송절(松節) 약침액이 자유기와 금속 이온으로 유도된 인체 저밀도 지단백질의 산화 반응에 미치는 효과

The Effect of Pinus Densiflora Gnarl Extract for Pharmacopuncture on Human LDL Oxidation Induced by Free Radical and Metal Ion

  • 임선희 (동국대학교 한의과대학 경혈학교실) ;
  • 이강파 (동국대학교 과학기술대학 생명과학과) ;
  • 문진영 (동국대학교 한의과대학 경혈학교실)
  • Leem, Sun-Hee (Dept. of Meridian & Acupoint, College of Oriental Medicine, Dong-Guk University) ;
  • Lee, Kang-Pa (Division of Bio Science, College of Science and Technology, Dong-Guk University) ;
  • Moon, Jin-Young (Dept. of Meridian & Acupoint, College of Oriental Medicine, Dong-Guk University)
  • 투고 : 2011.05.18
  • 심사 : 2011.06.16
  • 발행 : 2011.06.27

초록

목적 : 이 연구는 관절 및 심혈관계 질환 치료에 사용되는 송절(松節)(Pinus densiflora Gnarl)을 약침용 시료로 조제하여 본 약물의 항산화 효능을 규명하고자 하였으며 이를 다양한 시스템에서 검토하였다. 방법 : $FeCl_2$-ascorbic acid system에서 흰쥐 간조직의 지질과산화 반응을 관찰하였고, Fenton reaction system에서 자유기에 의한 plasmid DNA 분절을 유도하였다. 또한 deoxyribose assay를 통해 hydroxyl radical 소거능을 관찰하였고, NBT reduction assay로 superoxide radical 소거능을 검토하였다. 또한 human low-density lipoprotein(LDL)의 산화를 유도하기 위해 $CuSO_4$와 AAPH를 사용하였으며 relative electrophoretic mobility (REM) assay로 LDL 산화 억제 효능을 대조 항산화물질과 비교 검토하였다. 결과 : 송절 약침액은 자유기에 의한 간조직의 지질과산화(p < 0.01)및 DNA 분절을 현저하게 억제하였으며, hydroxyl radical, superoxide radical (p < 0.01), nitric oxide 및 peroxynitrite를 강하게 소거하였다. 또한 $CuSO_4$ ($IC_{50}=9.2{\pm}0.2\;{\mu}g/ml$)와 AAPH ($IC_{50}=34.8{\pm}5.1\;{\mu}g/ml$)에 의해 유도된 human LDL의 산화를 억제하였고, REM assay에서도 산화 억제 효능을 재확인할 수 있었다. 결론 : 송절 약침액은 활성산소종 및 활성질소종를 소거하였고, 지질과산화를 억제하였으며, 특히 human LDL의 산화적 손상을 방어하였다. 이에 본 약물은 자유기에 의한 심혈관의 산화적 손상을 효과적으로 보호할 것으로 판단된다.

키워드

참고문헌

  1. Kwak CS, Moon SC, Lee MS. Antioxidant, antimutagenic, and antitumor effects of pine needles (Pinus densiflora). Nutrition and cancer. 2006 ; 56 : 162-71. https://doi.org/10.1207/s15327914nc5602_7
  2. Choi EM. Antinociceptive and antiinflammatory activities of pine (Pinus densiflora) pollen extract. Phytotherapy research. 2007 ; 21 : 471-5. https://doi.org/10.1002/ptr.2103
  3. Hur J. Dong-eui-bo-gam. Seoul : Bupin Publishing. 2005 : 522-3.
  4. Minuz P, Fava C, Lechi A. Lipid peroxidation, isoprostanes and vascular damage. Pharmacological reports. 2006 ; 58 : 57-68.
  5. Dandona P, Ghanim H, Brooks DP. Antioxidant activity of carvedilol in cardiovascular disease. Journal of hypertension. 2007 ; 25 : 731-41. https://doi.org/10.1097/HJH.0b013e3280127948
  6. Trostchansky A, Batthyany C, Botti H, Radi R, Denicola A, Rubbo H. Formation of lipid-protein adducts in low-density lipoprotein by fluxes of peroxynitrite and its inhibition by nitric oxide. Archives of biochemistry and biophysics. 2001 ; 395 : 225-32. https://doi.org/10.1006/abbi.2001.2583
  7. Asmis R, Begley JG, Jelk J, Everson WV. Lipoprotein aggregation protects human monocyte-derived macrophages from OxLDL-induced cytotoxicity. Journal of lipid research. 2005 ; 46 : 1124-32. https://doi.org/10.1194/jlr.M400485-JLR200
  8. de Vries HE, Buchner B, van Berkel TJ, Kuiper J. Specific interaction of oxidized low-density lipoprotein with macrophage-derived foam cells isolated from rabbit atherosclerotic lesions. Arteriosclerosis, thrombosis, and vascular biology. 1999 ; 19 : 638-45. https://doi.org/10.1161/01.ATV.19.3.638
  9. Tabata T, Mine S, Kawahara C, Okada Y, Tanaka Y. Monocyte chemoattractant protein-1 induces scavenger receptor expression and monocyte differentiation into foam cells. Biochemical and biophysical research communications. 2003 ; 305 : 380-5. https://doi.org/10.1016/S0006-291X(03)00771-X
  10. Hafeman DG, Hoekstra WG. Lipid peroxidation in vivo during vitamin E and selenium deficiency in the rat as monitored by ethane evolution. The Journal of nutrition. 1977 ; 107 : 666-72.
  11. Halliwell B. Free radicals and antioxidants: a personal view. Nutrition reviews. 1994 ; 52 : 253-65.
  12. Suarna C, Wu BJ, Choy K, Mori T, Croft K, Cynshi O, Stocker R. Protective effect of vitamin E supplements on experimental atherosclerosis is modest and depends on preexisting vitamin E deficiency. Free radical biology & medicine. 2006 ; 41 : 722-30. https://doi.org/10.1016/j.freeradbiomed.2006.05.013
  13. Patro BS, Bauri AK, Mishra S, Chattopadhyay S. Antioxidant activity of Myristica malabarica extracts and their constituents. Journal of agricultural and food chemistry. 2005 ; 53 : 6912-8. https://doi.org/10.1021/jf050861x
  14. Lee JC, Kim HR, Kim J, Jang YS. Antioxidant property of an ethanol extract of the stem of Opuntia ficus-indica var. Saboten. Journal of agricultural and food chemistry. 2002 ; 50 : 6490-6. https://doi.org/10.1021/jf020388c
  15. Halliwell B, Gutteridge JM, Aruoma OI. The deoxyribose method: a simple "test-tube" assay for determination of rate constants for reactions of hydroxyl radicals. Analytical biochemistry. 1987 ; 165 : 215-9. https://doi.org/10.1016/0003-2697(87)90222-3
  16. Kweon MH, Hwang HJ, Sung HC. Identification and antioxidant activity of novel chlorogenic acid derivatives from bamboo (Phyllostachys edulis). Journal of agricultural and food chemistry. 2001 ; 49 : 4646-55. https://doi.org/10.1021/jf010514x
  17. Gotoh N, Niki E. Rates of interactions of superoxide with vitamin E, vitamin C and related compounds as measured by chemiluminescence. Biochimica et biophysica acta. 1992 ; 1115 : 201-7. https://doi.org/10.1016/0304-4165(92)90054-X
  18. Sutherland H, Khundkar R, Zolle O, McArdle A, Simpson AW, Jarvis JC, Salmons S. A fluorescence-based method for measuring nitric oxide in extracts of skeletal muscle. Nitric Oxide. 2001 ; 5 : 475-81. https://doi.org/10.1006/niox.2001.0374
  19. Crow JP. Dichlorodihydrofluorescein and dihydrorhodamine 123 are sensitive indicators of peroxynitrite in vitro: implications for intracellular measurement of reactive nitrogen and oxygen species. Nitric Oxide. 1997 ; 1 : 145-57. https://doi.org/10.1006/niox.1996.0113
  20. Xu MZ, Lee WS, Han JM, Oh HW, Park DS, Tian GR, Jeong TS, Park HY. Antioxidant and anti-inflammatory activities of N-acetyldopamine dimers from Periostracum Cicadae. Bioorganic & medicinal chemistry. 2006 ; 14 : 7826-34. https://doi.org/10.1016/j.bmc.2006.07.063
  21. Reid VC, Mitchinson MJ. Toxicity of oxidised low density lipoprotein towards mouse peritoneal macrophages in vitro. Atherosclerosis. 1993 ; 98 : 17-24. https://doi.org/10.1016/0021-9150(93)90219-K
  22. Amakura Y, Umino Y, Tsuji S, Tonogai Y. Influence of jam processing on the radical scavenging activity and phenolic content in berries. Journal of agricultural and food chemistry. 2000 ; 48 : 6292-7. https://doi.org/10.1021/jf000849z
  23. Lindqvist C, Nordstrom T. Generation of hydroxyl radicals by the antiviral compound phosphonoformic acid (foscarnet). Pharmacology & toxicology. 2001 ; 89 : 49-55. https://doi.org/10.1034/j.1600-0773.2001.d01-135.x
  24. Badisa VL, Latinwo LM, Odewumi CO, Ikediobi CO, Badisa RB, Ayuk-Takem LT, Nwoga J, West J. Mechanism of DNA damage by cadmium and interplay of antioxidant enzymes and agents. Environmental toxicology. 2007 ; 22 : 144-51. https://doi.org/10.1002/tox.20248
  25. Nageswara RM, Aleksandr V, Marschall SR. Oxidative stress and vascular disease. Arteriosclerosis, thrombosis, and vascular biology. 2005 ; 25 : 29-38.
  26. Kujala TS, Loponen JM, Klika KD, Pihlaja K. Phenolics and betacyanins in red beetroot (Beta vulgaris) root: distribution and effect of cold storage on the content of total phenolics and three individual compound. Journal of agricultural and food chemistry. 2000 ; 48 : 5338-42.
  27. Warnholtz A, Nickenig G, Schulz E, Macharzina R, Brasen JH, Skatchkov M, Heitzer T, Stasch JP, Griendling KK, Harrison DG, Bohm M, Meinertz T, Munzel T. Increased NADH-oxidase -mediated superoxide production in the early stages of atherosclerosis: evidence for involvement of the renin-angiotensin system. Circulation. 1999 ; 99 : 2027-33. https://doi.org/10.1161/01.CIR.99.15.2027
  28. Wassmann S, Wassmann K, Nickenig G. Modulation of oxidant and antioxidant enzyme expression and function in vascular cells. Hypertension. 2004 ; 44 : 381-6. https://doi.org/10.1161/01.HYP.0000142232.29764.a7
  29. Madamanchi NR, Vendrov A, Runge MS. Oxidative stress and vascular Disease. Arteriosclerosis, thrombosis, and vascular biology. 2005 ; 25 : 29-38.
  30. Madamanchi NR, Hakim ZS, Runge MS. Oxidative stress in atherogenesis and arterial thrombosis: the disconnect between cellular studies and clinical outcomes. Thrombosis and Haemostasis. 2004 ; 3 : 254-67.
  31. Hogg N, Darley-Usmar VM, Wilson MT, Moncada S. Production of hydroxyl radicals from the simultaneous generation of superoxide and nitric oxide. The Biochemical journal. 1992 ; 281 : 419-24.
  32. Steinbrecher UP, Parthasarathy S, Leake DS, Witztum JL, Steinberg D. Modification of low density lipoprotein by endothelial cells involves lipid peroxidation and degradation of low density lipoprotein phospholipids. Proceedings of the National Academy of Sciences of the United States of America. 1984 ; 81 : 3883-7. https://doi.org/10.1073/pnas.81.12.3883
  33. Goldstein JL, Ho YK, Basu SK, Brown MS. Binding site on macrophages mediated uptake and degradation of acetylated low density lipoprotein, producing massive cholesterol deposition. Proceedings of the National Academy of Sciences of the United States of America. 1979 ; 26 : 333-7.
  34. Liu SX, Zhou M, Chen Y, Wen WY, Sun MJ. Lipoperoxidative injury to macrophages by oxidatively modified low density lipoprotein may play an important role in foam cell formation. Atherosclerosis. 1996 ; 121 : 55-61. https://doi.org/10.1016/0021-9150(95)05683-1
  35. Holvoet P, Collen D. beta-VLDL hypercholesterolemia relative to LDL hypercholesterolemia is associated with higher levels of oxidized lipoproteins and a more rapid progression of coronary atherosclerosis in rabbits. Arteriosclerosis, thrombosis, and vascular biology. 1997 ; 17 : 2376-82. https://doi.org/10.1161/01.ATV.17.11.2376
  36. Yoon MA, Jeong TS, Park DS, Xu MZ, Oh HW, Song KB, Lee WS, Park HY. Antioxidant effects of quinoline alkaloids and 2,4-di-tertbutylphenol isolated fromScolopendra subspinipes. Biological & pharmaceutical bulletin. 2006 ; 29 : 735-9. https://doi.org/10.1248/bpb.29.735