Vol 61, No 4 (2023)
Original paper
Published online: 2023-11-28

open access

Page views 1006
Article views/downloads 623
Get Citation

Connect on Social Media

Connect on Social Media

Therapeutic effect of autophagy induced by rapamycin versus intermittent fasting in animal model of fatty liver

Sara Adel Hosny1, Mohammed Hafez Ahmed Moustafa1, Fatma Mahmoud Mehina1, Marwa Mohamed Sabry1
Pubmed: 38013515
Folia Histochem Cytobiol 2023;61(4):205-216.


Introduction. High-fructose, high-fat diet consumption (HFHF) is one of the primary causes of non-alcoholic fatty liver disease (NAFLD), which is due to impaired beta-oxidation or apolipoprotein secretion by hepatocytes. Activation of autophagy in hepatocytes could be a therapeutic method against hepatic complications. This study was designed to compare effects of rapamycin and intermittent fasting-inducing autophagy in rats with experimentally induced nonalcoholic fatty liver. Material and methods. Male rats were divided into five groups: C (control, n = 6), the experimental group (EX) subdivided, EXIa (HFHF, n = 6), EXIb (recovery, n = 6), EXII (rapamycin, n = 6) and EXIII (intermittent fasting, n = 6). All rats in the experimental group received HFHF diet for 8 weeks to induce nonalcoholic-fatty liver and obesity. Then, for the next 8 weeks the animals received either a daily oral dose of rapamycin (EXII group) or to intermittent fasting (IF) for 16 hours daily (EXIII group). Blood samples were drawn, and serum TG concentration as well as ALT and AST activities were determined. Hepatic sections were examined by light and electron microscopy. LC3B immunohistochemical staining, morphometric and statistical studies were performed. Results. Subgroups EXIa (HFHF subgroup) and EXIb (Recovery subgroup) showed marked increase in TG, ALT, and AST levels associated with loss of normal hepatic architecture, cytoplasmic vacuolations and faint LC3B immunoreactivity. Ultrathin sections exhibited many autophagosomes in hepatocytes. On the other hand, rapamycin (EXII) and IF (EXIII) groups showed significant improvement to a variable extent in comparison to EXI groups. Conclusions. It could be concluded that rapamycin and intermittent fasting significantly improved NAFLD-induced changes of liver structure and function by inducing autophagy in hepatocytes.

Article available in PDF format

View PDF Download PDF file


  1. Mao YQ, Yu F, Wang J, et al. Autophagy: a new target for nonalcoholic fatty liver disease therapy. Hepat Med. 2016; 8: 27–37.
  2. Allaire M, Rautou PE, Codogno P, et al. Autophagy in liver diseases: Time for translation? J Hepat. 2019; 70(5): 985–998.
  3. Yu X, Zhang L, Lu H, et al. Rapamycin protects against hepatic injury in streptozotocin-induced diabetic rats by enhancing autophagy. Int J Clin Exp Med. 2019; 12(9): 11383–11393.
  4. Saxton R, Sabatini D. mTOR Signaling in Growth, Metabolism, and Disease. Cell. 2017; 168(6): 960–976.
  5. Hahn D, Hodson EM, Hamiwka LA, et al. Target of rapamycin inhibitors (TOR-I; sirolimus and everolimus) for primary immunosuppression in kidney transplant recipients. Cochrane Database Syst Rev. 2019; 12(12): CD004290.
  6. Tian T, Li X, Zhang J. mTOR Signaling in Cancer and mTOR Inhibitors in Solid Tumor Targeting Therapy. Int J Mol Sci. 2019; 20(3): 755.
  7. Palavra F, Robalo C, Reis F. Recent advances and challenges of mTOR inhibitors use in the treatment of patients with Tuberous Sclerosis Complex. Oxid Med Cell Longev. 2017; 2017: 9820181.
  8. Almeneessier AS, Pandi-Perumal S, BaHammam A. Intermittent fasting, insufficient sleep, and circadian rhythm: interaction and effects on the cardiometabolic system. Curr Sleep Medicine Rep. 2018; 4(3): 179–195.
  9. Li L, Su Y, Li F, et al. The effects of daily fasting hours on shaping gut microbiota in mice. BMC Microbiol. 2020; 20(1): 65.
  10. Mukai R, Zablocki D, Sadoshima J. Intermittent fasting reverses an advanced form of cardiomyopathy. J Am Heart Assoc. 2019; 8(4): e011863.
  11. Ragab SM, Omar HE, Kh S. Hypolipidemic and antioxidant effects of phytochemical compounds against hepatic steatosis induced by high-fat high sucrose diet in rats. Archives of Biomedical Sciences. 2014; 2(1): 1–10.
  12. van Herck M, Vonghia L, Francque S. Animal models of nonalcoholic fatty liver disease — a starter’s guide. Nutrients. 2017; 9(10): 1072.
  13. Aouichat S, Chayah M, Bouguerra-Aouichat S, et al. Time-restricted feeding improves body weight gain, lipid profiles, and atherogenic indices in cafeteria-diet-fed rats: role of browning of inguinal white adipose tissue. Nutrients. 2020; 12(8): 2185.
  14. Kiernan JK. Histological and histochemical methods. In: Theory and practice. 3rd ed. Arnold Publisher, London, New York, and New Delhy 2001: 111–162.
  15. Woods AE, Stirling JW. Electron microscopy. In: Bancroft JD, Gamble M. ed. Theory and practice of histological techniques, 6th ed. Churchill Livingstone Elsevier, Edinburgh 2008: 601–636.
  16. Ayoubi R, McPherson PS, Laflamme C. Antibody Screening by Immunoblot. https://zenodo.org/records/5717510 (6.05.2023).
  17. Cui F, Hu H, Guo J, et al. The effect of autophagy on chronic intermittent hypobaric hypoxia ameliorating liver damage in metabolic syndrome rats. Front Physiol. 2020; 11: 13.
  18. Khambu B, Yan S, Huda N, et al. Autophagy in non-alcoholic fatty liver disease and alcoholic liver disease. Liver Res. 2018; 2(3): 112–119.
  19. Gao H, Zhou L, Zhong Y, et al. Kindlin-2 haploinsufficiency protects against fatty liver by targeting Foxo1 in mice. Nat Commun. 2022; 13(1): 1025.
  20. Qian Ll, Wu L, Zhang L, et al. Serum biomarkers combined with ultrasonography for early diagnosis of non-alcoholic fatty liver disease confirmed by magnetic resonance spectroscopy. Acta Pharmacol Sin. 2019; 41(4): 554–560.
  21. Albadawy R, Hasanin A, Agwa S, et al. Rosavin Ameliorates Hepatic Inflammation and Fibrosis in the NASH Rat Model via Targeting Hepatic Cell Death. Int J Mol Sci. 2022; 23(17): 10148.
  22. Tomizawa M, Kawanabe Y, Shinozaki F, et al. Triglyceride is strongly associated with nonalcoholic fatty liver disease among markers of hyperlipidemia and diabetes. Biomed Rep. 2014; 2(5): 633–636.
  23. Ore A, Ugbaja R, Adeogun A, et al. An albino mouse model of nonalcoholic fatty liver disease induced using high-fat liquid “Lieber-DeCarli” diet: a preliminary investigation. Porto Biomed J. 2020; 5(4): e071.
  24. Reddy JK, Rao MS. Lipid Metabolism and Liver Inflammation. II. Fatty liver disease and fatty acid oxidation. Am J Physiol Gastrointest Liver Physiol. 2006; 290(5): G852–G858.
  25. Rives C, Fougerat A, Ellero-Simatos S, et al. Oxidative Stress in NAFLD: Role of Nutrients and Food Contaminants. Biomolecules. 2020; 10(12): 1702.
  26. Okasha E, Hassan N, Soliman G, et al. Histological, immunohistochemical, and biochemical study of experimentally induced fatty liver in adult male albino rat and the possible protective role of pomegranate. J Microsc Ultrastruct. 2018; 6(1): 44–55.
  27. Kořínková L, Pražienková V, Černá L, et al. Pathophysiology of NAFLD and NASH in experimental models: the role of food intake regulating peptides. Front Endocrinol (Lausanne). 2020; 11: 597583.
  28. Ao Na, Yang J, Wang X, et al. Glucagon‐like peptide‐1 preserves non‐alcoholic fatty liver disease through inhibition of the endoplasmic reticulum stress‐associated pathway. Hepatol Res. 2016; 46(4): 343–353.
  29. Abdeen M, Zaghloul D, Saleh R, et al. Histomorphometric evidence of hepatic recovery of rats fed repeatedly heated palm oil. Egyptian Journal of Histology. 2022; 45(1): 115–124.
  30. Martinet W, Schrijvers DM, Timmermans JP, et al. Immunohistochemical analysis of macroautophagy. Autophagy. 2014; 9(3): 386–402.
  31. Korovila I, Höhn A, Jung T, et al. Reduced liver autophagy in high-fat diet induced liver steatosis in New Zealand obese mice. Antioxidants. 2021; 10(4): 501.
  32. Zhang Z, Qian Q, Li M, et al. The unfolded protein response regulates hepatic autophagy by sXBP1-mediated activation of TFEB. Autophagy. 2020; 17(8): 1841–1855.
  33. Chang GR, Chiu YS, Wu YY, et al. Rapamycin protects against high fat diet–induced obesity in C57BL/6J mice. J Pharmacol Sci. 2009; 109(4): 496–503.
  34. Pena-Leon V, Perez-Lois R, Seoane L. mTOR pathway is involved in energy homeostasis regulation as a part of the gut–brain axis. Int J Mol Sci. 2020; 21(16): 5715.
  35. Zhao R, Zhu M, Zhou S, et al. Rapamycin-Loaded mPEG-PLGA nanoparticles ameliorate hepatic steatosis and liver injury in non-alcoholic fatty liver disease. Front Chem. 2020; 8: 407.
  36. Alers S, Löffler A, Wesselborg S, et al. Role of AMPK-mTOR-Ulk1/2 in the regulation of autophagy: cross talk, shortcuts, and feedbacks. Molecular and Cellular Biology. 2023; 32(1): 2–11.
  37. Ma N, Wang YK, Xu S, et al. PPDPF alleviates hepatic steatosis through inhibition of mTOR signaling. Nat Commun. 2021; 12(1): 3059.
  38. Feng J, Qiu S, Zhou S, et al. mTOR: a potential new target in nonalcoholic fatty liver disease. Int J Mol Sci. 2022; 23(16): 9196.
  39. Gosis B, Wada S, Thorsheim C, et al. Inhibition of nonalcoholic fatty liver disease in mice by selective inhibition of mTORC1. Science. 2022; 376(6590).
  40. Periyasamy-Thandavan S, Jiang M, Schoenlein P, et al. Autophagy: molecular machinery, regulation, and implications for renal pathophysiology. Am J Physiol Renal Physiol. 2009; 297(2): F244–F256.
  41. Diao C, Wang L, Liu H, et al. Aged kidneys are refractory to autophagy activation in a rat model of renal ischemia-reperfusion injury. Clin Interv Aging. 2019; Volume 14: 525–534.
  42. Harahap H, Esa A, Erny K. The Effect of Intermitent Fasting (Time Restriction Feeding) on Body Weight, Aspartate Transaminase and Alkaline Transaminase in Sprague Dawley Rats. https://www.researchgate.net/publication/345039438_The_Effect_of_Intermitent_Fasting_Time_Restriction_Feeding_on_Body_Weight_Aspartate_Transaminase_and_Alkaline_Transaminase_in_Sprague_Dawley_Rats (6.05.2023).
  43. Yin C, Li Z, Xiang Y, et al. Effect of intermittent fasting on non-alcoholic fatty liver disease: systematic review and meta-analysis. Front Nutr. 2021; 8: 709683.
  44. Ahmed K, Arisha A, Sharsher S. The influence of intermittent fasting regimens on the regulatory mechanisms of metabolic health. Zagazig Veterinary Journal. 2021; 49(1): 56–66.
  45. Castro-de-Paiva Pde, Marinho T, Mandarim-de-Lacerda C, et al. Intermittent fasting, high-intensity interval training, or a combination of both have beneficial effects in obese mice with nonalcoholic fatty liver disease. J Nutr Biochem. 2022; 104: 108997.
  46. Ma YN, Jiang X, Tang W. Influence of intermittent fasting on autophagy in the liver. Biosci Trends. 2023; 17(5): 335–355.
  47. Yuan W, He X, Morin D, et al. Autophagy induction contributes to the neuroprotective impact of intermittent fasting on the acutely injured spinal cord. J Neurotrauma. 2021; 38(3): 373–384.
  48. Ebrahim HA, El-Gamal R, Sherif RN. Intermittent fasting attenuates high-fat diet-induced cerebellar changes in rats: involvement of tnf-α, autophagy, and oxidative stress. Cells Tissues Organs. 2021; 210(5-6): 351–367.