Skip to main content

Advertisement

SpringerLink
Log in

PNPLA3 I148M is involved in the variability in anti-NAFLD response to exenatide

  • Original Article
  • Published:
Endocrine Aims and scope Submit manuscript

Abstract

Purpose

GLP-1 receptor agonists, such as exenatide, have been proven to attenuate nonalcoholic fatty liver disease (NAFLD) in vivo and in vitro. However, the efficiency of exenatide had interindividual differences. PNPLA3 is a major susceptibility gene for NAFLD and its I148M polymorphism increases the risk of all disorders of the NAFLD spectrum. Whether this variant contributes to variability in exenatide response is still unclear.

Methods

PNPLA3 148I knockin HepG2 cells were constructed using the Cas9/sgRNA system. Oil Red O staining combined with TG quantification was used to evaluate lipid accumulation. Western blotting and qRT-qPCR were conducted, respectively, to measure the protein and mRNA expression of lipid metabolic and endoplasmic reticulum (ER) stress-related inflammatory markers. PNPLA3 I148M was genotyped in type 2 diabetics using Sanger sequencing. The exenatide-induced changes in liver fat content and other clinical parameters were compared between PNPLA3 I148M genotypes.

Results

Lipid deposition increased in both PNPLA3 148I/I and 148M/M HepG2 cells treated with palmitoleic acid, while cells with 148M/M had a higher TG content than those with 148I/I. Exendin-4 treatment was showed to be more significant in 148I/I cells than in 148M/M cells in terms of reducing the intrahepatic fat content, inhibiting SREBP-1c and ER stress-related inflammation, and activating AMPK-ACC lipid oxidation pathway. In patients with type 2 diabetes, 24-week treatment with exenatide improved liver fat content in patients carrying PNPLA3 148I/I better than in patients with 148M/M.

Conclusions

PNPLA3 I148M might modify the anti-NAFLD response to exenatide.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

Abbreviations

NAFLD:

Nonalcoholic fatty liver disease

T2DM:

Type 2 diabetes mellitus

PNPLA3:

Patatin-like phospholipase domain protein 3

PA:

palmitate

TG:

Triglyceride

ER:

Endoplasmic reticulum

SREBP-1c:

Sterol regulatory element binding protein 1

Bip:

Binding protein

JNK:

Jun N-terminal kinase

ACC:

Acetyl CoA carboxylase

AMPK:

AMP-activated protein kinase

References

  1. H. Tilg, A.R. Moschen, M. Roden, NAFLD and diabetes mellitus. Nat. Rev. Gastroenterol. Hepatol. 14(1), 32–42 (2017). https://doi.org/10.1038/nrgastro.2016.147

    Article  CAS  PubMed  Google Scholar 

  2. B. Fruci, S. Giuliano, A. Mazza, R. Malaguarnera, A. Belfiore, Nonalcoholic fatty liver: a possible new target for type 2 diabetes prevention and treatment. Int. J. Mol. Sci. 14(11), 22933–22966 (2013). https://doi.org/10.3390/ijms141122933

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. A.A. Alsabaani, A.A. Mahfouz, N.J. Awadalla, M.J. Musa, S.M. Al Humayed, Non-Alcoholic Fatty Liver Disease among Type-2 Diabetes Mellitus Patients in Abha City, South Western Saudi Arabia. Int. J. Environ. Res. Public Health 15(11), (2018). https://doi.org/10.3390/ijerph15112521

  4. G. Targher, C.D. Byrne, A. Lonardo, G. Zoppini, C. Barbui, Non-alcoholic fatty liver disease and risk of incident cardiovascular disease: a meta-analysis. J. Hepatol. 65(3), 589–600 (2016). https://doi.org/10.1016/j.jhep.2016.05.013

    Article  PubMed  Google Scholar 

  5. E. Vilar-Gomez, Y. Martinez-Perez, L. Calzadilla-Bertot, A. Torres-Gonzalez, B. Gra-Oramas, L. Gonzalez-Fabian, S.L. Friedman, M. Diago, M. Romero-Gomez, Weight loss through lifestyle modification significantly reduces features of nonalcoholic steatohepatitis. Gastroenterology 149(2), 367–378.e365 (2015). https://doi.org/10.1053/j.gastro.2015.04.005. quiz e314-365

    Article  PubMed  Google Scholar 

  6. P. Farzanegi, A. Dana, Z. Ebrahimpoor, M. Asadi, M.A. Azarbayjani, Mechanisms of beneficial effects of exercise training on non-alcoholic fatty liver disease (NAFLD): Roles of oxidative stress and inflammation. Eur. J. Sport Sci. 19(7), 994–1003 (2019). https://doi.org/10.1080/17461391.2019.1571114

    Article  PubMed  Google Scholar 

  7. X.C. Wang, A.M. Gusdon, H. Liu, S. Qu, Effects of glucagon-like peptide-1 receptor agonists on non-alcoholic fatty liver disease and inflammation. World J. Gastroenterol. 20(40), 14821–14830 (2014). https://doi.org/10.3748/wjg.v20.i40.14821

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. L. Iogna Prat, E.A. Tsochatzis, The effect of antidiabetic medications on non-alcoholic fatty liver disease (NAFLD). Hormones 17(2), 219–229 (2018). https://doi.org/10.1007/s42000-018-0021-9

    Article  PubMed  Google Scholar 

  9. P. Pingitore, K. Sasidharan, M. Ekstrand, S. Prill, D. Lindén, S. Romeo, Human multilineage 3D spheroids as a model of liver steatosis and fibrosis. Int. J. Mol. Sci. 20(7), (2019). https://doi.org/10.3390/ijms20071629

  10. D.L. Williams, D.G. Baskin, M.W. Schwartz, Leptin regulation of the anorexic response to glucagon-like peptide-1 receptor stimulation. Diabetes 55(12), 3387–3393 (2006). https://doi.org/10.2337/db06-0558

    Article  CAS  PubMed  Google Scholar 

  11. M. Kalogirou, E. Sinakos, Treating nonalcoholic steatohepatitis with antidiabetic drugs: Will GLP-1 agonists end the struggle? World J. Hepatol. 10(11), 790–794 (2018). https://doi.org/10.4254/wjh.v10.i11.790

    Article  PubMed  PubMed Central  Google Scholar 

  12. M.J. Armstrong, P. Gaunt, G.P. Aithal, D. Barton, D. Hull, R. Parker, J.M. Hazlehurst, K. Guo, G. Abouda, M.A. Aldersley, D. Stocken, S.C. Gough, J.W. Tomlinson, R.M. Brown, S.G. Hübscher, P.N. Newsome, Liraglutide safety and efficacy in patients with non-alcoholic steatohepatitis (LEAN): a multicentre, double-blind, randomised, placebo-controlled phase 2 study. Lancet 387(10019), 679–690 (2016). https://doi.org/10.1016/s0140-6736(15)00803-x

    Article  CAS  PubMed  Google Scholar 

  13. Y. Bi, B. Zhang, W. Xu, H. Yang, W. Feng, C. Li, G. Tong, M. Li, X. Wang, S. Shen, B. Zhu, J. Weng, D. Zhu, Effects of exenatide, insulin, and pioglitazone on liver fat content and body fat distributions in drug-naive subjects with type 2 diabetes. Acta Diabetol. 51(5), 865–873 (2014). https://doi.org/10.1007/s00592-014-0638-3

    Article  CAS  PubMed  Google Scholar 

  14. C. Pirazzi, M. Adiels, M.A. Burza, R.M. Mancina, M. Levin, M. Stahlman, M.R. Taskinen, M. Orho-Melander, J. Perman, A. Pujia, L. Andersson, C. Maglio, T. Montalcini, O. Wiklund, J. Boren, S. Romeo, Patatin-like phospholipase domain-containing 3 (PNPLA3) I148M (rs738409) affects hepatic VLDL secretion in humans and in vitro. J. Hepatol. 57(6), 1276–1282 (2012). https://doi.org/10.1016/j.jhep.2012.07.030

    Article  CAS  PubMed  Google Scholar 

  15. S. Sookoian, C.J. Pirola, Meta-analysis of the influence of I148M variant of patatin-like phospholipase domain containing 3 gene (PNPLA3) on the susceptibility and histological severity of nonalcoholic fatty liver disease. Hepatology 53(6), 1883–1894 (2011). https://doi.org/10.1002/hep.24283

    Article  CAS  PubMed  Google Scholar 

  16. Y.L. Liu, G.L. Patman, J.B. Leathart, A.C. Piguet, A.D. Burt, J.F. Dufour, C.P. Day, A.K. Daly, H.L. Reeves, Q.M. Anstee, Carriage of the PNPLA3 rs738409 C>G polymorphism confers an increased risk of non-alcoholic fatty liver disease associated hepatocellular carcinoma. J. Hepatol. 61(1), 75–81 (2014). https://doi.org/10.1016/j.jhep.2014.02.030

    Article  CAS  PubMed  Google Scholar 

  17. E. Smagris, S. BasuRay, J. Li, Y. Huang, K.M. Lai, J. Gromada, J.C. Cohen, H.H. Hobbs, Pnpla3I148M knockin mice accumulate PNPLA3 on lipid droplets and develop hepatic steatosis. Hepatology 61(1), 108–118 (2015). https://doi.org/10.1002/hep.27242

    Article  CAS  PubMed  Google Scholar 

  18. D. Lindén, A. Ahnmark, P. Pingitore, E. Ciociola, I. Ahlstedt, A.C. Andréasson, K. Sasidharan, K. Madeyski-Bengtson, M. Zurek, R.M. Mancina, A. Lindblom, M. Bjursell, G. Böttcher, M. Ståhlman, Y.M. Bohlooly, W.G. Haynes, B. Carlsson, M. Graham, R. Lee, S. Murray, L. Valenti, S. Bhanot, P. Åkerblad, S. Romeo, Pnpla3 silencing with antisense oligonucleotides ameliorates nonalcoholic steatohepatitis and fibrosis in Pnpla3 I148M knock-in mice. Mol. Metab. 22, 49–61 (2019). https://doi.org/10.1016/j.molmet.2019.01.013

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. A. Moravcová, Z. Červinková, O. Kučera, V. Mezera, D. Rychtrmoc, H. Lotková, The effect of oleic and palmitic acid on induction of steatosis and cytotoxicity on rat hepatocytes in primary culture. Physiol. Res. 64(Suppl 5), S627–S636 (2015). https://doi.org/10.33549/physiolres.933224

    Article  PubMed  Google Scholar 

  20. W. Xu, Y. Bi, Z. Sun, J. Li, L. Guo, T. Yang, G. Wu, L. Shi, Z. Feng, L. Qiu, Q. Li, X. Guo, Z. Luo, J. Lu, Z. Shan, W. Yang, Q. Ji, L. Yan, H. Li, X. Yu, S. Li, Z. Zhou, X. Lv, Z. Liang, S. Lin, L. Zeng, J. Yan, L. Ji, J. Weng, Comparison of the effects on glycaemic control and beta-cell function in newly diagnosed type 2 diabetes patients of treatment with exenatide, insulin or pioglitazone: a multicentre randomized parallel-group trial (the CONFIDENCE study). J. Intern. Med. 277(1), 137–150 (2015). https://doi.org/10.1111/joim.12293

    Article  CAS  PubMed  Google Scholar 

  21. M.F. Xia, Y. Ling, H. Bian, H.D. Lin, H.M. Yan, X.X. Chang, X.M. Li, H. Ma, D. Wang, L.S. Zhang, S.S. Wang, B.J. Wu, W.Y. He, N.Q. Zhao, X. Gao, I148M variant of PNPLA3 increases the susceptibility to non-alcoholic fatty liver disease caused by obesity and metabolic disorders. Aliment. Pharmacol. Ther. 43(5), 631–642 (2016). https://doi.org/10.1111/apt.13521

    Article  CAS  PubMed  Google Scholar 

  22. G. Aragones, T. Auguet, S. Armengol, A. Berlanga, E. Guiu-Jurado, C. Aguilar, S. Martinez, F. Sabench, J.A. Porras, M.D. Ruiz, M. Hernandez, J.J. Sirvent, D. Del Castillo, C. Richart, PNPLA3 expression is related to liver steatosis in morbidly obese women with non-alcoholic fatty liver disease. Int. J. Mol. Sci. 17(5) (2016). https://doi.org/10.3390/ijms17050630

  23. J.Z. Li, Y. Huang, R. Karaman, P.T. Ivanova, H.A. Brown, T. Roddy, J. Castro-Perez, J.C. Cohen, H.H. Hobbs, Chronic overexpression of PNPLA3I148M in mouse liver causes hepatic steatosis. J. Clin. Invest. 122(11), 4130–4144 (2012). https://doi.org/10.1172/jci65179

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. W. Chen, B. Chang, L. Li, L. Chan, Patatin-like phospholipase domain-containing 3/adiponutrin deficiency in mice is not associated with fatty liver disease. Hepatology 52(3), 1134–1142 (2010). https://doi.org/10.1002/hep.23812

    Article  CAS  PubMed  Google Scholar 

  25. Y.G. Wang, T.L. Yang, Liraglutide reduces oxidized LDL-induced oxidative stress and fatty degeneration in Raw 264.7 cells involving the AMPK/SREBP1 pathway. J. Geriatr. Cardiol. 12(4), 410–416 (2015). https://doi.org/10.11909/j.issn.1671-5411.2015.04.013

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. J. Lee, S.W. Hong, S.E. Park, E.J. Rhee, C.Y. Park, K.W. Oh, S.W. Park, W.Y. Lee, Exendin-4 inhibits the expression of SEPP1 and Fetuin-A via improvement of palmitic acid-induced endoplasmic reticulum stress by AMPK. Endocrinol. Metab. 30(2), 177–184 (2015). https://doi.org/10.3803/EnM.2015.30.2.177

    Article  CAS  Google Scholar 

  27. S. Sharma, J.E. Mells, P.P. Fu, N.K. Saxena, F.A. Anania, GLP-1 analogs reduce hepatocyte steatosis and improve survival by enhancing the unfolded protein response and promoting macroautophagy. PLoS ONE 6(9), (2011). https://doi.org/10.1371/journal.pone.0025269

  28. C.M. Jenkins, D.J. Mancuso, W. Yan, H.F. Sims, B. Gibson, R.W. Gross, Identification, cloning, expression, and purification of three novel human calcium-independent phospholipase A2 family members possessing triacylglycerol lipase and acylglycerol transacylase activities. J. Biol. Chem. 279(47), 48968–48975 (2004). https://doi.org/10.1074/jbc.M407841200

    Article  CAS  PubMed  Google Scholar 

  29. S. Romeo, A. Sanyal, L. Valenti, Leveraging human genetics to identify potential new treatments for fatty liver disease. Cell Metab. 31(1), 35–45 (2020). https://doi.org/10.1016/j.cmet.2019.12.002

    Article  CAS  PubMed  Google Scholar 

  30. M. Kumari, G. Schoiswohl, C. Chitraju, M. Paar, I. Cornaciu, A.Y. Rangrez, N. Wongsiriroj, H.M. Nagy, P.T. Ivanova, S.A. Scott, O. Knittelfelder, G.N. Rechberger, R. Birner-Gruenberger, S. Eder, H.A. Brown, G. Haemmerle, M. Oberer, A. Lass, E.E. Kershaw, R. Zimmermann, R. Zechner, Adiponutrin functions as a nutritionally regulated lysophosphatidic acid acyltransferase. Cell Metab. 15(5), 691–702 (2012). https://doi.org/10.1016/j.cmet.2012.04.008

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. S. Yuan, H. Liu, D. Yuan, J. Xu, Y. Chen, X. Xu, F. Xu, H. Liang, PNPLA3 I148M mediates the regulatory effect of NF-kB on inflammation in PA-treated HepG2 cells. J. Cell. Mole. Med. 24(2), 1541–1552 (2020). https://doi.org/10.1111/jcmm.14839

    Article  CAS  Google Scholar 

  32. F.V. Bruschi, T. Claudel, M. Tardelli, A. Caligiuri, T.M. Stulnig, F. Marra, M. Trauner, The PNPLA3 I148M variant modulates the fibrogenic phenotype of human hepatic stellate cells. Hepatology 65(6), 1875–1890 (2017). https://doi.org/10.1002/hep.29041

    Article  CAS  PubMed  Google Scholar 

  33. H. Liang, J. Xu, F. Xu, H. Liu, D. Yuan, S. Yuan, M. Cai, J. Yan, J. Weng, The SRE Motif in the human PNPLA3 promoter (-97 to -88 bp) mediates transactivational effects of SREBP-1c. J. Cell. Physiol. 230(9), 2224–2232 (2015). https://doi.org/10.1002/jcp.24951

    Article  CAS  PubMed  Google Scholar 

  34. F. Xu, Z. Li, X. Zheng, H. Liu, H. Liang, H. Xu, Z. Chen, K. Zeng, J. Weng, SIRT1 mediates the effect of GLP-1 receptor agonist exenatide on ameliorating hepatic steatosis. Diabetes 63(11), 3637–3646 (2014). https://doi.org/10.2337/db14-0263

    Article  CAS  PubMed  Google Scholar 

  35. S. Suzuki, J.R. Iben, S.L. Coon, T. Kino, SIRT1 is a transcriptional enhancer of the glucocorticoid receptor acting independently to its deacetylase activity. Mol. Cell. Endocrinol. 461, 178–187 (2018). https://doi.org/10.1016/j.mce.2017.09.012

    Article  CAS  PubMed  Google Scholar 

  36. Y. Li, K. Wong, A. Giles, J. Jiang, J.W. Lee, A.C. Adams, A. Kharitonenkov, Q. Yang, B. Gao, L. Guarente, M. Zang, Hepatic SIRT1 attenuates hepatic steatosis and controls energy balance in mice by inducing fibroblast growth factor 21. Gastroenterology 146(2), 539–549.e537 (2014). https://doi.org/10.1053/j.gastro.2013.10.059

    Article  CAS  PubMed  Google Scholar 

  37. Y.M. Liu, M. Moldes, J.P. Bastard, E. Bruckert, N. Viguerie, B. Hainque, A. Basdevant, D. Langin, J. Pairault, K. Clement, Adiponutrin: a new gene regulated by energy balance in human adipose tissue. J. Clin. Endocrinol. Metab. 89(6), 2684–2689 (2004). https://doi.org/10.1210/jc.2003-031978

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank Professor Yan Bi for her kind help in collecting clinical data.

Funding

The study was supported by the National Natural Science Foundation of China (No. 81370909); Guangdong Natural Science Foundation (No. 2020A1515010226) and Local Innovative and Research Teams Project of Guangdong Pearl River Talents Program (No. 2017BT01S131).

Author information

Author notes

  1. These authors contributed equally: Yunzhi Chen, Xuemei Yan

Authors and Affiliations

  1. Department of Endocrinology and Metabolism, Third Affiliated Hospital of Sun Yat-Sen University, and Guangdong Provincial Key Laboratory of Diabetology, Guangzhou, China

    Yunzhi Chen, Xiao Xu, Fen Xu & Hua Liang

  2. Department of Endocrinology and Metabolism, No. 903 Hospital of PLA Joint Logistic Support Force, Hangzhou, China

    Xuemei Yan

  3. Department of Endocrinology, Guangzhou Panyu Central Hospital, Guangzhou, China

    Shuhua Yuan

Authors
  1. Yunzhi Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Xuemei Yan
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Xiao Xu
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Shuhua Yuan
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Fen Xu
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Hua Liang
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

Y.C. and X.Y. performed the experiments. X.X. and S.Y. participated in the analyzed the data. F.X. interpreted the data. H.L. conceived and designed the experiments and drafted the manuscript.

Corresponding author

Correspondence to Hua Liang.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Chen, Y., Yan, X., Xu, X. et al. PNPLA3 I148M is involved in the variability in anti-NAFLD response to exenatide. Endocrine 70, 517–525 (2020). https://doi.org/10.1007/s12020-020-02470-7

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12020-020-02470-7

Keywords

Search

Navigation