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Anti-inflammatory effect of sweetfish-derived protein and its enzymatic hydrolysate on LPS-induced RAW264.7 cells via inhibition of NF-κB transcription

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  • Chemistry and Biochemistry
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Abstract

The present study examined the anti-inflammatory effect of a sweetfish-derived protein and its hydrolysates on lipopolysaccharide (LPS)-induced RAW264.7 macrophage cells. Hydrolysates of the sweetfish-derived protein were obtained on enzymatic hydrolysis by pepsin, trypsin, and α-chymotrypsin. The anti-inflammatory activity was determined based on the production of nitric oxide (NO), inflammatory cytokine [tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6)] and prostaglandin E2 (PGE2), mRNA expression levels of inflammation mediated proteins, and the inhibition of nuclear factor (NF)-κB. The fish protein and its enzymatic hydrolysates were not found to exert a cytotoxic effect on RAW264.7 macrophage cells; they inhibited the production of NO and cytokines in LPS-induced RAW264.7 cells. In particular, hydrolysates prepared by trypsin and α-chymotrypsin remarkably decreased the production of NO, cytokines, and PGE2. Reverse transcription-polymerase chain reaction analysis demonstrated that the decrease in NO and cytokine production was related to the inhibition of the mRNA expression of inducible nitric oxide synthase and cytokine (TNF-α, IL-6, and IL-1β) genes. Moreover, a reporter gene assay showed that the hydrolysates inhibited NF-κB nuclear translocation, and this inhibition was re-confirmed through inhibition of the mitogen-activated protein kinase pathway. The results suggested that the sweetfish-derived protein hydrolysates obtained using trypsin and α-chymotrypsin can be used as anti-inflammatory agents.

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References

  1. Encyclopaedia Britannica, Inc. (1998) Sweetfish. In: New encyclopaedia Britannica. Encyclopaedia Britannica, Inc., Chicago, 11:441

  2. Rietschel ET, Brade H (1992) Bacterial endotoxins. Sci Am 267:54–61

    Article  PubMed  CAS  Google Scholar 

  3. Palmer RM, Ashton DS, Moncada S (1988) Vascular endothelial cells synthesize nitric oxide from l-arginine. Nature 333:664–666

    Article  PubMed  CAS  Google Scholar 

  4. Hibbs JB Jr, Taintor RR, Vavrin Z (1987) Macrophage cytotoxicity: role for l-arginine deiminase and imino nitrogen oxidation to nitrite. Science 235:473–476

    Article  PubMed  CAS  Google Scholar 

  5. Lowenstein CJ, Alley EW, Raval P, Snowman AM, Snyder SH, Russell SW, Murphy WJ (1993) Macrophage nitric oxide synthase gene: two upstream regions mediate induction by interferon gamma and lipopolysaccharide. Proc Natl Acad Sci USA 90(20):9730–9734

    Article  PubMed  CAS  Google Scholar 

  6. Xie QW, Kashiwabara Y, Nathan C (1994) Role of transcription factor NF-kappa B/Rel in induction of nitric oxide synthase. J Biol Chem 269:4705–4708

    PubMed  CAS  Google Scholar 

  7. Hartmann R, Meisel H (2007) Food-derived peptides with biological activity: from research to food applications. Curr Opin Biotech 18:163–169

    Article  PubMed  CAS  Google Scholar 

  8. Kitts DD, Weiler K (2003) Bioactive proteins and peptides from food sources applications of bioprocesses used in isolation and recovery. Curr Pharm Design 9:1309–1323

    Article  CAS  Google Scholar 

  9. Korhonen H, Pihlanto A (2003) Food-derived bioactive peptides—opportunities for designing future foods. Curr Pharm Design 9:1297–1308

    Article  CAS  Google Scholar 

  10. Gildberg A, Bøgwald J, Johansen A, Stenberg E (1996) Isolation of acidic peptide fractions from a fish protein hydrolyzate with strong stimulatory effect on Atlantic salmon (Salmo salar) head kidney leukocytes. Comp Biochem Physiol 114B:97–101

    CAS  Google Scholar 

  11. Bøgwald J, Dalmo RA, Leifson RM, Stenberg E, Gildberg A (1996) The stimulatory effect of a muscle protein hydrolysate from Atlantic cod. Gadus morhua L., on Atlantic salmon, Salmo salar L., head kidney leucocytes. Fish Shellfish Immun 6:3–16

    Article  Google Scholar 

  12. Zhang J, Yan Q, Ji R, Zou W, Guo G (2009) Isolation and characterization of a hepcidin peptide from the head kidney of large yellow croaker, Pseudosciaena crocea. Fish Shellfish Immun 26:864–870

    Article  CAS  Google Scholar 

  13. Byun HG, Lee JK, Park HG, Jeon JK, Kim SK (2009) Antioxidant peptides isolated from the marine rotifer, Brachionus rotundiformis. Process Biochem 44:842–846

    Article  CAS  Google Scholar 

  14. Han EH, Hwang YP, Kim HG, Jeong HG (2007) Inflammatory effect of endosulfan via NF-κB activation in macrophages. Biochem Biophys Res Commun 355:860–865

    Article  PubMed  CAS  Google Scholar 

  15. Kim YW, Zhao RJ, Park SJ, Lee JR, Cho IJ, Yang CH, Kim SG, Kim SC (2008) Anti-inflammatory effects of liquiritigenin as a consequence of the inhibition of NF-κB-dependent iNOS and proinflammatory cytokines production. Brit J Pharmacol 154:165–173

    Article  CAS  Google Scholar 

  16. Rhule A, Navarro S, Smith JR, Shepherd DM (2006) Panax notoginseng attenuates LPS-induced pro-inflammatory mediators in RAW264.7 cells. J Ethnopharmacol 106:121–128

    Article  PubMed  Google Scholar 

  17. Choi CY, Kim JY, Kim YS, Chung YC, Hahm KS, Jeong HG (2001) Augmentation of macrophage functions by an aqueous extract isolated from Platycodon grandiflorum. Cancer Lett 166:17–25

    Article  PubMed  CAS  Google Scholar 

  18. Baeuerle PA, Baichwal VR (1997) NF-kappa B as a frequent target for immunosuppressive and anti-inflammatory molecules. Adv Immunol 65:111–137

    Article  PubMed  CAS  Google Scholar 

  19. Ghosh S, May MJ, Kopp EB (1998) NF-kB and Rel proteins: evolutionary conserved mediators of immune responses. Annu Rev Immunol 16:225–260

    Article  PubMed  CAS  Google Scholar 

  20. Chang YC, Li PC, Chen BC, Chang MS, Wang JL, Chiu WT, Lin CH (2006) Lipoteichoic acid-induced nitric oxide synthase expression in RAW264.7 macrophages is mediated by cyclooxygenase-2, prostaglandin E2, protein kinase A, p38 MAPK, and nuclear factor-κB pathways. Cell Signal 18:1235–1243

    Article  PubMed  CAS  Google Scholar 

  21. Weckmann AL, Alcocer-Varela J (1996) Cytokine inhibitors in autoimmune disease. Semin Arthr Rheum 26:539–557

    Article  CAS  Google Scholar 

  22. Elenkov IJ, Chrousos GP (2002) Stress hormones, proinflammatory and antiinflammatory cytokines, and autoimmunity. Ann N Y Acad Sci 966:290–303

    Article  PubMed  CAS  Google Scholar 

  23. Eigler A, Sinha B, Hartmann G, Endres S (1997) Taming TNF: strategies to restrain this proinflammatory cytokine. Immunology 18:487–492

    CAS  Google Scholar 

  24. Ohkubo K, Ikeda M, Pawankar R, Gotoh M, Yagi T, Okuda M (1998) Mechanisms of IL-6, IL-8, and GM-CSF release in nasal secretions of allergic patients after nasal challenge. Rhinology 36:156–161

    PubMed  CAS  Google Scholar 

  25. MacMicking J, Xie QW, Nathan C (1997) Nitric oxide and macrophage function. Annu Rev Immunol 15:323–350

    Article  PubMed  CAS  Google Scholar 

  26. Grassl C, Luckow B, Schlondorff D, Dendorfer U (1999) Transcriptional regulation of the interleukin-6 gene in mesangial cells. J Am Soc Nephrol 10:1466–1477

    PubMed  CAS  Google Scholar 

  27. Matsusaka T, Fujikawa K, Nishio Y, Mukaida N, Matsushima K, Kishimoto T, Akira S (1993) Transcription factors NF-IL6 and NF-κB synergistically activate transcription of the inflammatory cytokines, interleukin 6 and interleukin 8. Proc Natl Acad Sci USA 90:10193–10197

    Article  PubMed  CAS  Google Scholar 

  28. Hsiao HY, Mak OT, Yang CS, Liu YP, Fang KM, Tzeng SF (2007) TNF-a/IFN-c-Induced iNOS expression increased by prostaglandin E2 in rat primary astrocytes via EP2-evoked cAMP/PKA and intracellular calcium signaling. GLIA 55:214–223

    Article  PubMed  Google Scholar 

  29. Surh YJ, Chun KS, Cha HH, Han SS, Keum YS, Park KK (2001) Lee Sang Sup. Molecular mechanisms underlying chemopreventive activities of anti-inflammatory phytochemicals: down-regulation of COX-2 and iNOS through suppression of NF-κB activation. Mutat Res 480:243–268

    Article  PubMed  Google Scholar 

  30. Sautebin L (2000) Prostaglandins and nitric oxide as molecular targets for anti-inflammatory therapy. Fitoterapia 71:S48–S57

    Article  PubMed  CAS  Google Scholar 

  31. Barnes PJ, Karin M (1997) Nuclear factor-κB—a pivotal transcription factor in chronic inflammatory diseases. N Engl J Med 336:1066–1071

    Article  PubMed  CAS  Google Scholar 

  32. Bäuerle PA, Baltimore D (1996) NF-κB: ten years after. Cell 87:13–20

    Article  Google Scholar 

  33. Baldwin AS Jr (1996) The NF-κB and IκB proteins: new discoveries and insights. Annu Rev Immunol 14:649–681

    Article  PubMed  CAS  Google Scholar 

  34. Bonizzi G, Karin M (2004) The two NF-kappaB activation pathways and their role in innate and adaptive immunity. Trends Immunol 25:280–288

    Article  PubMed  CAS  Google Scholar 

  35. Pando MP, Verma IM (2000) Signal-dependent and -independent degradation of free and NF-kappa B-bound IkappaBalpha. J Biol Chem 275:21278–21286

    Article  PubMed  CAS  Google Scholar 

  36. Karin M, Ben-Neriah Y (2000) Phosphorylation meets ubiquitination: the control of NF-(kappa)B activity. Annu Rev Immunol 18:621–663

    Google Scholar 

  37. Brown K, Gerstberger S, Carlson L, Franzoso G, Siebenlist U (1995) Control of I kappa B-alpha proteolysis by site-specific, signal induced phosphorylation. Science 267:1485–1488

    Article  PubMed  CAS  Google Scholar 

  38. Traenckner EB, Pahl HL, Henkel T, Schmidt KN, Wilk S, Baeuerle PA (1995) Phosphorylation of human I kappa B-alpha on serines 32 and 36 controls I kappa B-alpha proteolysis and NF-kappa B activation in response to diverse stimuli. EMBO J 14:2876–2883

    PubMed  CAS  Google Scholar 

  39. Wu C, Ghosh S (1999) Beta-TrCP mediates the signal-induced ubiquitination of IkappaBbeta. J Biol Chem 274:29591–29594

    Article  PubMed  CAS  Google Scholar 

  40. Kroll M, Margottin F, Kohl A, Renard P, Concordet JP, Bachelerie F, Arenzana-Seisdedos F, Benarous R (1999) Inducible degradation of IkappaBalpha by the proteasome requires interaction with the F-box protein h-betaTrCP. J Biol Chem 274:7941–7945

    Article  PubMed  CAS  Google Scholar 

  41. Scherer DC, Brockman JA, Chen Z, Maniatis T, Ballard DW (1995) Signal-induced degradation of I kappa B alpha requires site-specific ubiquitination. Proc Natl Acad Sci USA 92:11259–11263

    Article  PubMed  CAS  Google Scholar 

  42. Baldi L, Brown K, Franzoso G, Siebenlist U (1996) Critical role for lysines 21 and 22 in signal-induced, ubiquitin-mediated proteolysis of I kappa B-alpha. J Biol Chem 271:376–379

    Article  PubMed  CAS  Google Scholar 

  43. Jung WK, Inhak Choi, Lee DY, Yea SS, Choi YH, Kim MM, Park SG, Seo SK, Lee SW, Lee CM, Park YM, Choi IW (2008) Caffeic acid phenethyl ester protects mice from lethal endotoxin shock and inhibits lipopolysaccharide-induced cyclooxygenase-2 and inducible nitric oxide synthase expression in RAW264.7 macrophages via the p38/ERK and NF-κB pathways. Int J Biochem Cell B 40:2572–2582

    Article  CAS  Google Scholar 

Download references

Acknowledgments

This research was co-supported by the National Research Foundation in the Nuclear Research & Development Program, and by the Basic Research Support Program of Korea Atomic Energy Research Institute.

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Correspondence to Jae-Hun Kim.

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Sung, NY., Jung, PM., Yoon, M. et al. Anti-inflammatory effect of sweetfish-derived protein and its enzymatic hydrolysate on LPS-induced RAW264.7 cells via inhibition of NF-κB transcription. Fish Sci 78, 381–390 (2012). https://doi.org/10.1007/s12562-011-0461-5

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  • DOI: https://doi.org/10.1007/s12562-011-0461-5

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