Chemical chaperon 4-phenylbutric acid improves cardiac function following isoproterenol-induced myocardial infarction in rats

Document Type : Original Article


1 Pharmacology and Toxicology Department, Faculty of Pharmacy, Tabriz University of Medical Sciences, Tabriz, Iran

2 Student Research Committee, Tabriz University of Medical Sciences, Tabriz, Iran


Objective(s): 4-Phenyl butyric acid (4-PBA) is a chaperone-mediated autophagy (CMA) inducer, which eliminates unnecessary and damaged cellular components through lysosomal enzymes. It could reduce misfolded and unfolded proteins produced after myocardial infarction (MI) and can improve cardiac function. We aimed to investigate the effect of 4-PBA on isoproterenol-induced MI in rats. 
Materials and Methods: Isoproterenol (100 mg/kg) was injected subcutaneously for two consecutive days simultaneous with an intraperitoneal (IP) injection of 4-PBA at 20, 40, or 80 mg/kg at 24-hr intervals for five days. On day 6, hemodynamic parameters, histopathological changes, peripheral neutrophil count, and total anti-oxidant capacity (TAC) were evaluated. The expression of autophagy proteins was measured by using western blotting. 4-PBA significantly improved post-MI changes in hemodynamic parameters.
Results: Histological improvement was found in 4-PBA 40 mg/kg (P<0.05). The neutrophil count in the peripheral blood significantly decreased in the treatment groups compared with isoproterenol. Furthermore, 4-PBA at 80 mg/kg significantly increased the serum TAC compared with isoproterenol (P<0.001). Western blotting showed a significant decrease in the P62 level (P<0.05) of 40 and 80 mg/kg 4-PBA treated groups.
Conclusion: This study demonstrated that 4-PBA could have a cardio-protective effect against isoproterenol-induced MI, which can be due to autophagy modulation and oxidative stress inhibition. Obtaining effective results in different doses shows the need for an optimum degree of cell autophagic activity.


1. Shepard D, VanderZanden A, Moran A, Naghavi M, Murray C, Roth G. Ischemic heart disease worldwide, 1990 to 2013: Estimates from the global burden of disease study 2013. Circ Cardiovasc Qual Outcomes 2015; 8:455-456. 
2. Frangogiannis NG, Smith CW, Entman ML. The inflammatory response in myocardial infarction. Cardiovasc Res 2002; 53:31-47. 
3. Thorp EB. The myocardial unfolded protein response during ischemic cardiovascular disease. Biochem Res Int 2012; 2012:583170. 
4. Thuerauf DJ, Marcinko M, Gude N, Rubio M, Sussman MA, Glembotski CC. Activation of the unfolded protein response in infarcted mouse heart and hypoxic cultured cardiac myocytes. Circ Res 2006; 99:275-282.
5. Levine B, Klionsky DJ. Development by self-digestion: Molecular mechanisms and biological functions of autophagy. Dev Cell 2004; 6:463-477. 
6. Gozuacik D, Kimchi A. Autophagy and cell death. Curr Top Dev Biol 2007; 78:217-245. 
7. Mizushima N. A brief history of autophagy from cell biology to physiology and disease. Nat Cell Biol 2018; 20:521-527. 
8. Klionsky DJ, Emr SD. Autophagy as a regulated pathway of cellular degradation. Science 2000; 290:1717-1721. 
9. Yu L, Chen Y, Tooze SA. Autophagy pathway: Cellular and molecular mechanisms. Autophagy. 2018; 14:207-215. 
10. Feng Y, He D, Yao Z, Klionsky DJ. The machinery of macroautophagy. Cell Research 2014; 24:24-41. 
11. Arias E, Cuervo AM. Chaperone-mediated autophagy in protein quality control. Curr Opin Cell Biol 2011; 23:184-189. 
12. Wu H, Chen S, Ammar A-B, Xu J, Wu Q, Pan K, et al. Crosstalk between macroautophagy and chaperone-mediated autophagy: Implications for the treatment of neurological diseases. Mol Neurobiol 2015; 52:1284-1296. 
13. Ott C, König J, Höhn A, Jung T, Grune TJRB. Macroautophagy is impaired in old murine brain tissue as well as in senescent human fibroblasts. Redox Biol 2016; 10:266-273.
14. Kang R, Zeh HJ, Lotze MT, Tang D. The Beclin 1 network regulates autophagy and apoptosis. Cell Death Differ 2011; 18:571-580. 
15. Rusten TE, Stenmark H. p62, an autophagy hero or culprit? Nature Cell Biology. 2010; 12:207-209.
16. Komatsu M, Ichimura Y. Physiological significance of selective degradation of p62 by autophagy. FEBS Lett 2010; 584:1374-1378. 
17. Komatsu M, Waguri S, Koike M, Sou YS, Ueno T, Hara T, et al. Homeostatic levels of p62 control cytoplasmic inclusion body formation in autophagy-deficient mice. Cell 2007; 131:1149-1163.
18. Bjørkøy G, Lamark T, Pankiv S, Øvervatn A, Brech A, Johansen T. Monitoring autophagic degradation of p62/SQSTM1. Methods Enzymol 2009; 452:181-197.
19. Massey AC, Kaushik S, Sovak G, Kiffin R, Cuervo AM. Consequences of the selective blockage of chaperone-mediated autophagy. Proc Natl Acad Sci U S A 2006; 103:5805-5810. 
20. Kaushik S, Massey AC, Mizushima N, Cuervo AM. Constitutive activation of chaperone-mediated autophagy in cells with impaired macroautophagy. 2008; 19:2179-2192.
21. Tanida I, Ueno T, Kominami E. LC3 and Autophagy. Methods Mol Biol 2008; 445:77-88.
22. Tanida I, Minematsu-Ikeguchi N, Ueno T, Kominami E. Lysosomal turnover, but not a cellular level, of endogenous LC3 is a marker for autophagy. Autophagy 2005; 1:84-91.
23. Taylor SC, Berkelman T, Yadav G, Hammond M. A defined methodology for reliable quantification of Western blot data. Mol Biotechnol 2013; 55:217-226.
24. Galluzzi L, Bravo-San Pedro JM, Levine B, Green DR, Kroemer G. Pharmacological modulation of autophagy: Therapeutic potential and persisting obstacles. Nat Rev Drug Discov 2017; 16:487-511.
25. Cortez L, Sim V. The therapeutic potential of chemical chaperones in protein folding diseases. Prion 2014; 8:197-202. 
26. Perlmutter DH. Chemical chaperones: A pharmacological strategy for disorders of protein folding and trafficking. Pediatr Res 2002; 52:832-836. 
27. Mimori S, Ohtaka H, Koshikawa Y, Kawada K, Kaneko M, Okuma Y, et al. 4-Phenylbutyric acid protects against neuronal cell death by primarily acting as a chemical chaperone rather than histone deacetylase inhibitor. Bioorg Med Chem Lett 2013; 23:6015-6018. 
28. Kim SW, Hooker JM, Otto N, Win K, Muench L, Shea C, et al. Whole-body pharmacokinetics of HDAC inhibitor drugs, butyric acid, valproic acid and 4-phenylbutyric acid measured with carbon-11 labeled analogs by PET. Nucl Med Biol 2013; 40:912-918. 
29. National Center for Biotechnology Information. PubChem Patent Summary for WO-2006059237-A1. Accessed Jan. 4, 2023.
30. Park CS, Cha H, Kwon EJ, Sreenivasaiah PK, Kim DH. The chemical chaperone 4-phenylbutyric acid attenuates pressure-overload cardiac hypertrophy by alleviating endoplasmic reticulum stress. Biochem Biophys Res Commun 2012; 421:578-584.
31. Ayala P, Montenegro J, Vivar R, Letelier A, Urroz PA, Copaja M, et al. Attenuation of endoplasmic reticulum stress using the chemical chaperone 4-phenylbutyric acid prevents cardiac fibrosis induced by isoproterenol. Exp Mol Pathol 2012; 92:97-104.
32. Kilkenny C, Browne WJ, Cuthill IC, Emerson M, Altman DG. Improving bioscience research reporting: the ARRIVE guidelines for reporting animal research. PLoS Biol 2010; 8:e1000412.
33. Luo ZF, Feng B, Mu J, Qi W, Zeng W, Guo YH, et al. Effects of 4-phenylbutyric acid on the process and development of diabetic nephropathy induced in rats by streptozotocin: Regulation of endoplasmic reticulum stress-oxidative activation. Toxicol Appl Pharmacol. 2010; 246:49-57. 
34. Innovation OotVPfRa. Anesthesia for Laboratory Animals: University of Oregon. Available from:
35. Garjani A, Andalib S, Biabani S, Soraya H, Doustar Y, Garjani A, et al. Combined atorvastatin and coenzyme Q10 improve the left ventricular function in isoproterenol-induced heart failure in rat. Eur J Pharmacol 2011; 666:135-141.
36. Yousefi K, Soraya H, Fathiazad F, Khorrami A, Hamedeyazdan S, Maleki-Dizaji N, et al. Cardioprotective effect of methanolic extract of Marrubium vulgare L. on isoproterenol-induced acute myocardial infarction in rats. Indian J Exp Biol. 2013; 51:653-660.
37. Benjamin IJ, Jalil JE, Tan LB, Cho K, Weber KT, Clark WA. Isoproterenol-induced myocardial fibrosis in relation to myocyte necrosis. Circ res 1989; 65:657-670. 
38. Michaud K, Basso C, d’Amati G, Giordano C, Kholová I, Preston SD, et al. Diagnosis of myocardial infarction at autopsy: AECVP reappraisal in the light of the current clinical classification. Virchows Arch. 2020; 476:179-194.
39. Benzie IF, Strain JJ. The ferric reducing ability of plasma (FRAP) as a measure of “anti-oxidant power”: the FRAP assay. Anal Biochem. 1996; 239:70-76. 
40. Upaganlawar A, Gandhi H, Balaraman R. Isoproterenol induced myocardial infarction: Protective role of natural products. J Pharmacol Toxicol 2011; 6:1-17.
41. Benjamin IJ, McMillan DR. Stress (heat shock) proteins: molecular chaperones in cardiovascular biology and disease. Circ Res 1998; 83:117-132.
42. Ringe D, Petsko GA. What are pharmacological chaperones and why are they interesting? J Biol. 2009; 8:80-Last page. 
43. Martinet W, Agostinis P, Vanhoecke B, Dewaele M, De Meyer GR. Autophagy in disease: a double-edged sword with therapeutic potential. Clin Sci (Lond). 2009; 116:697-712.
44. Shintani T, Klionsky DJ. Autophagy in health and disease: a double-edged sword. Science 2004; 306:990-995. 
45. Humeres C, Montenegro J, Varela M, Ayala P, Vivar R, Letelier A, et al. 4-Phenylbutyric acid prevent cytotoxicity induced by thapsigargin in rat cardiac fibroblast. Toxicol In vitro 2014; 28:1443-1448. 
46. Ono K, Nimura S, Nishinakagawa T, Hideshima Y, Enjyoji M, Nabeshima K, et al. Sodium 4-phenylbutyrate suppresses the development of dextran sulfate sodium-induced colitis in mice. Exp Ther Med 2014; 7:573-578. 
47. Zhou SX, Zhou Y, Zhang YL, Lei J, Wang JF. Anti-oxidant probucol attenuates myocardial oxidative stress and collagen expressions in post-myocardial infarction rats. J Cardiovasc Pharmacol 2009; 54:154-162. 
48. Massey AC, Kaushik S, Sovak G, Kiffin R, Cuervo AM. Consequences of the selective blockage of chaperone-mediated autophagy. Proceedings of the National Academy of Sciences of the United States of America 2006; 103:5805-5810. 
49. Park H-J, Son H-J, Sul O-J, Suh J-H, Choi H-S. 4-Phenylbutyric acid protects against lipopolysaccharide-induced bone loss by modulating autophagy in osteoclasts. Biochem Pharmacol 2018; 151:9-17. 
50. Kim DS, Li B, Rhew KY, Oh HW, Lim HD, Lee W, et al. The regulatory mechanism of 4-phenylbutyric acid against ER stress-induced autophagy in human gingival fibroblasts. Arch Pharm Res 2012; 35:1269-1278.
51. Liang S, Ping Z, Ge J. Coenzyme Q10 regulates anti-oxidative stress and autophagy in acute myocardial ischemia-reperfusion injury. Oxid Med Cell Longev 2017; 2017:9863181.
52. Wu H, Chen S, Ammar AB, Xu J, Wu Q, Pan K, et al. Crosstalk between macroautophagy and chaperone-mediated autophagy: implications for the treatment of neurological diseases. Mol Neurobiol 2015; 52:1284-1296.