Chloride channel protein 2 prevents glutamate-induced apoptosis in retinal ganglion cells

Document Type : Original Article

Authors

1 Department of Ophthalmology, The Second Hospital of Jilin University, Jilin University, Changchun, Jilin, 130041, P.R. China

2 Department of Ophthalmology, The China-Japan Union Hospitial of Jilin University, Jilin University, Changchun, Jilin, 130033, Xiantai Street No. 126, Jilin Province, China

3 Department of Colon and Anal Surgery, the First hospital of Jilin University, Jilin University, Changchun, Jilin, 130021, P.R. China

Abstract

Objective(s): The purpose of this study was to investigate the role of chloride channel protein 2 (ClC-2) in glutamate-induced apoptosis in the retinal ganglion cell line (RGC-5).
Materials and Methods: RGC-5 cells were treated with 1 mM glutamate for 24 hr. The expression of ClC-2, Bax, and Bcl-2 was detected by western blot analysis. Cell survival and apoptosis were measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and flow cytometry assays, respectively. Caspase-3 and -9 activities were determined by a colorimetric assay. The roles of ClC-2 in glutamate-induced apoptosis were examined by using ClC-2 complementary deoxyribonucleic acid (cDNA) and small inference ribonucleic acid (RNA) transfection technology.
Results: Overexpression of ClC-2 in RGC-5 cells significantly decreased glutamate-induced apoptosis and increased cell viability, whereas silencing of ClC-2 with short hairpin (sh) RNA produced opposite effects. ClC-2 overexpression increased the expression of Bcl-2, decreased the expression of Bax, and decreased caspase-3 and -9 activation in RGC-5 cells treated with glutamate, but silencing of ClC-2 produced opposite effects.
Conclusion: Our data suggest that ClC-2 chloride channels might play a protective role in glutamate-induced apoptosis in retinal ganglion cells via the mitochondria-dependent apoptosis pathway.

Keywords

References

1. Nickells RW: Apoptosis of retinal ganglion cells in glaucoma: an update of the molecular pathways involved in cell death. Surv Ophthalmol 1999, 43 Suppl 1:S151-161.
2. Sucher NJ, Lipton SA, Dreyer EB: Molecular basis of glutamate toxicity in retinal ganglion cells. Vision Res 1997, 37:3483-3493.
3. Fang JH, Wang XH, Xu ZR, Jiang FG. Neuroprotective effects of bis(7)-tacrine against glutamate-induced retinal ganglion cells damage. BMC Neurosci 2010; 11:31.
4. Krishnamoorthy RR, Agarwal P, Prasanna G, Vopat K, Lambert W, Sheedlo HJ, et al. Characterization of a transformed rat retinal ganglion cell line. Brain Res Mol Brain Res 2001; 86:1-12.
5. Pang IH, Clark AF. Rodent models for glaucoma retinopathy and optic neuropathy. J Glaucoma 2007; 16:483-505.
6.Ju WK, Kim KY, Angert M, Duong-Polk KX, Lindsey JD, Ellisman MH, et al. Memantine blocks mitochondrial OPA1 and cytochrome c release and subsequent apoptotic cell death in glaucomatous retina. Invest Ophthalmol Vis Sci 2009; 50:707-716.
7.Souktani R, Ghaleh B, Tissier R, d'Anglemont de Tassigny A, Aouam K, Bedossa P, et al. Inhibitors of swelling-activated chloride channels increase infarct size and apoptosis in rabbit myocardium. Fundam Clin Pharmacol 2003; 17:555-561.
8. Wei L, Xiao AY, Jin C, Yang A, Lu ZY, Yu SP. Effects of chloride and potassium channel blockers on apoptotic cell shrinkage and apoptosis in cortical neurons. Pflugers Arch 2004; 448:325-334.
9. Lui VC, Lung SS, Pu JK, Hung KN, Leung GK. Invasion of human glioma cells is regulated by multiple chloride channels including ClC-3. Anticancer Res 2010; 30:4515-4524.
10.Qian Y, Du YH, Tang YB, Lv XF, Liu J, Zhou JG, et al. ClC-3 chloride channel prevents apoptosis induced by hydrogen peroxide in basilar artery smooth muscle cells through mitochondria dependent pathway. Apoptosis 2011; 16:468-477.
11. Yu L, Han N, Jiang LG, Zheng YJ, Liu LF. Neuroprotective effects of ClC-3 chloride channel in glutamate-induced retinal ganglion cell RGC-5 apoptosis. Neural Regen Res 2011; 6:450-456.
12. Britton FC, Hatton WJ, Rossow CF, Duan D, Hume JR, Horowitz B. Molecular distribution of volume-regulated chloride channels (ClC-2 and ClC-3) in cardiac tissues. Am J Physiol Heart Circ Physiol 2000; 279:H2225-2233.
13. Walshe TE, Leach LL, D'Amore PA. TGF-beta signaling is required for maintenance of retinal ganglion cell differentiation and survival. Neuroscience 2011; 189:123-131.
14. Fan W, Agarwal N, Kumar MD, Cooper NG. Retinal ganglion cell death and neuroprotection: Involvement of the CaMKIIalpha gene. Brain Res Mol Brain Res 2005; 139:306-316.
15. Kumar DM, Perez E, Cai ZY, Aoun P, Brun-Zinkernagel AM, Covey DF, et al. Role of nonfeminizing estrogen analogues in neuroprotection of rat retinal ganglion cells against glutamate-induced cytotoxicity. Free Radic Biol Med 2005; 38:1152-1163.
16. Wang SJ, Xie LH, Heng B, Liu YQ. Classification of potassium and chlorine ionic currents in retinal ganglion cell line (RGC-5) by whole-cell patch clamp. Vis Neurosci 2012; 29:275-282.
17. Enz R, Ross BJ, Cutting GR. Expression of the voltage-gated chloride channel ClC-2 in rod bipolar cells of the rat retina. J Neurosci 1999; 19:9841-9847.
18. Lemonnier L, Shuba Y, Crepin A, Roudbaraki M, Slomianny C, Mauroy B, et al. Bcl-2-dependent modulation of swelling-activated Cl- current and ClC-3 expression in human prostate cancer epithelial cells. Cancer Res 2004; 64:4841-4848.
19. Harder JM, Fernandes KA, Libby RT. The Bcl-2 family member BIM has multiple glaucoma-relevant functions in DBA/2J mice. Sci Rep 2012; 2:530.
20. Meng Q, Lv J, Ge H, Zhang L, Xue F, Zhu Y, et al. Overexpressed mutant optineurin(E50K) induces retinal ganglion cells apoptosis via the mitochondrial pathway. Mol Biol Rep 2012; 39:5867-5873.
21. Varma R, Peeples P, et al. Disease progression and the need for neuroprotection in glaucoma manage-ment. Am J Manage Care. 2008; 14(1 Suppl): S15-19.
22. Nickells RW. Ganglion cell death in glaucoma: from mice to men. Vet Ophthalmol. 2007; Suppl 1:88-94.
23. Xu Y, Zheng H, Kang JS, Zhang L, Su J, Li HY, et al. 5-Nitro-2-(3-phenylpropylamino) benzoic acid induced drug resistance to cisplatin in human erythroleukemia cell lines. Anat Rec (Hoboken) 2011; 294:945-952.
24. Lang F, Shumilina E, Ritter M, Gulbins E, Vereninov A, Huber SM. Ion channels and cell volume in regulation of cell proliferation and apoptotic cell death. Contrib Nephrol 2006; 152:142-160.
25. Martinou JC, Youle RJ. Mitochondria in apoptosis: Bcl-2 family members and mitochondrial dynamics. Dev Cell 2011; 21:92-101.
26. Youle RJ, Strasser A. The BCL-2 protein family: opposing activities that mediate cell death. Nat Rev Mol Cell Biol 2008; 9:47-59.
27. XJ Jiang, XD Wang. Cytochrome c-mediated apoptosis, Annu. Rev. Biochem. 73 (2004) 87-106.
28. Li Z, Jo J, Jia JM, Lo SC, Whitcomb DJ, Jiao S, et al. Caspase-3 activation via mitochondria is required for long-term depression and AMPA receptor inter-nalization. Cell 2010; 141:859-871.
29. Yoshida A, Pommier Y, Ueda T. Endonuclease activation and chromosomal DNA fragmentation during apoptosis in leukemia cells. Int J Hematol 2006; 84:31-37.
Iranian Journal of Basic Medical Sciences
Volume 19, Issue 7
July 2016
Pages 705-711

Files

Share

How to cite

Statistics