Inhibition of ALDH2 expression aggravates renal injury in a rat sepsis syndrome model

Transcription

1 EXPERIMENTAL AND THERAPEUTIC MEDICINE 14: , 2017 Inhibition of ALDH2 expression ggrvtes renl injury in rt sepsis syndrome model JUN FENG HU 1*, HUA XUE WANG 2*, HUI HUI LI 3, JIE HU 4, YING YU 4 nd QIN GAO 4 Deprtments of 1 Respirtory Disese nd 2 Intensive Cre Unit, The First Affilited Hospitl of Bengbu Medicl College, Bengbu, Anhui ; Deprtments of 3 Histology nd Embryology nd 4 Physiology, Bengbu Medicl College, Bengbu, Anhui , P.R. Chin Received Februry 28, 2016; Accepted Mrch 10, 2017 DOI: /etm Abstrct. Mitochondril ldehyde dehydrogense 2 (ALDH2) is closely ssocited with orgn injury. The im of the present study ws to investigte the chnge of ALDH2 expression in rt model of sepsis induced cute renl injury, nd to observe the effect of ALDH2 inhibition on the kidney. A model of sepsis syndrome ws estblished in Sprgue Dwley (SD) rts by cecl ligtion nd puncture (CLP). The rts were divided into shm, CLP nd CLP + cynmide (CYA, n ALDH2 inhibitor) groups. The hemodynmic prmeters hert rte (HR) nd men rteril blood pressure (MABP) were mesured. Plsm cretinine (CRE) nd ure nitrogen (BUN) levels were mesured using n utomtic biochemicl nlyzer. Mlondildehyde (MDA) content nd superoxide dismutse (SOD) ctivity in the kidney tissue were mesured. Histologicl chnges of the kidney tissue were observed using hemtoxylin nd eosin stining nd NF κb p65 expression ws observed by n immunohistochemicl stining method. The expression of renl ALDH2 t the mrna nd protein levels ws detected by reverse trnscription polymerse chin rection nd western blotting. In the CLP compred with the shm group fter 24 h, the MABP ws decresed, plsm CRE nd BUN levels were elevted, the renl MDA level ws incresed nd SOD ctivity ws decresed. In ddition, glomerulr trophy occurred, the renl protein expression of NF κb p65 ws incresed, nd the mrna nd protein expression levels of ALDH2 were decresed. In contrst with the CLP group, in the CLP + CYA group, the MABP nd ALDH2 expression were further decresed while glomerulr trophy ws ggrvted. Furthermore, CRE, BUN, MDA Correspondence to: Dr Qin Go, Deprtment of Physiology, Bengbu Medicl College, 2600 Dong Hi Avenue, Bengbu, Anhui , P.R. Chin E mil: bbmcgq@126.com * Contributed eqully Key words: sepsis, renl injury, mitochondril ldehyde dehydrogense 2, nucler fctor κb levels nd NF κb p65 expression were further incresed nd SOD ctivity ws further reduced. In this rt model of sepsis syndrome, the reduction of renl ALDH2 expression ws ccompnied by kidney injury. Inhibition of ALDH2 with CYA ggrvted the renl injury, nd ws ssocited with the overproduction of rective oxygen species nd inflmmtory rection. Introduction Sepsis is the leding cuse of mortlity in the intensive cre unit (ICU), with mortlity rte rnging from 20% in sepsis to >60% in septic shock (1). In prticulr, sepsis induced cute kidney injury (AKI) ccounts for ~50% of ll cses of AKI. Furthermore, ICU mortlity is s high s 70% in ptients with sepsis nd concomitnt AKI (2 4). The pthophysiology of septic AKI hs not been completely estblished, but is likely to involve immunologicl, toxic nd inflmmtory fctors tht my ffect the microvsculture nd tubulr cells, subsequently inducing renl necrosis nd poptosis (5). Renl mitochondril dysfunction lso occurs during the course of sepsis induced AKI (6). Mitochondril ldehyde dehydrogense 2 (ALDH2) is member of the ALDH gene fmily, which is highly expressed in the hert, liver, kidney nd muscle (7). ALDH2 ctlyzes the oxidtion of cetldehyde to cetic cid in the metbolism of ethnol, nd detoxifies rective ldehydes (8). With regrd to the role of ALDH2 in the pthogenesis of sepsis, Chen et l reported tht heptic ALDH2 ctivity ws downregulted in septic rts following cecl ligtion nd puncture (CLP) (9). It hs been reported tht preconditioning with ethnol to stimulte ALDH2 ctivity prevented kidney ischemi nd reperfusion injury in mice (10); however, the role of renl ALDH2 in sepsis induced AKI is uncler. It remins to be determined whether ALDH2 is lso key fctor ssocited with kidney injury. In the present study, rt model of septic injury ws creted by the CLP method, with the im of observing whether the septic injury dmged kidney morphology nd function, nd evluting the effects of the septic injury on ALDH2 in the kidney. Furthermore, the effects of tretment with n ALDH2 inhibitor were investigted.

2 2250 HU et l: INHIBITION OF ALDH2 AGGRAVATES SEPSIS-INDUCED RENAL INJURY Mterils nd methods Animls. A totl of 18 Mle Sprgue Dwley rts ( g; 7 8 weeks) were obtined from the Animl Center of Bengbu Medicl College (Bengbu, Chin). The rts were housed in controlled environment ( constnt 12 h light/drk cycle with temperture of 21+1 C nd 50 60% humidity), fed norml chow nd hd free ccess to tp wter. All niml procedures were in conducted ccordnce with the 8th Edition of the Ntionl Institutes of Helth Guide for the Cre nd Use of Lbortory Animls, 2010, nd were pproved by the Animl Reserch Ethics Committee of Bengbu Medicl College (Bengbu, Chin). Chemicls nd regents. Cynmide (CYA) ws purchsed from Sigm Aldrich (Merck KGA, Drmstdt, Germny). Mouse nti ALDH2 monoclonl ntibody (sc ) nd nti β ctin monoclonl ntibody (sc 81178) were purchsed from Snt Cruz Biotechnology, Inc. (Dlls, TX, USA). Rbbit nti nucler fctor (NF) κb p65 polyclonl ntibody (PB0321) nd horserdish peroxidse (HRP) linked nti mouse IgG secondry ntibody (BA1050) were purchsed from Wuhn Boster Biologicl Technology, Ltd, Wuhn, Chin. Mlondildehyde (MDA; ct. no. A003 1) nd superoxide dismutse (SOD; ct. no. A001 1) detection kits were purchsed from Jincheng Bioengineering Institute (Nnjing, Chin). The primers for ALDH2 were 5' GTG TTC GGA GAC GTC AAA GA 3' nd 5' GCA GAG CTT GGG ACA GGT AA 3', with n mplified frgment length of 187 bp; the primers for β ctin were 5' GAT GGT GGG TAT GGG TCA GAA GGA C 3' nd 5' GCT CAT TGC CGA TAG TGA TGA CT 3', with n mplified frgment length of 630 bp. All other chemicls were of the highest purity vilble. Induction of sepsis nd experimentl protocol. Animls were rndomly divided into shm, CLP nd CLP + CYA groups (ech n=6). Sepsis ws induced in overnight fsted rts by the CLP modeling method (11). All rts were injected with 0.4 g/kg chlorl hydrte intrperitonelly to induce nesthesi. In the CLP group, under sterile surgicl conditions, 2 cm bdominl incision ws mde long the ventrl surfce of the bdomen to expose the cecum. The cecum ws then punctured twice with 22 guge needle nd fecl contents were llowed to lek into the peritoneum by gently squeezing cecum. The bowel ws then returned to the bdomen nd the bdominl cvity ws closed. In the shm group, the nimls were submitted to lprotomy nd the cecum ws mnipulted but neither ligted nor punctured. In the CLP + CYA group, rts were subjected to CLP nd then injected intrvenously with CYA t dose of 25 mg/kg body weight 30 min fter the CLP procedure. Hemodynmic prmeter mesurement. At 24 h fter CLP, rts were injected with 0.4 g/kg chlorl hydrte intrperitonelly to induce nesthesi. Hemodynmic prmeters including the hert rte (HR) nd men rteril blood pressure (MABP) were mesured continuously for 50 min by intubtion of the left common crotid rtery using Medlb biologicl signl collecting nd processing system (Nnjing, Chin). Plsm cretinine (CRE) nd blood ure nitrogen (BUN) mesurement. After the hemodynmic prmeters of the rts hd been tken, 2 ml blood ws obtined from the left common crotid rtery, nd the plsm CRE nd BUN levels were mesured using n utomtic biochemicl nlyzer (AU5400; Olympus Corportion, Tokyo, Jpn). MDA content nd SOD ctivity mesurement. All rts were injected with 0.4 g/kg chlorl hydrte intrperitonelly to induce nesthesi. Under sterile surgicl conditions, n bdominl incision ws mde nd kidneys were perfused with 0.01 mol/l PBS vi the bdominl ort, until the blood ws wshed out, nd the prenchym ws ple in ppernce. Then the kidneys were removed s quickly s possible, nd the right kidney ws stored t 80 C for MDA content nd SOD ctivity mesurement. Kidney tissue (100 mg) ws homogenized in ice cold sline solution. The MDA content nd SOD ctivity were mesured using commercilly vilble kits, ccording to the mnufcturer's instructions (12). Renl morphology observtion nd NF κb p65 protein expression mesurement. The renl tissues of left kidney were immersed in 10% buffered formlin. The tissues were grdully dehydrted, embedded in prffin, cut into 5 µm sections nd stined with hemtoxylin nd eosin (H&E) for histologicl evlution. Renl NF κb p65 protein expression ws detected by n immunohistochemistry method. Immunohistochemicl stining ws performed using n Elivision Plus detection kit ccording to the mnufcturer's protocol (KIT 9902, LbVision AB, Torsby, Sweden). Kidney sections (4 µm) were deprffinized with xylene nd rehydrted with decresing percentges of ethnol, then wshed with PBS (ph 7.2) for 10 min. The endogenous peroxidse ctivity ws blocked by incubtion with 3% H 2 O 2 in methnol for 10 min t room temperture. Sections were subsequently plced in citrte buffer (ph 6.0) t 95 C for ntigen repir for 30 min, nd the slides were rinsed with PBS 3 times. All sections were blocked with 10% got serum ( ; Zhejing Tinhng Biotechnology Co., Ltd., Huzhou, Chin) for 30 min t room temperture. Kidney sections were incubted overnight t 4 C with rbbit nti NF κb p65 polyclonl ntibody (1:200). All sections were counterstined with hemtoxylin for 30 sec t 60 C, dehydrted, ir dried, nd mounted. Histologicl sections were observed nd imges were cptured using n Olympus CKX41 Inverted light microscope (Olympus Corportion). Detection of ALDH2 mrna by reverse trnscription quntittive polymerse chin rection (RT qpcr). RT qpcr ws used to detect the renl expression of ALDH2 mrna (12). Briefly, totl RNA ws extrcted from rt renl tissue with TRIzol regent (Invitrogen; Thermo Fisher Scientific, Inc. Wlthm, MA, USA) ccording to the mnufcturer's protocol. Totl RNA ws treted with DNse I (D8071; Beijing Solrbio Science nd Technology Co., Ltd., Beijing, Chin) to remove the genomic DNA. RNA ws solubilized in ribonuclese free wter nd quntified by mesuring the bsorbnce t 260 nm by UV spectrophotometer (BioPhotometer plus; Eppendorf, Hmburg, Germny). The purity of RNA ws confirmed by exmining the opticl density (OD) 260/280 s Totl RNA (2 µg) ws reverse trnscribed to cdna using the

3 EXPERIMENTAL AND THERAPEUTIC MEDICINE 14: , RevertAid First Strnd cdna Synthesis kit (K1622; Thermo Fisher Scientific Inc.) ccording to the mnufcturer's protocol. Briefly, the mix (2 µg totl RNA, 1 µl OligodT 18 primer, 4 µl 5X Rection Buffer, 1 µl 20 U/µl RiboLock RNse Inhibitor, 2 µl 10 mmol/l dntps Mix, 1 µl 200 U/µl RevertAid M MuLV RT nd ribonuclese free wter to give finl colume of 20 µl) ws incubted for 60 min t 42 C, nd the rection ws terminted by heting the mixture to 70 C for 5 min. PCR ws performed using PCR Mster Mix (2X; K0171; Thermo Fisher Scientific, Inc.) using the forementioned primers ccording to the mnufcturer's protocol. The rection mixture contined 25 µl 2X PCR Mster Mix (0.05 U/µl Tq DNA polymerse, rection buffer, 4 mmol/l MgCl 2, nd 0.4 mmol/l of ech dntp), 2 µl templte DNA (50 ng), 0.5 µl of ech 20 pmol/µl primer, nd 22 µl of double distilled H 2 O to give totl volume of 50 µl. The PCR conditions were s follows: 95 C for 3 min, then 30 cycles of 95 C for 50 sec, 60 C for 45 sec, nd 72 C for 60 sec, followed by finl extension step t 72 C for 10 min. PCR products were seprted on 1% grose gel. Densitometry results for ALDH2 were nlyzed with Tnon Gel Imge System 1D (version 4.1.2; Tnon, Shnghi, Chin) nd compred with the corresponding β ctin levels to ccount for loding differences. All experiments were repeted three times. Detection of ALDH2 protein by western blotting. Rt renl tissue (0.1 g) ws homogenized in lysis buffer (P0013; Beyotime Institute of Biotechnology, Himen, Chin) supplemented with 1% protese inhibitor cocktil. Following centrifugtion t 12,000 x g for 30 min t 4 C, the superntnt ws collected, nd the protein concentrtion ws determined using n enhnced bicinchoninic cid protein ssy kit ccording to the mnufcturer's protocol (Nnjing Jincheng Bioengineering Institute, Nnjing, Chin). Protein (80 µg) ws seprted on sodium dodecyl sulfte polycrylmide gels in minigel pprtus nd then trnsferred electrophoreticlly to polyvinylidene difluoride membrne. The membrnes were blocked with 5% non ft dry milk in Tris buffered sline prior to overnight incubtion t 4 C with nti ALDH2 (1:500) nd nti β ctin (1:500) ntibody. Membrnes were then incubted for 1 h t 37 C with HRP linked nti mouse IgG (1:5,000). Autordiogrphs were scnned using the ChemiDoc XRS Gel Imge System nd nlyzed with Imge Lb softwre (version 3.0; Bio Rd Lbortories, Inc., Hercules, CA, USA) (12). Sttisticl nlysis. All vlues re expressed s the men ± stndrd error of the men. Sttisticl comprisons were performed by one wy nlysis of vrince nd the Newmn Keuls test. P<0.05 ws considered to indicte sttisticlly significnt difference. Results Chnges of hemodynmic prmeters. The MABP in the CLP group ws decresed significntly compred with tht in the shm group, nd the MABP ws decresed further in the CLP + CYA group t 24 h fter the CLP procedure. No significnt difference in HR ws observed mong the different groups (Tble I). Tble I. Hemodynmic dt in the three groups (n=6). Group HR (bets/min) MABP (mmhg) Shm ± ±3.84 CLP ± ±4.79 CLP + CYA ± ±6.33,b P<0.01 vs. the shm group; b P<0.01 vs. the CLP group. HR, hert rte; MABP, men rteril blood pressure; CLP, cecl ligtion nd puncture; CYA, cynmide. Tble II. Renl functionin the three groups (n=6). Group BUN (mmol/l) CRE (µmol/l) Shm 9.17± ±2.58 CLP 19.25± ±7.50 b CLP + CYA 32.24±8.43 b,c 82.66±10.07 b,c P<0.05 nd b P<0.01 vs. the shm group; c P<0.01 vs. the CLP group. BUN, blood ure nitrogen; CRE, cretinine; CLP, cecl ligtion nd puncture; CYA, cynmide. Tble III. Renl SOD ctivity nd MDA content in the three groups (n=6). Group SOD (U/mg.protein) MDA (nmol/mg.protein) Shm ± ±0.42 CLP ± ±1.33 CLP + CYA ±10.81,b 11.42±2.56,b P<0.01 vs. the shm group; b P<0.01 vs. the CLP group. SOD, superoxide dismutse; MDA, mlondildehyde; CLP, cecl ligtion nd puncture; CYA, cynmide. Chnges of plsm CRE nd BUN levels. In comprison with the plsm CRE nd BUN levels in the shm group, those in the CLP group were incresed, nd those in the CLP + CYA group were further incresed (Tble II). Chnges of renl MDA content nd SOD ctivity. The renl MDA content ws incresed nd SOD ctivity ws decresed in the CLP group compred with the shm group. Furthermore, the MDA content ws higher nd SOD ctivity ws lower in the CLP + CYA group compred with the CLP group (Tble III). Histologicl nlysis of the renl tissue. In the shm group, the renl corpuscles ppered norml with dense, rounded structures, nd the Bowmn's spces surrounding the glomeruli were nrrow. In the CLP group, the glomeruli ppered wrinkled nd the Bowmn's spces were wider thn those in the shm group. In the CLP + CYA group, the wrinkling of the glomeruli ws ggrvted compred with tht in the CLP group (Fig. 1).

4 2252 HU et l: INHIBITION OF ALDH2 AGGRAVATES SEPSIS-INDUCED RENAL INJURY Tble IV. Renl ALDH2 expression t the mrna nd protein levels in the three groups (n=6). Group ALDH2/β ctin mrna ALDH2/β ctin protein Shm 0.86± ±0.17 CLP 0.52± ±0.08 b CLP + CYA 0.45±0.08,c 0.38±0.09,c P<0.01 nd b P<0.05 vs. the shm group; c P<0.05 vs. the CLP group. ALDH2, ldehyde dehydrogense 2; CLP, cecl ligtion nd puncture; CYA, cynmide. Chnge of NF κb p65 in renl tissue. In the shm group, the renl expression of NF κb p65 protein ws low. Compred with tht in the shm group, renl NF κb p65 protein ws highly expressed in the CLP nd CLP + CYA groups, nd ws prticulrly high in the CLP + CYA group (Fig. 2). Figure 1. Histologicl observtion of the kidney tissue in the three groups with hemtoxylin nd eosin stining. (A) Mgnifiction, x100 nd (B) mgnifiction, x400. In the shm group, renl corpuscles ppered norml with dense, rounded structures, nd the glomeruli (indicted by rrows) were surrounded by nrrow Bowmn's spces. In the CLP group, the glomerulus ws wrinkled with wider Bowmn's spces. In the CLP + CYA group, the wrinkling of the glomerulus ws ggrvted. CLP, cecl ligtion nd puncture; CYA, cynmide. Chnges of renl ALDH2 t the mrna nd protein levels. Renl ALDH2 expression ws decresed t the mrna nd protein levels in the rts of the CLP nd CLP + CYA groups compred with the shm group, nd ws prticulrly low in the CLP + CYA group (Tble IV nd Fig. 3). Discussion In the current study, it ws identified tht in the CLP induced rt sepsis model, renl expression of ALDH2 t the mrna nd protein levels ws decresed, which ws ccompnied by reduction of renl function, the overproduction of rective oxygen species nd incresed NF κb protein expression when compred with rts subjected to shm procedure. When the septic rts were treted with CYA, n ALDH2 inhibitor, renl injury, oxidtive stress nd inflmmtion were ggrvted s the renl ALDH2 ctivity ws reduced, which suggests tht ALDH2 could be key fctor regulting the pthologicl chnges of sepsis induced renl injury. Despite dvnces in modern medicine, sepsis nd its sequele remin mjor cuse of ptient mortlity. CLP in rts cretes relistic sepsis model. Sepsis is systemic inflmmtory response tht induces tissue dmge nd multiple orgn dysfunction syndrome, including AKI, septic shock, septic lethlity nd even deth (13). The ctivtion of NF κb plys centrl role in the pthophysiology of septic shock. During sepsis, NF κb is ctivted in mny orgns nd hs potent effect on downstrem signling pthwys nd tissue injury. Inhibiting NF κb expression restores systemic hypotension, meliortes septic myocrdil dysfunction nd vsculr derngement, inhibits the expression of multiple proinflmmtory genes, diminishes intrvsculr cogultion, reduces tissue neutrophil influx nd prevents microvsculr endothelil lekge (14). In study of ptients with sepsis, NF κb ctivity ws found to be mrkedly incresed in vrious orgns, nd higher NF κb ctivity exhibited n ssocition with higher mortlity rtes nd worse clinicl outcomes (15), which indictes tht inflmmtory pthwys ply n importnt role Figure 2. Protein expression of nucler fctor κb p65 in the kidney tissue of rts from the three groups, stined using the Elivision plus two step System nd counterstined with hemtoxylin (mgnifiction, x400). CLP, cecl ligtion nd puncture; CYA, cynmide. Figure 3. Expressions of renl ALDH2 t the mrna nd protein levels in the three groups. Representtive (A) reverse trnscription polymerse chin rection nd (B) western blotting results re shown. ALDH2, ldehyde dehydrogense 2; CLP, cecl ligtion nd puncture; CYA, cynmide. in sepsis induced orgn dysfunction. In the present study, it ws observed tht in the rt model of sepsis, renl injury ws

5 EXPERIMENTAL AND THERAPEUTIC MEDICINE 14: , ssocited incresed renl NF κb expression, suggesting tht sepsis induced renl injury is ssocited with n inflmmtory response. In the present study, it ws observed tht the inhibition of renl ALDH2 using CYA ws ccompnied by oxidtive stress in the septic rts. Furthermore, renl function injury, oxidtive stress nd the inflmmtory response were further ggrvted when ALDH2 ctivity ws reduced. Sepsis is ssocited with mitochondril dysfunction nd impired oxygen consumption (16,17), ALDH2 is loclized in the mitochondril frction, nd mitochondril dysfunction chnges ALDH2 function. A number of studies hve reported tht ALDH2 protects ginst orgn injury, such s tht ffecting the kidneys, hert nd neurons (10,18 20). Our previous studies showed tht crdic ALDH2 expression ws decresed in rts with dibetes, or myocrdil ischemi nd reperfusion injury, nd tht upregultion of ALDH2 protected ginst myocrdil injury nd protected the lungs ginst dibetes induced lung injury, ll of which indicte tht ALDH2 could be n intrinsic fctor regulting the occurrence of disese (12,21,22). Whether ALDH2 is lso n intrinsic fctor for the regultion of sepsis induced orgn injury ws thus considered. It hs previously been reported tht ALDH2 ctivity is reduced in the livers of rts with CLP induced sepsis (9). ALDH2 hs lso been demonstrted to be protective ginst LPS induced crdic dysfunction (23), nd incresed ALDH2 expression ws observed to reduce renl cell poptosis during ischemi nd reperfusion injury (24). However, the role of ALDH2 in sepsis induced renl injury remins poorly elucidted. ALDH2 hs been reported to ttenute oxidtive stress. Sepsis induces mitochondril dysfunction, nd inflmmtion ssocited with the overproduction of rective oxygen species (17). The scvenging of mitochondril rective oxygen species protects mitochondril function, ttenutes tissue inflmmtion, nd improves whole orgn ctivities in the hert during sepsis (25). Since ALDH2 cn rpidly detoxify toxic rective ldehydes nd ttenute the oxidtive stress response, when ALDH2 ctivtion in sepsis rt ws inhibited, the kidney injury ws ggrvted, suggesting tht ALDH2 plys key protective role ginst sepsis induced orgn injury. The present study hs certin limittions. The study simply mesured the chnges of ALDH2 in rts with sepsis induced renl injury, nd observed the chnges of renl function when the rts were treted with CYA, non specific ALDH2 inhibitor. However, this is preliminry study conducted to verify whether ALDH2 prticiptes in sepsis induced kidney injury. In future studies, we will firstly select specific ALDH2 inhibitor such s didzin to observe its effects on the kidney functions of the septic rts, then use the ALDH2 ctivtor ld 1 to observe whether the ctivtion of ALDH2 ttenutes kidney injury. The likely mechnism for the protective effect of ALDH2 in sepsis induced renl injury will be investigted further in the future. From the present study, it ws concluded tht in sepsis induced renl injury, renl ALDH2 expression ws decresed, renl function ws ttenuted, nd oxidtive stress nd inflmmtion occurred. Inhibition of ALDH2 expression in the kidney ggrvted the renl injury nd inflmmtion, which suggests tht ALDH2 might be n intrinsic regultor of sepsis induced kidney dysfunction. Further studies re necessry to confirm the role of ALDH2. Acknowledgements This study ws supported by the Nturl Science Foundtion of Anhui, Chin (grnt nos MH169 nd QH150). References 1. Angus DC, Pereir CA nd Silv E: Epidemiology of severe sepsis round the world. Endocr Metb Immune Disord Drug Trgets 6: , Sriswt N nd Kellum JA: Acute kidney injury: Definition, epidemiology, nd outcome. Curr Opin Crit Cre 17: , Oppert M: Acute kidney injury in sepsis: More thn just brief encounter. Crit Cre Med 41: , White LE, Chudhry R, Moore LJ, Moore FA nd Hssoun HT: Surgicl sepsis nd orgn crosstlk: The role of the kidney. J Surg Res 167: , Wn L, Bgshw SM, Lngenberg C, Sotome T, My C nd Bellomo R: Pthophysiology of septic cute kidney injury: Wht do we relly know? Crit Cre Med 36 (Suppl 4): S198 S203, Regueir T, Andresen M, Mercdo M nd Downey P: Physiopthology of cute renl filure during sepsis. Med Intensiv 35: , 2011 (In Spnish). 7. Stewrt MJ, Mlek K nd Crbb DW: Distribution of messenger RNAs for ldehyde dehydrogense 1, ldehyde dehydrogense 2, nd ldehyde dehydrogense 5 in humn tissues. J Invest Med 44: 42 46, Buds GR, Distnik MH nd Mochly Rosen D: Aldehyde dehydrogense 2 in crdic protection: A new therpeutic trget? Trends Crdiovsc Med 19: , Chen HW, Kuo HT, Hwng LC, Kuo MF nd Yng RC: Proteomic ltertion of mitochondril ldehyde dehydrogense 2 in sepsis regulted by het shock response. Shock 28: , Yun Q, Hong S, Hn S, Zeng L, Liu F, Ding G, Kng Y, Mo J, Ci M, Zhu Y nd Wng QX: Preconditioning with physiologicl levels of ethnol protect kidney ginst ischemi/reperfusion Injury by modulting oxidtive stress. PLoS One 6: e25811, Mttick JS, Yng Q, Ormn MA, Ierpetritou MG, Berthiume F nd Androulkis IP: Long term gene expression profile dynmics following cecl ligtion nd puncture in the rt. J Surg Res 178: , Go Q, Wng HJ, Wng XM, Kng PF, Yu Y, Ye HW, Zhou H nd Li ZH: Activtion of ALDH2 with ethnol ttenutes dibetes induced myocrdil injury in rts. Food Chem Toxicol 56: , Bellomo R, Wn L, Lngenberg C nd My C: Septic cute kidney injury: New concepts. Nephron Exp Nephrol 109: e95 e100, Liu SF nd Mlik AB: NF kppb ctivtion s pthologicl mechnism of septic shock nd inflmmtion. Am J Physiol Lung Cell Mol Physiol 290: L622 L645, Pterson RL, Glley HF, Dhillon JK nd Webster NR: Incresed nucler fctor kpp B ctivtion in criticlly ill ptients who die. Crit Cre Med 28: , Grrbou G, Morén C, López S, Tobís E, Crdellch F, Miró O nd Csdemont J: The effects of sepsis on mitochondri. J Infect Dis 205: , Glley HF: Oxidtive stress nd mitochondril dysfunction in sepsis. Br J Anesth 107: 57 64, Chen CH, Buds GR, Churchill EN, Distnik MH, Hurley TD nd Mochly Rosen D: Activtion of ldehyde dehydrogense 2 reduces ischemic dmge to the hert. Science 321: , Kod K, Slzr Rodriguez M, Corti F, Chn NY, Estephn R, Silver RB, Mochly Rosen D nd Levi R: Aldehyde dehydrogense ctivtion prevents reperfusion rrhythmis by inhibiting locl renin relese from crdic mst cells. Circultion 122: , Wey MC, Fernndez E, Mrtinez PA, Sullivn P, Goldstein DS nd Strong R: Neurodegenertion nd motor dysfunction in mice lcking cytosolic nd mitochondril ldehyde dehydrogenses: Implictions for Prkinson's disese. PLoS One 7: e31522, 2012.

6 2254 HU et l: INHIBITION OF ALDH2 AGGRAVATES SEPSIS-INDUCED RENAL INJURY 21. Yu Y, Ji XJ, Zong QF, Zhng GJ, Ye HW, Hu J, Go Q nd Gun SD: Remote ischemic postconditioning protects the hert by upregulting ALDH2 expression levels through the PI3K/Akt signling pthwy. Mol Med Rep 10: , Hu JF, Zhng GJ, Wng L, Kng PF, Li J, Wng HJ, Go Q nd Chen YQ: Ethnol t low concentrtion ttenutes dibetes induced lung injury in rts model. J Dibetes Res 2014: , Hu Y, Yn JB, Zheng MZ, Song XH, Wng LL, Shen YL nd Chen YY: Mitochondril ldehyde dehydrogense ctivity protects ginst lipopolyscchride induced crdic dysfunction in rts. Mol Med Rep 11: , Zhong Z, Hu Q, Fu Z, Wng R, Xiong Y, Zhng Y, Liu Z, Wng Y nd Ye Q: Incresed expression of ldehyde dehydrogense 2 reduces renl cell poptosis during ischemi/reperfusion injury fter hypothermic mchine perfusion. Artif Orgns 40: , Zng QS, Sdek H, Mss DL, Mrtinez B, M L, Kilgore JA, Willims NS, Frntz DE, Wigginton JG, Nwriku FE, et l: Specific inhibition of mitochondril oxidtive stress suppresses inflmmtion nd improves crdic function in rt pneumoni relted sepsis model. Am J Physiol Hert Circ Physiol 302: H1847 H1859, 2012.