Acute lung injury induced by acute uremia and renal ischemic-reperfusion injury: The role of toll-like receptors 2 and 4, and oxidative stress

Document Type : Original Article


1 Burn and Wound Healing Research Center, Shiraz University of Medical Sciences, Shiraz, Iran

2 Shiraz Nephro-Urology Research Center, Shiraz University of Medical Sciences, Shiraz, Iran



Objective(s): Acute lung injury (ALI) is a common complication of distant organ dysfunction induced by acute kidney injury (AKI). Toll-like receptors (TLRs) have a critical role in progression of AKI. The main goal of this study was to determine whether lung gene expression of TLR2 and TLR4 change by ischemic (renal bilateral ischemic-reperfusion; BIR) and uremic (bilateral nephrectomy; BNX) AKI.
Materials and Methods: Forty male rats were divided into five groups. Two kidneys were removed in BNX, and renal pedicles were clamped in BIR for 45 min. The kidney and lung tissue, and blood samples were collected and saved after 24 hr in all groups. The bone marrow mesenchymal stem cells were immediately injected (1×106,IV) into the treated groups. The expression of TLR2, TLR4, TNF-α, and VEGF was checked by RT-PCR in the tissue samples. MDA level, SOD, and CAT activity were evaluated in the tissue samples.
Results: Structural disturbance of ALI was detected as alveolar hemorrhage and vascular congestion after BIR and BNX.  Lung TLR2 and TLR4 but not TNF-α and VEGF up-regulated in these groups. Oxidative stress stabilized after the BIR and BNX in the tissue samples. BMSCs reduce the expression of TLR2 and TLR4 and oxidative stress in the treated groups. 
Conclusion: Acutely gathering systemic mediators after renal ischemic or uremic injury induce ALI through overexpression of TLR2 and TLR4 and oxidative stress. Therefore, the Lung protective effect of BMSCs may be related to modulation of TLR2 and TLR4 and oxidative stress in the kidney and lung tissue.


1. Grams ME, Rabb H. The distant organ effects of acute kidney injury. Kidney Int 2012; 81:942-948.
2. Basu RK, Wheeler DS. Kidney-lung cross-talk and acute kidney injury. Pediatr Nephrol 2013; 28:2239-2248.
3. Karimi Z, Ketabchi F, Alebrahimdehkordi N, Fatemikia H, Owji SM, Moosavi SM. Renal ischemia/reperfusion against nephrectomy for induction of acute lung injury in rats. Ren Fail 2016; 38:1503-1515.
4. Hoke TS, Douglas IS, Klein CL, He Z, Fang W, Thurman JM, et al. Acute renal failure after bilateral nephrectomy is associated with cytokine-mediated pulmonary injury. J Am Soc Nephrol 2007; 18:155-164.
5. Rabb H, Wang Z, Nemoto T, Hotchkiss J, Yokota N, Soleimani M. Acute renal failure leads to dysregulation of lung salt and water channels. Kidney Int 2003; 63:600-606.
6. Klein CL, Hoke TS, Fang WF, Altmann CJ, Douglas IS, Faubel S. Interleukin-6 mediates lung injury following ischemic acute kidney injury or bilateral nephrectomy. Kidney Int 2008; 74:901-909.
7. Kimbrell DA, Beutler B. The evolution and genetics of innate immunity. Nat Rev Genet 2001; 2:256-267.
8. Wu H, Chen G, Wyburn KR, Yin J, Bertolino P, Eris JM, et al. TLR4 activation mediates kidney ischemia/reperfusion injury. J Clin Invest 2007; 117:2847-2859.
9. Ioannou S, Voulgarelis M. Toll-like receptors, tissue injury, and tumourigenesis. Mediators Inflamm 2010; 581837:1-9.
10. Dasari P, Nicholson IC, Hodge G, Dandie GW, Zola H. Expression of toll-like receptors on B lymphocytes. Cell Immunol 2005; 236:140-145.
11. Delneste Y, Beauvillain C, Jeannin P. Innate immunity: structure and function of TLRs. Med Sci 2007; 23:67-73.
12. Song Y, Shou LM, Ai LY, Bei Y, Chen MT. Mini-review: the non-immune functions of toll-like receptors. Crit Rev Eukaryot Gene Expr 2019; 29:37-45.
13. Kumar H, Kawai T, Akira S. Toll-like receptors and innate immunity. Biochem Biophys Res Commun 2009; 388:621-625.
14. Shimamoto A, Pohlman TH, Shomura S, Tarukawa T, Takao M, Shimpo H. Toll-like receptor 4 mediates lung ischemia-reperfusion injury. Ann Thorac Surg 2006; 82:2017-2023.
15. Imai Y, Kuba K, Neely GG, Yaghubian-Malhami R, Perkmann T, van Loo G, et al. Identification of oxidative stress and toll-like receptor 4 signaling as a key pathway of acute lung injury. Cell 2008; 133:235-249.
16. Klein CL, Hoke TS, Fang W-F, Altmann CJ, Douglas IS, Faubel S. Interleukin-6 mediates lung injury following ischemic acute kidney injury or bilateral nephrectomy. Kidney Int 2008; 74:901-909.
17. Kehrer JP. Free radicals as mediators of tissue injury and disease. Crit Rev Toxicol 1993; 23:21-48.
18. Aragno M, Cutrin JC, Mastrocola R, Perrelli MG, Restivo F, Poli G, et al. Oxidative stress and kidney dysfunction due to ischemia/reperfusion in rat: attenuation by dehydroepiandrosterone. Kidney Int 2003; 64:836-843.
19. Caplan AI, Dennis JE. Mesenchymal stem cells as trophic mediators. J Cell Biochem 2006; 98:1076-1084.
20. Pourfath MR, Behzad-Behbahani A, Hashemi SS, Derakhsahnfar A, Taheri MN, Salehi S. Monitoring wound healing of burn in rat model using human wharton’s jelly mesenchymal stem cells containing cGFP integrated by lentiviral vectors. Iran J Basic Med Sci 2018; 21:70-76.
21. Aggarwal S, Pittenger MF. Human mesenchymal stem cells modulate allogeneic immune cell responses. Blood 2005; 105:1815-1822.
22. Abdel Aziz M, Wassef M, Rashed L, Mhfouz S, Omar N, Elsebaie M. Mesenchymal stem cells therapy in acute renal failure: possible role of hepatocyte growth factor. J Stem Cell Res Ther 2011; 1:1-7.
23. Hashemi SS, Mohammadi AA, Kabiri H, Hashempoor MR, Mahmoodi M, Amini M, et al. The healing effect of wharton’s jelly stem cells seeded on biological scaffold in chronic skin ulcers: arandomized clinical trial. J Cosmet Dermatol 2019; 18:1961-1967.
24. Aebi H. Catalase. Methods of Enzymatic Analysis 1983.
25. Caprioli I, O’Sullivan M, Monahan FJ. Interference of sodium caseinate in the TBARS assay. Food Chem 2011; 124:1284-1287.
26. Najafi H, Owji SM, Kamali-Sarvestani E, Moosavi SM. A1 -adenosine receptor activation has biphasic roles in development of acute kidney injury at 4 and 24 hr of reperfusion following ischaemia in rats. Exp Physiol 2016; 101:913-931.
27. Sutton TA, Fisher CJ, Molitoris BA. Microvascular endothelial injury and dysfunction during ischemic acute renal failure. Kidney Int 2002; 62:1539-1549.
28. Le Dorze M, Legrand M, Payen D, Ince C. The role of the microcirculation in acute kidney injury. Curr Opin Crit Care 2009; 15:503-508.
29. Fatemikia H, Ketabchi F, Karimi Z, Moosavi SMS. Distant effects of unilateral renal ischemia/reperfusion on contralateral kidney but not lung in rats: the roles of ROS and iNOS. Can J Physiol Pharmacol 2015; 94:477-487.
30. Moosavi SM, Bayat G, Owji SM, Panjehshahin MR. Early renal post-ischaemic tissue damage and dysfunction with contribution of A1-adenosine receptor activation in rat. Nephrology (Carlton) 2009; 14:179-188.
31. Ishii T, Doi K, Okamoto K, Imamura M, Dohi M, Yamamoto K, et al. Neutrophil elastase contributes to acute lung injury induced by bilateral nephrectomy. Am J Pathol 2010; 177:1665-1673.
32. Devarajan P. Update on mechanisms of ischemic acute kidney injury. J Am Soc Nephrol 2006; 17:1503-1520.
33. Lempiäinen J. Ischaemia-reperfusion-induced kidney injury: Experimental studies on the effects of caloric restriction, AMPK activator AICAR and α2-adrenoceptor agonists in the rat. Dissertationes Scholae Doctoralis Ad Sanitatem Investigandam Universitatis Helsinkiensis 2014; 9:1-77.
34. Jang HR, Ko GJ, Wasowska BA, Rabb HJJomm. The interaction between ischemia–reperfusion and immune responses in the kidney. J Mol Med 2009; 87:859-864.
35. Shigeoka AA, Holscher TD, King AJ, Hall FW, Kiosses WB, Tobias PS, et al. TLR2 is constitutively expressed within the kidney and participates in ischemic renal injury through both MyD88-dependent and -independent pathways. J Immunol 2007; 178:6252-6258.
36. Gholampour F, Roozbeh J, Janfeshan S, Karimi Z. Remote ischemic per-conditioning protects against renal ischemia–reperfusion injury via suppressing gene expression of TLR4 and TNF-α in rat model. Can J Physiol Pharmacol 2019; 97:112-119.
37. Neufeld G, Cohen T, Gengrinovitch S, Poltorak Z. Vascular endothelial growth factor (VEGF) and its receptors. FASEB J 1999; 13:9-22.
38. Basile DP, Fredrich K, Chelladurai B, Leonard EC, Parrish AR. Renal ischemia reperfusion inhibits VEGF expression and induces ADAMTS-1, a novel VEGF inhibitor. Am J Physiol Renal Physiol 2008; 294:928-936.
39. Ma ZG, Xia HQ, Cui SL, Yu J. Attenuation of renal ischemic reperfusion injury by salvianolic acid B via suppressing oxidative stress and inflammation through PI3K/Akt signaling pathway. Braz J Med Biol Res 2017; 50:5954-5962.
40. Alkadi H. A review on free radicals and antioxidants. Infect Disord Drug Targets 2020; 20:16-26.
41. Husain-Syed F, Slutsky AS, Ronco C. Lung-kidney cross-talk in the critically Ill patient. Am J Respir Crit Care Med 2016; 194:402-414.
42. Hoke TS, Douglas IS, Klein CL, He Z, Fang W, Thurman JM, et al. Acute renal failure after bilateral nephrectomy is associated with cytokine-mediated pulmonary injury. J Am Soc Nephrol 2007; 18:155-164.
43. Donnahoo KK, Meng X, Ayala A, Cain MP, Harken AH, Meldrum DR. Early kidney TNF-alpha expression mediates neutrophil infiltration and injury after renal ischemia-reperfusion. Am J Physiol 1999; 277:922-929.
44. Wolfs TG, Buurman WA, van Schadewijk A, de Vries B, Daemen MA, Hiemstra PS, et al. In vivo expression of toll-like receptor 2 and 4 by renal epithelial cells: IFN-gamma and TNF-alpha mediated up-regulation during inflammation. J Immunol 2002; 168:1286-1293.
45. Barratt S, Medford AR, Millar AB. Vascular endothelial growth factor in acute lung injury and acute respiratory distress syndrome. Respiration 2014; 87:329-342.
46. Fink MP. Role of reactive oxygen and nitrogen species in acute respiratory distress syndrome. Curr Opin Crit Care 2002; 8:6-11.
47. Sousa RH, Carvalho FE, Ribeiro CW, Passaia G, Cunha JR, Lima-Melo Y, et al. Peroxisomal APX knockdown triggers antioxidant mechanisms favourable for coping with high photorespiratory H2 O2 induced by CAT deficiency in rice. Plant Cell Environ 2015; 38:499-513.
48. Dong Z, Yuan Y. Accelerated inflammation and oxidative stress induced by LPS in acute lung injury: iotanhibition by ST1926. Int J Mol Med 2018; 41:3405-3421.
49. Lei J, Wei Y, Song P, Li Y, Zhang T, Feng Q, et al. Cordycepin inhibits LPS-induced acute lung injury by inhibiting inflammation and oxidative stress. Eur J Pharmacol 2018; 818:110-114.
50. Gill R, Tsung A, Billiar T. Linking oxidative stress to inflammation: toll-like receptors. Free Radic Biol Med 2010; 48:1121-1132.
51. Malek M, Nematbakhsh M. Renal ischemia/reperfusion injury; from pathophysiology to treatment. J Renal Inj Prev 2015; 4:20-27.
52. Hashemi S, Rafati A. Comparison between human cord blood serum and platelet-rich plasma supplementation for human wharton’s jelly stem cells and dermal fibroblasts culture. Health Sci 2016; 5:191-196.
53. Liu H, McTaggart SJ, Johnson DW, Gobe GC. Original article anti-oxidant pathways are stimulated by mesenchymal stromal cells in renal repair after ischemic injury. Cytotherapy 2012; 14:162-172.
54. Lee KH, Tseng WC, Yang CY, Tarng DC. The anti-inflammatory, anti-oxidative, and anti-apoptotic benefits of stem cells in acute ischemic kidney injury. Int J Mol Sci 2019; 20:3529-3542.
55. Horie S, Laffey JG. Recent insights: mesenchymal stromal/stem cell therapy for acute respiratory distress syndrome. F1000Res 2016; 5:1532-1539.