cell biochemistry and function Cell Biochem Funct 2015; 33: 113–120. Published online 10 March 2015 in Wiley Online Library (wileyonlinelibrary.com) DOI: 10.1002/cbf.3092

Combination therapy with human umbilical cord mesenchymal stem cells and angiotensin-converting enzyme 2 is superior for the treatment of acute lung ischemia–reperfusion injury in rats Xiaomiao Zhang1, Fengying Gao2, Yunqi Yan3, Zheng Ruan1 and Zhenwei Liu4* 1

Department of Department of 3 Department of 4 Department of 2

Thoracic Surgery, First People’s Hospital Affiliated to Shanghai Jiaotong University, Shanghai, China Respiratory Medicine, Shanghai Jiangong Hospital, Shanghai, China Anesthesia, Renji Hospital Affiliated to Shanghai Jiaotong University School of Medicine, Shanghai, China Respiratory Medicine, First People’s Hospital Affiliated to Shanghai Jiaotong University, Shanghai, China

Acute lung ischemia–reperfusion injury (ALIRI) is a serious disease that seriously affects human’s life. In this study, we aimed to explore a more effective treatment method by combining human umbilical cord mesenchymal stem cells (HUMSCs) and angiotensin-converting enzyme 2 (ACE2) for ALIRI. Fifty rats were firstly divided into five groups, namely sham surgery group (sham) and four model groups (model, ACE2, HUMSCs and HUMSCs + ACE2) that were reperfused with 0.1 ml physiological saline (PS), 0.1 ml PS containing 1 × 106 lentiviral-ACE2/HUMSCs/ACE2 + UMSCs, respectively. Quantitative reverse transcription-PCR (qRT-PCR) and western blot assays were then conducted to detect the messenger RNA (mRNA) and protein levels of inflammatory cytokines [intercellular adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule 1 (VCAM-1), tumour necrosis factor α (TNF-α), nuclear factor κB (NF-κB), platelet-derived growth factor (PDGF) and angiotensin II (Ang II)], antioxidant proteins [NAD(P)H quinone oxidoreductase 1 (NQO1), heme oxygenase 1 (HO-1)], DNA damage and apoptotic indicators [BCL2-associated X (Bax), cleaved caspase-3 (C-Csp 3), cleaved-poly(ADP-ribose) polymerase (C-PARP), Y-H2AX], anti-apoptotic indicator (Bcl-2) and smooth muscle cell proliferation indicator [connexin 43 (Cx43)]. According to the qRT-PCR and western results, the mRNA and protein expression levels of ICAM-1, VCAM-1, TNF-α, NF-κB, PDGF, Bax, C-Csp 3, C-PARP and Y-H2AX were significantly higher in model group than those in sham group and they were significantly reduced by HUMSCs or ACE2 treatment (P < 0.05). On the contrary, Bcl-2 showed an opposite expression trend with the previous proteins. The mRNA and protein levels of NQO1 and HO-1 were sequentially increased in sham, model, ACE2, HUMSCs and HUMSCs + ACE2 groups. Besides, HUMSCs combined with ACE2 exhibited a better inhibition effect on ALIRI than HUMSCs or ACE2 alone (P < 0.05). In summary, HUMSCs combined with ACE2 was demonstrated to have the best therapeutic effect on ALIRI through anti-inflammation, oxidative stress and anti-apoptotic processes. Copyright © 2015 John Wiley & Sons, Ltd. key words—acute lung ischemia–reperfusion injury; human umbilical cord mesenchymal stem cells; angiotensin-converting enzyme; inflammation; oxidative stress; apoptosis

INTRODUCTION Acute lung ischemia–reperfusion injury (ALIRI) is a serious disease that will lead to acute lung injury (ALI). In clinical, ALIRI is a lesion process, in which the lung transplantation, extracorporeal circulation, circulatory arrest and recovery, hypoxaemic respiratory failure, smoke inhalation, pulmonary embolism and septicaemia are involved. 1–6 Nowadays, the main therapy method for ALIRI is protective ventilation, and the related researches on ALIRI are mostly focused on the interactions and expressional regulations of inflammatoryrelated cells, cytokines and cell adhesion molecules. However, the underlying pathogenic mechanism of ALIRI remains not *Correspondence to: Zhenwei Liu, Department of Respiratory Medicine, First People’s Hospital Affiliated to Shanghai Jiaotong University, No. 100 Haining Road, Shanghai 200080, China. E-mail: [email protected]

Copyright © 2015 John Wiley & Sons, Ltd.

completely clear, and more effective treatment methods are still needed. 1,2 Mesenchymal stem cells (MSCs) are pluripotent stem cells with high self-renewal, proliferation and differentiation potential, and they are derived from mesoderm of early phase. MSCs have been identified to be involved in the reparation and regeneration of lung tissue through inflammation suppression, oxidative stress and anti-apoptotic process. 7–9 As reported, transplantation of different sources of MSCs could relieve ALI in rodents. 10–12 Besides, MSCs could also serve as gene vectors in the treatment of lung disease. Aguilar et al. reported that MSCs carrying keratinocyte growth factor was an effective therapy for pulmonary fibrosis. 13 Therefore, MSCs integrated with critical gene might become an effective treatment method on ALIRI. Furthermore, angiotensinconverting enzyme 2 (ACE2), a specific exopeptidase belonged to renin–angiotensin system, could degrade several Received 24 November 2014 Revised 25 December 2014 Accepted 5 January 2015

114

x. zhang ET AL.

proinflammatory peptides, such as angiotensin II (Ang II), Ang IV and des-Arg9-BK .14–17 Ever since the discovery of ACE2 being a critical receptor of severe acute respiratory syndrome virus in 2003, the protective roles of ACE2 in acute and chronic lung diseases have been revealed in the mechanisms of anti-inflammatory, anti-oxidation and anti-apoptosis. 18,19 As previously reported, ALI was more serious in ACE2 knockout mice, which indicated that ACE2 played a critical role in ALI .20 Fortunately, ACE2 was easy to be carried and transferred, which made ACE2 as a great potential to be utilized in gene therapy. 19 ALIRI could lead to serious inflammation, enhance oxidative stress and increase reactive oxygen species (ROS), which would further damage the pulmonary parenchyma, gas exchange system and macrocirculation and microcirculation in the lung. 21–23 Therefore, preventing the production and amplification of inflammation, ROS and oxidative stress may become an effective treatment strategy in ALIRI. 22,24,25 The specific mechanisms of HUMSCs and ACE2 on the treatment of ALI were far from completely clear, and there were still no reports about the effects of HUMSCs integrated with ACE2 on ALI. We suspected that the HUMSCs combined with ACE2 might exhibit a more effective treatment on ALI. In order to verify our conjecture and extend our understanding about the treatment mechanisms of HUMSCs and ACE2 in ALI, HUMSCs harbouring ACE2 treatment was used on ALIRI rat model. Several inflammatory cytokines, antioxidant proteins, DNA damage and apoptotic indicators were detected. We suspected that HUMSCs harbouring ACE2 would provide informative evidences and promising insights for the treatment of ALIRI.

tightly for 45 min after left thoracotomy and then reperfused for 72 h. The construction of ALIRI model was successful according to our previous studies. 26 These four groups (model, ACE2, HUMSCs and HUMSCs + ACE2) were reperfused with 0.1 ml PS, 0.1 ml PS containing lentiviralACE2, 0.1 ml PS containing 1 × 106 HUMSCs and 0.1 ml PS containing 1 × 106 ACE2 + UMSCs, respectively. The sham group with only left thoracotomy was considered as control group. Real-time quantitative reverse transcription-PCR (qRT-PCR) The total RNAs of these five groups were isolated with Trizol reagent (BD Biosciences, Franklin Lakes, NJ, USA), respectively. Reverse transcriptase and oligo (dT) (PrimeScript RT reagent Kit, Code No. 9160) were utilized to conduct reverse transcription. The real-time qRT-PCR procedure was performed as follows: 95 °C for 5 min; 40 cycles of 95 °C for 20 s, 60 °C for 30 s and 72 °C for 20 s. Levels of messenger RNA (mRNA) were conducted on iQ5 Multicolor Real-Time PCR Detection System (Bio-Rad, Hercules, CA, USA) by using a SYBR Green Real-time PCR Master Mix (TaKaRa, Shiga, Japan). The expression level of Glyceraldehyde 3phosphate dehydrogenase (GAPDH) was used as an internal control. Relative mRNA expression level was calculated using the 2 ΔΔCt method. Three parallels were set for each sample, and all the experiments were repeated three times. The primers are listed in Table 1. Table 1. Sequences of primers used for real-time quantitative reverse transcription-PCR (qRT-PCR) Gene symbol

MATERIALS AND METHODS

NF-κB

Construction and treatment of ALIRI rat model

TNF-α

Rattus norregiucs (No. 50; SD; female; 8 weeks; weight 250 ± 20 g) were provided by the Department of Laboratory Animal Science, School of Medicine, Shanghai Jiaotong University. Rats were divided in ten cages and fed with regular feed in well-ventilated room (free drinking; temperature: 21 ± 1 °C; 60% humidity; 0.05, Figure 1), the protein expression of PDGF was lower in HUMSCs + ACE2 group than that in HUMSCs group (P < 0.05, Figure 2B). Then, the mRNA and protein levels of another three inflammatory indicators Ang II, VCAM-1 and ICAM-1 were detected. Similarly, their mRNA and protein levels were significantly increased in model group compared with controls, and they could be most effectively decreased by HUMSCs + ACE2 treatment (P < 0.05, Figure 1 and Figure 2D–F). Cell Biochem Funct 2015; 33: 113–120.

EFFECTS OF HUMSCS AND ACE2 ON ALIRI

117

Figure 5. The mRNA levels of DNA damage and apoptotic indicators in left lung parenchyma of acute lung ischemia–reperfusion injury (ALIRI) model (n = 6 each group). Sham: sham surgery group; model: reperfused with 0.1 ml physiological saline (PS); ACE2: reperfused with 0.1 ml PS containing 6 6 lentiviral-ACE2; HUMSCs: reperfused with 0.1 ml PS containing 1 × 10 HUMSCs; HUMSCs + ACE2: reperfused with 0.1 ml PS containing 1 × 10 ACE2 + UMSCs. Different lowercase letters (a–e) indicate statistical difference (P < 0.05)

The effects of ACE2 and HUMSCs on protein oxidation of ALIRI As an indicator of oxidative stress, the expression of protein carbonyls was analysed in ALIRI model (Figure 3A). As shown in Figure 3B, the oxidative index was highest in model group and lowest in sham group. In addition, the oxidative index in ACE2 and HUMSCs group was significantly lower than in model group (P < 0.05). However, lower expression levels were exhibited after HUMSCs + ACE2 treatment compared with HUMSCs or ACE2 treatment alone. Besides, the expression of Cx43, an indicator of smooth muscle cell proliferation, was found to be just consistent with oxidative index (Figure 4 and Figure 6A). On the other hand, NQO1 and HO-1 as typical antioxidant biomarkers were then detected in ALIRI model by qRTPCR and western blot. We noted that the mRNA (Figure 4) and protein (Figure 6B and C) levels of NQO1 and HO-1 were sequentially increased in sham, model, ACE2, HUMSCs and HUMSCs + ACE2 groups with significantly different (P < 0.05). The effects of ACE2 and HUMSCs on DNA damage and apoptosis of ALIRI Y-H2AX was a typical indicator of DNA damage. The results of qRT-PCR and western blot analyses showed that the expression level of Y-H2AX was highest in model group and lowest in sham group. In addition, Y-H2AX expressed higher in ACE2 group and lower in HUMSCs + ACE2 group when compared with HUMSCs group (P < 0.05, Figure 5 and Figure 6H). However, apoptotic indicator C-Csp 3 had a similar expression trend with Y-H2AX in different groups (Figure 5 and Figure 6E). Different with C-Csp 3, the expression of another apoptotic indicator C-PARP was similar and exhibited the lowest level in both sham and HUMSCs + ACE2 groups (Figure 5 and Figure 6F). Besides, no significant difference was found in the expression of Bax that was also identified as an apoptotic indicator among HUMSCs, ACE2 and HUMSCs + ACE2 groups Copyright © 2015 John Wiley & Sons, Ltd.

(P > 0.05, Figure 5 and Figure 6D). On the contrary, as an anti-apoptotic indicator, Bcl-2 exhibited an opposite expression trend with C-Csp 3. Although there was significant difference of Bcl-2 mRNA expression between HUMSCs and ACE2 groups, the protein expression level of Bcl-2 was without statistical difference in these two groups (P > 0.05). However, the mRNA and protein levels of Bcl2 were significantly increased in HUMSCs + ACE2 group than that in HUMSCs or ACE2 groups (P > 0.05, Figure 5 and Figure 6G).

DISCUSSION ALIRI is a complex pathogenetic situation involving several biochemical, cellular and molecular alterations. As reported, acute ischemia-related organ damage in ALIRI could induce inflammation, oxidative stress, increased ROS and apoptosis. 23,27 Consistent with the previous studies, we found that the expressions of inflammatory cytokines, oxidative stress and apoptosis were significantly higher and the anti-apoptotic factor (Bcl-2) was significantly lower in ALIRI model than normal group (P < 0.05), which indicated that ALIRI model was successfully constructed. Nowadays, stem cell therapy in clinical has been widely used in ischemia-related organ dysfunction with optimistic prospect. 28,29 In this study, an easily to be carried and transferred gene ACE2 was skilfully combined with pluripotent vector HUMSCs, and then, a combination therapy of cell and gene was achieved. Meanwhile, both HUMSCs and ACE2 treatment could reduce the expressions of these indicators and increase the expression of Bcl-2. What is encouraged, the treatment effects of HUMSCs combined with ACE2 were better than these two alone in ALIRI. This indicated that ACE2 and HUMSCs might play synergistically therapeutic effects on ALIRI. Luciano et al. has reported that ALI is correlated with impaired NF-κB activation, 30 which is a major regulator of Cell Biochem Funct 2015; 33: 113–120.

118

x. zhang ET AL.

Figure 6. The expressions of antioxidant indicators, DNA damage and apoptotic indicators in left lung parenchyma of acute lung ischemia–reperfusion injury (ALIRI) model (n = 6 each group). Sham: sham surgery group; model: reperfused with 0.1 ml physiological saline (PS); ACE2: reperfused with 0.1 ml PS con6 6 taining lentiviral-ACE2; HUMSCs: reperfused with 0.1 ml PS containing 1 × 10 HUMSCs; HUMSCs + ACE2: reperfused with 0.1 ml PS containing 1 × 10 ACE2 + UMSCs. (A) Connexin 43 (CX43); (B) NAD(P)H quinone oxidoreductase 1 (NQO 1); (C) heme oxygenase 1 (HO 1); (D) BCL2-associated X (Bax); (E) cleaved caspase-3 (C-Csp 3); (F) cleaved-poly(ADP-ribose) polymerase (C-PARP); (G) B cell leukemia/lymphoma 2 (Bcl-2); (H) Y-H2AX. Actin was considered to be the control. Different lowercase letters (a–e) indicate statistical difference (P < 0.05)

innate and adaptive immunity and inflammatory. 31 Meanwhile, some drugs play anti-inflammatory effects in the treatment of cerebral ischemia–reperfusion injury by influencing the functional expression of NF-κB and TNF-α. 32 Huang et al. reported that rutin could improve ALI via inhibiting the expression of VCAM-1. 33 The overexpression of ICAM-1 is found in ALI according to multiple studies. 34,35 In addition, ICAM-1, VCAM-1, PDGF and Ang II were all related with the inflammatory process and depended on the activation of NF-κB signalling pathway. 36–39 Thus, we speculated that HUMSCs harbouring ACE2 might improve the inflammatory situation of ALIRI by regulating the activation of NF-κB signalling pathway and then influence the expressions of related inflammatory factors, including NF-κB, TNF-α, ICAM-1, VCAM-1, PDGF and Ang II. Copyright © 2015 John Wiley & Sons, Ltd.

HO-1 can catalyse the degradation of free cellular heme to iron and the conversion of carbon monoxide (CO) and biliverdin to bilirubin in the lung. 40 Nowadays, HO-1 is considered as a general protective mechanism for the lung in response to stress stimuli, especially oxidative. 41 NQO1 plays a pivotal role in the oxidative process and is downregulated in ALI and some human lung cancer cells. Increasing NQO1 activity and expression can be used to increase the anti-cancer effects of β-lapachone in lung cancer. 42 In this study, HO-1 and NQO1 were significantly increased after HUMSCs and ACE2 treatment, and the effects were best in HUMSCs and ACE2 group (P < 0.05). Therefore, HUMSCs and ACE2 might enhance the antioxidant response in ALIRI by regulating the expressions of related antioxidant proteins, including HO-1 and NQO1. Cell Biochem Funct 2015; 33: 113–120.

EFFECTS OF HUMSCS AND ACE2 ON ALIRI Apoptosis is a biological process that produces timely death in the senescent cells. Apoptosis is critical in the developmental biology and in tissues remodelling during repair. Many apoptosis-related pathways converge in the intracellular protease cascades that lead to DNA cleavage and cell death. 43 Increasing evidences reveal that apoptosis plays a pivotal role and Fas and Fas ligand are upregulated in ALI. 44,45 The apoptosis-related factors, Bax, C-Csp 3, C-PARP and Y-H2AX, were significantly decreased, and the anti-apoptosis indicator, Bcl-2, was significantly increased after HUMSCs and ACE2 treatment, and the effects were most obvious in ACE2 + UMSCs group (P < 0.05). It was reasonable to conclude that ACE2 and HUMSCs might suppress the apoptosis process in by ALIRI by regulating the expressions of Bax, C-Csp 3, CPARP, Y-H2AX and Bcl-2, and the Fas signalling pathway might be involved. In conclusion, both ACE2 and HUMSCs could improve ALIRI through anti-inflammation, oxidative stress and anti-apoptotic. Moreover, HUMSCs combined with ACE2 was demonstrated to have better therapeutic effects on ALIRI than either HUMSCs or ACE2 alone. Based on the previous analysis, the NF-κB and Fas signalling pathways might be involved in the treatment process and might help to exploit the underlying mechanism. Meanwhile, this combination method would provide informative evidences and promising insights for the treatment of ALIRI in future. However, there was still no clinical application of this combination carried out, and the efficacy and safety in humans were still not certified. Therefore, more prospective randomized clinical trials and related researches were still needed.

9.

10.

11.

12.

13.

14.

15.

16.

17.

18. 19.

20.

CONFLICT OF INTEREST The authors have declared that there is no conflict of interest.

REFERENCES 1. Matthay MA, Zimmerman GA. Acute lung injury and the acute respiratory distress syndrome: four decades of inquiry into pathogenesis and rational management. Am J Respir Cell Mol Biol 2005; 33: 319. 2. Rubenfeld GD, Caldwell E, Peabody E, et al. Incidence and outcomes of acute lung injury. N Engl J Med 2005; 353: 1685–1693. 3. Ailawadi G, Lau CL, Smith PW, et al. Does reperfusion injury still cause significant mortality after lung transplantation? J Thorac Cardiovasc Surg 2009; 137: 688–694. 4. Phua J, Badia JR, Adhikari NK, et al. Has mortality from acute respiratory distress syndrome decreased over time? A systematic review. Am J Respir Cell Mol Biol 2009; 179: 220–227. 5. Fiser SM, Tribble CG, Long SM, et al. Ischemia-reperfusion injury after lung transplantation increases risk of late bronchiolitis obliterans syndrome. Ann Thorac Surg 2002; 73: 1041–1048. 6. Ware LB, Matthay MA. The acute respiratory distress syndrome. N Engl J Med 2000; 342: 1334–1349. 7. Sueblinvong V, Weiss DJ. Stem cells and cell therapy approaches in lung biology and diseases. Transl Res 2010; 156: 188–205. 8. Mei SH, Mccarter SD, Deng Y, Parker CH, Liles WC, Stewart DJ. Prevention of LPS-induced acute lung injury in mice by Copyright © 2015 John Wiley & Sons, Ltd.

21.

22.

23.

24. 25.

26.

27.

28.

119

mesenchymal stem cells overexpressing angiopoietin 1. PLoS Med 2007; 4: e269. Qin ZH, Xu JF, Qu JM, et al. Intrapleural delivery of MSCs attenuates acute lung injury by paracrine/endocrine mechanism. J Cell Mol Med 2012; 16: 2745–2753. Gupta N, Su X, Popov B, Lee JW, Serikov V, Matthay MA. Intrapulmonary delivery of bone marrow-derived mesenchymal stem cells improves survival and attenuates endotoxin-induced acute lung injury in mice. J Immunol 2007; 179: 1855–1863. Kim ES, Chang YS, Choi SJ, et al. Intratracheal transplantation of human umbilical cord blood-derived mesenchymal stem cells attenuates Escherichia coli-induced acute lung injury in mice. Respir Res 2011; 12: 108. Liang Z, Sun J, Wang P, Tian Q, Yang Z, Chen L. Bone marrowderived mesenchymal stem cells protect rats from endotoxin-induced acute lung injury. Chin Med J (Engl) 2011; 124: 2715–2722. Aguilar S, Scotton CJ, Mcnulty K, et al. Bone marrow stem cells expressing keratinocyte growth factor via an inducible lentivirus protects against bleomycin-induced pulmonary fibrosis. PLoS One 2009; 4: e8013. Donoghue M, Hsieh F, Baronas E, et al. A novel angiotensinconverting enzyme-related carboxypeptidase (ACE2) converts angiotensin I to angiotensin 1–9. Circ Res 2000; 87: e1–e9. Zhong JC, Yu XY, Lin QX, et al. Enhanced angiotensin converting enzyme 2 regulates the insulin/Akt signalling pathway by blockade of macrophage migration inhibitory factor expression. Br J Pharmacol 2008; 153: 66–74. Warner F, Smith A, Hooper N, Turner A. Whats new in the reninangiotensin system? Angiotensin-converting enzyme-2: a molecular and cellular perspective. Cell Mol Life Sci 2004; 61: 2704–2713. Zhong JC, Huang DY, Yang YM, et al. Upregulation of angiotensinconverting enzyme 2 by all-trans retinoic acid in spontaneously hypertensive rats. Hypertension 2004; 44: 907–912. Imai Y, Kuba K, Penninger JM. The renin–angiotensin system in acute respiratory distress syndrome. Drug Discov Today 2006; 3: 225–229. Kaparianos A, Argyropoulou E. Local renin-angiotensin II systems, angiotensin-converting enzyme and its homologue ACE2: their potential role in the pathogenesis of chronic obstructive pulmonary diseases, pulmonary hypertension and acute respiratory distress syndrome. Curr Med Chem 2011; 18: 3506–3515. Imai Y, Kuba K, Rao S, et al. Angiotensin-converting enzyme 2 protects from severe acute lung failure. Nature 2005; 436: 112–116. Oyaizu T, Fung SY, Shiozaki A, et al. Src tyrosine kinase inhibition prevents pulmonary ischemia–reperfusion-induced acute lung injury. Intensive Care Med 2012; 38: 894–905. Bittner HB, Binner C, Dahlberg P, Mohr FW. Reducing ischemiareperfusion injury in clinical lung transplantation. Transplantation Proceedings 2007; 39: 489–492. Sun CK, Yen CH, Lin YC, et al. Autologous transplantation of adipose-derived mesenchymal stem cells markedly reduced acute ischemia-reperfusion lung injury in a rodent model. J Transl Med 2011; 9: 1–13. Botha P, Jeyakanthan M, Rao JN, et al. Inhaled nitric oxide for modulation of ischemia–reperfusion injury in lung transplantation. J Heart Lung Transplant 2007; 26: 1199–1205. Sun CK, Yen CH, Lin YC, et al. Autologous transplantation of adipose-derived mesenchymal stem cells markedly reduced acute ischemia-reperfusion lung injury in a rodent model. J Transl Med 2011; 9: 118. Liu F, Gao F, Li Q, Liu Z. The functional study of human umbilical cord mesenchymal stem cells harbouring angiotensin-converting enzyme 2 in rat acute lung ischemia-reperfusion injury model. Cell Biochem Funct 2014; 32: 580–589. Chen YT, Sun CK, Lin YC, et al. Adipose-derived mesenchymal stem cell protects kidneys against ischemia-reperfusion injury through suppressing oxidative stress and inflammatory reaction. J Transl Med 2011; 9: 51–57. Schächinger V, Erbs S, Elsässer A, et al. Intracoronary bone marrow– derived progenitor cells in acute myocardial infarction. N Engl J Med 2006; 355: 1210–1221. Cell Biochem Funct 2015; 33: 113–120.

120

x. zhang ET AL.

29. Lunde K, Solheim S, Aakhus S, et al. Intracoronary injection of mononuclear bone marrow cells in acute myocardial infarction. N Engl J Med 2006; 355: 1199–1209. 30. Filgueiras LR, Jr, Martins JO, Serezani CH, Capelozzi VL, Montes MB, Jancar S. Sepsis-induced acute lung injury (ALI) is milder in diabetic rats and correlates with impaired NFkB activation. PLoS One 2012; 7: e44987. 31. Karin M, Greten FR. NF-kappaB: linking inflammation and immunity to cancer development and progression. Nat Rev Immunol 2005; 5: 749–759. 32. Wang H, Zhang K, Zhao L, Tang J, Gao L, Wei Z. Anti-inflammatory effects of vinpocetine on the functional expression of nuclear factorkappa B and tumor necrosis factor-alpha in a rat model of cerebral ischemia-reperfusion injury. Neurosci Lett 2014; 566: 247–251. 33. Huang YC, Horng CT, Chen ST, et al. Rutin improves endotoxininduced acute lung injury via inhibition of iNOS and VCAM-1 expression. Environ Toxicol 2014. doi:10.1002/tox.22033. 34. Kira S, Daa T, Kashima K, Mori M, Noguchi T, Yokoyama S. Mild hypothermia reduces expression of intercellular adhesion molecule-1 (ICAM-1) and the accumulation of neutrophils after acid-induced lung injury in the rat. Acta Anaesthesiol Scand 2005; 49: 351–359. 35. Zhang JY, Ding CH, Suo XH, Dong CF, Ling YL. Effects of melatonin on anti-oxidative function of lung injury induced by lipopolysaccharide in rats and ICAM-1 expression. Zhongguo Ying Yong Sheng Li Xue Za Zhi 2008; 24: 464–467. 36. Baek JE, Yang WS, Chang JW, et al. Fatty acid-bearing albumin induces VCAM-1 expression through c-Src kinase-AP-1/NF-kB pathways: effect of L-carnitine. Kidney & Blood Pressure Research 2010; 33: 72–84. 37. Tsai CL, Chen WC, Hsieh HL, Chi PL, Hsiao LD, Yang CM. TNF-alpha induces matrix metalloproteinase-9-dependent soluble intercellular

Copyright © 2015 John Wiley & Sons, Ltd.

38.

39.

40. 41.

42.

43. 44.

45.

adhesion molecule-1 release via TRAF2-mediated MAPKs and NFkappaB activation in osteoblast-like MC3T3-E1 cells. J Biomed Sci 2014; 21: 12. Alexander MR, Murgai M, Moehle CW, Owens GK. Interleukin-1beta modulates smooth muscle cell phenotype to a distinct inflammatory state relative to PDGF-DD via NF-kappaB-dependent mechanisms. Physiol Genomics 2012; 44: 417–429. Liu GX, Li YQ, Huang XR, et al. Disruption of Smad7 promotes ANG II-mediated renal inflammation and fibrosis via Sp1-TGF-beta/Smad3NF.kappaB-dependent mechanisms in mice. PLoS One 2013; 8: e53573. Raval CM, Lee PJ. Heme oxygenase-1 in lung disease. Curr Drug Targets 2010; 11: 1532–1540. Inoue S, Suzuki M, Nagashima Y, et al. Transfer of heme oxygenase 1 cDNA by a replication-deficient adenovirus enhances interleukin 10 production from alveolar macrophages that attenuates lipopolysaccharide-induced acute lung injury in mice. Hum Gene Ther 2001; 12: 967–979. Kung HN, Weng TY, Liu YL, Lu KS, Chau YP. Sulindac compounds facilitate the cytotoxicity of beta-lapachone by up-regulation of NAD (P)H quinone oxidoreductase in human lung cancer cells. PLoS One 2014; 9: e88122. Kuwano K, Hara N. Signal transduction pathways of apoptosis and inflammation induced by the tumor necrosis factor receptor family. Am J Respir Cell Mol Biol 2000; 22: 147–149. Albertine KH, Soulier MF, Wang Z, et al. Fas and fas ligand are upregulated in pulmonary edema fluid and lung tissue of patients with acute lung injury and the acute respiratory distress syndrome. Am J Pathol 2002; 161: 1783–1796. Martin TR, Nakamura M, Matute-Bello G. The role of apoptosis in acute lung injury. Crit Care Med 2003; 31: S184–188.

Cell Biochem Funct 2015; 33: 113–120.