Mechanism of Xianlian Jiedu Prescription in Maintaining Cancer Relative Vascular Endothelial Cell Homeostasis and Inhibiting Tumor Neovascularization / 中国实验方剂学杂志

ObjectiveTo observe the effect of Xianlian Jiedu prescription （XLJDP） on the proliferation， apoptosis， and migration of cancer-relative endothelial （CRE） cells， and to decipher the mechanism of XLJDP in regulating angiopoietin2 （Ang2） to maintain CRE cell homeostasis and inhibit tumor neovascularization. MethodHuman umbilical vein endothelial cell line （HUVEC-c） was induced into CRE cells in the human colorectal cancer HCT-116 cell-conditioned medium. The CRE cells were assigned into the blank group， conditioned medium group， and XLJDP groups （1， 2， 3 g·L-1） and treated for 48 h. The proliferation of CRE cells was detected by methyl thiazolyl tetrazolium （MTT） colorimetry. The morphological changes of CRE cells were observed via an inverted microscope. The apoptosis rate was detected by flow cytometry. Wound healing test and Transwell migration assay were employed to detect the 2D/3D migration ability of CRE cells. The protein levels of vimentin， N-cadherin， matrix metalloproteinase-9 （MMP-9）， and Ang2 in CRE cells were measured by Western blot. ResultThe MTT results showed that the cell viability was higher in the conditioned medium group than in the blank group （P<0.05）. Compared with the conditioned medium group， XLJDP decreased the cell proliferation rate （P<0.01） and changed the cell morphology. The total apoptosis rates of all the XLJDP groups were higher than that of the conditioned medium group （P<0.01）. The 2D and 3D migration abilities of the conditioned medium group were higher than those of the blank group （P<0.05， P<0.01）. Compared with the conditioned medium group， XLJDP at all the concentrations weakened the 2D migration ability （P<0.01） and medium- and high-concentration XLJDP weakened the 3D migration ability （P<0.01）. The protein levels of N-cadherin， Vimentin， MMP-9， and Ang2 were up-regulated in the conditioned medium group compared with those in the blank group （P<0.05， P<0.01）. Compared with the conditioned medium group， XLJDP at all the concentrations down-regulated the protein level of Ang2 （P<0.05， P<0.01）， and medium- and high-concentration XLJDP down-regulated those of N-cadherin， vimentin， and MMP-9 protein （P<0.01）. ConclusionXLJDP may inhibit the proliferation， migration， differentiation， and apoptosis of CRE cells by down-regulating the expression of Ang2， inhibiting tumor neovascularization， and maintaining the cell homeostasis.

Effect and Mechanism of Xianlian Jiedu Prescription Against Proliferation of Colorectal Cancer Cells in Hypoxic Microenvironment / 中国实验方剂学杂志

ObjectiveTo study the effect of Xianlian Jiedu prescription （XLJDP） on the activation of nuclear transcription factor-κB （NF-κB） signaling pathway induced by bromodomain-containing protein 4 （Brd4） in hypoxic microenvironment and to explore its mechanism in inhibiting the proliferation of colorectal cancer HT-29 cells. MethodThe human colorectal cancer HT-29 cells were cultured in a hypoxic incubator or normoxia incubator and treated with XLJDP at 0.8，1，1.2，1.6，3.2，6.4，and 12.8 g·L-1 for 48 h， respectively. Following the detection of cell vitality using methyl thiazolyl tetrazolium （MTT） colorimetry， the effects of XLJDP （1.25，2.5，and 5 g·L-1） on the cell mitochondrial membrane potential were determined using a fluorescent probe （JC-1）， and the apoptosis of colorectal cancer HT-29 cells was detected by flow cytometry. The cell colony formation assay and 5-ethynyl-2'-deoxyuridine （EDU） staining were conducted to test the proliferation of colorectal cancer HT-29 cells. The Western blot was carried out to measure the expression levels of Brd4 and its downstream relevant proteins such as c-Myc and hexamethylene bisacetamide-inducible protein 1 （HEXIM1）， as well as the effects of XLJDP on related proteins in the NF-κB signaling pathway. ResultCompared with the blank control group， XLJDP at 0.8，1，1.2，1.6，3.2，6.4，and 12.8 g·L-1 inhibited the vitality of colorectal cancer HT-29 cells （P<0.05 ， P<0.01）， with the median inhibitory concentration （IC50） under the hypoxic condition higher than that under the normoxia condition. Compared with the blank control group， XLJDP at 1.25，2.5，and 5 g·L-1 significantly decreased the mitochondria membrane potential， enhanced the apoptosis （P<0.05，P<0.01）， and lowered the number of cell colonies and also the EDU-positive cells （P<0.05， P<0.01）. The results of Western blot showed that compared with the blank control group， XLJDP at 1.25，2.5，and 5 g·L-1 down-regulated Brd4， c-Myc， p-NF-κB p65， and p-IκBα protein expression to varying degrees and up-regulated the expression of HEXIM1 （P<0.05， P<0.01）. ConclusionIn the hypoxic microenvironment， XLJDP inhibits the proliferation of colorectal cancer HT-29 cells regulated by Brd4， which may be related to its inhibition of the activation of NF-κB signaling pathway.

Multi-index components of Scutellariae Barbatae Herba according to UHPLC-QTRAP-MS coupled with chemometrics / 中国中药杂志

The present study developed an ultra-fast liquid chromatography coupled with triple quadrupole-linear ion trap composite mass spectrometry(UHPLC-QTRAP-MS) to simultaneously determine the content of potential active components in Scutellariae Barbatae Herba and also to provide a reference approach for screening out the differential quality control components among different batches of Scutellariae Barbatae Herba. Chromatographic separations were conducted on a Thermo Acclaim~(TM) RSLC 120 C_(18) column(3.0 mm×100 mm, 2.2 μm) in a gradient program. The mobile phase consisted of 0.1% aqueous formic acid and acetonitrile, and the column temperature was maintained at 40 ℃. The flow rate was 0.4 mL·min~(-1) and the injection volume was 2 μL. The targeted compounds were monitored in the multiple reaction monitoring(MRM) mode. The acquired data were processed by hierarchical cluster analysis(HCA) and partial least square discriminant analysis(PLS-DA). Sixteen compounds all showed good linear relationship within the corresponding linear ranges and the R~2 values were all higher than 0.993 2. The RSDs of precision, repeatability, and stability were less than or equal to 3.7%. Mean recovery rates were in the range of 95.67% and 104.8% with RSDs≤3.2%. According to HCA and PLS-DA, all samples were clustered into four categories. Scutellarin, acteoside, scutellarein, and scutebarbatine X(VIP>1) were considered as differential chemical markers in the four categories. In conclusion, the developed method can be used for the simulta-neous determination of the multiple components and quality control of Scutellariae Barbatae Herba.

Sub-anesthesia Dose of Isoflurane in 60% Oxygen Reduces Inflammatory Responses in Experimental Sepsis Models / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Sepsis is a major cause of mortality in Intensive Care Units. Anesthetic dose isoflurane and 100% oxygen were proved to be beneficial in sepsis; however, their application in septic patients is limited because long-term hyperoxia may induce oxygen toxicity and anesthetic dose isoflurane has potential adverse consequences. This study was scheduled to find the optimal combination of isoflurane and oxygen in protecting experimental sepsis and its mechanisms.</p><p><b>METHODS</b>The effects of combined therapy with isoflurane and oxygen on lung injury and sepsis were determined in animal models of sepsis induced by cecal ligation and puncture (CLP) or intraperitoneal injection of lipopolysaccharide (LPS) or zymosan. Mouse RAW264.7 cells or human peripheral blood mononuclear cells (PBMCs) were treated by LPS to probe mechanisms. The nuclear factor kappa B (NF-κB) signaling molecules were examined by Western blot and cellular immunohistochemistry.</p><p><b>RESULTS</b>The 0.5 minimum alveolar concentration (MAC) isoflurane in 60% oxygen was the best combination of oxygen and isoflurane for reducing mortality in experimental sepsis induced by CLP, intraperitoneal injection of LPS, or zymosan. The 0.5 MAC isoflurane in 60% oxygen inhibited proinflammatory cytokines in peritoneal lavage fluids (tumor necrosis factor-alpha [TNF-β]: 149.3 vs. 229.7 pg/ml, interleukin [IL]-1β: 12.5 vs. 20.6 pg/ml, IL-6: 86.1 vs. 116.1 pg/ml, and high-mobility group protein 1 [HMGB1]: 323.7 vs. 449.3 ng/ml; all P< 0.05) and serum (TNF-β: 302.7 vs. 450.7 pg/ml, IL-1β: 51.7 vs. 96.7 pg/ml, IL-6: 390.4 vs. 722.5 pg/ml, and HMGB1: 592.2 vs. 985.4 ng/ml; all P< 0.05) in septic animals. In vitro experiments showed that the 0.5 MAC isoflurane in 60% oxygen reduced inflammatory responses in mouse RAW264.7 cells, after LPS stimulation (all P< 0.05). Suppressed activation of NF-κB pathway was also observed in mouse RAW264.7 macrophages and human PBMCs after LPS stimulation or plasma from septic patients. The 0.5 MAC isoflurane in 60% oxygen also prevented the increases of phospho-IKKβ/β, phospho-IκBβ, and phospho-p65 expressions in RAW264.7 macrophages after LPS stimulation (all P< 0.05).</p><p><b>CONCLUSION</b>Combined administration of a sedative dose of isoflurane with 60% oxygen improves survival of septic animals through reducing inflammatory responses.</p>

Analysis on chemical components from water extract of paeoniae radix alba by high performance liquid chromatography-electrospray ionization-quadrupole-time of flight-mass spectrometry / 中国中药杂志

This experiment was performed to establish a qualitative analysis on chemical composition in water extract of Paeoniae Radix Alba by HPLC-ESI-Q-TOF-MS. The analysis was conducted on a C18 (Hanbon Lichrospher, 4.6 mm x 250 mm, 5 microm) column with methanol-0.1% formic acid as the mobile phase for gradient elution; ESI ion source was used for mass spectra, and data were collected in both positive and negative modes. The results showed that eleven compounds from water extract of Paeoniae Radix Alba had been identified by analyzing positive and negative ion mass data including element composition and by comparing with data from literatures. Since efficient separation of HPLC and the high sensitive detection of MS was used, this experiment, it will provide evidences for elucidation of the effective substance in the water extract of Paeoniae Radix Alba.

Prevention of anterior glottic stenosis after CO2 laser cordectomy with chitosan / 中华耳鼻咽喉头颈外科杂志

<p><b>OBJECTIVE</b>To observe the effect of immediate topical application of chitosan on preventing anterior glottic stenosis (AGS) after microsurgical resection of both vocal fold with CO2 laser, including the anterior commissure, in a canine model.</p><p><b>METHODS</b>Sixteen canine larynges were injured by microresecting procedure of both vocal folds with CO2 laser. The dogs were randomly divided into two groups, chitosan group and control group. The chitosan and isotonic sodium chloride solution (control) were used for 5 minutes immediately after surgery. One week after the initial surgery, three dogs in each group were randomly selected , ultrastructure of fibroblast were examined with transmission electronic microscope and expression of basic fibroblast growth factor (bFGF) and transforming growth factor beta1 (TGF-beta1) were evaluated by enzyme-linked immunosorbent assay (ELISA). Three weeks after surgery, the rest dogs' glottic web were lysed and repeatedly treated with chitosan and isotonic sodium chloride solution respectively. The glottic wound healing and AGS formation were examined every week, and all larynges were harvested and examined histologically six weeks after the initial surgery.</p><p><b>RESULTS</b>Transmission electronic microscope examination of the ultrastructure of fibroblast indicated that chitosan inhibited the proliferation of fibroblast. Chitosan increased the expression of bFGF and TGF-beta1, and bFGF and TGF-beta1 in chitosan group, which was significantly higher than that in control group (z=-2.887 and -2.005, P=0.002 and 0.041). Chitosan decreased the extent of AGS formation. Three weeks after the surgery, the AGS lesion in the control group affected mean 49% of the length of the vocal folds from the anterior commissure to the vocal process, while chitosan group affected mean 7%, which was significantly less than the extent of web formation in the control group, (z=-2.619, P=0.008). The grade of collagen content in chitosan group was significantly lower than that in control group (P=0.003).</p><p><b>CONCLUSION</b>Chitosan is effective in preventing AGS after CO2 laser cordectomy.</p>

Chemical constituents from water extract of Radix isatidis / 药学学报

<p><b>AIM</b>To study the chemical constituents from water extract of Radix isatidis. (Isatis indigotica Fort. ).</p><p><b>METHODS</b>The water extract was underwent absorption by D101 macroporous resin, the portion eluted by ethanol of different concentrations was isolated and purified on silica gel column repeatedly. The obtained compounds were identified and structurally elucidated by their physico-chemical properties and spectral analysis.</p><p><b>RESULTS</b>Five compounds were isolated from water extract of Radix isatidis, and were partly identified separately: 3-[2'-(5'-hydroxymethyl) furyl] -1 (2H) -isoquinolinone-7-O-beta-D-glucoside (I), lariciresinol-4,4'-di-O-beta-D-glucopyranoside (II), lariciresinol-4-O-beta-D-glucopyranoside (III), 2-hydroxy-1, 4-benzenedicarboxylic acid (IV), mannitol (V).</p><p><b>CONCLUSION</b>Compound I is a new compound and compounds IV and V were isolated from the plant for the first time.</p>