Ulinastatin protects cardiomyocytes against ischemia‑reperfusion injury by regulating autophagy through mTOR activation

Xiao,Jian; Zhu,Xiaoyan; Ji,Guangyu; Yang,Qian; Kang,Bo; Zhao,Jianquan; Yao,Feng; Wu,Lihui; Ni,Xin; Wang,Zhinong

doi:10.3892/mmr.2014.2450

Ulinastatin protects cardiomyocytes against ischemia‑reperfusion injury by regulating autophagy through mTOR activation

Authors:
- Jian Xiao
- Xiaoyan Zhu
- Guangyu Ji
- Qian Yang
- Bo Kang
- Jianquan Zhao
- Feng Yao
- Lihui Wu
- Xin Ni
- Zhinong Wang
View Affiliations

Affiliations: Department of Cardiothoracic Surgery, Changzheng Hospital, The Second Military Medical University, Shanghai 200003, P.R. China, Department of Physiology, The Second Military Medical University, Shanghai 200433, P.R. China, Department of Anesthesiology, Shanghai Chest Hospital Affiliated to Shanghai Jiaotong University, Shanghai 200030, P.R. China
Published online on: August 5, 2014 https://doi.org/10.3892/mmr.2014.2450
Pages: 1949-1953

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

Autophagy is significant in myocardial ischemia‑reperfusion (IR) injury. Ulinastatin has been demonstrated to protect cardiomyocytes against IR through inducing anti‑inflammatory effects. However, whether ulinastatin has an anti‑autophagic effect is yet to be elucidated. The present study aimed to investigate the effect of ulinastatin on the regulation of autophagy during IR injury. Cardiomyocytes of neonatal rats were randomly divided into control, hypoxia‑reoxygenation (HR) and ulinastatin groups. In order to investigate whether mammalian target of rapamycin (mTOR) is involved in mediating the protective effect of ulinastatin, cells were treated with the mTOR inhibitor, rapamycin 30 min prior to ulinastatin treatment. To demonstrate the anti‑autophagic effect of ulinastatin in vivo, a rat IR model was established. Ulinastatin (1x104 U/kg body weight) was administered 30 min prior to the induction of IR via peritoneal injection. Light chain 3 (LC3), phosphorylated (p)‑mTOR, p‑protein kinase B (Akt) and p‑P70S6 kinase (p‑P70S6K) protein expression were assessed using western blot analysis. In addition, cell vitality, myocardial infarct size and lactate dehydrogenase (LDH) levels were measured. LC3‑Ⅱ protein expression was found to be downregulated, while p‑Akt, p‑mTOR and p‑P70S6K protein expression were observed to be upregulated by ulinastatin. In addition, cell vitality was found to increase and LDH was observed to decrease in the ulinastatin group compared with the HR group in vitro. Furthermore, rapamycin was found to attenuate the myocardial protective effect that is induced by ulinastatin. In vivo, ulinastatin was found to downregulate LC3‑Ⅱ protein expression, and reduce myocardium infarct size and LDH serum levels. These findings indicate that ulinastatin exhibits a myocardial protective effect against IR injury by regulating autophagy through mTOR activation.

View Figures

View References

1	Shin IW, Jang IS, Lee SM, et al: Myocardial protective effect by ulinastatin via an anti-inflammatory response after regional ischemia/reperfusion injury in an in vivo rat heart model. Korean J Anesthesiol. 61:499–505. 2011. View Article : Google Scholar : PubMed/NCBI
2	Vassalli G, Milano G and Moccetti T: Role of mitogen-activated protein kinases in myocardial ischemia-reperfusion injury during heart transplantation. J Transplant. 2012:9289542012.
3	Ishii D, Schenk AD, Baba S and Fairchild RL: Role of TNFalpha in early chemokine production and leukocyte infiltration into heart allografts. Am J Transplant. 10:59–68. 2010. View Article : Google Scholar : PubMed/NCBI
4	Li C, Capan E, Zhao Y, et al: Autophagy is induced in CD4⁺ T cells and important for the growth factor-withdrawal cell death. J Immunol. 177:5163–5168. 2006.PubMed/NCBI
5	Meléndez A, Tallóczy Z, Seaman M, Eskelinen EL, Hall DH and Levine B: Autophagy genes are essential for dauer development and life-span extension in C. elegans. Science. 301:1387–1391. 2003.PubMed/NCBI
6	Stromhaug PE and Klionsky DJ: Approaching the molecular mechanism of autophagy. Traffic. 2:524–531. 2001. View Article : Google Scholar : PubMed/NCBI
7	Otto GP, Wu MY, Kazgan N, Anderson OR and Kessin RH: Macroautophagy is required for multicellular development of the social amoeba Dictyostelium discoideum. J Biol Chem. 278:17636–17645. 2003. View Article : Google Scholar : PubMed/NCBI
8	Gurusamy N and Das DK: Is autophagy a double-edged sword for the heart? Acta Physiol Hung. 96:267–276. 2009. View Article : Google Scholar : PubMed/NCBI
9	De Meyer GR and Martinet W: Autophagy in the cardiovascular system. Biochim Biophys Acta. 1793:1485–1495. 2009.
10	Inoue K, Takano H, Shimada A, et al: Urinary trypsin inhibitor protects against systemic inflammation induced by lipopolysaccharide. Mol Pharmacol. 67:673–680. 2005. View Article : Google Scholar : PubMed/NCBI
11	Xu CE, Zhang MY, Zou CW and Guo L: Evaluation of the pharmacological function of ulinastatin in experimental animals. Molecules. 17:9070–9080. 2012. View Article : Google Scholar : PubMed/NCBI
12	Cao YZ, Tu YY, Chen X, Wang BL, Zhong YX and Liu MH: Protective effect of ulinastatin against murine models of sepsis: inhibition of TNF-α and IL-6 and augmentation of IL-10 and IL-13. Exp Toxicol Pathol. 64:543–547. 2012.PubMed/NCBI
13	Jian X, Xiao-yan Z, Bin H, et al: MiR-204 regulate cardiomyocyte autophagy induced by hypoxia-reoxygenation through LC3-II. Int J Cardiol. 148:110–112. 2011. View Article : Google Scholar : PubMed/NCBI
14	He B, Xiao J, Ren AJ, et al: Role of miR-1 and miR-133a in myocardial ischemic postconditioning. J Biomed Sci. 18:222011. View Article : Google Scholar : PubMed/NCBI
15	Zhuo Y, Chen PF, Zhang AZ, Zhong H, Chen CQ and Zhu YZ: Cardioprotective effect of hydrogen sulfide in ischemic reperfusion experimental rats and its influence on expression of survivin gene. Biol Pharm Bull. 32:1406–1410. 2009. View Article : Google Scholar : PubMed/NCBI
16	Masuda T, Sato K, Noda C, et al: Protective effect of urinary trypsin inhibitor on myocardial mitochondria during hemorrhagic shock and reperfusion. Crit Care Med. 31:1987–1992. 2003. View Article : Google Scholar : PubMed/NCBI
17	Tanaka R, Fujita M, Tsuruta R, et al: Urinary trypsin inhibitor suppresses excessive generation of superoxide anion radical, systemic inflammation, oxidative stress, and endothelial injury in endotoxemic rats. Inflamm Res. 59:597–606. 2010. View Article : Google Scholar
18	Koga Y, Fujita M, Tsuruta R, et al: Urinary trypsin inhibitor suppresses excessive superoxide anion radical generation in blood, oxidative stress, early inflammation, and endothelial injury in forebrain ischemia/reperfusion rats. Neurol Res. 32:925–932. 2010. View Article : Google Scholar
19	Wu YJ, Ling Q, Zhou XH, et al: Urinary trypsin inhibitor attenuates hepatic ischemia-reperfusion injury by reducing nuclear factor-kappa B activation. Hepatobiliary Pancreat Dis Int. 8:53–58. 2009.PubMed/NCBI
20	Chen CC, Liu ZM, Wang HH, He W, Wang Y and Wu WD: Effects of ulinastatin on renal ischemia-reperfusion injury in rats. Acta Pharmacol Sin. 25:1334–1340. 2004.PubMed/NCBI
21	Ren B, Wu H, Zhu J, et al: Ulinastatin attenuates lung ischemia-reperfusion injury in rats by inhibiting tumor necrosis factor alpha. Transplant Proc. 38:2777–2779. 2006. View Article : Google Scholar : PubMed/NCBI
22	Deretic V: Autophagy as an immune defense mechanism. Curr Opin Immunol. 18:375–382. 2006. View Article : Google Scholar : PubMed/NCBI
23	Nishida K, Kyoi S, Yamaguchi O, Sadoshima J and Otsu K: The role of autophagy in the heart. Cell Death Differ. 16:31–38. 2009. View Article : Google Scholar : PubMed/NCBI
24	Matsui Y, Takagi H, Qu X, et al: Distinct roles of autophagy in the heart during ischemia and reperfusion: roles of AMP-activated protein kinase and Beclin 1 in mediating autophagy. Circ Res. 100:914–922. 2007. View Article : Google Scholar : PubMed/NCBI