Cell Stress and Chaperones

, Volume 23, Issue 5, pp 857–869 | Cite as

Scopoletin intervention in pancreatic endoplasmic reticulum stress induced by lipotoxicity

  • Kalaivanan Kalpana
  • Emayavaramban Priyadarshini
  • S. Sreeja
  • Kalivarathan Jagan
  • Carani Venkatraman Anuradha
Original Paper


Endoplasmic reticulum (ER), a dynamic organelle, plays an essential role in organizing the signaling pathways involved in cellular adaptation, resilience, and survival. Impairment in the functions of ER occurs in a variety of nutritive disorders including obesity and type 2 diabetes. Here, we hypothesize that (scopoletin) SPL, a coumarin, has the potential to alleviate ER stress induced in vitro and in vivo models by lipotoxicity. To test this hypothesis, the ability of SPL to restore the levels of proteins of ER stress was analyzed. Rat insulinoma 5f (RIN5f) cells and Sprague Dawley rats were the models used for this study. Groups of control and high-fat, high-fructose diet (HFFD)-fed rats were treated with either SPL or 4-phenylbutyric acid. Status of ER stress was enumerated by quantitative RT-PCR, Western blot, electron microscopic, and immunohistochemical studies. Proximal proteins of ER stress inositol requiring enzyme 1 (IRE1), protein kinase like endoplasmic reticulum kinase (PERK), and activating transcription factor 6 (ATF6) were reduced in the β-cells by SPL. The subsequent signaling proteins X-box binding protein 1, eukaryotic initiation factor2α, activating transcription factor 4, and C/EBP homologous protein were also suppressed in their expression levels when treated with SPL. IRE1, PERK signaling leads to c-Jun-N-terminal kinases phosphorylation, a kinase that interrupts insulin signaling, which was also reverted upon scopoletin treatment. Finally, we confirm that SPL has the ability to suppress the stress proteins and limit pancreatic ER stress which might help in delaying the progression of insulin resistance.


Scopoletin ER stress 4-Phenylbutyric acid Lipotoxicity 



The authors wish to thank DST-FIST and UGC-SAP for the infrastructure facilities developed in the Department of Biochemistry and Biotechnology, Annamalai University, for executing the present study. We also thank Dr. Pushpa Viswanathan, Professor and Head, Dept. of Electron Microscope, Adyar Cancer Research Institute, Chennai, for conducting the electron microscopic studies successfully.

Funding information

This work was supported by Department of Science and Technology, Women Scientists Scheme-A, New Delhi, India under “Disha Programme for Women in Science” (SR/WOS-A/LS-1170/2014).

Compliance with ethical standards

Experimental procedures were approved by the Institutional Animal Ethics Committee (IAEC), Annamalai University, and conducted according to the guidelines by the Committee for the Purpose of Control and Supervision on Experiments on Animals (CPCSEA) (Proposal No. 1091).

Conflict of interest

The authors declare that they have no conflicts of interest.


  1. Adamopoulos C, Farmaki E, Spilioti E, Kiaris H, Piperi C, Papavassiliou AG (2014) Advanced glycation end-products induce endoplasmic reticulum stress in human aortic endothelial cells. Clin Chem Lab Med 52(1):151–160. CrossRefPubMedGoogle Scholar
  2. Back SH, Kaufman RJ (2012) Endoplasmic reticulum stress and type 2 diabetes. Annu Rev Biochem 81:767–793. CrossRefPubMedPubMedCentralGoogle Scholar
  3. Bailly-Maitre B, Belgardt BF, Jordan SD, Coornaert B, von Freyend MJ, Kleinridders A, Mauer J, Cuddy M, Kress CL, Willmes D, Essig M, Hampel B, Protzer U, Reed JC, Bruning JC (2010) Hepatic Bax inhibitor-1 inhibits IRE1alpha and protects from obesity-associated insulin resistance and glucose intolerance. J Biol Chem 285(9):6198–6207. CrossRefPubMedGoogle Scholar
  4. Bhuvaneswari S, Yogalakshmi B, Sreeja S, Anuradha CV (2014) Astaxanthin reduces hepatic endoplasmic reticulum stress and nuclear factor-kappaB-mediated inflammation in high fructose and high fat diet-fed mice. Cell Stress Chaperones 19(2):183–191. CrossRefPubMedGoogle Scholar
  5. Birkenfeld AL, Lee HY, Majumdar S, Jurczak MJ, Camporez JP, Jornayvaz FR, Frederick DW, Guigni B, Kahn M, Zhang D, Weismann D, Arafat AM, Pfeiffer AF, Lieske S, Oyadomari S, Ron D, Samuel VT, Shulman GI (2011) Influence of the hepatic eukaryotic initiation factor 2alpha (eIF2alpha) endoplasmic reticulum (ER) stress response pathway on insulin-mediated ER stress and hepatic and peripheral glucose metabolism. J Biol Chem 286(42):36163–36170. CrossRefPubMedPubMedCentralGoogle Scholar
  6. Boden G, Duan X, Homko C, Molina EJ, Song W, Perez O, Cheung P, Merali S (2008) Increase in endoplasmic reticulum stress-related proteins and genes in adipose tissue of obese, insulin-resistant individuals. Diabetes 57(9):2438–2444. CrossRefPubMedPubMedCentralGoogle Scholar
  7. Castro G, MF CA, Weissmann L, Quaresma PG, Katashima CK, Saad MJ, Prada PO (2013) Diet-induced obesity induces endoplasmic reticulum stress and insulin resistance in the amygdala of rats. FEBS Open Bio 3:443–449. CrossRefPubMedPubMedCentralGoogle Scholar
  8. Cnop M, Ladriere L, Igoillo-Esteve M, Moura RF, Cunha DA (2010) Causes and cures for endoplasmic reticulum stress in lipotoxic beta-cell dysfunction. Diabetes Obes Metab 12(Suppl 2):76–82. CrossRefPubMedGoogle Scholar
  9. Cnop M, Foufelle F, Velloso LA (2012) Endoplasmic reticulum stress, obesity and diabetes. Trends Mol Med 18(1):59–68. CrossRefPubMedGoogle Scholar
  10. Eizirik DL, Cardozo AK, Cnop M (2008) The role for endoplasmic reticulum stress in diabetes mellitus. Endocr Rev 29(1):42–61. CrossRefPubMedGoogle Scholar
  11. Fonseca SG, Urano F, Weir GC, Gromada J, Burcin M (2012) Wolfram syndrome 1 and adenylyl cyclase 8 interact at the plasma membrane to regulate insulin production and secretion. Nat Cell Biol 14(10):1105–1112. CrossRefPubMedPubMedCentralGoogle Scholar
  12. Gao D, Bambang IF, Putti TC, Lee YK, Richardson DR, Zhang D (2012) ERp29 induces breast cancer cell growth arrest and survival through modulation of activation of p38 and upregulation of ER stress protein p58IPK. Lab Investig J Tech Methods Pathol 92(2):200–213. CrossRefGoogle Scholar
  13. Gregor MF, Hotamisligil GS (2007) Thematic review series: adipocyte biology. Adipocyte stress: the endoplasmic reticulum and metabolic disease. J Lipid Res 48(9):1905–1914. CrossRefPubMedGoogle Scholar
  14. Gregor MF, Yang L, Fabbrini E, Mohammed BS, Eagon JC, Hotamisligil GS, Klein S (2009) Endoplasmic reticulum stress is reduced in tissues of obese subjects after weight loss. Diabetes 58(3):693–700. CrossRefPubMedPubMedCentralGoogle Scholar
  15. Ham JR, Lee HI, Choi RY, Sim MO, Choi MS, Kwon EY, Yun KW, Kim MY, Lee MK (2016) Anti-obesity and anti-hepatosteatosis effects of dietary scopoletin in high-fat diet fed mice. J Funct Foods 25:433–446. CrossRefGoogle Scholar
  16. Harding HP, Zeng H, Zhang Y, Jungries R, Chung P, Plesken H, Sabatini DD, Ron D (2001) Diabetes mellitus and exocrine pancreatic dysfunction in perk-/- mice reveals a role for translational control in secretory cell survival. Mol Cell 7(6):1153–1163CrossRefPubMedGoogle Scholar
  17. Hou ZQ, Li HL, Gao L, Pan L, Zhao JJ, Li GW (2008) Involvement of chronic stresses in rat islet and INS-1 cell glucotoxicity induced by intermittent high glucose. Mol Cell Endocrinol 291(1–2):71–78. CrossRefPubMedGoogle Scholar
  18. Jaeschke A, Davis RJ (2007) Metabolic stress signaling mediated by mixed-lineage kinases. Mol Cell 27(3):498–508. CrossRefPubMedPubMedCentralGoogle Scholar
  19. Jung IR, Choi SE, Jung JG, Lee SA, Han SJ, Kim HJ, Kim DJ, Lee KW, Kang Y (2015) Involvement of iron depletion in palmitate-induced lipotoxicity of beta cells. Mol Cell Endocrinol 407:74–84. CrossRefPubMedGoogle Scholar
  20. Kadowaki H, Nishitoh H (2013) Signaling pathways from the endoplasmic reticulum and their roles in disease. Genes 4(3):306–333. CrossRefPubMedPubMedCentralGoogle Scholar
  21. Kalivarathan J, Chandrasekaran SP, Kalaivanan K, Ramachandran V, Carani Venkatraman A (2017) Apigenin attenuates hippocampal oxidative events, inflammation and pathological alterations in rats fed high fat, fructose diet. Biomed Pharmacother 89:323–331. CrossRefPubMedGoogle Scholar
  22. Karaskov E, Scott C, Zhang L, Teodoro T, Ravazzola M, Volchuk A (2006) Chronic palmitate but not oleate exposure induces endoplasmic reticulum stress, which may contribute to INS-1 pancreatic beta-cell apoptosis. Endocrinology 147(7):3398–3407. CrossRefPubMedGoogle Scholar
  23. Kawasaki N, Asada R, Saito A, Kanemoto S, Imaizumi K (2012) Obesity-induced endoplasmic reticulum stress causes chronic inflammation in adipose tissue. Sci Rep 2:799. CrossRefPubMedPubMedCentralGoogle Scholar
  24. Kharroubi I, Ladriere L, Cardozo AK, Dogusan Z, Cnop M, Eizirik DL (2004) Free fatty acids and cytokines induce pancreatic beta-cell apoptosis by different mechanisms: role of nuclear factor-kappaB and endoplasmic reticulum stress. Endocrinology 145(11):5087–5096. CrossRefPubMedGoogle Scholar
  25. Kim I, Xu W, Reed JC (2008) Cell death and endoplasmic reticulum stress: disease relevance and therapeutic opportunities. Nat Rev Drug Discov 7(12):1013–1030. CrossRefPubMedGoogle Scholar
  26. Laybutt DR, Preston AM, Akerfeldt MC, Kench JG, Busch AK, Biankin AV, Biden TJ (2007) Endoplasmic reticulum stress contributes to beta cell apoptosis in type 2 diabetes. Diabetologia 50(4):752–763. CrossRefPubMedGoogle Scholar
  27. Lee AS (2005) The ER chaperone and signaling regulator GRP78/BiP as a monitor of endoplasmic reticulum stress. Methods 35(4):373–381. CrossRefPubMedGoogle Scholar
  28. Lee AH, Iwakoshi NN, Glimcher LH (2003) XBP-1 regulates a subset of endoplasmic reticulum resident chaperone genes in the unfolded protein response. Mol Cell Biol 23(21):7448–7459CrossRefPubMedPubMedCentralGoogle Scholar
  29. Lee AH, Heidtman K, Hotamisligil GS, Glimcher LH (2011) Dual and opposing roles of the unfolded protein response regulated by IRE1alpha and XBP1 in proinsulin processing and insulin secretion. Proc Natl Acad Sci U S A 108(21):8885–8890. CrossRefPubMedPubMedCentralGoogle Scholar
  30. Lee HI, Yun KW, Seo KI, Kim MJ, Lee MK (2014) Scopoletin prevents alcohol-induced hepatic lipid accumulation by modulating the AMPK-SREBP pathway in diet-induced obese mice. Metab Clin Exp 63(4):593–601. CrossRefPubMedGoogle Scholar
  31. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real time quantitative PCR and the 2-ΔΔCT method. Methods 25:402–408. CrossRefPubMedGoogle Scholar
  32. Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with the Folin phenol reagent. J Biol Chem 193(1):265–275PubMedGoogle Scholar
  33. Meusser B, Hirsch C, Jarosch E, Sommer T (2005) ERAD: the long road to destruction. Nat Cell Biol 7(8):766–772. CrossRefPubMedGoogle Scholar
  34. Mogana R, Teng-Jin K, Wiart C (2013) Anti-inflammatory, anticholinesterase, and antioxidant potential of scopoletin isolated from Canarium patentinervium Miq. (Burseraceae Kunth). Evid Based Complement Alternat Med 2013:734824. CrossRefPubMedPubMedCentralGoogle Scholar
  35. Moon PD, Lee BH, Jeong HJ, An HJ, Park SJ, Kim HR, Ko SG, Um JY, Hong SH, Kim HM (2007) Use of scopoletin to inhibit the production of inflammatory cytokines through inhibition of the IkappaB/NF-kappaB signal cascade in the human mast cell line HMC-1. Eur J Pharmacol 555(2–3):218–225. CrossRefPubMedGoogle Scholar
  36. Mosmann T (1983) Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J Immunol Methods 65(1–2):55–63CrossRefPubMedGoogle Scholar
  37. Nam H, Kim MM (2015) Scopoletin has a potential activity for anti-aging via autophagy in human lung fibroblasts. Phytomedicine 22(3):362–368. CrossRefPubMedGoogle Scholar
  38. Okuda-Shimizu Y, Hendershot LM (2007) Characterization of an ERAD pathway for nonglycosylated BiP substrates, which require Herp. Mol Cell 28(4):544–554. CrossRefPubMedPubMedCentralGoogle Scholar
  39. Oyadomari S, Mori M (2004) Roles of CHOP/GADD153 in endoplasmic reticulum stress. Cell Death Differ 11(4):381–389. CrossRefPubMedGoogle Scholar
  40. Ozcan U, Cao Q, Yilmaz E, Lee AH, Iwakoshi NN, Ozdelen E, Tuncman G, Gorgun C, Glimcher LH, Hotamisligil GS (2004) Endoplasmic reticulum stress links obesity, insulin action, and type 2 diabetes. Science 306(5695):457–461. CrossRefPubMedGoogle Scholar
  41. Ozcan L, Ergin AS, Lu A, Chung J, Sarkar S, Nie D, Myers MG Jr, Ozcan U (2009) Endoplasmic reticulum stress plays a central role in development of leptin resistance. Cell Metab 9(1):35–51. CrossRefPubMedGoogle Scholar
  42. Panda S, Kar A (2006) Evaluation of the antithyroid, antioxidative and antihyperglycemic activity of scopoletin from Aegle marmelos leaves in hyperthyroid rats. Phytother Res 20(12):1103–1105. CrossRefPubMedGoogle Scholar
  43. Ron D, Walter P (2007) Signal integration in the endoplasmic reticulum unfolded protein response. Nat Rev Mol Cell Biol 8(7):519–529. CrossRefPubMedGoogle Scholar
  44. Rutkowski DT, Kaufman RJ (2007) That which does not kill me makes me stronger: adapting to chronic ER stress. Trends Biochem Sci 32(10):469–476. CrossRefPubMedGoogle Scholar
  45. Scheuner D, Vander Mierde D, Song B, Flamez D, Creemers JW, Tsukamoto K, Ribick M, Schuit FC, Kaufman RJ (2005) Control of mRNA translation preserves endoplasmic reticulum function in beta cells and maintains glucose homeostasis. Nat Med 11(7):757–764. CrossRefPubMedGoogle Scholar
  46. Seo HY, Kim YD, Lee KM, Min AK, Kim MK, Kim HS, Won KC, Park JY, Lee KU, Choi HS, Park KG, Lee IK (2008) Endoplasmic reticulum stress-induced activation of activating transcription factor 6 decreases insulin gene expression via up-regulation of orphan nuclear receptor small heterodimer partner. Endocrinology 149(8):3832–3841. CrossRefPubMedPubMedCentralGoogle Scholar
  47. Siatka T, Kasparova M (2008) Effects of auxins on growth and scopoletin accumulation in cell suspension cultures of Angelica archangelica L. Ceska Slov Farm 57(1):17–20PubMedGoogle Scholar
  48. Verma A, Dewangan P, Kesharwani D, Kela PS (2013) Hypoglycemic and hypolipidemic activity of scopoletin (coumarin derivative) in streptozotocin induced diabetic rats. Int J Pharm Sci Rev Res 22(1):79–83Google Scholar
  49. Wang Y, Vera L, Fischer WH, Montminy M (2009) The CREB coactivator CRTC2 links hepatic ER stress and fasting gluconeogenesis. Nature 460(7254):534–537. CrossRefPubMedPubMedCentralGoogle Scholar
  50. Yao X, Ding Z, Xia Y, Wei Z, Luo Y, Feleder C, Dai Y (2012) Inhibition of monosodium urate crystal-induced inflammation by scopoletin and underlying mechanisms. Int Immunopharmacol 14(4):454–462. CrossRefPubMedGoogle Scholar
  51. Ye J, Rawson RB, Komuro R, Chen X, Dave UP, Prywes R, Brown MS, Goldstein JL (2000) ER stress induces cleavage of membrane-bound ATF6 by the same proteases that process SREBPs. Mol Cell 6(6):1355–1364CrossRefPubMedGoogle Scholar
  52. Yogalakshmi B, Bhuvaneswari S, Sreeja S, Anuradha CV (2014) Grape seed proanthocyanidins and metformin act by different mechanisms to promote insulin signaling in rats fed high calorie diet. J Cell Commun Signal 8(1):13–22. CrossRefPubMedGoogle Scholar
  53. Yoshida H, Matsui T, Yamamoto A, Okada T, Mori K (2001) XBP1 mRNA is induced by ATF6 and spliced by IRE1 in response to ER stress to produce a highly active transcription factor. Cell 107(7):881–891CrossRefPubMedGoogle Scholar
  54. Zhang P, McGrath B, Li S, Frank A, Zambito F, Reinert J, Gannon M, Ma K, McNaughton K, Cavener DR (2002) The PERK eukaryotic initiation factor 2 alpha kinase is required for the development of the skeletal system, postnatal growth, and the function and viability of the pancreas. Mol Cell Biol 22(11):3864–3874CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Cell Stress Society International 2018

Authors and Affiliations

  • Kalaivanan Kalpana
    • 1
  • Emayavaramban Priyadarshini
    • 1
  • S. Sreeja
    • 1
  • Kalivarathan Jagan
    • 1
  • Carani Venkatraman Anuradha
    • 1
  1. 1.Department of Biochemistry and BiotechnologyAnnamalai UniversityAnnamalai NagarIndia

Personalised recommendations