO-1 and NQO1 in myocardium. Results showed that latifolin significantly inhibited ST-segment changes induced by pituitrin or isoproterenol, and increased heart rate. Further mechanism study showed that latifolin reduced cardiac troponin I(cTnI) level, aspartate transaminase(AST) and lactate dehydrogenase(LDH) activities in serum, increased myocardial superoxide dismutase(SOD) activity and reduced myocardial malondialdehyde(MDA) level, and protected myocardium with less necrosis, infiltration of inflammatory cells and fracture of myocardial fibers. Furthermore, latifolin obviously reduced ROS level in myocardium, inhibited the expression of Kelch-like ECH-associated protein-1(Keap1), increased the nuclear translocation of nuclear factor erythroid 2 related factor 2(Nrf2), and promoted the expression of Heme oxygenase-1(HO-1) and NAD(P)H quinone oxidoreductase-1 (NQO1) in myocardial tissues. Our data suggest that latifolin has a potent protective effect against pituitrin or isoproterenol-induced myocardial injury, which may be related to inhibition of oxidative stress by activating Nrf2 signaling pathway.