• Korean Journal of Veterinary Research
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• v.63 no.3
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• pp.30.1-30.8
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• 2023

Metformin is a treatment used widely for non-insulin-dependent diabetes mellitus with few side effects and acts by inhibiting hepatic gluconeogenesis and glucose absorption from the gastrointestinal tract. Lymphoma is one of the most common hematological malignancies in dogs. Chemotherapy is used mainly on lymphoma, but further research on developing anticancer drugs for lymphoma is needed because of its severe side effects. This study examined the anticancer effects of metformin alone and in combination with 2-deoxy-D-glucose (2-DG), a glucose analog, on EL4 cells (mouse T cell lymphoma). Metformin reduced the metabolic activity of EL4 cells and showed an additive effect when combined with 2-DG. In addition, cell death was confirmed using a trypan blue exclusion test, Hochest 33342/propidium iodide (PI) staining, and Annexin V/PI staining. An analysis of the cell cycle and mitochondria membrane potential (MMP) to investigate the mechanism of action showed that metformin stopped the G2/M phase of EL4 cells, and metformin + 2-DG decreased MMP. Metformin exhibited anticancer effects as a G2/M phase arrest mechanism in EL4 cells and showed additive effects when combined with 2-DG via MMP reduction. Unlike cytotoxic chemotherapeutic anticancer drugs, metformin and 2-DG are related to cellular glucose metabolism and have little toxicity. Therefore, metformin and 2-DG can be an alternative to reduce the toxicity caused by chemotherapeutic anticancer drugs. Nevertheless, research is needed to verify the in vivo efficacy of metformin and 2-DG before they can be used in lymphoma treatments.

• Journal of Physiology & Pathology in Korean Medicine
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• v.23 no.3
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• pp.581-589
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• 2009

The water extract of Dohongsamul-tang(DHSMT) has been traditionally used in treatment of ischemic heart and brain diseases in Oriental Medicine. However, little is known about the mechanism by which DHSMT protects C6 glial cells from glucose deprevation induced damages. Therefore, this study was designed to evaluate the protective effects of DHSMT on 2-deoxy-D-glucose induced autophagy of C6 glial cells. Autophagic phenotype is evaluated by fluorescence microscopy and flow cytometry with specific biological staining dyes, including monodansylcadaverine and acridine orange, as well as Western blot analysis with microtubule-associated protein 1 light chain 3(LC3) and Beclin-1. Treatment with 2-deoxy-D-glucose significantly resulted in a decrease of the viability of C6 glial cells and increase of the extracellular LDH release in a dose and time-dependent manner. However, pretreatment with DHSMT protected C6 glial cells from glucose deprivation with 2-deoxy-D-glucose. The author also observed the fact that autophagy phenotype occurred by 2-deoxy-D-glucose in C6 glial cells. Pretreatment with 3-MA, a pharmacological inhibitior of autophagy, abolished the formation of acidic vesicle organelle in C6 glial cells treated with 2-deoxy-D-glucose. However, pretreatment with DHSMT inhibited the formation of autophagic phenotypes, including formation of acidic vesicle organelle, and increase of the expression of LC-3 II Beclin-1 proteins in C6 glial cells treated with 2-deoxy-D-glucose. Taken together, these data suggest that DHSMT is able to protect C6 glial cells from glucose deprivation with marked inhibition of autophagy formation.

• BMB Reports
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• v.45 no.5
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• pp.317-322
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• 2012

2-deoxy-D-glucose(2DG)-caused endoplasmic reticulum (ER) stress inhibits protein phosphorylation at tyrosine residues. However, the accurate regulatory mechanisms, which determine the inflammatory response of chondrocytes to ER stress via protein tyrosine phosphorylation, have not been systematically evaluated. Thus, in this study, we examined whether protein phosphorylation at tyrosine residues can modulate the expression and glycosylation of COX-2, which is reduced by 2DG-induced ER stress. We observed that protein tyrosine phosphatase (PTP) inhibitors, sodium orthovanadate (SOV), and phenylarsine oxide (PAO) significantly decreased expression of ER stress inducible proteins, glucose-regulated protein 94 (GRP94), and CCAAT/ enhancer-binding-protein- related gene (GADD153), which was induced by 2DG. In addition, we demonstrated that SOV and PAO noticeably restored the expression and glycosylation of COX-2 after treatment with 2DG. These results suggest that protein phosphorylation of tyrosine residues plays an important role in the regulation of expression and glycosylation during 2DG-induced ER stress in rabbit articular chondrocytes.

• Imaging Science in Dentistry
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• v.39 no.3
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• pp.121-132
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• 2009

Purpose : To investigate the effects of 2-deoxy-D-glucose (2-DG) and quercetin (QCT) on gene expression of bone sialoprotein (BSP) and osteocalcin (OC) during the differentiation in irradiated MC3T3-E1 osteoblastic cells. Materials and Methods : When MC3T3-E1 osteoblastic cells had reached 70-80% confluence, cultures were transferred to a differentiating medium supplemented with 5 mM 2-DG or $10{\mu}M$ QCT, and then irradiated with 2, 4, 6, and 8 Gy. At various times after irradiation, the cells were analyzed for the synthesis of type I collagen, and expression of BSP and OC. Results : The synthesis of type I collagen in cells exposed to 2 Gy of radiation in the presence of 2-DG or QCT showed no significant difference compared with the control group within 15 days post-irradiation. When the cells were irradiated with 8 Gy, 2-DG facilitated the irradiation mediated decrease of type I collagen synthesis, whereas such decrease was inhibited by treating with QCT. During MC3T3-E1 osteoblastic cell differentiation, the mRNA expression of BSP and OC showed the peak value at 14 days and 21 days, respectively. 2-DG or QCT treatment alone decreased the level of BSP mRNA, but increased the OC mRNA level only at early time of differentiation (day 7). In the cells irradiated with 2, 4, 8 Gy, the mRNA expression of BSP and OC decreased at 7 days after the irradiation. The cells were treated with various dose of radiation in the presence of 2-DG or QCT, the mRNA level of both BSP and OC increased although this increase was observed at low dose of radiation (2 Gy) and at the early stage of differentiation. However, when the cells were exposed to 4, 6, or 8 Gy, the increase of BSP and OC mRNAs was detected only in cells co-incubated with QCT. Conclusion : This study demonstrates that 2-DG and QCT affect differently the expression of bone formation related factors, type I collagen, BSP, and OC in the irradiated MC3T3-E1 osteoblasic cells, according to the dose of radiation and the times of differentiation. Overall, the present findings suggest that 2-DG and QCT could have the regulatory roles as radiation-sensitizer and -protector, respectively.

• Imaging Science in Dentistry
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• v.36 no.4
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• pp.189-198
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• 2006

Purpose: To investigate the in vitro response of MC3T3-E1 osteoblastic cells to X-ray in the presence and absence of 2 deoxy-D-glucose (2-DG) and quercetin (QCT). Materials and Methods: The MC3T3-E1 cells were cultured in an ${\alpha}-MEM$ supplemented with 5 mM 2-DG or $10{\mu}M$ QCT and then the cells were incubated for 12 h prior to irradiation with 2, 4, 6, and 8Gy using a linear accelerator (Mevaprimus, Germany) delivered at a rate of 1.5 Gy/min. At various times after the irradiation, the cells were processed for the analyses of proliferation, viability, cytotoxicity, and mineralization. Results: Exposure of the cells to X-ray inhibited the tritium incorporation, 3-(4, 5-dimethylthiazol-2yl-)-2, 5-diphenyl tetrazolium bromide (MTT)-reducing activity, and alkaline phosphatase (ALP) activity, and caused cytotoxicity and apoptosis in a dose-dependent manner of the X-ray. This effect was further apparent on day 3 and 7 after the irradiation. RA+2-DG showed the decrease of DNA content, cell viability, and increase of cytotoxicity rather than RA. ALP activity increased on day 7 and subsequently its activity dropped to a lower level. 2-DG suppressed the calcium concentration, but visual difference of number of calcified nodules between RA and RA+2-DG was not noticed. RA+QCT showed the increase of DNA content, cell viability, but decrease of cytotoxicity and subG1 stage cells in the cell cycle, and increased calcified nodules in von Kossa staining rather than the RA. ALP activity showed significant increases on day 7 and subsequently its activity dropped to a lower level. Conclusion: The results showed that the 2-DG acted as a radiosensitizing agent and QCT acted as a radiosensitizing agent respectively in the irradiated MC3T3-E1 osteoblast-like cells.

• BMB Reports
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• v.37 no.2
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• pp.239-245
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• 2004

We have previously demonstrated that the redox reactant pyruvate prevents apoptosis in the oxidant model of bovine pulmonary artery endothelial cells (BPAEC), and that the anti-apoptotic mechanism of pyruvate is mediated in part via the mitochondrial matrix compartment. However, cytosolic mechanisms for the cytoprotective feature of pyruvate remain to be elucidated. This study investigated the pyruvate protection against endothelial cytotoxicity when the glycolysis inhibitor 2-deoxy-D-glucose (2DG) was applied to BPAEC. Millimolar 2DG blocked the cellular glucose uptake in a concentration- and time-dependent manner with >85% inhibition at $\geq$5 mM within 24 h. The addition of 2DG evoked BPAEC cytotoxicity with a substantial increase in lipid peroxidation and a marked decrease in intracellular total glutathione. Exogenous pyruvate partially prevented the 2DG-induced cell damage with increasing viability of BPAEC by 25-30%, and the total glutathione was also modestly increased. In contrast, 10 mM L-lactate, as a cytosolic reductant, had no effect on the cytotoxicity and lipid peroxidation that are evoked by 2DG. These results suggest that 2DG toxicity may be a consequence of the diminished potential of glutathione antioxidant, which was partially restored by exogenous pyruvate but not L-lactate. Therefore, pyruvate qualifies as a cytoprotective agent for strategies that attenuate the metabolic dysfunction of the endothelium, and cellular glucose oxidation is required for the functioning of the cytosolic glutathione/NADPH redox system.

• Imaging Science in Dentistry
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• v.38 no.4
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• pp.195-202
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• 2008

Purpose : To characterize the effects of 2-deoxy-D-glucose (2-DG) and quercetin (QCT) on gene expression of osteonectin (ON) and osteopontin (OP) in irradiated MC3T3-El cells. Materials and Methods : When MC3T3-El osteoblastic cells had reached 70-80% confluence, cultures were transferred to a differentiating medium supplemented with 5 mM 2-DG or 10 ${\mu}M$ QCT and then irradiated with 2, 4, 6, and 8 Gy. At various times after irradiation, the cells were analyzed for the expression of bone mineralization genes such as ON and OP. Results : The mRNA expression of both ON and OP was increased according to the culture time in the differentiation medium, and the increase of the genes peaked at 14 days after the differentiation induction. In the case of OP, the increase of mRNA expression was maintained to 28 days after the differentiation, while the mRNA level of ON was reduced to the basal level at the same time. Irradiation adding 2-DG showed a significant peak value in the expression pattern of ON at 4 Gy 7 days after irradiation. Irradiation adding QCT increased the mRNA expression of ON and OP in a dose-dependant manner, but irradiation adding 2-DG did not show any differences between the control and experiments 14 days after irradiation. Irradiation adding QCT increased significantly the expression patterns of ON 21 days after irradiation. Conclusion : The results showed that QCT acted as a radiosensitizer in the gene expression of ON and OP during differentiation of the late stage of irradiated MC3T3-E1 osteoblastic cells in vitro. (Korean J Oral Maxillofac Radiol 2008; 38: 195-202)

• Asian Pacific Journal of Cancer Prevention
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• v.16 no.18
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• pp.8431-8438
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• 2016

Doxorubicin (DOX) was introduced as an effective chemotherapeutic for a wide range of cancers but with some severe side effects especially on myocardia. 2-Deoxy-D-glucose (2DG) enhances the damage caused by chemotherapeutics and ionizing radiation (IR) selectively in cancer cells. We have studied the effects of $1{\mu}M$ DOX and $500{\mu}M$ 2DG on radiation induced cell death, apoptosis and also on the expression levels of p53 and PTEN genes in T47D and SKBR3 breast cancer cells irradiated with 100, 150 and 200 cGy x-rays. DOX and 2DG treatments resulted in altered radiation-induced expression levels of p53 and PTEN genes in T47D as well as SKBR3 cells. In addition, the combination along with IR decreased the viability of both cell lines. The radiobiological parameter (D0) of T47D cells treated with 2DG/DOX and IR was 140 cGy compared to 160 cGy obtained with IR alone. The same parameters for SKBR3 cell lines were calculated as 120 and 140 cGy, respectively. The sensitivity enhancement ratios (SERs) for the combined chemo-radiotherapy on T47D and SKBR3 cell lines were 1.14 and 1.16, respectively. According to the obtained results, the combination treatment may use as an effective targeted treatment of breast cancer either by reducing the single modality treatment side effects.

• Biomedical Science Letters
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• v.15 no.2
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• pp.113-118
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• 2009

Actin cytoskeletal architecture is believed to be a crucially important modulator of chondrocyte phenotype. 2DG(2-Dexoy-D-glucose) induces reorganization of actin cytoskeletal architecture in chondrocytes. In this study, we have investigated the effects of 2DG on dedifferentiation and inflammation via reorganization of cytoskeletal architecture in rabbit articular chondrocytes, with a focus on p38 kinase pathway. Treatment of 2DG alone reduced type II collagen and COX-2 expression in chondrocytes. But, 2DG reduced type II collagen was recovered by CD, disruptor of actin cytoskeletal architecture, whereas did not affect on COX-2 expression and production of $PGE_2$ compared with 2DG alone treated cells. Treatment of 2DG with JAS, inducer of cytoskeletal architecture polymerization, accelerated reduction of type II collagen expression and synthesis of proteoglycan but did not affect on COX-2 expression and production of $PGE_2$. Also, 2DG stimulated activation of p38 kinase. This result showed that 2DG regulates type II collagen but not cyclooxygenase-2 expression through reorganization of cytoskeletal architecture via p38 kinase pathway in rabbit articular chondrocytes.

• Imaging Science in Dentistry
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• v.35 no.4
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• pp.191-198
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• 2005

Purpose: To characterize the effects of 2-deoxy-D-glucose (2DG) and quercetin (QCT) on cytokine secretion of IL-6, $TGF-\beta$ and gene expression of Col I in irradiated MC3T3-E1 cells Materials and Methods: The MC3T3-El cells were cultured in an a-MEM supplemented with 5mM 2DG or 10mM QCT and then the cells were incubated 12h before irradiation with 2, 4, 6, and 8Gy X-ray using a linear accelerator delivered at a dose rate of 1.5Gy/min. Level of IL-6 and $TGF-\beta$ was determined by ELISA. Also expression of Col I was examined by RT-PCR. Results: In accordance with the radiation dose, the amount of $TGF-\beta$ was not different in RA + QCT, but it showed a peak value in control and RA + 2DG at 4Gy on the 3rd day. However, all groups showed a decreasing tendency dose-dependently in RA+QCT on the 7th day (p<0.01). In accordance with the radiation dose, the amount of IL-6 increased dose-dependently in all groups on the 3rd day. On the 7th and 21st day, all groups showed peak values at 4Gy. RA+QCT showed a slightly increased amount of IL-6 at 2Gy, but it showed a slightly decreased amount at 4, 6, and 8Gy. In accordance with the period of culture after irradiation, the expression of Col I increased dose-dependently in RA+QCT. Conclusion: The result showed that QCT acted as radiosensitizer in the secretion of $TGF-\beta$ and gene expression of Col I during differentiation in irradiated MC3T3-E1 cells at the cellular level.