Construction and Identification of an Antisense Glucose Transporter-1 Plasmid

2008 ◽  
Vol 36 (5) ◽  
pp. 1001-1007
Author(s):  
XM Chen ◽  
SH Zhou ◽  
J Fan ◽  
MJ Hu ◽  
SQ Wang ◽  
...  
Keyword(s):  
Laryngeal Carcinoma ◽  
Glucose Transporter ◽  
Therapeutic Potential ◽  
Restriction Analysis ◽  
Expression System ◽  
Carcinoma Cells ◽  
Glucose Transporter 1 ◽  
Glut 1 ◽  
Firm Basis ◽  
Polymerase Chain

Our aim was to construct a pcDNA3.1(+) eucaryotic expression system vector containing the antisense glucose transporter-1 ( Glut-1) gene. Total RNA was isolated from human Hep-2 laryngeal carcinoma cells, and the Glut-1 and antisense Glut-1 sequences were amplified by polymerase chain reaction. Expression plasmids containing the sense and antisense cDNA were constructed using the pcDNA3.1(+) vector. The resulting sense and antisense vectors, pcDNA3.1(+)-Glut-1 and pcDNA3.1(+)-antiGlut-1, respectively, were examined by restriction analysis and DNA sequencing. The pcDNA3.1(+)-antiGlut-1 was subsequently transfected into Hep-2 cells. Anti Glut-1 mRNA expression was detected, indicating the successful construction of an antisense Glut-1 plasmid capable of transfecting Hep-2 laryngeal carcinoma cells. These data provide a firm basis for additional studies using the plasmid pcDNA3.1(+)-antiGlut-1 to determine its therapeutic potential for the treatment of laryngeal carcinoma.

scholarly journals Effect of Inhibition of GLUT1 Expression and Autophagy Modulation on the Growth and Migration of Laryngeal Carcinoma Stem Cells under Hypoxic and Low-Glucose Conditions

2020 ◽  
Author(s):  
Shui-Hong Zhou ◽  
Xiao-Hong Chen ◽  
Jia Liu ◽  
Jiang-Tao Zhong ◽  
Jun Fan
Keyword(s):  
Stem Cells ◽  
Glucose Uptake ◽  
Laryngeal Carcinoma ◽  
Glucose Transporter ◽  
Cell Counting ◽  
Glucose Transporter 1 ◽  
Glut 1 ◽  
Low Glucose ◽  
And Migration ◽  
Proliferation And Migration

Abstract Background: Enhanced glucose uptake and autophagy are means by which cells adapt to stressful microenvironments. We investigated the roles of glucose transporter-1 (GLUT-1) and autophagy in laryngeal carcinoma stem cells under hypoxic and low-glucose conditions.Methods: CD133+ Tu212 laryngeal carcinoma stem cells were purified by magnetic-activated cell sorting and subjected to hypoxic and/or low-glucose conditions. Proliferation was evaluated using a cell-counting kit and a clone-formation assay, and migration was evaluated through a Transwell assay. Autophagy was assessed via transmission electron microscopy. GLUT-1 and beclin-1 expression were silenced using an shRNA and autophagy was manipulated using rapamycin, 3-MA, or chloroquine. Gene expression levels were evaluated by quantitative reverse transcription-polymerase chain reaction and protein concentrations were assessing via Western blotting.Results: Compared to CD133– stem cells, CD133+ cells showed increased proliferation and migration, and reduced apoptosis, under hypoxic or low-glucose conditions. They also showed increased expression of GLUT-1 and autophagy markers. Finally, GLUT-1 knockdown or autophagy inhibition reduced their proliferation and migration.Conclusions: Enhanced glucose uptake and autophagy maintain the functions of CD133+ laryngeal carcinoma stem cells under hypoxic and low-glucose conditions.

scholarly journals Effect of Inhibition of GLUT1 Expression and Autophagy Modulation on the Growth and Migration of Laryngeal Carcinoma Stem Cells Under Hypoxic and Low-Glucose Conditions

2020 ◽  
Author(s):  
Xiao-Hong Chen ◽  
Jia Liu ◽  
Jiang-Tao Zhong ◽  
Shui-Hong Zhou ◽  
Jun Fan
Keyword(s):  
Stem Cells ◽  
Glucose Uptake ◽  
Laryngeal Carcinoma ◽  
Glucose Transporter ◽  
Cell Counting ◽  
Glucose Transporter 1 ◽  
Glut 1 ◽  
Low Glucose ◽  
And Migration ◽  
Proliferation And Migration

Abstract Background: Enhanced glucose uptake and autophagy are means by which cells adapt to stressful microenvironments. We investigated the roles of glucose transporter-1 (GLUT-1) and autophagy in laryngeal carcinoma stem cells under hypoxic and low-glucose conditions.Methods: CD133+ Tu212 laryngeal carcinoma stem cells were purified by magnetic-activated cell sorting and subjected to hypoxic and/or low-glucose conditions. Proliferation was evaluated using a cell-counting kit and a clone-formation assay, and migration was evaluated through a Transwell assay. Autophagy was assessed via transmission electron microscopy. GLUT-1 and beclin-1 expression were silenced using an shRNA and autophagy was manipulated using rapamycin, 3-MA, or chloroquine. Gene expression levels were evaluated by quantitative reverse transcription-polymerase chain reaction and protein concentrations were assessing via Western blotting. Results: Compared to CD133– stem cells, CD133+ cells showed increased proliferation and migration, and reduced apoptosis, under hypoxic or low-glucose conditions. They also showed increased expression of GLUT-1 and autophagy markers. Finally, GLUT-1 knockdown or autophagy inhibition reduced their proliferation and migration.Conclusions: Enhanced glucose uptake and autophagy maintain the functions of CD133+ laryngeal carcinoma stem cells under hypoxic and low-glucose conditions.

Differential glycolytic and glycogenogenic transduction pathways in male and female bovine embryos produced in vitro

2012 ◽  
Vol 24 (2) ◽  
pp. 344 ◽  
Author(s):  
M. Garcia-Herreros ◽  
I. M. Aparicio ◽  
D. Rath ◽  
T. Fair ◽  
P. Lonergan
Keyword(s):  
Glucose Metabolism ◽  
Protein Expression ◽  
Glucose Uptake ◽  
Glucose Transporter ◽  
Glycogen Synthase ◽  
Bovine Embryos ◽  
High Concentration ◽  
Glucose Transporter 1 ◽  
Male And Female ◽  
Glut 1

Previous studies have shown that developmental kinetic rates following IVF are lower in female than in male blastocysts and that this may be related to differences in glucose metabolism. In addition, an inhibition of phosphatidylinositol 3-kinase (PI3-K) inhibits glucose uptake in murine blastocysts. Therefore, the aim of this study was to identify and compare the expression of proteins involved in glucose metabolism (hexokinase-I, HK-I; phosphofructokinase-1, PFK-1; pyruvate kinase1/2, PK1/2; glyceraldehyde-3-phosphate dehydrogenase, GAPDH; glucose transporter-1, GLUT-1; and glycogen synthase kinase-3, GSK-3) in male and female bovine blastocysts to determine whether PI3-K has a role in the regulation of the expression of these proteins. Hexokinase-I, PFK-1, PK1/2, GAPDH and GLUT-1 were present in bovine embryos. Protein expression of these proteins and GSK-3 was significantly higher in male compared with female blastocysts. Inhibition of PI3-K with LY294002 significantly decreased the expression of HK-I, PFK-1, GAPDH, GSK-3 A/B and GLUT-1. Results showed that the expression of glycolytic proteins HK-I, PFK-1, GAPDH and PK1/2, and the transporters GLUT-1 and GSK-3 is regulated by PI3-K in bovine blastocysts. Moreover, the differential protein expression observed between male and female blastocysts might explain the faster developmental kinetics seen in males, as the expression of main proteins involved in glycolysis and glycogenogenesis was significantly higher in male than female bovine embryos and also could explain the sensitivity of male embryos to a high concentration of glucose, as a positive correlation between GLUT-1 expression and glucose uptake in embryos has been demonstrated.

Sign in / Sign up

Export Citation Format

Share Document