생명과학회지 (Journal of Life Science)

  • 제27권11호
  • /
  • Pages.1349-1356
  • /
  • 2017
  • /
  • 1225-9918(pISSN)
  • /
  • 2287-3406(eISSN)
한국생명과학회 (Korean Society of Life Science)
권호 표지
DOI QR코드

DOI QR Code

망막 색소상피세포에서 산화성 세포 손상과 미토콘드리아기능 저해에 미치는 NS 1619의 보호 효과

The Ca2+-activated K+ (BK) Channel-opener NS 1619 Prevents Hydrogen Peroxide-induced Cell Death and Mitochondrial Dysfunction in Retinal Pigment Epithelial Cells

  • 강재훈 (부산대학교 의학전문대학원 생리학교실) ;
  • 우재석 (부산대학교 의학전문대학원 생리학교실)
  • Kang, Jae Hoon (Department of Physiology, Pusan National University School of Medicine) ;
  • Woo, Jae Suk (Department of Physiology, Pusan National University School of Medicine)
  • 투고 : 2017.09.13
  • 심사 : 2017.10.23
  • 발행 : 2017.11.30
PDF 다운로드
⟨ 이전 논문 다음 논문 ⟩

초록

$K^+$ 통로 개방제들은 심근, 뇌, 골격근 등에서 세포막 혹은 미토콘드리아 내막에 존재하는 큰 전도성의 $Ca^{2+}$-의 존성 $K^+$ (BK) 통로 및 ATP-조절성 $K^+$ 통로(ATP-sensitive $K^+$ channels, $K_{ATP}$)에 작용하여 허혈성 혹은 산화성 세포 손상을 완화하는 효과가 있는 것으로 보고되어 있다. 본 연구에서는 망막 색소 상피세포주인 ARPE-19 세포를 실험 모델로 하여 큰 전도성의 BK 통로 개방제인 NS 1619가 유사한 보호 효과를 나타낼 수 있는지, 또한 그 작용기전이 무엇인지를 확인하고자 하였다. AREE-19 세포를 여러 형태의 산화 스트레스에 노출시켜 세포 손상을 유발하고 그 손상의 정도 및 이에 미치는 NS 1619의 효과를 trypan blue 배출능, Tunel 염색 분석을 통하여 측정하였다. NS 1619는 여러 형태의 산화 스트레스에 의한 괴사성 및 apoptosis에 의한 세포 손상을 효과적으로 방지하였으며 그 보호 효과는 BK 통로 봉쇄제인 paxilline 의해 차단되었다. NS 1619는 $H_2O_2$에 의한 세포내 ATP 고갈을 현저히 완화시켰으며, 또한 MTT 환원능으로 측정한 미토콘드리아의 기능을 보호하는 효과를 보였다. 유세포형광 분석법을 이용한 실험에서 NS 1619는 $H_2O_2$에 의한 미토콘드리아 막전압의 소실을 유의하게 방지하였다. 이상의 결과들을 종합하면 NS 1619는 망막 색소 상피세포에서 산화성 세포 손상을 방지하는 효과를 나타내며 그 기전에 미토콘드리아 기능에 대한 보호 작용이 연관되어 있는 것으로 사료된다.

Potassium channel openers (KCOs) produce physiological and pharmacological defense mechanisms against cell injuries caused by oxidative stress of diverse origins. Openings of mitochondrial and plasmalemmal $K^+$ channels are involved in the defense mechanisms. This study tested whether NS 1619, an opener of large-conductance BK channels, has a similar beneficial influence on the pigment epithelial cells of retinas. The human retinal pigment epithelial cell line ARPE-19 was exposed to $H_2O_2$-induced oxidative stress in the absence and presence of NS 1619. The degrees of the cells' injuries were assessed by analyzing the cells' trypan-blue exclusion abilities and TUNEL staining. NS 1619 produced remarkable protections against cell injuries caused by $H_2O_2$. It prevented apoptotic and necrotic cell deaths. The protective effect of NS 1619 was significantly diminished when the cells were treated with NS 1619 in combination with the BK channel-blocker paxilline. NS 1619 significantly ameliorated cellular ATP deprivations in $H_2O_2$-treated cells. It helped mitochondria preserve their functional integrity, which was estimated by their MTT reduction abilities and mitochondrial membrane potential. In conclusion, it was suggested that NS 1619 had a beneficial effect on mitochondria in regards to preserving their functional integrity under oxidative stress, and it produces defense mechanisms against oxidant-induced cell injuries in ARPE-19 cells.

키워드

참고문헌

  1. Bentzen, B. H., Olesen, S. P., Ronn, L. C. and Grunnet, M. 2014. BK channel activators and their therapeutic perspectives. Front. Physiol. 5, 389.
  2. Bjorkerud, S. and Bondjers, G. 1972. Endothelial integrity and viability in the aorta of the normal rabbit and rat as evaluated with dye exclusion tests and interference contrast microscopy. Atherosclerosis 15, 285-300. https://doi.org/10.1016/0021-9150(72)90019-6
  3. Clarke, P. G. 1990. Developmental cell death: morphological diversity and multiple mechanisms. Anat. Embryol. 181, 195-213.
  4. Cole, W. C., McPherson, C. D. and Sontag, D. 1991. ATPregulated $K^{+}$ channels protect the myocardium against ischemia/reperfusion damage. Circ. Res. 69, 571-581. https://doi.org/10.1161/01.RES.69.3.571
  5. Daut, J., Maier-Rudolph, W., von Beckerath, N., Mehrke, G., Gunther, K. and Goedel-Meinen, L. 1990. Hypoxic dilation of coronary arteries is mediated by ATP-sensitive potassium channels. Science 247, 1341-1344. https://doi.org/10.1126/science.2107575
  6. Duty, S. and Weston, A. H. 1990. Potassium channel openers. Pharmacological effects and future uses. Drugs 40, 785-791. https://doi.org/10.2165/00003495-199040060-00002
  7. Edwards, G., Niederste-Hollenberg, A., Schneider, J., Noack, T., and Weston, A. H. 1994. Ion channel modulation by NS 1619, the putative BKCa channel opener, in vascular smooth muscle. Br. J. Pharmacol. 113, 1538-1547. https://doi.org/10.1111/j.1476-5381.1994.tb17171.x
  8. Edwards, G. and Weston, A. H. 1990. Structure-activity relationships of $K^{+}$ channel openers. Trends. Pharmacol. Sci. 11, 417-422. https://doi.org/10.1016/0165-6147(90)90149-3
  9. Farber, J. L., Kyle, M. E. and Coleman, J. B. 1990. Mechanisms of cell injury by activated oxygen species. Lab. Invest. 62, 670-679.
  10. Foster, C. D., Fujii, K., Kingdon, J. and Brading, A. F. 1989. The effect of cromakalim on the smooth muscle of the guinea-pig urinary bladder. Br. J. Pharmacol. 97, 281-291. https://doi.org/10.1111/j.1476-5381.1989.tb11952.x
  11. Gaspar, T., Katakam, P., Snipes, J. A., Kis, B., Domoki, F., Bari, F. and Busija, D. W. 2008. Delayed neuronal preconditioning by NS1619 is independent of calcium activated potassium channels. J. Neurochem. 105, 1115-1128. https://doi.org/10.1111/j.1471-4159.2007.05210.x
  12. Gunter, T. E., Gunter, K. K., Sheu, S. S. and Gavin, C. E. 1994 Mitochondrial calcium transport: physiological and pathological relevance. Am. J. Physiol. 267, C313-C339. https://doi.org/10.1152/ajpcell.1994.267.2.C313
  13. Hagar, H., Ueda, N. and Shah, S. V. 1996. Endonuclease induced DNA damage and cell death in chemical hypoxic injury to LLC-PK1 cells. Kidney Int. 49, 355-361. https://doi.org/10.1038/ki.1996.52
  14. Inoue, I., Nagase, H., Kishi, K. and Higuti, T. 1991. ATP-sensitive $K^{+}$ channel in the mitochondrial inner membrane. Nature 352, 244-247. https://doi.org/10.1038/352244a0
  15. Kersten, J. R., Gross, G. J., Pagel, P. S. and Warltier, D. C. 1998. Activation of adenosine triphosphate-regulated potassium channels: mediation of cellular and organ protection. Anesthesiology 88, 495-513. https://doi.org/10.1097/00000542-199802000-00029
  16. Latorre, R., Castillo, K., Carrasquel-Ursulaez, W., Sepulveda, R. V., Gonzalez-Nilo, F., Gonzalez, C. and Alvarez, O. 2017. Molecular determinants of BK channel functional diversity and functioning. Physiol. Rev. 97, 39-87. https://doi.org/10.1152/physrev.00001.2016
  17. Lemasters, J. J., Nieminen, A. L., Qian, T., Trost, L. C., Elmore, S. P., Nishimura, Y., Crowe, R. A., Cascio, W. E., Bradham, C. A., Brenner, D. A. and Herman, B. 1998. The mitochondrial permeability transition in cell death: a common mechanism in necrosis, apoptosis and autophagy. Biochim. Biophys. Acta 1366, 177-196. https://doi.org/10.1016/S0005-2728(98)00112-1
  18. Lyman, G. E. and DeVincenzo, J. P. 1967. Determination of picogram amounts of ATP using the luciferin-luciferase enzyme system. Anal. Biochem. 21, 435-443. https://doi.org/10.1016/0003-2697(67)90318-1
  19. Morgan, D. M. 1998. Tetrazolium (MTT) assay for cellular viability and activity. Methods. Mol. Biol. 79, 179-183.
  20. Murray, M. A., Boyle, J. P. and Small, R. C. 1989. Cromakalim-induced relaxation of guinea-pig isolated trachealis: antagonism by glibenclamide and by phentolamine. Br. J. Pharmacol. 98, 865-874. https://doi.org/10.1111/j.1476-5381.1989.tb14615.x
  21. Murry, C. E., Jennings, R. B. and Reimer, K. A. 1986. Preconditioning with ischemia: a delay of lethal cell injury in ischemic myocardium. Circulation 74, 1124-1136. https://doi.org/10.1161/01.CIR.74.5.1124
  22. Robertson, D. W. and Steinberg, M. I. 1990. Potassium channel modulators: scientific applications and therapeutic promise. J. Med. Chem. 33, 1529-1541. https://doi.org/10.1021/jm00168a001
  23. Schwartzman, R. A. and Cidlowski, J. A. 1993. Apoptosis: the biochemistry and molecular biology of programmed cell death. Endocr. Rev. 14, 133-151.
  24. Sparrow, J. R., Hicks, D. and Hamel, C. P. 2010. The retinal pigment epithelium in health and disease. Curr. Mol. Med. 10, 802-823. https://doi.org/10.2174/156652410793937813
  25. Standen, N. B., Quayle, J. M., Davies, N. W., Brayden, J. E., Huang, Y. and Nelson, M. T. 1989. Hyperpolarizing vasodilators activate ATP-sensitive $K^{+}$ channels in arterial smooth muscle. Science 245, 177-180. https://doi.org/10.1126/science.2501869
  26. Stern, J. and Temple, S. 2015. Retinal pigment epithelial cell proliferation. Exp. Biol. Med (Maywood). 240, 1079-1986. https://doi.org/10.1177/1535370215587530
  27. Szewczyk, A. and Marban, E. 1999. Mitochondria: a new target for $K^{+}$ channel openers? Trends Pharmacol. Sci. 20, 157-161. https://doi.org/10.1016/S0165-6147(99)01301-2
  28. Testai, L., Rapposelli, S. and Calderone, V. 2007. Cardiac ATP- sensitive potassium channels: a potential target for an anti-ischaemic pharmacological strategy. Cardiovasc. Hematol. Agents. Med. Chem. 5, 79-90. https://doi.org/10.2174/187152507779315831
  29. Testai, L., Rapposelli, S., Martelli, A., Breschi, M. C. and Calderone, V. 2015. Mitochondrial potassium channels as pharmacological target for cardioprotective drugs. Med. Res. Rev. 35, 520-553. https://doi.org/10.1002/med.21332
  30. Vakifahmetoglu-Norberg, H. Ouchida, A. T. and Norberg, E. 2017. The role of mitochondria in metabolism and cell death. Biochem Biophys. Res. Commun. 482, 426-431. https://doi.org/10.1016/j.bbrc.2016.11.088
  31. Wimmers, S., Karl, M. O. and Strauss, O. 2007. Ion channels in the RPE. Prog. Retin. Eye. Res. 26, 263-301. https://doi.org/10.1016/j.preteyeres.2006.12.002