Optimization of Reference Genes for Normalization of the Quantitative Polymerase Chain Reaction in Tissue Samples of Gastric Cancer

Zhao, Lian-Mei,Zheng, Zhao-Xu,Zhao, Xiwa,Shi, Juan,Bi, Jian-Jun,Pei, Wei,Feng, Qiang Asian Pacific Journal of Cancer Prevention 2014 Asian Pacific journal of cancer prevention Vol.15 No.14

For an exact comparison of mRNA transcription in different samples or tissues with real time quantitative reverse transcription-polymerase chain reaction (qRT-PCR), it is crucial to select a suitable internal reference gene. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and beta-actin (ACTB) have been frequently considered as house-keeping genes to normalize for changes in specific gene expression. However, it has been reported that these genes are unsuitable references in some cases, because their transcription is significantly variable under particular experimental conditions and among tissues. The present study was aimed to investigate which reference genes are most suitable for the study of gastric cancer tissues using qRT-PCR. 50 pairs of gastric cancer and corresponding peritumoral tissues were obtained from patients with gastric cancer. Absolute qRT-PCR was employed to detect the expression of GAPDH, ACTB, RPII and 18sRNA in the gastric cancer samples. Comparing gastric cancer with corresponding peritumoral tissues, GAPDH, ACTB and RPII were obviously upregulated 6.49, 5.0 and 3.68 fold, respectively. Yet 18sRNA had no obvious expression change in gastric cancer tissues and the corresponding peritumoral tissues. The expression of GAPDH, ${\beta}$-actin, RPII and 18sRNA showed no obvious changes in normal gastric epithelial cells compared with gastric cancer cell lines. The carcinoembryonic antigen (CEA), a widely used clinical tumor marker, was used as a validation gene. Only when 18sRNA was used as the normalizing gene was CEA obviously elevated in gastric cancer tissues compared with peritumoral tissues. Our data show that 18sRNA is stably expressed in gastric cancer samples and corresponding peritumoral tissues. These observations confirm that there is no universal reference gene and underline the importance of specific optimization of potential reference genes for any experimental condition.

Study of Antimicrobial Effects of Different Types of Glycyrrhiza Extracts by Microcalorimetry

Wei, Ting,Lin, Guimei,Liu, Lian,Zhao, Zhongxi Korean Chemical Society 2014 Bulletin of the Korean Chemical Society Vol.35 No.8

Recently studies indicate that the microcalorimetry is a suitable measurement for metabolic activities of organisms by recording the rate of heat outputs. In this work, we investigated the growth thermogenic curves of Escherichia coli (E. coli) affected by three kinds of Glycyrrhiza extracts by microcalorimetry. The power-time and exponential phase power-time curves of the E. coli growth at various concentrations of extracts were generated. The kinetic parameters such as the growth rate constants (C), maximum power outputs (Pm), peak times (Tm), and inhibition ratios were calculated and the relationships between Pm or Tm and C were established. Also, the clear correlations among the antimicrobial effects, Pm and C were obtained. The results revealed the Glycyrrhiza extracts had inhibitory activities towards E. coli while the Glycyrrhiza polysaccharides showed the most potent effects.

Vibration test method of aero-engine 3D printing pre-swirl nozzle based on equivalent installation stiffness

Yujie Zhao,Yeda Lian,Lei Li,Xu Gong,Xianghai Chai,Wei Liu 대한기계학회 2023 JOURNAL OF MECHANICAL SCIENCE AND TECHNOLOGY Vol.37 No.2

Pre-swirl nozzle is an important cooling component that is located inside of aero-engine and exposed to complex vibration excitation, and exploring its vibration characteristics and durability is of significant importance, especially for 3D printing pre-swirl nozzle. The present study provides a novel vibration test method based on equivalent installation stiffness. Firstly, the vibration characteristics of pre-swirl nozzle in real engine assembly state is calculated by the finite element method, and the equivalent constraint fixture is designed and checked based on this. The check results indicate that the designed fixture can well restore the installation stiffness of engine and the strength of the test fixture elements is enough in the vibration test process. Then, the modal characteristics of pre-swirl nozzle are measured by modal test, based on which the accuracy of the finite element model is verified. Finally, a test program of vibration durability is established to evaluate the vibration durability of pre-swirl nozzle, in which the excitation frequency is determined by combining logarithmic frequency sweep and linear frequency sweep and the location of maximum stress is determined based on a novel method (density increasing method). Besides, the calibration of relation between vibration stress and excitation amplitude is completed. Based on the proposed test program, vibration durability test is performed. The results show that the natural frequency of pre-swirl nozzle only decreases by 0.62 % after 5×10 7 cycles, which indicates that the specimen is always in the stage of fatigue crack initiation over the test duration.

Identification of a Lead small-molecule inhibitor of anthrax Lethal toxin by using fluorescence-based high-throughput screening

( Dong Wei ),( Zhao Yun Bu ),( Ai Lian Yu ),( Feng Li ) 생화학분자생물학회(구 한국생화학분자생물학회) 2011 BMB Reports Vol.44 No.12

Inhalational anthrax is caused by B. anthracis, a virulent spore- forming bacterium which secretes anthrax toxins consisting of protective antigen (PA), lethal factor (LF) and edema factor (EF). LF is a Zn-dependent metalloprotease and is the main determinant in the pathogenesis of anthrax. Here we report the identification of a lead small-molecule inhibitor of anthrax lethal factor by screening an available synthetic small-molecule inhibitor library using fluorescence-based high-throughput screening (HTS) approach. Seven small molecules were found to have inhibitory effect against LF activity, among which SM157 had the highest inhibitory activity. All theses small molecule inhibitors inhibited LF in a noncompetitive inhibition mode. SM157 and SM167 are from the same family, both having an identical group complex, which is predicted to insert into S1` pocket of LF. More potent small-molecule inhibitors could be developed by modifying SM157 based on this identical group complex. [BMB reports 2011; 44(12): 811-815]

Study of Antimicrobial Effects of Different Types of Glycyrrhiza Extracts by Microcalorimetry

Ting Wei,Guimei Lin,Lian Liu,Zhongxi Zhao 대한화학회 2014 Bulletin of the Korean Chemical Society Vol.35 No.8

Recently studies indicate that the microcalorimetry is a suitable measurement for metabolic activities of organisms by recording the rate of heat outputs. In this work, we investigated the growth thermogenic curves of Escherichia coli (E. coli) affected by three kinds of Glycyrrhiza extracts by microcalorimetry. The powertime and exponential phase power-time curves of the E. coli growth at various concentrations of extracts were generated. The kinetic parameters such as the growth rate constants (C), maximum power outputs (Pm), peak times (Tm), and inhibition ratios were calculated and the relationships between Pm or Tm and C were established. Also, the clear correlations among the antimicrobial effects, Pm and C were obtained. The results revealed the Glycyrrhiza extracts had inhibitory activities towards E. coli while the Glycyrrhiza polysaccharides showed the most potent effects.

Endophytic Trichoderma gamsii YIM PH30019: a promising biocontrol agent with hyperosmolar, mycoparasitism, and antagonistic activities of induced volatile organic compounds on root-rot pathogenic fungi of Panax notoginseng

Chen, Jin-Lian,Sun, Shi-Zhong,Miao, Cui-Ping,Wu, Kai,Chen, You-Wei,Xu, Li-Hua,Guan, Hui-Lin,Zhao, Li-Xing The Korean Society of Ginseng 2016 Journal of Ginseng Research Vol.40 No.4

Background: Biocontrol agents are regarded as promising and environmental friendly approaches as agrochemicals for phytodiseases that cause serious environmental and health problems. Trichoderma species have been widely used in suppression of soil-borne pathogens. In this study, an endophytic fungus, Trichoderma gamsii YIM PH30019, from healthy Panax notoginseng root was investigated for its biocontrol potential. Methods: In vitro detached healthy roots, and pot and field experiments were used to investigate the pathogenicity and biocontrol efficacy of T. gamsii YIM PH30019 to the host plant. The antagonistic mechanisms against test phytopathogens were analyzed using dual culture, scanning electron microscopy, and volatile organic compounds (VOCs). Tolerance to chemical fertilizers was also tested in a series of concentrations. Results: The results indicated that T. gamsii YIM PH30019 was nonpathogenic to the host, presented appreciable biocontrol efficacy, and could tolerate chemical fertilizer concentrations of up to 20%. T. gamsii YIM PH30019 displayed antagonistic activities against the pathogenic fungi of P. notoginseng via production of VOCs. On the basis of gas chromatography-mass spectrometry, VOCs were identified as dimethyl disulfide, dibenzofuran, methanethiol, ketones, etc., which are effective ingredients for antagonistic activity. T. gamsii YIM PH30019 was able to improve the seedlings' emergence and protect P. notoginseng plants from soil-borne disease in the continuous cropping field tests. Conclusion: The results suggest that the endophytic fungus T. gamsii YIM PH30019 may have a good potential as a biological control agent against notoginseng phytodiseases and can provide a clue to further illuminate the interactions between Trichoderma and phytopathogens.

An Efficient PEG/CaCl2-Mediated Transformation Approach for the Medicinal Fungus Wolfiporia cocos

( Qiao Sun ),( Wei Wei ),( Juan Zhao ),( Jia Song ),( Fang Peng ),( Shao Peng Zhang ),( Yong Lian Zheng ),( Ping Chen ),( Wen Jun Zhu ) 한국미생물 · 생명공학회 2015 Journal of microbiology and biotechnology Vol.25 No.9

Sclerotia of Wolfiporia cocos are of medicinal and culinary value. The genes and molecular mechanisms involved in W. cocos sclerotial formation are poorly investigated because of the lack of a suitable and reproducible transformation system for W. cocos. In this study, a PEG/ CaCl2-mediated genetic transformation system for W. cocos was developed. The promoter Pgpd from Ganoderma lucidum effectively drove expression of the hygromycin B phosphotransferase gene in W. cocos, and approximately 30 transformants were obtained per 10 μg DNA when the protoplast suspension density was 106 protoplasts/ml. However, no transformants were obtained under the regulation of the PtrpC promoter from Aspergillus nidulans.

Three-dimensional cone beam computed tomography analysis of temporomandibular joint response to the Twin-block functional appliance

Yuan-yuan Jiang,Lian Sun,Hua Wang,Chun-yang Zhao,Wei-Bing Zhang 대한치과교정학회 2020 대한치과교정학회지 Vol.50 No.2

Objective: To propose a three-dimensional (3D) method for evaluating temporomandibular joint (TMJ) changes during Twin-block treatment. Methods: Seventeen patients with Class II division 1 malocclusion treated using Twinblock and nine untreated patients with a similar malocclusion were included in this research. We collected their cone beam computed tomography (CBCT) data from before and 8 months after treatment. Segmentations were constructed using ITK-SNAP. Condylar volume and superficial area were measured using 3D Slicer.The 3D landmarks were identified on CBCT images by using Dolphin software to assess the condylar positional relationship. 3D models of the mandible and glenoid fossa of the patients were constructed and registered via voxel-based superimposition using 3D Slicer. Thereafter, skeletal changes could be visualized using 3DMeshMetric in any direction of the superimposition on a color-coded map. All the superimpositions were measured using the same scale on the distance color-coded map, in which red color represents overgrowth and blue color represents resorption. Results: Significant differences were observed in condylar volume, superficial area, and condylar position in bothgroups after 8 months. Compared with the control group (CG), the Twin-block group exhibited more obvious condyle-fossa modifications and joint positional changes. Moreover, on the color-coded map, more obvious condyle-fossa modifications could be observed in the posterior and superior directions in the Twin-block group than in the CG. Conclusions: We successfully established a 3D method for measuring and evaluating TMJ changes caused by Twin-block treatment. The treatment produced a larger condylar size and caused condylar positional changes. [Korean J Orthod 2020;50(2):86-97]

Effect of Trichostatin A on Anti HepG2 Liver Carcinoma Cells: Inhibition of HDAC Activity and Activation of Wnt/β-Catenin Signaling

Shi, Qing-Qiang,Zuo, Guo-Wei,Feng, Zi-Qiang,Zhao, Lv-Cui,Luo, Lian,You, Zhi-Mei,Li, Dang-Yang,Xia, Jing,Li, Jing,Chen, Di-Long Asian Pacific Journal of Cancer Prevention 2014 Asian Pacific journal of cancer prevention Vol.15 No.18

Purpose: To investigate the effect of deacetylase inhibitory trichostatin A (TSA) on anti HepG2 liver carcinoma cells and explore the underlying mechanisms. Materials and Methods: HepG2 cells exposed to different concentrations of TSA for 24, 48, or 72h were examined for cell growth inhibition using CCK8, changes in cell cycle distribution with flow cytometry, cell apoptosis with annexin V-FTIC/PI double staining, and cell morphology changes under an inverted microscope. Expression of ${\beta}$-catenin, HDAC1, HDAC3, H3K9, CyclinD1 and Bax proteins was tested by Western blotting. Gene expression for ${\beta}$-catenin, HDAC1and HDAC3 was tested by q-PCR. ${\beta}$-catenin and H3K9 proteins were also tested by immunofluorescence. Activity of Renilla luciferase (pTCF/LEF-luc) was assessed using the Luciferase Reporter Assay system reagent. The activity of total HDACs was detected with a HDACs colorimetric kit. Results: Exposure to TSA caused significant dose-and time-dependent inhibition of HepG2 cell proliferation (p<0.05) and resulted in increased cell percentages in G0/G1 and G2/M phases and decrease in the S phase. The apoptotic index in the control group was $6.22{\pm}0.25%$, which increased to $7.17{\pm}0.20%$ and $18.1{\pm}0.42%$ in the treatment group. Exposure to 250 and 500nmol/L TSA also caused cell morphology changes with numerous floating cells. Expression of ${\beta}$-catenin, H3K9and Bax proteins was significantly increased, expression levels of CyclinD1, HDAC1, HDAC3 were decreased. Expression of ${\beta}$-catenin at the genetic level was significantly increased, with no significant difference in HDAC1and HDAC3 genes. In the cytoplasm, expression of ${\beta}$-catenin fluorescence protein was not obvious changed and in the nucleus, small amounts of green fluorescence were observed. H3K9 fluorescence protein were increased. Expression levels of the transcription factor TCF werealso increased in HepG2 cells following induction by TSA, whikle the activity of total HDACs was decreased. Conclusions: TSA inhibits HDAC activity, promotes histone acetylation, and activates Wnt/${\beta}$-catenin signaling to inhibit proliferation of HepG2 cell, arrest cell cycling and induce apoptosis.

Downregulation of LINC01508 contributes to cisplatin resistance in ovarian cancer via the regulation of the Hippo-YAP pathway

Lan Xiao,Xiao-Yan Shi,Ze-Lian Li,Min Li,Min-Min Zhang,Shi-Jie Yan,Zhao-Lian Wei 대한부인종양학회 2021 Journal of Gynecologic Oncology Vol.32 No.5

Background: Some long non-coding RNAs (lncRNAs) have been found to contribute to cisplatin resistance. Here, we identified a novel lncRNA that was downregulated in cisplatin- resistant to ovarian cancer (OC) cells and aimed to examine the contribution of LINC01508 to cisplatin resistance in OC cells. Methods: Differences in the lncRNA expression profile between OV2008 and C13K cells were assessed by lncRNA expression microarray. The expression of LINC01508 in ovarian epithelial cells, four OC cells, and OC, benign ovary tumor and normal ovary, cisplatin- resistant and non-resistant OC specimens were evaluated by quantitative real-time polymerase chain reaction (qPCR). The role of LINC01508 in OC cisplatin-resistant was evaluated by cell counting kit-8 (CCK-8), flow cytometry, colony formation, wound healing, Transwell, and tumor growth inhibition study in vivo. The clinical associations of LINC01508 in OC were evaluated using correlation analysis. The effects of verteporfin (VP) on cisplatin were explored to reveal the function of the hippo-YAP pathway on the cisplatin tolerance of C13K. Results: LINC01508 was downregulated in cisplatin-resistant OC cells and platinum- resistant OC tissue (p<0.01). LINC01508 downregulation was correlated with tumor size, residual tumor, and platinum resistance. The overexpression of LINC01508 improves in vitro and in vivo sensitivity to cisplatin while predicts the poor overall survival which need further follow-up research. The increased level of LINC01508 could suppress the cisplatin resistance of OC cells through the inhibition of the hippo-YAP pathway. Conclusions: The study proposes that dysregulation of LINC01508 expression results in resistance of OC to cisplatin through the inhibition of the hippo-YAP pathway.