Xiaotan Sanjie recipe, a compound Chinese herbal medicine, inhibits gastric cancer metastasis by regulating GnT-V-mediated E-cadherin glycosylation / 中西医结合学报

OBJECTIVE@#Xiaotan Sanjie recipe (XTSJ), a Chinese herbal compound medicine, exerts a significant inhibitory effect on gastric cancer (GC) metastasis. This work investigated the mechanism underlying the XTSJ-mediated inhibition of GC metastasis.@*METHODS@#The effect of XTSJ on GC metastasis and the associated mechanism were investigated in vitro, using GC cell lines, and in vivo, using a GC mouse model, by focusing on the expression of Glc-N-Ac-transferase V (GnT-V; encoded by MGAT5).@*RESULTS@#The migration and invasion ability of GC cells decreased significantly after XTSJ administration, which confirmed the efficacy of XTSJ in treating GC in vitro. XTSJ increased the accumulation of E-cadherin at junctions between GC cells, which was reversed by MGAT5 overexpression. XTSJ administration and MGAT5 knockdown alleviated the structural abnormality of the cell-cell junctions, while MGAT5 overexpression had the opposite effect. MGAT5 knockdown and XTSJ treatment also significantly increased the accumulation of proteins associated with the E-cadherin-mediated adherens junction complex. Furthermore, the expression of MGAT5 was significantly lower in the lungs of BGC-823-MGAT5 + XTSJ mice than in those of BGC-823-MGAT5 + solvent mice, indicating that the ability of gastric tumors to metastasize to the lung was decreased in vivo following XTSJ treatment.@*CONCLUSION@#XTSJ prevented GC metastasis by inhibiting the GnT-V-mediated E-cadherin glycosylation and promoting the E-cadherin accumulation at cell-cell junctions. Please cite this article as: Huang N, He HW, He YY, Gu W, Xu MJ, Liu L. Xiaotan Sanjie recipe, a compound Chinese herbal medicine, inhibits gastric cancer metastasis by regulating GnT-V-mediated E-cadherin glycosylation. J Integr Med. 2023; 21(6): 561-574.

Study on Expression Condition Optimization of Two Recombinant Methioninases and Their Inhibitory Effects on Human Lung Adenocarcinoma Cells GLC / 中国药房

OBJECTIVE： To optimize the expression induction condition of two recombinant methioninases， and to investigate their inhibitory effects on the proliferation of human lung adenocarcinoma cells GLC. METHODS： Recombinant methioninases expression plasmid PGEX-4T1-4A1-MGL and PGEX-4T1-3B8-MGL were transfected into competent Escherichia coli Dh5α， and induced by isopropyl-β-D-thiogalactoside. Using the expression level of target protein as index， the initial OD600 nm value before induction， culture temperature and induction time were optimized by single factor test. The recombinant methioninase 4A1-MGL and 3B8-MGL were purified by affinity chromatography. The concentration of recombinant methioninase was detected by Coomassie blue method. The purity of the product was detected by sodium lauryl benzene sulfonate-polyacrylamide gel electrophoresis； its activity was detected by spectrophotometry. The proliferation of cells was detected by MTT assay after treated with low-dose， medium-dose and high-dose of recombinant methioninases （4A1-MGL or 3B8-MGL was 0.1， 0.2， 0.4 U/mL） for 24， 48， 74 h. Inhibitory rate of cells were calculated. RESULTS： The optimal induction condition of two recombinant methioninases included that initial OD600 nm of 0.9， culture temperature of 37 ℃， induction time of 5 h. The results of validation test showed that protein expression level of 4A1-MGL was 1.52±0.04， that of 3B8-MGL was 1.28±0.03 （RSD＜3％，n＝3）. After purification， the concentration， purity and activity of 4A1-MGL were （0.70±0.02）mg/mL， （96.42±3.15）％ and （0.45±0.02）    U/mg； and those of 3B8-MGL were （0.56±0.02）mg/mL， （97.43±2.96）％ and （0.91±0.03）U/mg. After treated with low-dose and medium-dose of 4A1-MGL and 3B8-MGL for 48 and 72 h， treated with high-dose of 4A1-MGL and 3B8-MGL for 24， 48 and 72 h， inhibitory rate of GLC cell was increased significantly， and high-dose group for 72 h was significantly higher than low-dose and medium-dose groups at same time point （P＜0.05）. CONCLUSIONS： The induction conditions of recombinant methioninase expression are successfully optimized in this study. The obtained 4A1-MGL and 3B8-MGL could inhibit the proliferation of GLC cells in a dose-dependent manner.

GLC-MS profiling of non-polar extracts from Phlomis bucharica and P. salicifolia and their cytotoxicity / Perfiles GLC-MS de extractos no polares de Phlomis bucharica y P. salicifolia y su citotoxicidad

Phlomis species (Phlomis bucharica Regel and P. salicifolia Regel) have been traditionally used by Uzbek people as stimulant, tonic, diuretic, and in the treatment of ulcers, hemorrhoids, wounds and gynecological problems. In the present study, we characterized the chemical composition of non-polar extracts from P. bucharica and P. salicifolia by high resolution GLC-MS and evaluated their cytotoxicity. Concentrations of hexadecanoic acid in hexane and chloroform extracts were higher in P. bucharica than in P. salicifolia. 1,8- Cineol, camphor, borneol, alfa-terpinol, thymol, and isobornyl acetate were detected in P. bucharica but not in P. salicifolia. About 45 components were identified in P. bucharica and 40 in P. salicifolia. The chloroform extract from P. bucharica showed cytotoxicity in HeLa and HL-60 cells, with IC50 values of 26.07 and 29.42 ug/ml, respectively.

Las especies Phlomis (Phlomis bucharica Regel y P. salicifolia Regel) se han utilizado tradicionalmente por la gente de Uzbekistán como estimulante, tónico, diurético, y en el tratamiento de las úlceras, hemorroides, heridas y problemas ginecológicos. En el presente estudio, hemos caracterizado la composición química de los extractos no polares de P. bucharica y P. salicifolia por GLC-MS de alta resolución y se evaluó su citotoxicidad. Las concentraciones de ácido hexadecanoico en extractos de hexano y cloroformo fueron mayores en P. bucharica que en P. salicifolia. 1,8-cineol, alcanfor, borneol, se detectaron alfa-terpinol, timol, y acetato de isobornilo en P. bucharica pero no en P. salicifolia. Cerca de 45 componentes fueron identificados en P. bucharica y 40 en P. salicifolia. El extracto de cloroformo a partir de P. bucharica mostró citotoxicidad en células HL-60 y HeLa, con valores de CI 50 de 26,07 y 29,42 ug/ml, respectivamente.

Effect of ampelopsin sodium combined with carboplatin on the proliferation and apoptosis of human lung adenocarcinoma cell GLC-82 / 中国药理学通报

Aim To investigate the cytotoxic effect and mechanism of ampelopsin sodium ( AMP-Na ) on hu-man lung adenocarcinoma cell line GLC-82 alone or combined with carboplatin ( CBP ) . Methods The cytotoxic effect of human lung adenocarcinoma cell line GLC-82 was investigated by 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide ( MTT ) colori-metric assay. Ultrastructure change of apoptotic GLC-82 cells was observed with transmission electron micro-scope. The changes of the cell apoptosis and the ex-pression of caspase-3 were analyzed with flow cytome-ter. Results Combined with AMP-Na, the IC50 of CBP decreased from (17. 10 ± 4. 78) mg·L-1 to tron microscope and flow cytometric analysis, the apop-tosis and necrosis ratios also increased in the combina-tion group. The necrosis ratios increased from (2. 56 ± 0. 41 )% to ( 71. 83 ± 5. 43 )% ( P<0. 01 ) . The ex-pression of caspase-3 was increased significantly after treated with AMP-Na or combined with CBP. Conclu-sions There is a synergistic cytotoxic effect on GLC-82 cells treated with AMP-Na combined with CBP. Ap-optotic cells and necrotic cells are found in GLC-82 cells treated with AMP-Na alone or combined with CBP. One of the mechanisms to induce apoptosis is probably that activation of caspase-3 mediates signal transduction pathway in cells.

Evaluation of the Urinary Glucose Tetrasaccharide Assay Using Ultra-Performance Liquid Chromatography-Tandem Mass Spectrometry for Diagnosis of Pompe Disease

We evaluated the urinary glucose tetrasaccharide (Glc4) assay using ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The calibration curve was linear over a range of 5-500 micromol/L. Performance parameters such as intra- and inter-day imprecision CVs were 6.52-14.6% and 11.5-13.2%, respectively. The mean concentrations of urinary Glc4 in 27 normal controls and 3 pseudodeficiency patients were 1.5 and 12.1 mmol/mol creatinine, respectively. Urinary Glc4 concentration in a patient with Pompe disease was 171.3 mmol/mol creatinine, which decreased to 130.9 mmol/mol following enzyme replacement therapy. Based on our results, we suggest that the urinary Glc4 assay using UPLC-MS/MS can be a reliable diagnostic tool for identification of patients with Pompe disease.

Inhibitory Effect and Its Mechanism of Dihydrotanshinone on Human Lung Cancer GLC-82 Cell / 中国药房

OBJECTIVE:To study the inhibitory effect of dihydrotanshinone(DTS)on human lung cancer GLC-82 cell and its mechanism. METHODS:After treated with 0(blank control),5,10,20,40,80 and 100 μg/ml DTS for 24 and 48 h,MTT as-say was used to measure the inhibition rates and IC50 of cells;cell apoptosis was detected by flow cytometry after treated with 17.85 μg/ml DTS for 12,24 and 48 h to calculate apoptotic rate;Western blot was used to detect the protein expressions of Bcl-2, Bax and Caspase-3. RESULTS:Compared with blank control group,different concentrations of DTS inhibited the proliferation of cells;24 and 48 h maximal inhibition rate were 54.48% and 64.95%,respectively;IC50 were 62.36 and 33.94 μg/ml. DTS could induce cell apoptosis in positive time dependent manner,and the range of inhibition rate was 5.6%-29.6%;Western blot showed DTS could down-regulate the expression of Bcl-2 protein and up-regulate the expression of Caspase-3 protein (P<0.01 or P<0.05). CONCLUSIONS:DTS have significant inhibitory effect on GLC-82 cells and also induce cell apoptosis,by a possible mech-anism of down-regulating the expression of Bcl-2 protein and up-regulating the expression of Caspase-3 protein.

Three novel polymorphic microsatellite markers for the glaucoma locus GLC1B by datamining tetranucleotide repeats on chromosome 2p12-q12

In order to identify new markers around the glaucoma locus GLC1B as a tool to refine its critical region at 2p11.2-2q11.2, we searched the critical region sequence obtained from the UCSC database for tetranucleotide (GATA)n and (GTCT)n repeats of at least 10 units in length. Three out of four potential microsatellite loci were found to be polymorphic, heterozygosity ranging from 64.56 percent to 79.59 percent. The identified markers are useful not only for GLC1B locus but also for the study of other disease loci at 2p11.2-2q11.2, a region with scarcity of microsatellite markers.

Nicotinamide enhances the effect of none focused ultrasound to kill human pulmonary carcinoma cell line GLC-82 / 基础医学与临床

Objective To determine the sensitization of Nicotinamide (NA) on None Focused Ultrasound (NFU) on GLC-82 cells in vitro. Methods The subjectives were divided into A、B、C and D four groups, group A was negative control, group B was treated with NA, group C was irradiated with NFU, group D was treated with both NFU and NA; MTT assay was used to evaluate the cytotoxicity of NA and its sensitivity for NFU on GLC-82 cells quantified by calculating the sensitive enhancement ratios (SER); Morphologic change of cells was observed with light microscope, fluorescence microscope and scanning electron microscope;Flow cytometry was performed to determine the percentage changes of apoptosis and distributional percentage of the cell cycle of GLC-82 cells in response to NFU, NA and the effect of its intervention to NFU. Results The cytotoxicity of NA increased in a dose-dependent manner after 24-hour treatment, with the optimal dose range of 1 mg/mL～5 mg/mL. A sub-toxic dose of NA at 3 mg/ml was used in the subsequent experiments; After treatment with NA plus NFU, microvilli and filipodium of GLC-82 cells reduced, shortened and shrunk into corpuscule-shape; The apoptosis rate of group D was more than that in group B and C, with cell cycle arrested at phase S and G2.Conclusion NA can significantly enhance the effect of NFU on GLC-82 cells, which provides evidence for NA as a sensitization to NFU in clinic.

The Role of Recombinant p53 Adenovirus Vector in the Growth of Lung Adenocarcinoma Cell Line GLC-82 / 中国肿瘤生物治疗杂志

Wild-type p53 has been shown to be involved in several aspects of cell growth control. In order to study the role of wild-type p53 in the growth of lung adenocarcinoma cell line GLC-82, recombinant p53 adenovirus vector and the control LacZ gene were constructed. Infection conditions were detected by biochemistry staining for LacZ s expression product ?-gal, immunohistochemical analysis for p53 protein expression, and PCR technique for the infected cells. The cell growth rate and cell cycle were analysed with flow cytometry. The results showed that the constructed recombinant adenovirus vector could infect cells with high level expression of (?-gal and p53 protein and p53 cDNA could be found in the infected cells. The wild-type p53 could inhibit GLC-82 cell proliferation and cause cellular death. These results indicated that wild-type p53 gene mediated by adenovirus vector might be used in the treatment of lung adenocarcinoma.

Gas-Liquid Chromamgraphic Analysis of Residual Ethylene Oxide and MMA Monomer in Various lntraocular Lenses

Recently, we performed an ezperi nlent to detect residual ethylene oxide(EO) gas and methylmethacrylate(MMA) monomer in the intraocular lenses from eleven different makers using gas-liquid chromatography. The residual EO gas over 25ppm was seen in 2 makers and also the MMA monomer over 2% was present in two makers. As a result, we recommend that the precise product quality-control of the IOLes is necessary especially for the lenses from foreign manufacturers.

Gas-Liquid Chromamgraphic Analysis of Residual Ethylene Oxide and MMA Monomer in Various lntraocular Lenses

Recently, we performed an ezperi nlent to detect residual ethylene oxide(EO) gas and methylmethacrylate(MMA) monomer in the intraocular lenses from eleven different makers using gas-liquid chromatography. The residual EO gas over 25ppm was seen in 2 makers and also the MMA monomer over 2% was present in two makers. As a result, we recommend that the precise product quality-control of the IOLes is necessary especially for the lenses from foreign manufacturers.