Influence of ataxia-telangiectasia mutated activation on cellular oxidative stress induced by high glucose in bovine retinal capillary endothelial cells_Chinese Journal of Ocular Fundus Diseases

Authors：

LuLi , ZhengZhi , LiJingwen , 肖文玮 ,  李春霞

Bengbu Medical College, Bengbu 233030, China;

Corresponding author：

李春霞, Email: cxli-66@hotmail.com

Keywords：

Oxidative Stress/immunology; Endothelial cells/enzymology; Animal experimentation

DOI：

10.3760/cma.j.issn.1005-1015.2014.06.016

Video：

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Objective To investigate the influence of Ataxia-telangiectasia mutated (ATM) activation on cellular oxidative stress induced by high glucose in bovine retinal capillary endothelial cells(BRECs). Methods The BRECs were treated by different culture medium with various glucose concentrations (5 mmol/L glucose, 30 mmol/L glucose, 30 mmol/L glucose+10 μmol/L KU55933) as normal glucose group, high glucose group and treatment group respectively.After the cells incubated for 48 hours, the protein expression of ATM, P-ATM, Mitogen-Activated Protein Kinase P38(P38), P-P38, Extracellular signal-regulated kinases(ERKs), P-ERKs was detected by Western blot; cellular ROS level was detected by Reactive Oxygen Species Assay Kit; propidium iodide/Hoechst staining was used for analysis of apoptosis; the expression of vascular endothelial growth factor (VEGF) in the supernatant was determined by Enzyme-Linked Immunosorbent Assay (ELISA); the paracellular permeability between endothelium cells was detected by FITC-dextran. Results Compared with the protein level of P-ATM, P-P38 and P-ERKs in high glucose group increased. Especially, P-P38, P-ERKs expressed much more than in high glucose group. The secretion of VEGF in high glucose group was higher than that in the normal glucose group but less than that in treatment group. The same tendency existed in ROS assay, apoptosis assay and paracellular permeability measuring. Conclusions High glucose induced altered activation of ATM which might play a protective role in cellular oxidative stress. Deficiency of ATM might lead to ROS explosion, cell apoptosis and dysfunction of endothelial barrier. The mechanism might be associated with P38, ERKs and VEGF.

Citation： LuLi, ZhengZhi, LiJingwen. Influence of ataxia-telangiectasia mutated activation on cellular oxidative stress induced by high glucose in bovine retinal capillary endothelial cells. Chinese Journal of Ocular Fundus Diseases, 2014, 30(6): 604-607. doi: 10.3760/cma.j.issn.1005-1015.2014.06.016 Copy

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7.	Allegra M, D'Acquisto F, Tesoriere L, et al. Cross-talk between minimally primed HL-60 cells and resting HUVEC reveals a crucial role for adhesion over extracellularly released oxidants[J]. Biochem Pharmacol, 2011, 81:396-401.
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9.	Tian X, Sui S, Huang J, et al. Neuroprotective effects of Arctium lappa L. roots against glutamate-induced oxidative stress by inhibiting phosphorylation of p38, JNK and ERK 1/2 MAPKs in PC12 cells[J]. Environ Toxicol Pharmacol, 2014, 38:189-198.
10.	Kim J, Wong PKY. Loss of ATM impairs proliferation of neural stem cells through oxidative stress-mediated p38 MAPK signaling[J]. Stem Cells, 2009, 27:1987-1998.
11.	Kim J, Wong PKY. Oxidative stress is linked to ERK1/2-p16 signaling-mediated growth defect in ATM-deficient astrocytes[J]. J Biol Chem, 2009, 284:14396-14404.
12.	Cui Y, Xu X, Bi H, et al. Expression modification of uncoupling proteins and MnSOD in retinal endothelial cells and pericytes induced by high glucose:the role of reactive oxygen species in diabetic retinopathy[J]. Exp Eye Res, 2006, 83:807-816.
13.	Weis SM, Cheresh DA. Pathophysiological consequences of VEGF-induced vascular permeability[J]. Nature, 2005, 437:497-504.

1. Kowluru RA, Chan PS. Oxidative stress and diabetic retinopathy. Exp Diabetes Res, 2007, 2007:43603.[2007-02-08] http://www.hindawi.com/journals/jdr/2007/043603/abs/.
2. Kanwar M, Chan PS, Kern TS, et al. Oxidative damage in the retinal mitochondria of diabetic mice:possible protection by superoxide dismutase[J]. Invest Ophthalmol Vis Sci, 2007, 48:3805-3811.
3. Le Bras M, Clement M, Pervaiz S, et al. Reactive oxygen species and the mitochondrial signaling pathway of cell death[J]. Histol Histopathol, 2005, 20:205-219.
4. Guo Z, Kozlov S, Lavin MF, et al. ATM activation by oxidative stress[J]. Science, 2010, 330:517-521.
5. Ousset M, Bouquet F, Fallone F, et al. Loss of ATM positively regulates the expression of hypoxia inducible factor 1 (HIF-1) through oxidative stress:role in the physiopathology of the disease[J]. Cell Cycle, 2010, 9:2886-2894.
6. Pesant M, Sueur S, Dutartre P, et al. Peroxisome proliferator-activated receptor δ (PPARδ) activation protects H9c2 cardiomyoblasts from oxidative stress-induced apoptosis[J]. Cardiovasc Res, 2006, 69:440-449.
7. Allegra M, D'Acquisto F, Tesoriere L, et al. Cross-talk between minimally primed HL-60 cells and resting HUVEC reveals a crucial role for adhesion over extracellularly released oxidants[J]. Biochem Pharmacol, 2011, 81:396-401.
8. Lu TH, Hsieh SY, Yen CC, et al. Involvement of oxidative stress-mediated ERK1/2 and p38 activation regulated mitochondria-dependent apoptotic signals in methylmercury-induced neuronal cell injury[J]. Toxicol Lett, 2011, 204:71-80.
9. Tian X, Sui S, Huang J, et al. Neuroprotective effects of Arctium lappa L. roots against glutamate-induced oxidative stress by inhibiting phosphorylation of p38, JNK and ERK 1/2 MAPKs in PC12 cells[J]. Environ Toxicol Pharmacol, 2014, 38:189-198.
10. Kim J, Wong PKY. Loss of ATM impairs proliferation of neural stem cells through oxidative stress-mediated p38 MAPK signaling[J]. Stem Cells, 2009, 27:1987-1998.
11. Kim J, Wong PKY. Oxidative stress is linked to ERK1/2-p16 signaling-mediated growth defect in ATM-deficient astrocytes[J]. J Biol Chem, 2009, 284:14396-14404.
12. Cui Y, Xu X, Bi H, et al. Expression modification of uncoupling proteins and MnSOD in retinal endothelial cells and pericytes induced by high glucose:the role of reactive oxygen species in diabetic retinopathy[J]. Exp Eye Res, 2006, 83:807-816.
13. Weis SM, Cheresh DA. Pathophysiological consequences of VEGF-induced vascular permeability[J]. Nature, 2005, 437:497-504.

Chinese Journal of Ocular Fundus Diseases

Influence of ataxia-telangiectasia mutated activation on cellular oxidative stress induced by high glucose in bovine retinal capillary endothelial cells

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