Patrinia herba aqueous extract on the proliferation, apoptosis, invasion and migration of hepatocellular carcinoma cells.

This study aims to study the effect and mechanism of Patrinia herba aqueous extract on proliferation, apoptosis, invasion and migration of hepatocellular carcinoma cells. Hepatocellular carcinoma cells MHCC97-H were treated with 2.5, 5, 10 mg/mL P. herba aqueous extract, cell counting kit 8 (CCK-8), flow cytometry, plate cloning experiments, and Transwell measured cell survival, apoptosis, colony formation, invasion, and migration, respectively. Real-time quantitative PCR (qPCR) and western blot were used to detect long non-coding RNA (lncRNA) HTR2A-AS1 and expression of proteins P21, Caspase-3, E-cadherin and matrix metalloproteinase-2 (MMP-2), respectively. Transfected pcDNA3.1-HTR2A-AS1 in MHCC97-H cells, or transfected si-HTR2A-AS1 and treat with 10 mg/mL P. herba aqueous extract to evaluate their roles in cell proliferation, apoptosis, invasion, and migration. Different concentrations of P. herba aqueous extract significantly reduced the survival rate, colony formation, number of migrating cells, number of invading cells, and MMP-2 protein expression of MHCC97-H cells, and obviously increased the cell apoptosis rate, the expression levels of Caspase-3, E-cadherin protein and HTR2A-AS1 (P<0.05), which were all concentration-dependent. Overexpression of HTR2A-AS1 evidently decreased the survival rate, colony formation, number of migrating cells, number of invading cells, and MMP-2 protein levels in MHCC97-H cells, while remarkably enhanced the apoptosis rate of cells, P21, Caspase-3, and E-cadherin protein levels and HTR2A-AS1 expression level (P<0.05). Inhibition of HTR2A-AS1 greatly improved the survival rate, the number of clone formation, the number of migrating cells, the number of invading cells and the expression of MMP-2 protein of MHCC97-H cells treated with P. herba aqueous extract, dramatically reducing the cell apoptosis rate, P21, Caspase-3, E-cadherin protein levels and HTR2A-AS1 expression levels (P<0.05). P. herba aqueous extract may inhibit the proliferation, invasion and migration of hepatocellular carcinoma cells by up-regulating the expression of HTR2A-AS1 in hepatocellular carcinoma cells and induce apoptosis.

[Effect of neoflavonoid latifolin isolated from Dalbergia odorifera on acute myocardial ischemia in rats and its mechanism of Nrf2 signaling pathway].

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.

Anticancer Effects of Baicalein in Pancreatic Neuroendocrine Tumors In Vitro and In Vivo.

OBJECTIVES: Baicalein is a Chinese traditional medicine that inhibits tumor migration and growth. Pancreatic neuroendocrine tumors (pNETs) have a high incidence in China, but there are still no effective treatments. The aim of our study was to investigate whether baicalein could inhibit pNETs. METHODS: In vitro, we used BON1-a cell line of pNETs-to analyze the apoptosis and migration and invasion after baicalein treatment via flow cytometry and Western blot. In vivo, we used a xenograft tumors model to evaluate the size of tumors after baicalein treatment. Western blot was used to analyze the expression of apoptosis and migration-related protein. RESULTS: In vitro, the Cell Counting Kit 8 assay showed that baicalein decreased BON1 viability, and flow cytometry demonstrated that baicalein induced BON1 apoptosis and protein changes. In addition, baicalein inhibited BON1 migration and invasion as shown via a Transwell assay. In vivo, baicalein inhibited tumor growth and migration and also increased apoptosis-related protein expression. CONCLUSIONS: Baicalein could increase caspase-3 and Bax expression and decrease survivin and Bcl-2 to induce apoptosis. It inhibits migration and invasion by decreasing expression of vascular endothelial growth factor and matrix metalloproteinases 2 and 9.

An integrated strategy using UPLC-QTOF-MSE and UPLC-QTOF-MRM (enhanced target) for pharmacokinetics study of wine processed Schisandra Chinensis fructus in rats.

Currently the pharmacokinetic (PK) research of herbal medicines is still limited and facing critical technical challenges on quantitative analysis of multi-components from biological matrices which often accompanied by lacking of authentic standards and low concentration. This present work contributes to the development of an integrated strategy for extensive pharmacokinetics assessments, and a selective and sensitive method independent of authentic standards for multi-components analysis based on the use of ultra-performance liquid chromatography/quadrupole-time-of-flight/MSE (UPLC-TOF-MSE) and UPLC-TOF-MRM (rnhanced target). Initially, phytochemicals were identified by UPLC-TOF-MSE analysis, subsequently the identified components were matched with authentic standards and pre-classified, and UPLC-QTOF-MRM method optimized and developed. To guarantee reliable results, three rules are necessary: (1) detection with a mass error of less than 5ppm; (2) same class chemical compositions with structural high similarity between analytes with and without authentic reference substance; (3) a matching retention time between TOF-MRM mode and TOF-MSE within 0.2min. The developed and validated method was applied for the simultaneous determination of 12 lignans in rat plasma after administered with wine processed Schisandra Chinensis fructus (WPSCF) extract. Such an approach was found capable of providing extensive pharmacokinetic profiles of multi-components absorbed into blood after oral administrated with WPSCF extract. The results also indicated that significant difference in pharmacokinetics parameters of dibenzocyclooctadiene lignans was observed between schizandrin and gomisin compounds. For lignans, the absorption via gastrointestinal tract were all rapid and maintained relatively long retention time, especially for schisantherin A and schisantherin B with higher plasma exposure.

An alternative to current therapies of functional dyspepsia: self-administrated transcutaneous electroacupuncture improves dyspeptic symptoms.

Functional dyspepsia is of high prevalence with little treatment options. The aim of this study was to develop a new treatment method using self-management transcutaneous electroacupuncture (TEA) for functional dyspepsia (FD). Twenty-eight patients with FD were enrolled and underwent a crossover clinical trial with 2-week TEA at ST36 and PC6 and 2-week sham-TEA at nonacupuncture sham-points. Questionnaires were used to assess symptoms of dyspepsia and quality of life. Physiological testing included gastric emptying and electrogastrography. It was found that (1) TEA but not sham-TEA significantly improved dyspeptic symptoms and 4 domains in quality of life; improvement was also noted in self-rated anxiety and depression scores; (2) gastric emptying was significantly and substantially increased with 2-week TEA but not sham-TEA; and (3) gastric accommodation was also improved with TEA but not sham-TEA, reflected as increased ingested nutrient volumes at the levels of satiety and maximum tolerance. These findings suggest a therapeutic potential of self-administrated TEA method for functional dyspepsia, possibly attributed to improvement in gastric motility.

Astragalus mongholicus regulate the Toll-like-receptor 4 meditated signal transduction of dendritic cells to restrain stomach cancer cells.

BACKGROUND: According to the traditional view, we depend on three methods to treat tumors; surgery, chemotherapy and radiotherapy. However, these methods have its own limitations in application. Traditional Chinese Medicine (TCM) is one of the oldest healing systems. Astragalus mongholicus (AMs) that is the common herbal medicine, the biggest part of TCM, have been proved to be effective in treating cancers from lots of clinical cases. However, we have not fully understood the anti-tumor mechanism of AMs, and this has lead to some doubt for some Western-Medicine scholars and restricts its wide use. The main objective of this research is to discuss the effect and mechanism of AMs to human stomach cancer. MATERIALS AND METHODS: To observe the effect and mechanism of tumor treatment by AMs, we have done the research from three major aspects, the influence of DCs, the inhibition of tumor in vitro as well as the animal studies in vivo after treatment. First, we culture the mouse dendritic cells (DCs) from bone marrow of mouse hind legs according to the method using Interleukin-4(IL-4) and Granulocyte-macrophage colony stimulating factor (GM-CSF), which refer to the way established by Inaba (Inaba K, 1992). And then we investigate the growth-rate of the DCs co-cultured with AMs injection. We analyze the expression of the Toll-like-receptor 4 (TLR4), with SYBR-Green I Real-time PCR and the I-kappa-B-alpha (IκB-α) with Western-Blot, the main regulatory protein to control nuclear factor NFκB-p65 nuclear translocation. Second, we choose the human gastric cancer cell lines MKN 45 as the target cell, which was co-cultured with DCs, T cells from spleen of mouse and AMs injection, and use MTT assay to judge the amount of cell lines and Immnunoflurescene to analyze the expression of anti-active caspase 3 pAb anti-PARP P85 fragment pAb, the mark of apoptosis of cells. Third, we have conducted the animal studies beside the basic experiment in vitro. The nude mouse developed stomach cancer, due to intra-preritoneal injection with MKN45 have been divided into two groups: the treatment group challenged with AMs injection and the control group with saline injection. We took the average of the diameter of each group as the y axis and the days after administered with AMs as x axis. After 40 days, all animals were killed by detruncation, and the tumor were removed and measured. We compare the diameter (<40 days) and weight (>40 days) of the tumor as well as the survival days between different groups to investigate the effect of inhibition of cancer. RESULTS: All results show that AMs is effective in treating human stomach cancer and the mechanism might be regulated by TLR4 mediated signal transduction of DCs. The results are briefly introduced as follows: First, we succeed in culturing the DCs induced by IL-4 and GM-CSF and find the positive rate of CD11c expression, the mark of DCs, is beyond 90% (Fig-1). We detect AMs can precipitate DCs maturation by upregulating TLR4 in SYBR-Green I Real-time PCR (Fig-2) and suppressing I.B-aby Western-Blot (Fig-3). Second, after the MKN45 co-cultured with DCs, T cells and AMs injection, the result show that AMs can great reduce the amount of cell lines by MTT assay (Fig-4) and induce apoptosis with Immunofluorescence (Fig-5). Finally, we have conducted animal studies beside the experiment in vitro, and the result in vivo show that AMs can delay tumor development from the diameter and weight of the tumor (Fig-6, Fig-7), prolong life-span and improve life-quality. Figure 1the morphology and phenotypic identification of DCs.The form of DCs observed by microscope with field 20*.The isotype antibody control using FCM.The positive rate of CD11c expression.Figure 2the melting curve and the chart of TLR4 expressiona) the melting curve of beta-actin; b)the melting curve of TLR4;c)the TLR4 expression of DCs stimulated with AM at different dose. There is significant statistic difference between the 60ng/mL and 80ng/mL group and other group (P<0.05 by rank test)Figure 3the IκB-α expression of DCs with different dose of AMsL0: 0ng/mL; L1:20ng/mL; L2:40ng/mL; L3:60ng/mL; L4:80ng/mLFigure 4MTT assay to analyse the viability and proliferation of the two cell lines (P<0.05 between the group with the dose of 60ng/mL and 80ng/mL and other group). the horizontal axis is the group treated with AM and saline at different dose, the vertical axis is the cell number.Figure 5the anti-active caspase-3 pAb (a) and anti-PARP P85 fragment pAb (b) actived by immunofluorescence.The cell mix were treated with 100uL anti-active caspase-3 pAb at a 1:250 dilution and anti-PARP P85 fragment pAb at a 1:100 dilution, and the secondary Ab was donkey anti-rabbit Cy®3 conjugate diluted 1:500 in PBS (Jackson Cat#711-165-152). From the photo, we find that anti-active caspase-3 pAb and anti-PARP P86 fragment pAb can express which is very important to indicate cell-apoptosis.Figure 6The difference of tumor between treatment group and control group. CONCLUSION: Ams Can play a great role in treating human stomach cancers as a good Chinese herbal medicine by precipitating DCs maturation, which is probably due to its effects by regulating the TLR4 mediated signal transduction.

[Predictors of efficacy of biofeedback therapy in patients with chronic constipation].

OBJECTIVE: To explore the anorectal physiology, psychological state, quality of life, lifestyle of patients with chronic constipation (CC) and evaluate the factors which potentially predict the efficacy of biofeedback therapy (BF). METHODS: Seventy CC patients receiving BF training were enrolled in this study. Anorectal physiology, Zung's Self-Rating Depression Scale (SDS), Zung's Self-Rating Anxiety Scale (SAS) and Chinese version of the MOS 36-item short form healthy survey (SF-36), lifestyle scale were recorded before BF training. A bowel symptom measurement including five major symptoms was recorded before and after BF training. The improvement in the symptom score was considered as criteria of clinical efficacy of BF therapy. Thirty-two possible influencing factors of efficacy of BF therapy were selected. Univariate analysis and multivariate analysis were conducted to assess the independent predictors. RESULTS: The results of univariate analysis showed that efficacy of BF therapy was positively correlated to the role physical (r = 0.256, P = 0.031), negatively correlated to the score of SDS (r = -0.315, P = 0.007) and the first sensation threshold (r = -0.278, P = 0.020). The multivariate analysis showed the score of SDS (ß = -0.263, P = 0.033) and the first sensation volume (ß = -0.281, P = 0.013) were the independent predicator of efficacy of BF therapy. CONCLUSION: CC patients who tended to depression state and rectal hyposensitivity have poor response to BF therapy. To treat these patients purposely could optimize the efficacy of BF therapy.