The Effects of Haedoksamul-tang on Oxidative Stress and Hyperlipidemia in LPS-induced ICR Mouse

Article information

J Korean Med. 2016;37(1):77-89
Publication date (electronic) : 2016 March 31
doi : https://doi.org/10.13048/jkm.16008
Department of Internal Medicine of Korean Medicine, College of Korean Medicine, Dae-gu Haany University
Correspondence to: 신현철, 대구한의대학교 한의과대학 내과학교실, Tel: +82-54-281-0055, Fax: +82-54-281-7464, E-mail: ungaeshin@naver.com
Received 2016 February 15; Revised 2016 March 21; Accepted 2016 March 28.

Abstract

Objectives:

The present study was conducted to examine whether Haedoksamul-tang (HS), a traditional oriental herbal medicine, have beneficail effects on anti-inflammation and dyslipidemia in lipopolysaccharide (LPS)-induced ICR mouse.

Methods:

Twenty four 8-week old male ICR mouse were divided into four groups: normal untreated; LPS treatment only; HS 10 mg/kg plus LPS treatment; and HS 30 mg/kg plus LPS treatment. HS was orally administered per day for 2days. Twenty-four hours after LPS injection (10 mg/kg/day, i.p.), all the mice were sacrificed, and serological changes were evaluated. The levels of nuclear factor-κB (NF-κB), sterol regulatory element-binding transcription protein 1 (SREBP-1) activity and cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), tumor necrosis factor a (TNF-a), monocyte chemotactic protein 1 (MCP-1), acetyl-CoA carboxylase a (ACCa) expression were analyzed in Western blot analysis.

Results:

HS inhibited oxidative stress in the liver of LPS-induced ICR mice. The LPS-induced ICR mice exhibited the increase of NF-κB activity and COX-2, iNOS, TNF-a, MCP-1 expressions in the liver, while HS treatment significantly inhibited them. Moreover, The administration of HS significantly decreased the elevated serum triglyceride and down-regulated the levels of SREBP-1, ACCa in the liver of LPS-induced ICR mice.

Conclusions:

In conclusion, HS could have hepato-protective effects against the oxidative stress-related inflammation and abnormal lipid metabolism.

Fig. 1.

Inhibition effects of HS on serum and hepatic oxidative stress in LPS-induced ICR mice. Serum ROS (A), hepatic ROS (B), hepatic TBARS (C) levels. N: normal group, Veh: vehicle-treated mice, HS10: HS 10 mg/kg treated mice, HS30: HS 30 mg/kg treated mice. Bars represent means ± SD. ** P < 0.01 versus vehicle-treated mice values.

Fig. 2.

Effects of HS on NF-κBp65 activity in LPS-induced ICR mice liver. N: normal group, Veh: vehicle-treated mice, HS10: HS 10 mg/kg treated mice, HS30: HS 30 mg/kg treated mice. Histone was used for loading control. Bars represent means ± SD. * P < 0.05, *** P < 0.001 versus vehicle-treated mice values.

Fig. 3.

Effects of HS on COX-2 and iNOS expressions in LPS-induced ICR mice liver. COX-2 (A) and iNOS (B) protein expressions in liver. N: normal group, Veh: vehicle -treated mice, HS10: HS 10 mg/kg treated mice, HS30: HS 30 mg/kg treated mice. ß-actin was used for loading control. Bars represent means ± SD. * P < 0.05, ** P < 0.01, *** P < 0.001 versus vehicle-treated mice values.

Fig. 4.

Effects of HS on TNF-a and MCP-1 expressions in LPS-induced ICR mice liver. TNF-a (A) and MCP-1 (B) protein expressions in liver. N: normal group, Veh: vehicle-treated mice, HS10: HS 10 mg/kg treated mice, HS30: HS 30 mg/kg treated mice. ß-actin was used for loading control. Bars represent means ± SD. * P < 0.05, ** P < 0.01, *** P < 0.001 versus vehicle-treated mice values.

Fig. 5.

Effects of HS on serum and hepatic triglyceride levels in LPS-induced ICR mice. Serum triglyceride (A), hepatic triglyceride (B) levels. N: normal group, Veh: vehicle-treated mice, HS10: HS 10 mg/kg treated mice, HS30: HS 30 mg/kg treated mice. Bars represent means ± SD. * P < 0.05, ** P < 0.01, *** P < 0.001 versus vehicle-treated mice values.

Fig. 6.

Effects of HS on SREBP-1 activity and ACCa expression in LPS-induced ICR mice liver. SREBP-1 (A) activity and ACCa (B) protein expression in liver. N: normal group, Veh: vehicle-treated mice, HS10: HS 10 mg/kg treated mice, HS30: HS 30 mg/kg treated mice. Histone or ß-actin was used for loading control. Bars represent means ± SD. * P < 0.05, ** P < 0.01, *** P < 0.001 versus vehicle-treated mice values.

Composition of Haedoksamul-tang

References

1. Johan F. Immunity, atherosclerosis and cardiovascular disease. BMC med 2013;11:117–29.
2. Feng X, Zhang Y, Xu R, Xie X, Tao L, Gao H, et al. Lipopolysaccharide up-regulates the expression of Fcalpha/mu receptor and promotes the binding of oxidized low-density lipoprotein and its IgM antibody comples to activated human macrophages. Atherosclerosis 2010;208:396–405.
3. Wiesner P, Choi SH, Almazan F, Benner C, Huang W, Diehl CJ, et al. Low doses of lipopolysaccharide and minimally oxidized low-density lipoprotein cooperatively activate macrophages via nuclear factor kappa B and activator protein-1: possible mechanism for acceleration of atherosclerosis by subclinical endotoxemia. Circ Res 2010;107:56–65.
4. Maitra U, Li L. Molecular mechanisms responsible for the ruduced expression of cholesterol transporters from macrophages by low-dose endotoxin. Arterioscler Thromb Vasc Biol 2013;33:24–33.
5. Vikatmaa P, Lajunen T, Ikonen TS, Pussinen PJ, Leinonen M, Saikku P, et al. Chlamydial lipopolysaccharide (cLPS) is present in atherosclerotic and aneurysmal arterial wall--cLPS levels depend on disease manifestation. Cardiovasc Pathol 2010;19(1):48–54.
6. Jean L, Bret D, Chistophe C. Systemic capsaicin pretreatment fails to bolck the decrease in food-motivated behavior induced by lipopolysaccharide and interleukin-1ß. Brain Research Bul 1997;42:443–9.
7. Sharifov OF, Nawar G, Ternovov W, Mishra VK, Litovsky SH, Palqunachare MN, et al. Cationic peptide mR18L with lipid lowering properties inhibits LPS-induced systemic and liver inflammation in rats. Biochem Biophys Res Commun 2013;436(4):705–10.
8. Kim ID, Kang KS, Kwon RH, Yang JO, Lee JS, Ha BJ. The effect of Rubus coreanum Miquel against lipopolysaccharide-induced oxidative stress and lipid metaobolism. J. Fd Hyg. Safety 2007;22(3):213–7.
9. Ju JH. Dan-gye-sim-beop-bu-yeo Seoul: Daesung Publishing Co. Ltd; 1982. p. 714.
10. Lee SI. Cheon-jin-cheo-bang-hea-sheol Seoul: Sheong-bo-sa; 1995. p. 289–91.
11. Kim ES. A experimental study of Hwangryeonheadock-Tang and Onchung-Eum on hyperlipidemia & hypertension. J. of Korean Medicine 1999;20(1):185–96.
12. Kim YH, Jo HB. Anti-inflammatory effects of Haedoksamul-tang in RAW 264.7 cells. J. of Oriental Obstetrics & Gynecology 2008;21(2):166–83.
13. Beom HB. Effects of Onchungeum and Gamionchungeum on the antiallergic response and blood coagulation. J. of Kyung Hee University 1990;6(4):490–9.
14. Jeon YG. Effects of Haedoksamul-tang on Trimelliticanhydride-induced contact hypersensitivity in a mouse model 2010;:1–41.
15. Seo M, Jeon BH, Woo WH, Jeong WY. Effect of Onchengyeum on the damaged liver cell by carbon tetrachloride in rats. Korean J. Oriental Physiology & Pathology 1989;11(2):27–36.
16. Im HJ, Hwang CY, Gang HC, Kim NG, Gwon IH. Inhibitory effects of Onchumgeum on cytokine production from phytohaemagglutin-stimulated peripheral blood mononuclear cells of behcets patients. Korean J. Oriental Physiology & Pathology 2002;16(4):768–73.
17. Zhu GF, Guo HJ, Huang Y, Wu CT, Zhang XF. Eriodictyol, a plant flavonoid, attenuates LPS-induced acute lung injury through its antioxidative and anti-inflammatory activity. Exp Ther Med 2015;10(6):2259–66.
18. Heo J. Dong-ui-bo-gam Seoul: Bubin Publishing Co. Ltd; 2007. 416p. 455.
19. Rush GF, Gorski JR, Ripple MG, Sowinski J, Bugelski P. Organic hydroperoxide -induced lipid peroxidation and cell death in isolated hepatocytes. Toxicol Appl Pharmacol 1985;78:473–83.
20. Izeboud CA, Hoebe KH, Grootendorst AF, Nijmeijer SM, van Miert AS, Witkamp RF, et al. Endotoxin-induced liver damage in rats is minimized by beta 2-adrenoceptor stimulation. Inflamm Res 2004;53:93–9.
21. Munford RS. Severe sepsis and septic sock: the role of gram-negative bacteremia. Annu Rev Pathol 2006;1:467–96.
22. Kleemann R, Verschuren L, van Erk M, Nikolsky Y, Verheij ER, Smilde AK, et al. Atherosclerosis and liver inflammation induced by increased dietary cholesterol intake: a combined transcriptomics and metabolomics analysis. Genome Biol 2007;8:R200.
23. Xu X, Hu J, McGrath BC, Cavener DR. GCN2 in the brain programs PPARγ2 and triglyceride storage in the liver during perinatal development in response to maternal dietary fat. PLos One 2013;8(10):e75917.
24. Liu RH, Hotchkiss JH. Potential genotoxicity of chronically elevated nitirc oxide: a review. Mutat. Res 1996;339:73–89.
25. Collins T, Cybulsky MI. NF-kappaB: pivotal mediator or innocent bystander in atherogenesis. J Clin Invest 2001;107:255–64.
26. Thuberg BL, Collins T. The nuclear factor-B/inhibitor of B autoregulatory system and atherosclerosis. Curr Opin Lipidol 1998;9:387–96.
27. Wang QS, Xiang Y, Cui YL, Lin KM, Zhang XF. Dietary blue pigments derived from Genipin, attenuate inflammation by inhibiting LPS-Induced iNOS and COX-2 expression via the NF-κB inactivation. PLos One 2012;7(3):1–11.
28. Kwon HJ, Sung BK, Kim JW, Lee JH, Kim ND, Yoo MA, et al. The effect of lipopolysaccharide on enhanced inflammatory process with age: modulation of NF-κB. J. Amer. Aging Assoc 2001;24:163–72.
29. Napoli C, de Nigris F, Williams-Ignarro S, Pignalosa O, Sica V, Ignarro LJ. Nitric oxide and atherosclerosis: an update. Nitric Oxide 2006;15:265–79.
30. Baldwin AS Jr. The NF-kappa B and I kappa B proteins: new discoveries and insights. Annu Rev Immunol 1996;14:649–83.
31. DeGraba TJ. Expression of inflammatory mediators and adhesion molecules in human atherosclerotic plaque. Neurology 1997;49(Supple 4):S15–9.
32. Fassbender K, Rossol S, Kammer T, Daffertshofer M, Wirth S, Dollman M, et al. Proinflammatory cytokines in serum of patients with acute cerebral ischemia: kinetics of secretion and relation to the extent of brain damage and outcome of disease. J Neurol Sci 1994;122(2):135–9.
33. Adams DH, Lioyd AF. Chemokines: leukocyte recruitment and activation cytokines. Lancet 1997;349:490–5.
34. Brown MS, Goldstein JL. The SREBP pathway: regulation of cholesterol metabolism by proteolysis of a membrane-bound transcription factor. Cell 1997;89:331–40.
35. Weber LW, Boll M, Stampfl A. Maintaining cholesterol homeostasis: sterol regulatory element-binding proteins. World J Gastroenterol 2004;10(21):3081–7.
36. Horton JD, Goldstein JL, Brown MS. SREBPs: activators of the complete program of cholesterol and fatty acid synthesis in the liver. J Clin Invest 2002;109:1125–31.
37. Foufelle F, Ferre P. New perspectives in the regulation of hepatic glycolytic and lipogenic genes by insulin and glucose: a role for the transcription factor sterol regulatory element binding protein-1c. Biochem J 2002;366:377–91.
38. Ferre P, Foretz M, Azzout-Marniche D, Becard D, Foufelle F. Sterol-regulatory-element-binding protein 1c mediates insulin action on hepatic gene expression. Biochemical Society Transactions 2001;29(4):547–52.
39. Horton JD. Sterol regulatory element-binding proteins: transcriptional activators of lipid synthesis. Biochemical Society Transactions 2001;30(6):1091–5.
40. Tong L. Acetyl-coenzyme A carboxylase: crucial metabolic enzyme and attractive target for drug discovery. Cell Mol Life Sci 2005;62(16):1784–803.
41. Munday MR. Regulation of mammalian acetyl-CoA carboxylase. Biochem Soc Trans 2002;30(6):1059–64.

Article information Continued

Fig. 1.

Inhibition effects of HS on serum and hepatic oxidative stress in LPS-induced ICR mice. Serum ROS (A), hepatic ROS (B), hepatic TBARS (C) levels. N: normal group, Veh: vehicle-treated mice, HS10: HS 10 mg/kg treated mice, HS30: HS 30 mg/kg treated mice. Bars represent means ± SD. ** P < 0.01 versus vehicle-treated mice values.

Fig. 2.

Effects of HS on NF-κBp65 activity in LPS-induced ICR mice liver. N: normal group, Veh: vehicle-treated mice, HS10: HS 10 mg/kg treated mice, HS30: HS 30 mg/kg treated mice. Histone was used for loading control. Bars represent means ± SD. * P < 0.05, *** P < 0.001 versus vehicle-treated mice values.

Fig. 3.

Effects of HS on COX-2 and iNOS expressions in LPS-induced ICR mice liver. COX-2 (A) and iNOS (B) protein expressions in liver. N: normal group, Veh: vehicle -treated mice, HS10: HS 10 mg/kg treated mice, HS30: HS 30 mg/kg treated mice. ß-actin was used for loading control. Bars represent means ± SD. * P < 0.05, ** P < 0.01, *** P < 0.001 versus vehicle-treated mice values.

Fig. 4.

Effects of HS on TNF-a and MCP-1 expressions in LPS-induced ICR mice liver. TNF-a (A) and MCP-1 (B) protein expressions in liver. N: normal group, Veh: vehicle-treated mice, HS10: HS 10 mg/kg treated mice, HS30: HS 30 mg/kg treated mice. ß-actin was used for loading control. Bars represent means ± SD. * P < 0.05, ** P < 0.01, *** P < 0.001 versus vehicle-treated mice values.

Fig. 5.

Effects of HS on serum and hepatic triglyceride levels in LPS-induced ICR mice. Serum triglyceride (A), hepatic triglyceride (B) levels. N: normal group, Veh: vehicle-treated mice, HS10: HS 10 mg/kg treated mice, HS30: HS 30 mg/kg treated mice. Bars represent means ± SD. * P < 0.05, ** P < 0.01, *** P < 0.001 versus vehicle-treated mice values.

Fig. 6.

Effects of HS on SREBP-1 activity and ACCa expression in LPS-induced ICR mice liver. SREBP-1 (A) activity and ACCa (B) protein expression in liver. N: normal group, Veh: vehicle-treated mice, HS10: HS 10 mg/kg treated mice, HS30: HS 30 mg/kg treated mice. Histone or ß-actin was used for loading control. Bars represent means ± SD. * P < 0.05, ** P < 0.01, *** P < 0.001 versus vehicle-treated mice values.

Table 1.

Composition of Haedoksamul-tang

Herb name Scientific name Amounts (g)
黃芩 Scutellaria baicalensis George 4
黃連 Coptis chinensis Franch. 4
黃柏 Phellodendron amurense Ruprecht 4
梔子 Gardenia jasminoides Ellis 4
當歸 Angelica acutiloba (S. et Z.) Kitagawa 4
川芎 Cnidium officinale Makino 4
白芍藥 Paeonia latiflora Pall. 4
生乾地黃 Rehmannia glutinosa (Gaetner) Libosch. 4

Total 32