research article propofol attenuates small intestinal...

Research ArticlePropofol Attenuates Small Intestinal IschemiaReperfusion Injury through Inhibiting NADPH OxidaseMediated Mast Cell Activation

Xiaoliang Gan12 Dandan Xing1 Guangjie Su1 Shun Li1 Chenfang Luo1 Michael G Irwin3

Zhengyuan Xia3 Haobo Li3 and Ziqing Hei1

1 Department of Anesthesiology TheThird Affiliated Hospital Sun Yat-sen University Guangzhou 510630 China2 Zhongshan Ophthalmic Center Department of Anesthesiology Sun Yat-sen University Guangzhou 510060 China3Department of Anesthesiology University of Hong Kong Hong Kong

Correspondence should be addressed to Haobo Li haobolihkuhk and Ziqing Hei heiziqingsinacom

Received 18 July 2014 Accepted 7 September 2014

Academic Editor Mengzhou Xue

Copyright copy 2015 Xiaoliang Gan et al This is an open access article distributed under the Creative Commons Attribution Licensewhich permits unrestricted use distribution and reproduction in any medium provided the original work is properly cited

Both oxidative stress and mast cell (MC) degranulation participate in the process of small intestinal ischemia reperfusion (IIR)injury and oxidative stress induces MC degranulation Propofol an anesthetic with antioxidant property can attenuate IIR injuryWe postulated that propofol can protect against IIR injury by inhibiting oxidative stress subsequent fromNADPHoxidasemediatedMC activation Cultured RBL-2H3 cells were pretreated with antioxidant N-acetylcysteine (NAC) or propofol and subjected tohydrogen peroxide (H

2O2) stimulation without or with MC degranulator compound 4880 (CP) H

2O2significantly increased

cells degranulation which was abolished by NAC or propofol MC degranulation by CP further aggravated H2O2induced cell

degranulation of small intestinal epithelial cell IEC-6 cells stimulated by tryptase Rats subjected to IIR showed significant increasesin cellular injury and elevations of NADPH oxidase subunits p47phox and gp91phox protein expression increases of the specificlipid peroxidation product 15-F

2t-Isoprostane and interleukin-6 and reductions in superoxide dismutase activity with concomitantenhancements in tryptase and 120573-hexosaminidase MC degranulation by CP further aggravated IIR injury And all these changeswere attenuated by NAC or propofol pretreatment which also abrogated CP-mediated exacerbation of IIR injury It is concludedthat pretreatment of propofol confers protection against IIR injury by suppressing NADPH oxidase mediated MC activation

1 Introduction

Small intestinal ischemia reperfusion (IIR) injury has beenemerged in many pathophysiological settings including sep-tic and hemorrhagic shock induced hypoperfusion [1] aswell as acute mesenteric ischemia [2] and small intestinetransplantation or liver resection [3] IIR remains to be acritical problem associated with high mortality [4]

During IIR oxidative stress is increased due to burstproduction of reactive oxygen species (ROS)which is amajormechanism of IIR injury [5] Numerous studies have revealedthat increased ROS production resulted from overactivationof the prooxidant enzyme NADPH oxidase which abun-dantly exists in intestine tissue [6] and plays critical roles inmediating tissue injury related to a range of inflammatory

diseases [7] including ischemia reperfusion injury [8 9]Inhibition of NADPH oxidase by N-acetylcysteine (NAC)a scavenger of oxygen radicals has been shown to greatlyattenuate myocardial ischemia reperfusion injury [10 11]However the mechanism governing ROS production specif-ically from overactivation of NADPH oxidase during IIR isyet to be explored

Activation of NADPHoxidase can lead tomast cells (MCcells that widely present throughout small intestine) degran-ulationactivation [12] wherever MCs activation has beendemonstrated to be a key mediator in the pathogenesis of IIRand contributing to many disorders as a result of increasedrelease a diverse range of mediators including histaminetryptase and inflammatory cytokines such as interleukin-6(IL-6) [13] and that inhibition of MCs from degranulation

Hindawi Publishing CorporationOxidative Medicine and Cellular LongevityVolume 2015 Article ID 167014 15 pageshttpdxdoiorg1011552015167014

2 Oxidative Medicine and Cellular Longevity

can alleviate IIR injury [14 15] We have previously foundthat ROS production was significantly increased after IIRwhich is parallel with the enhancements inMCdegranulation[16] but it is unknown whether a causal relationship existsbetween increased ROS production and MC degranulationin the setting of IIR in vivo These findings [6 17] togetherwith reports showing that MC can be activated in a ROS-dependent pathway [18] both in vitro [18] and in vivo [16]prompted us to postulate that during IIR increased ROSproduction initiates andor exacerbates IIR injury primarilyvia activating MC and that NADPH oxidase activation isincreased during IIR which may be a major source of ROSoverproduction during IIR

Propofol an intravenous anesthetic with antioxidantproperty that we widely used in intensive care unit andoperation theatre has been shown to dose-dependentlyattenuate myocardial ischemia reperfusion injury in patients[19] Propofol has also been shown to inhibit mast cellexocytosis in a dose-dependent manner in vitro [20] A mostrecent study shows that propofol attenuates brain traumainduced cerebral injury through inhibiting NADPH oxidaseactivation [21] We therefore hypothesized that inhibition ofROS mediated MC activation subsequent to attenuation ofintestinal NADPH oxidase activation may represent a majormechanism by which propofol attenuates IIR injury Thishypothesis was tested in a rat model of mesenteric ischemiareperfusion in vivo and a rat cell line of mast cell exposed toROS in vitro

2 Materials and Methods

21 Cell Culture Ratmast cell line (RBL-2H3) was purchasedfrom the cell bank of the Chinese Academy of Sciences(Shanghai China) and was cultured in Eaglersquos minimumessential medium containing 10 fetal bovine serum sup-plemented with 100UmL penicillin and 100 120583gmL strep-tomycin at 37∘C in a humidified atmosphere of 5 CO

2

as described [22] A rat small intestinal epithelial cell line(IEC-6) was also obtained from the cell bank of the ChineseAcademy of Sciences (Shanghai China) This cell line wascultured in 1640 RPMI medium with 10 fetal bovine serumat 37∘C in a 5 CO

2incubator

22 Measurement of Cell Degranulation Degranulation ofRBL-2H3 cells was measured by determining the activityof released 120573-hexosaminidase in the culture supernatantsand cell lysates [23] The RBL-2H3 cells were incubatedwith different concentrations of NAC [24] or propofol [20]for 12 h after the cells were washed with 1 times PBS for 3times RBL-2H3 cells were incubated in a 24-well plate (1 times106 cellswell) at 37∘C overnightThe above cells were washedwith 1timesPBS and then incubatedwith different concentrationsof lipopolysaccharides (LPS) [25] or H

2O2[26] or Com-

pound 4880 [27] for 1 h or 4 h To measure the amount of120573-hexosaminidase activity released from the cells the cul-tured media were transferred and centrifuged at 4∘C Thesupernatant (25 120583L) was mixed with 50120583L p-NAG (10mM)in 01M sodium citrate buffer (pH 45) into a 96-well plate

and incubated for 1 h at 37∘CThe reaction was terminated bythe addition of stop buffer (01M sodium carbonate bufferpH 100) The 120573-hexosaminidase activity was determinedby measuring the difference of absorbance at wavelength405 nm

23 Measurement of Cell Viability IEC-6 cells were seededinto 96-well flat bottom culture plates at a density of 5 times104 cellswell and incubated for 24 h After exposure totryptase (0ndash1000 ngmL) for 12 h in RPMI 1640 culturemedium the cell viability was measured by modified MTTassay as described [28] The assay detected the reduction ofthe tetrazolium to formazan product The cell viability wasevaluated using the following formula Survival rate () =(119860 sample ndash 119860blank)(119860control ndash 119860blank) times 100

24 Assessment of Cell Cytotoxicity Cytotoxicity (cell necro-sis) was assessed by measuring the release of lactate dehy-drogenase (LDH) After exposure of the cells to various con-centrations of tryptase (0ndash1000 ngmL) for 2 h in RPMI 1640culture medium cell necrosis was assessed by LDH-releaseassay using a commercial kit (Jiancheng China) according tothe manufacturerrsquos instruction

25 Animals Female Sprague-Dawley (SD) rats weighing180ndash200 g purchased from Animal Center of GuangdongProvince (Guangzhou China) were housed individually inwire-bottomed cages and were placed under pathogen freecondition illuminated from 800 AM to 800 PM (1212 hlight-dark cycle) for one week before use The experimentalprotocol and design was approved by the Sun Yat-sen Uni-versity Animal Experimentation Committee and performedaccording to Sun Yat-sen University Guidelines for AnimalExperimentation

26 In Vivo Rat IIR Model and Treatments All the animals were fasted for 16 h (while free access to water wasallowed) before surgery Rats were respectively injectedwithN-acetylcysteine (NAC 05 gkg from Sigma company)propofol (50mgkg commercial product Diprivan fromAstraZeneca) intralipid (50mgkg 20 emulsion fromSigma) or normal saline (05mL100 g) which served as thecontrol group intraperitoneally at 600 PM for 3 successivedays The dosages of NAC were chosen based on the resultsshowing that treatment of rats with ip NAC (500mg kgminus1per day for 9 days) improved the renal hemodynamic changestriggered by cisplatin-mediated nephrotoxicity [29]The doseof propofol was chosen based on the finding that propofol50mgkg given intraperitoneally provided sedative effect butnot anesthetic effect [30] and that propofol when used at thisdosage attenuated IIR injury in rats [31] At the 4th day partsof the rats were sacrificed by overdose of anesthetic chloralhydrate and the intestinal mucous was obtained and scrapedfor further determination and the intestinal morphologicalchanges were assessed

Oxidative Medicine and Cellular Longevity 3

27 Experimental Groups The other rats were divided intothe following groups

(1) Sham-operated group (SHAM) (119899 = 6) rats pre-treated with normal saline (10mLkg ip) weresubjected to identical surgical procedures exceptfor superior mesenteric artery (SMA) occlusion for75min and were kept under anesthesia during theexperiment and were administrated with the samevolume of normal saline (1mLkg iv) as reagentsolvent control

(2) Sole IIR group (IIR) (119899 = 6) rats pretreated withnormal saline (10mLkg ip) were subjected to smallintestinal ischemia by occluding SMA (75min) fol-lowed by reperfusion (2 h) plus administration of nor-mal saline (1mLkg iv) 5min immediately beforereperfusion

(3) IIR + Compound 4880 group (IIR + CP) (119899 = 12)rats pretreated with normal saline (10mLkg ip)were subjected to small intestinal ischemia by occlud-ing SMA (75min) followed by reperfusion (2 h) plusadministration of Compound 4880 (075mgkg iv)dissolved in normal saline (1mLkg) 5min immedi-ately before reperfusion

(4) NAC+ IIR group (NAC+ IIR) (119899 = 6) rats pretreatedwith NAC (05 gkg ipday) dissolved in normalsaline (10mLkg) for 3 successive days were sub-jected to small intestinal ischemia by occluding SMA(75min) followed by reperfusion (2 h) plus adminis-tration of normal saline (1mLkg iv)

(5) NAC + IIR + Compound 4880 group (NAC + IIR +CP) (119899 = 6) rats pretreated with NAC (05 gkg ip)were subjected to small intestinal ischemia by occlud-ing SMA (75min) followed by reperfusion (2 h) plusadministration of Compound 4880 (075mgkg iv)

(6) Propofol + IIR group (Pro + IIR) (119899 = 6) rats pre-treatedwith propofol (50mgkg ipday) dissolved inintralipid (10mLkg) for 3 successive days were sub-jected to small intestinal ischemia by occluding SMA(75min) followed by reperfusion (2 h) plus adminis-tration of normal saline (1mLkg iv)

(7) Propofol + IIR + Compound 4880 group (Pro +IIR + CP) (119899 = 6) rats pretreated with propofol(50mgkg ip) were subjected to small intestinalischemia by occluding SMA (75min) followed byreperfusion (2 h) plus administration of Compound4880 (075mgkg iv)

(8) Intralipid + IIR group (Lip + IIR) (119899 = 12) ratspretreated with intralipid (50mgkg ip) (with a vol-ume of 10mLkg) were subjected to small intestinalischemia by occluding SMA (75min) followed byreperfusion (2 h) plus administration of normal saline(1mLkg iv)

(9) Intralipid + IIR + Compound 4880 group (Lip +IIR + CP) (119899 = 12) rats pretreated with intralipid(50mgkg ip) were subjected to small intestinal

ischemia by occluding SMA (75min) followed byreperfusion (2 h) plus administration of Compound4880 (075mgkg iv)

It is of notice that with respect to the low survival rate in ratsthat underwent IIR in the presence of CP experiments wereperformed on a total of 12 animals in IIR + CP and IIR + CP+ intralipid groups In all the experimental groups the ratswere anesthetized by intraperitoneal injection of 10 chloralhydrate (35mLkg) after fasting for 16 h The Compound4880 (Sigma USA 075mgkg) or the same volume ofphysiological saline was intravenously injected via the tailvein at 5min before reperfusion The doses of agents wereadjusted in accordance with our previous study [15] Duringthe surgery all the rat body temperature was maintained at38∘C using heated pad And 10mLkg 37∘C normal salinewas injected subcutaneously to avoid dehydration after theabdomen had been closed

28 Collection of Intestinal Mucosa Upon the completion ofthe abovementioned treatments and experiments the ratswere anesthetized and then euthanized The whole smallintestine was removed carefully and a segment of 10 cmintestine (located at 10 cm away from the terminal ileum)was cut and fixed in 10 formaldehyde and then embeddedin paraffin for section The remaining small intestine waswashed thoroughly with 0∘C normal saline and then openedlongitudinally to expose the intestinal epithelium after beingrinsed completely with 0∘C normal saline and dried withsuction paper The mucosal layer was harvested by gentlescraping of the epitheliumwith a glass slidewith a plate on theice and then was stored at minus70∘C for further measurements

29 Intestinal Histology Five 120583m thick sections were pre-pared from paraffin-embedded intestine tissue the segmentof small intestine was stained with hematoxylin-eosin Andthe damage of intestinal mucosa was evaluated by two histol-ogists blinded to the experiment according to Chiursquos standard[32] Grade 0 normal mucosa Grade 1 development ofsubepithelial Gruenhagenrsquos space at the tip of villus Grade 2extension of the space with moderate epithelial lifting Grade3 massive epithelial lifting with a few denuded villi Grade 4denuded villi with exposed capillariesGrade 5 disintegrationof the lamina propria ulceration and hemorrhage

210 Detection of 15-F2119905-Isoprostane Content in the Small

Intestinal Mucosa Small intestinal mucosa was homoge-nized with normal saline The tissue content of free 15-F

2t-isoprostane an index of in vivo oxidative stress-induced lipidperoxidation was measured using commercial immunoassaykits (CaymanChemical AnnArborMI) aswedescribed [33]

211 Assay of Superoxide Dismutase (SOD) Activity andHydrogen Peroxide Content in Small Intestinal Mucosa Smallintestinal mucosa was made into a homogenate with nor-mal saline frozen at minus20∘C for 5min and centrifuged for15min at 4000 rmin Supernatants were transferred intofresh tubes for evaluation of SOD activity and hydrogen


peroxide content SOD activity and hydrogen peroxide con-tent were assessed by SOD and hydrogen peroxide detectionkits according to the manufacturerrsquos instructions (JianchengBioengineering Ltd Nanjing China) Presented data werenormalized to tissue weight

212 Measurement of 120573-Hexosaminidase Level in Serum 120573-hexosaminidase level in the serum was determined usingmodification of a previously described method [34] Briefly50120583L of serum was incubated with 50 120583L of 1mM p-nitrophenyl-N-acetyl-120573-D-glucosaminide dissolved in 01Mcitrate buffer (pH 5) in a 96-well plate at 37∘C for 1 h Thereaction was terminated with 200 120583Lwell of 01M carbonatebuffer (pH 105) The absorbance at 405 nm was measuredusing a Microplate reader

213 Western Blotting Intestinal mucosa samples werehomogenized with lysis buffer for 30 seconds in a mortar andpestle with liquid nitrogen Homogenates were centrifuged at13000 rpm for 10min at 4∘Cand the supernatantwas collectedas the source of protein sample The proteins were processedwith standardmethods forWestern blot analysis as described[10] Rat monoclonal anti-tryptase antibody rat monoclonalanti-gp91phox and anti-p47phox antibodies rat monoclonalanit-P-selectin and anti-ICAM-1 antibodies and 120572-tubulinantibody were obtained from Santa Cruz (Santa Cruz CAUSA) The secondary antibody conjugated to horseradishperoxidase was diluted at 1 2000 (Santa Cruz USA)Immunoblots were incubated with an enhanced chemilumi-nescence detection system (KeyGen Biotech China) and thedensitometry analysis was performed using Quantity Onesoftware

214 Determination of IL-6 Production in Small IntestinalMucosa by Enzyme Immunoassay Intestinal mucosa tissueswere made into homogenates with frozen normal saline andspun at 4000 rmin for 15 minutes Supernatants were thentransferred into fresh tubes for detection of IL-6 contentsBriefly intestinal protein was measured by BCA ProteinAssay Kit provided by KenGen Biotech Company NanjingChina and the protein contents were expressed as gL Thelevels of IL-6 were measured using commercial ELISA kitsfollowing manufacturerrsquos instructions (RampD systems IncUSA) The absorbance was read at 450 nm by a Biokineticsmicroplate reader Model EL340 (Biotek Instruments USA)and the results were expressed as ngL then the final levels ofIL-6 in the intestine were calculated as ngg protein

215 Determination of Myeloperoxidase (MPO) Activity inSmall Intestinal Mucosa Myeloperoxidase (MPO) activitywas determined with the O-dianisidine method [35] usinga MPO detection kit (Nanjing Jiancheng BioengineeringInstitute) as we described [36] MPO activity was definedas the quantity of enzyme degrading 1120583mol of peroxide perminute at 37∘C and was expressed in units per gram weightof wet tissue

216 Statistical Analysis The data (except for the survivalrates) were expressed as Mean plusmn SEM All biochemicalassays were performed in duplicate Analysis of variance wasperformedusingGraphpadPrism softwareOne-way analysisof variance was used for multiple comparisons followed byBonferronirsquos Studentrsquos 119905-test for unpaired values The survivalrate was expressed as the percentage of live animals and theMantel Cox log rank test was used to determine differencesbetween groups Differences were considered significantwhen 119875 values were less than 005

3 Results

31 Oxidative Stress but Not LPS-Induced RBL-2H3 CellDegranulation Can Be Reversed by NAC and Propofol Pre-treatments Bacterial translocation and oxidative stress arethe two characterizations of small IIR [37] and our previousstudy showed that mast cell (MC) degranulation couldexacerbate the injury [14] Therefore in the current studywe initially sought to explore whether bacteria and oxidativestress can induce MC degranulation As shown in Figure 1exposure of the RBL-2H3 to different concentrations ofLPS for 1 or 4 h did not cause significant changes in therelease of 120573-hexosaminidase activities as compared with thecontrol group (Figure 1(a)) whereasMC activator Compound4880 significantly increased the released 120573-hexosaminidaseactivities as compared with the control group (Figure 1(b))This result indicates that LPS per se has no effects on MCdegranulation at least in this experimental setting Howevertreatment of RBL-2H3 cell with H

2O2dose-dependently

increased the activity of the released 120573-hexosaminidase(Figure 1(c)) Pretreatments with NAC a scavenger of oxy-gen radicals abolished H

2O2mediated elevations of the

released 120573-hexosaminidase activities (Figure 1(d)) Similarlypretreatment with propofol but not intralipid (propofolsolvent) abrogated the enhancements of the released 120573-hexosaminidase activities induced by H

2O2(Figure 1(e))

indicating that oxidative stress contributes to MC degran-ulation and propofol inhibits MC degranulation via itsantioxidant property

32 TryptaseContributed to Small Intestinal Epithelium InjuryTryptase the unique mediator released from MC degranu-lation [38] plays a critical role in the IIR mediated acutelung injury in vivo [15] Therefore we sought to define thedirect role of tryptase in the small intestinal epithelium injuryin vitro A small intestinal epithelial cell line IEC-6 wasemployed and exposed to different concentrations of tryptaseThe results showed that tryptase could directly cause IEC-6injury in a dose-dependent manner manifested as dramaticincreases in LDH activities and concomitant reductions incell viability as compared with the control group (Figures 1(f)and 1(g))

33 NAC Propofol and Intralipid PretreatmentsDidNotAffectSmall Intestinal Structure before IIR First we sought to assessif pretreatment of rats with NAC propofol or intralipid maycause significant intestinal injury At the dosages used neither


0

5

10

15

20120573

-Hex

osam

inid

ase r

eleas

ed (

)

Control LPS10ngmL

LPS100ngmL

LPS1000ngmL

1h4h

(a)

0

20

40

60

80

Compound 4880

120573-H

exos

amin

idas

e rele

ased

()

Control10120583gmL100120583gmL

lowast

lowast

(b)

0

10

20

30

120573-H

exos

amin

idas

e rele

ased

()

Control

H2O2

5120583M50120583M500120583M

lowast

lowast

(c)

0

10

20

30

40(

)

H2O2 500120583MNAC 10mM

NAC 20mM

minus

minus

minus

minus minus

minus minus

+ + +

+

+

lowast

(d)

0

10

20

30

40

()

lowast

minusminusminusminus

minusminus minus minus

minusminus minusminus minus

+ ++

+

+

+

+

lowast

lowast

lowast$

H2O2 500120583MPropofol 5120583gmL

Propofol 05 120583gmLIntralipid 05 120583gmL

(e)

0

20

40

60

80

100

Tryptase

Control

10ngmL1ngmL

100ngmL1000ngmL

lowast

lowast

Cel

l via

bilit

y (

)

lowast

(f)

Figure 1 Continued


0

200

400

600

Tryptase

Control

10ngmL1ngmL

100ngmL1000ngmL

lowastlowast lowast

lowast

LDH

(UL

)

(g)

Figure 1 Effects of LPS Compound 4880 and hydrogen peroxide (H2O2) on the degranulation of RBL-2H3 cells with or without NAC or

propofol treatment (a) 120573-Hexosaminidase release after LPS treatment for 1 h or 4 h (b) 120573-hexosaminidase release after Compound 4880stimulation for 1 h and (c) 120573-hexosaminidase release after H

2O2treatment for 1 h (d) and (e) quantified the degranulation of RBL-2H3 cells

pretreated with NAC or propofol for 12 h induced by H2O2stimulation for 1 h respectively (f) Cell viability of IEC-6 cells treated with

tryptase for 12 h Results were expressed as percentage of control group error bars represented the standard error of the mean (g) Lactatedehydrogenase (LDH) activity of IEC-6 cells treated with tryptase for 2 h Results were expressed as Mean plusmn SEM lowast119875 lt 005 versus controlgroup 119875 lt 005 versus H2O2 500 120583M group $119875 lt 005 versus propofol 05120583gmL group

NACnor propofol or intralipid caused any challenges to smallintestinal structure as compared to that in rats treated withnormal saline (NS) As shown in Figure 2 all the layers ofintestine were normal in all the groups and there was nosignificant difference among groups in terms of injury score(Figure 2(d))

34 NAC and Propofol but Not Intralipid Pretreatment AlteredIntestinal Oxidant and Antioxidant Levels before IIR NACandpropofol arewell known for their antioxidative properties[10 39] As shown in Figure 3 NAC and propofol simi-larly attenuated oxidant level as demonstrated by significantdecreases in 15-F

2t-isoprostane (Figure 3(a)) contents andincreases in SOD activities (Figure 3(b)) in small intestinalmucosa as comparedwithNS treated group In additionNACand propofol pretreatment led to significant reductions inprotein expressions of gp91phox and p47phox the importantcomponents of NADPH enzymes in small intestinal mucosacompared with NS treated group before IIR (Figures 3(c)ndash3(e)) By contrast intralipid showed comparative results toNS treated groupThedata indicated that propofol at the doseused confers similar antioxidant effects to that of NAC

35 NAC Propofol and Intralipid Pretreatments Did Not Acti-vate Mast Cell before IIR Trypatse and 120573-hexosaminidaseare specific markers for the assessment of mast cells degran-ulation [40] As shown in Figures 3(f) and 3(g) NACpropofol or intralipid alone had no significant effect onmast cell degranulationThere were no significant differencesin tryptase protein expressions in small intestinal mucosa

and 120573-hexosaminidase levels among all pretreated groupsbefore IIRThe findings from the current study indicated thatNACandpropofol pretreatments can substantially amelioratesuperoxide productions without affecting mast cell and smallintestine morphology under normal condition

36 NACandPropofol Improved Reduction of Survival Rates inRats Challenged to IIR through InhibitingMast Cell ActivationWe next sought to investigate the effects of NAC and propo-fol in combating IIR and inhibiting mast cell in the processof IIR injury As illustrated in Figure 4(a) activation of mastcell by Compound 4880 resulted in significant decreases in2 h survival rates after the clamp releasing as compared withsole IIR group By contrast NAC and propofol pretreatmentsshowed similar promising benefits in reducing mast celldegranulation mediated injury during 2 h reperfusion periodevidenced as increased postischemic 2 h survival rates to100 (Figure 4(b)) Intralipid did not attenuate Compound4880 mediated exacerbation of postischemic survival ratein rats subjected to IIR (Figure 4(c)) These data suggestedthat oxidative stress is a major mechanism whereby mast celldegranulation exacerbated IIR injury

37 NAC and Propofol Attenuated Small Intestinal Injury inRats Undergoing IIR through Inhibiting Mast Cell ActivationAfter 2 h reperfusion the sections of small intestine wereevaluated by HE staining IIR induced severe damage tosmall intestine As depicted in Figures 4(d) and 4(e) multipleerosions and bleeding were observed in IIR group whileCompound 4880 further aggravated IIR injury manifested


(a) (b)

(c) (d)

00

02

04

NS NAC Pro Lip

Chiu

rsquos sc

ores

(e)

Figure 2 Morphological changes of intestine under light microscope in rats treated with NAC propofol or intralipid ((a)ndash(d))Representative images of rats treated with normal saline (NS) NAC (05 gkg ip) propofol (50mgkg ip) and intralipid (50mgkg ip)prior to ischemia reperfusion (HE staining times200) (e) quantified the intestine histological scores in normal saline pretreated group (NS)NAC pretreated group (NAC) propofol pretreated group (Pro) and intralipid pretreated group (Lip) Results are expressed as Mean plusmn SEM119899 = 3 per group

as moremultiple erosions and bleedings andmore inflamma-tory cell sequestrations seen in the IIR + CP group whereasthe villus and glands were normal and no inflammatorycell infiltration was observed in mucosal epithelial layerin Sham-operated group NAC and propofol similarly andsignificantly reduced the injuries in small intestine and onlyslight edema of mucosa villus and infiltration of few necroticepithelial inflammatory cells were found in mucosa epitheliallayer Moreover NAC and propofol pretreatments blockedCompound 4880 induced exacerbation in small intestinal

morphology changes after 2 h reperfusion In contrast pre-treatment with intralipid could not attenuate IIR and Com-pound 4880 induced injury Consistent with morphologicalchanges Chiursquos scores markedly increased in IIR group ascompared with Sham-operated group while treatment withmast cell degranulator Compound 4880 after ischemia led tofurther increases inChiursquos scoresNACandpropofol similarlydramatically lowered Chiursquos scores and significantly limitedthe changes induced by Compound 4880 (119875 lt 005 NAC +IIR + CP or propofol + IIR + CP versus IIR + CP group) Of


0

200

400

600

800

1000

NS NAC Pro Lip

lowast lowast15

-F2t

-isop

rost

ane (

pgm

g)

(a)

0

50

100

150

SOD

activ

ities

(uni

tsm

g)

NS NAC Pro Lip

lowast

lowast

(b)

NS NAC Propofol Intralipid

p47phox

gp91phox

120572-Tubulin

(c)

00

05

10

15

NS NAC Pro Lip

p47ph

oxex

pres

sions

(au

)

lowast lowast

(d)

00

05

10

15

NS NAC Pro Lip

gp91

phox

expr

essio

ns (a

u)

lowast lowast

(e)

00

05

10

15

Tryptase

Tryp

tase

expr

essio

ns (a

u)

NS NAC Pro Lip

120572-Tubulin

(f)

00

05

10

15

NS NAC Pro Lip

120573-H

exos

amin

idas

e (nM

)

(g)

Figure 3 Changes of 15-F2t-isoprostane contents SOD activities p47phox and gp91phox protein expressions in intestine mucosa and intestinal

tryptase protein expression and serum 120573-hexosaminidase levels in rats treated with NAC propofol and intralipid (a) 15-F2t-isoprostane

contents (b) SOD activities ((c)ndash(e)) intestinal p47phox and gp91phox protein expressions (f) tryptase protein expression and (g) 120573-hexosaminidase levels in normal saline pretreated group (NS) NAC pretreated group (NAC) propofol pretreated group (Pro) and intralipidpretreated group (Lip) (119899 = 3) Results are expressed as Mean plusmn SEM lowast119875 lt 005 versus NS


0 20 40 60 80 100 1200

20

40

60

80

100

IIRIIR + CP

P lt 00001

(min)

Surv

ival

rate

s (

)(a)

0 20 40 60 80 100 120(min)

0

20

40

60

80

100

P lt 00001

NAC + IIR + CPPro + IIR + CP

IIR + CPSurv

ival

rate

s (

)

(b)

0 20 40 60 80 100 120(min)

0

20

40

60

80

100

P lt 00001

P =

05

005

P =

00

384IIR

Lip + IIR

Lip + IIR + CP

IIR + CPSu

rviv

al ra

tes (

)

(c)

SHAM IIR

IIR + CP

IIR

IIR + CP

IIR

IIR + CP

IIR

IIR + CP

NAC Propofol Intralipid

(d)

0

2

4

6

Propofol

Intralipid

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

lowast lowast

Chiu

rsquos sc

ores NAC

lowast$ lowast$

lowast998771lowast

lowastlowast

(e)

Figure 4 Survival rates and morphological changes of intestine and intestinal histological score under light microscope after IIR SHAMgroup (Sham-operated group) IIR group (75min intestinal ischemia and 2 h reperfusion) and IIR + CP group (IIR group + Compound4880 1mgkg) in the absence or presence of NAC (05 gkg) or propofol (50mgkg) or intralipid (50mgkg) ((a)ndash(c)) Survival rats in ratssubjected to IIR with or without NAC propofol or intralipid pretreatment Results are expressed as percentage of live animals 119899 = 6 pergroup whereas 119899 = 12 in IIR + CP group and IIR + CP + intralipid group (d) Representative images of HE staining (times200) Black arrowlamina propria villus shedding Red arrow engorgement of capillary vessel White arrow top villus shedding (e) quantified the intestinehistological scores Results are expressed as Mean plusmn SEM 119899 = 6 per group whereas 119899 = 4 in IIR + CP group lowast119875 lt 005 versus SHAM group119875 lt 005 versus IIR group $119875 lt 005 versus IIR + CP group amp119875 lt 005 versus IIR with NAC pretreated group 119875 lt 005 versus IIR withpropofol pretreated group and 998771119875 lt 005 versus IIR with intralipid pretreated group


note there were no differences in Chiursquos scores between IIRand intralipid pretreated groups

38 NAC and Propofol Reduced Oxidative Stress in SmallIntestine in Rats Undergoing IIR through Inhibiting Mast CellActivation Ischemia reperfusion injury is characterized byupregulation of ROS [10] In linewith previous results [16] weobserved that 75min ischemia followed by 2 h reperfusion ledto substantial enhancements in 15-F

2t-isoprostane contentsin small intestinal mucosa and marked reductions in SODactivities as compared with Sham-operated group (Figures5(a) and 5(b)) Meanwhile IIR also caused great increases ingp91phox and p47phox protein expressionwhen comparedwithSham-operated group Moreover Compound 4880 furtheraggravated the changes of oxidative stress induced by IIR(119875 lt 005 IIR + CP versus IIR) Not surprisingly propofoland NAC similarly attenuated IIR mediated oxidative stressevidenced as downregulation of 15-F

2t-isoprostane contentsand gp91phox and p47phox protein expression and upregula-tion of SOD activities in propofol and NAC treated groups ascompared with group IIR Furthermore propofol and NACpretreatments also limited the further increases of oxidativestress induced by Compound 4880 As expected intralipiddid not show any antioxidative properties as the markers ofoxidative stress measured were comparable between groupIIR and group intralipid + IIR Intralipid had no protectiveeffect against Compound 4880 induced upregulation ofoxidative stress

39 NAC and Propofol Attenuated Inflammation in SmallIntestine in Rats Undergoing IIR through Inhibiting Mast CellActivation Interleukin 6 (IL-6) is one of the inflammatorycytokines that has been demonstrated to be implicated inthe pathogenesis of IIR injury [41] In the present studywe observed that IL-6 levels in small intestinal mucosain group IIR were significantly increased as compared tothat in the Sham-operated group (Figure 5(e)) Moreoveradministration with Compound 4880 further resulted indramatic enhancements in IL-6 levels in group IIR + CP(119875 lt 005 versus group IIR) Pretreatments with propofol andNAC not only similarly abrogated the increases in IL-6 levelsbut also block the enhancements in IL-6 levels resulting fromIIR in the presence of the MC activator Compound 4880(Figure 5(e))

310 NAC and Propofol Inhibited Neutrophil Rolling in SmallIntestine in Rats Undergoing IIR through Inhibiting Mast CellActivation IIR injury is also characterized by neutrophilsequestrating into the inflamed tissues [42] Consistent withprevious results [42] we also found that IIR led to substan-tial increases in MPO activities (Figure 5(f)) and ICAM-1(Figure 5(g)) and P-selectin (Figure 5(h)) protein expressionsas compared with Sham-operated group moreover Com-pound 4880 resulted in further increases in MPO activitiesand P-selectin protein expression in group IIR + CP than thatin group IIR Administrations with propofol andNAC signif-icantly inhibited neutrophil infiltrationactivation evidencedas downregulation of MPO activities and P-selectin protein

expression caused by IIR Furthermore propofol and NACalso blocked Compound 4880 mediated exacerbation of IIRinduced neutrophil infiltration

311 NAC and Propofol Reduced Mast Cell Degranulation inSmall Intestine in Rats Undergoing IIR Mast cell releases anumber ofmediators that contribute to the aggravation of IIRinjury and there are many factors that can induce mast celldegranulation during IIR Tryptase and 120573-hexosaminidaseare the unique markers released from mast cell and theirincreased release can be recognized as mast cell degranula-tion As shown in Figure 6 after 2 h reperfusion we foundthat tryptase protein expression and 120573-hexosaminidase levelwere greatly increased in group IIR as compared with Sham-operated group and Compound 4880 resulted in furthermast cell degranulation evidenced as dramatic increasesin tryptase protein expression and 120573-hexosaminidase levelobserved in group IIR + CP relative to that in group IIROf note propofol and NAC similarly inhibited mast celldegranulation by downregulating tryptase protein expressionand 120573-hexosaminidase level triggered by IIR and Compound4880 Collectively the findings from the current study indi-cated that oxidative stress mediated mast cell degranulationaggravated IIR injury and propofol through its antioxidativeproperties protects against IIR by stabilizing mast cell

4 Discussion

We have shown in the current study that increased ROSduring IIR exacerbated IIR injury primarily via activatingmast cells evidenced as concomitant increases of postis-chemic 15-F

2t-isoprostane and elevations in tryptase and 120573-hexosaminidase and increases in intestinal injury score lead-ing to significantly reduced postischemic survival Furtherwe showed that activation of intestinal NADPH oxidase is amajor source of postischemic ROS production AntioxidantNAC and propofol inhibited postischemic intestinal NADPHoxidase activation by reducing gp91phox and p47phox proteinoverexpression and reduced ROS and the subsequent mastcell activation in vivo and in vitro which may represent themajor mechanism whereby propofol attenuates IIR injuryand enhances postischemic survival

Mast cell (MC) which contains a diverse range of medi-ators resides throughout gastrointestinal tract and is alsoknown as intestinal mucosal mast cell (IMMC) Despite thefact thatMCmay function as host defense to prevent bacterialinvasion [43] enhancedMC activation may contribute to thedevelopment of a variety of disorders including IIR injuryby releasing histamine tryptase TNF-120572 and other factors Itshould be noted that MC is the main source of TNF-120572 in thetract [44] Ramos et al reported that mast cell degranulationplays a significant role in the development of sepsis by reg-ulating cell death which resulted in multiorgan dysfunction[45] Consistent with the previous studies showing that stabi-lizing MC from degranulation would be one of the potentialstrategies in combating IIR injury [14] our current studyfurther revealed that increased oxidative stress during IIRplays an important role in mast cell activationdegranulation


0

1000

2000

3000

4000

NAC Propofol

Intralipid

15-F

2t-is

opro

stan

e (pg

mg)

lowast lowast

$ $

lowast lowast998771

(a)

NAC Propofol

Intralipid

0

50

100

150

200

lowast lowast

lowast

SOD

activ

ities

(uni

tsm

g)

$

$

lowast998771

(b)

NAC Propofol

Intralipid

00

05

10

15

20

25

lowast lowast$ $

p47phox

120572-Tubulin

p47ph

oxex

pres

sions

(au

) lowast lowast998771

(c)

NAC Propofol

Intralipid

00

05

10

15

20

25

lowast lowast

$ $

gp91phox

120572-Tubulin

gp91

phox

expr

essio

ns (a

u) lowast lowast998771

(d)

NAC Propofol

Intralipid

0

5000

10000

15000

lowast lowast

lowast$ lowast$

IL-6

leve

ls (p

gm

g)

lowastlowast998771

(e)

NAC Propofol

Intralipid

0

10

20

30

40

lowast lowast

lowastlowast lowast$lowast$

MPO

activ

ities

(uni

tsg)

lowast lowast998771

(f)

$ $lowast lowastNAC Propofol

Intralipid

00

05

10

15

20

25

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

120572-Tubulin

ICAM-1

ICA

M-1

expr

essio

ns (a

u)

lowast lowast998771

(g)

$$

lowastlowast

lowast

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

NAC Propofol

Intralipid

00

05

10

15

20

25

120572-Tubulin

P-selectin

P-se

lect

in ex

pres

sions

(au

)

lowast998771

(h)

Figure 5 Changes of SOD activities 15-F2t-isoprostane contents p47phox protein expression gp91phox P-selectin and ICAM-1 protein

expressions IL-6 levels and MPO activities in intestine mucosa after IIR injury SHAM group (Sham-operated group) IIR group (75minintestinal ischemia and 2 h reperfusion) and IIR + CP group (IIR group + Compound 4880 1mgkg) in the absence or presence of NAC(05 gkg) propofol (50mgkg) intralipid (50mgkg) (a) SOD activities (b) 15-F

2t-isoprostane contents in intestine (119899 = 6 except 119899 = 4 inIIR + CP group) ((c) and (d)) p47phox and gp91phox protein expressions respectively (119899 = 3) ((e) and (f)) IL-6 levels and MPO activities inintestinal mucosa (119899 = 6 except 119899 = 4 in IIR + CP group) ((g) and (h)) ICAM-1 and P-selectin protein expressions respectively (119899 = 3)Results are expressed as Mean plusmn SEM lowast119875 lt 005 versus SHAM group

119875 lt 005 versus IIR group $119875 lt 005 versus IIR + CP group

amp119875 lt 005 versus IIR with NAC pretreated group 119875 lt 005 versus IIR with propofol pretreated group and 998771119875 lt 005 versus IIR with

intralipid pretreated group


NAC Propofol

Intralipidlowast

lowast998771

120572-Tubulin

lowast lowast

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

3

2

1

0

Tryptase120573

-Hex

osam

inid

ase (

nM)

lowast$$

(a)

NAC Propofol

Intralipid

lowast lowast998771

lowastlowast

3

2

1

0

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

Tryp

tase

expr

essio

ns (a

u)

$$

(b)

Figure 6 Changes of intestinal tryptase protein expression and serum120573-hexosaminidase levels after IIR injury SHAMgroup (Sham-operatedgroup) IIR group (75min intestinal ischemia and 2 h reperfusion) and IIR + CP group (IIR group + Compound 4880 1mgkg) in theabsence or presence of NAC (05 gkg) propofol (50mgkg) intralipid (50mgkg) (a) Intestinal tryptase protein expression (119899 = 3) (b)120573-hexosaminidase levels in serum (119899 = 6 except 119899 = 4 in IIR + CP group) Results are expressed as Mean plusmn SEM lowast119875 lt 005 versus SHAMgroup 119875 lt 005 versus IIR group $119875 lt 005 versus IIR + CP group amp119875 lt 005 versus IIR with NAC pretreated group 119875 lt 005 versus IIRwith propofol pretreated group and 998771119875 lt 005 versus IIR with intralipid pretreated group

evidenced as enhancements in tryptase protein expressionand 120573-hexosaminidase levels which causes or exacerbatesIIR injury This notion is further supported by the fact thatactivating mast cell by Compound 4880 further aggravatedIIR injury and tryptase directly damaged small intestinalepithelium evidenced by the reduced cell viability and theenhanced LDH activity in vitro The data from the currentstudy further proved that mast cell activation plays a centralrole in exacerbating IIR injury although the mechanismgoverningmast cell activation during IIR is yet to be explored

Mast cell can be activated by several distinct mecha-nisms in addition to IgEFc120576 the classical signal pathwayof activation there is a growing body of evidence to sug-gest that nonimmunological stimuli such as trauma andphysical stress also contribute to mast cell activation [46]Yoshimaru and colleagues demonstrated that silver mediatedmast cell activation by upregulation of NADPH oxidasemediated ROS production [18] Additionally Collaco et alfound that superoxide production contributes to mast celldegranulation and inhibiting ROS can stabilize mast cell invitro [12] Moreover our previous study in a rat model ofIIR revealed significant increase of mast cell activation withconcomitant elevations in ROS [16] These results providedus sound rationale to speculate that there exists a linkagebetween mast cell activation and superoxide productionOur current study extended findings of previous studies[16] by showing that NADPH oxidase is overexpressedduring IIR which contributed to increased oxidative stressand MC activation leading to exacerbation of IIR injuryFurther using antioxidant N-acetylcysteine (NAC) whichhas been shown to prevent myocardial NADPH oxidaseoverexpression in diabetes [47 48] and attenuate myocardial

ischemia reperfusion injury [10] we found that in in vivomodel of IIR pretreatment of rats with NAC not onlyattenuatedNADPHoxidase overexpression and reducedROSproduction but concomitantly reduced MC activation andconsequently attenuated IIR injury and increased survivalrate We also found that NAC reversed the oxidative stressmediated mast cell degranulation in vitro The results fromthe current study indicated that oxidative stress subsequentto NADPH oxidase overexpression during IIR plays a criticalrole in mast cell activation which in turn led to a deleteriousinjury Of note in the current study mast cell activation byCP also induced oxidative stress manifested as increased 15-F2t-isoprostane and reduced SOD associated with increased

p47phox and gp91phox protein expression in IIR + CP groupcompared to IIR group (Figures 5(a)ndash5(d))This is consistentwith study from Collaco et al showing that sodium sulfiteactivated mast cell degranulation and subsequently increasedintercellular oxidative stress in RBL-2H3 cells [12] Theseindicate that mast cell activation per se induces oxidativestress

Excessive production of ROS by NADPH oxidase isgenerally considered to be involved in the pathogenesis ofinflamed tissues [49] including ischemic tissues There area number of NADPH homologs presented in many diverseorgans such as Nox1 Nox2 (also named as gp91phox) andNox3-4 [50] It is important to appreciate that Nox2 is pre-dominantly expressed within epithelial cells In addition toROS scavenger NAC also inhibits NADPH oxidase activitiesour previous results showed that pretreatment of diabeticrats withNAC attenuated cardiac ischemia reperfusion injuryby downregulating NADPH subunits p67phox and p22phoxexpression and reducing oxidative stress [48] Inoue et al


have demonstrated that blockade of ROS generation byNADPH oxidase inhibition can limit mast cell degranulation[51] Guan et al demonstrated that intracellular NADPHconcentration in villus tip cells in intestine was significantlyrapidly increased even after short-term ischemia [52] Inthe current study we also found that elevations of gp91phox

and p47phox protein expression are the prominent features ofIIR Furthermore precondition of IIR-rats with antioxidantNAC or propofol not only significantly reduced gp91phox andp47phox protein expression but also played a significant rolein stabilizing mast cell

Propofol is a short-acting intravenously administeredhypnotic agent In addition to its sedationhypnotic prop-erties propofol displays protective effects in many organssubjected to ischemia reperfusion injury by inhibiting oxida-tive cellular damage [53 54] Hama-Tomioka et al recentlyreported that propofol can block superoxide productionoriginated from NADPH oxidase in vitro [55] Although asdescribed previously a single intraperitoneal injection of 50or 100mgkg propofol could significantly attenuate IIR injuryin acute rat models [31] propofol is the most commonlyused agent overall during short-term and intermediate-length sedation in ICU [56] thus in the current study weexplored propofol pretreatment for three successional daysand incorporated the investigation of postischemic mortalityin addition to observing its effect on IIR injury To ourknowledge our study is the first to show that propofolpretreatment given at a sedative dosage significantly inhibitedthe overexpression of NADPH oxidase and reduced oxidativemediated MC action during IIR or in vitro and reduced IIRinjury and subsequently enhanced postischemic mortality inrats

IIR injury is also characterized by uncontrollable inflam-mation and neutrophils sequestration in inflamed tissuesMigration of neutrophils to injurious site is mediated byselectins and intercellular adhesion molecules (ICAMs) bythe activated endothelium [57] Compton et al reviewed thattryptase released from mast cell degranulation can attractleukocytes infiltrate and migrate to ischemic tissues [58]And treatment of rats with mast cell stabilizer cromolynsodium has been shown to greatly reduce the expressions ofICAM-1 in the lungs in pancreatitis-associated lung injuryand decreased IL-6 release [59] These findings point to theimportance of mast cell degranulation through increasedrelease of proinflammatory mediator in inducing neutrophilmigration to inflamed tissues In the present study we foundthat during IIR mast cell degranulation resulted in moreneutrophils infiltration into small intestine evidenced assignificant increases in MPO activities and ICAM-1 and P-selectin protein expressions and that propofol and NAC sim-ilarly blocked the alterations induced by IIR and preventedMC stimulator Compound 4880 mediated exacerbation ofIIR injury Our finding that propofol can inhibit intestinalMC activation during IIR and reduce tissue neutrophilsinfiltration as well as the subsequent systemic inflammationmay have potential clinical importance given that IIR occursoften in patient who underwent major surgeries such ascardiac surgery with cardiopulmonary bypass [60] and is

a challenging and life-threatening clinical problemThe delayin diagnosis and treatment of IIR injury contributes to thecontinued high in-hospital mortality rate [61] Preventiveadministration of propofol may be a promising approach inattenuating postoperative intestinal ischemia and mortality

In a summary we have shown that mast cell activationthrough increased release of mediators contributed to dele-terious injury induced by IIR and that oxidative stress playsa central role in mast cell activation Propofol similar to theantioxidant NAC inhibited ROS mediated mast cell activa-tion and attenuated IIR injury and enhanced postischemicmortality Propofol mediated reduction of oxidative stressand MC activation during IIR is achieved at least in partthrough attenuating IIR induced NADPH oxidase overex-pression while the underlying mechanism merits furtherstudy Mast cell activation contributes to and exacerbatessmall intestinal ischemia reperfusion injury

Conflict of Interests

The authors declare that there is no conflict of interests

Authorsrsquo Contribution

Xiaoliang Gan Dandan Xing Guangjie Su Shun Li Chen-fang Luo and Michael Irwin performed the experiment andanalyzed the data XiaoliangGanwrote the paper ZhengyuanXia Haobo Li and Ziqing Hei conceived and designed thestudy and revised the paper Haobo Li and Ziqing Hei sharesenior authorship Dr Ziqing Hei takes full responsibility forthework as awhole including the study design access to dataand the decision to submit and publish the paper XiaoliangGan and Dandan Xing contributed equally to this study

Acknowledgments

This study was supported by National Natural Science Foun-dation of China (NSFC 30901408) and was partly supportedby the Fundamental Research Funds for the Central Univer-sities of China (12ykpy37)

References

[1] Q LuD Z Xu S Sharpe et al ldquoThe anatomic sites of disruptionof the mucus layer directly correlate with areas of traumahemorrhagic shock-induced gut injuryrdquo Journal of Trauma vol70 no 3 pp 630ndash635 2011

[2] O Corcos Y Castier A Sibert et al ldquoEffects of a multimodalmanagement strategy for acute mesenteric ischemia on srvivaland intestinal failurerdquoClinical Gastroenterology andHepatologyvol 11 no 2 pp 158e2ndash165e2 2013

[3] S A W G Dello K W Reisinger R M van Dam et al ldquoTotalintermittent pringlemaneuver during liver resection can induceintestinal epithelial cell damage and endotoxemiardquo PLoS ONEvol 7 no 1 Article ID e30539 2012

[4] A Siniscalchi A Cucchetti Z Miklosova et al ldquoPost-reper-fusion syndrome during isolated intestinal transplantationoutcome and predictorsrdquo Clinical Transplantation vol 26 no3 pp 454ndash460 2012


[5] P R Bertoletto A T Ikejiri F Somaio Neto et al ldquoOxidativestress gene expression profile in inbred mouse after ischemiareperfusion small bowel injuryrdquo Acta Cirurgica Brasileira vol27 no 11 pp 773ndash782 2012

[6] T Otamiri and R Sjodahl ldquoOxygen radicals their role inselected gastrointestinal disordersrdquoDigestiveDiseases vol 9 no3 pp 133ndash141 1991

[7] Q-G Zhang M D Laird D Han et al ldquoCritical role of nadphoxidase in neuronal oxidative damage and microglia activationfollowing traumatic brain injuryrdquo PLoS ONE vol 7 no 4Article ID e34504 2012

[8] J Shen X-Y Bai Y Qin et al ldquoInterrupted reperfusion reducesthe activation of NADPH oxidase after cerebral IR injuryrdquo FreeRadical Biology andMedicine vol 50 no 12 pp 1780ndash1785 2011

[9] G T Gang J H Hwang Y H Kim et al ldquoProtec-tion of NAD(P)H quinone oxidoreductase 1 against renalischemiareperfusion injury in micerdquo Free Radical Biology andMedicine vol 67 pp 139ndash149 2014

[10] T Wang S Qiao S Lei et al ldquoN-acetylcysteine and allopurinolsynergistically enhance cardiac adiponectin content and reducemyocardial reperfusion injury in diabetic ratsrdquo PLoS ONE vol6 no 8 Article ID e23967 2011

[11] T Wang X Mao H Li et al ldquoN-Acetylcysteine and allopuri-nol up-regulated the JakSTAT3 and PI3KAkt pathways viaadiponectin and attenuated myocardial postischemic injury indiabetesrdquo Free Radical Biology and Medicine vol 63 pp 291ndash303 2013

[12] C R Collaco D J Hochman R M Goldblum and E GBrooks ldquoEffect of sodium sulfite onmast cell degranulation andoxidant stressrdquo Annals of Allergy Asthma and Immunology vol96 no 4 pp 550ndash556 2006

[13] B Y de Winter R M van den Wijngaard and W J de JongeldquoIntestinal mast cells in gut inflammation and motility disturb-ancesrdquo Biochimica et Biophysica Acta vol 1822 no 1 pp 66ndash732012

[14] H Zi-Qing G Xiao-Liang H Pin-Jie W Jing S Ning andG Wan-Ling ldquoInfluence of Ketotifen Cromolyn Sodium andCompound 4880 on the survival rates after intestinal ischemiareperfusion injury in ratsrdquo BMCGastroenterology vol 8 article42 2008

[15] X Gan D Liu P Huang W Gao X Chen and Z Hei ldquoMast-cell-releasing tryptase triggers acute lung injury induced bysmall intestinal ischemia-reperfusion by activating PAR-2 inratsrdquo Inflammation vol 35 no 3 pp 1144ndash1153 2012

[16] Z Q Hei X L Gan G J Luo S R Li and J Cai ldquoPretreatmentof cromolyn sodium prior to reperfusion attenuates earlyreperfusion injury after the small intestine ischemia in ratsrdquoWorld Journal of Gastroenterology vol 13 no 38 pp 5139ndash51462007

[17] A T Canada R F Werkman C M Mansbach II and G MRosen ldquoBiochemical changes in the intestine associated withanoxia and reoxygenation in vivo and in vitro studiesrdquo Journalof Free Radicals in Biology andMedicine vol 2 no 5-6 pp 327ndash334 1986

[18] T Yoshimaru Y Suzuki T Inoue O Niide and C Ra ldquoSilveractivates mast cells through reactive oxygen species productionand a thiol-sensitive store-independent Ca2+ influxrdquo Free Rad-ical Biology and Medicine vol 40 no 11 pp 1949ndash1959 2006

[19] Z Xia Z Huang and D M Ansley ldquoLarge-dose propofolduring cardiopulmonary bypass decreases biochemicalmarkersofmyocardial injury in coronary surgery patients a comparison

with isofluranerdquo Anesthesia and Analgesia vol 103 no 3 pp527ndash532 2006

[20] T Fujimoto T Nishiyama and K Hanaoka ldquoInhibitory effectsof intravenous anesthetics on mast cell functionrdquo Anesthesiaand Analgesia vol 101 no 4 pp 1054ndash1059 2005

[21] T Luo J Wu S V Kabadi et al ldquoPropofol limits microglialactivation after experimental brain trauma through inhibi-tion of nicotinamide adenine dinucleotide phosphate oxidaserdquoAnesthesiology vol 119 no 6 pp 1370ndash1388 2013

[22] H Komiyama K Miyake K Asai K Mizuno and T ShimadaldquoCyclical mechanical stretch enhances degranulation and IL-4 secretion in RBL-2H3 mast cellsrdquo Cell Biochemistry andFunction vol 32 no 1 pp 70ndash76 2014

[23] J-I Onose Y Yoshioka Y Q Ye et al ldquoInhibitory effects ofvialinin A and its analog on tumor necrosis factor-120572 release andproduction fromRBL-2H3 cellsrdquoCellular Immunology vol 279no 2 pp 140ndash144 2012

[24] N A Kretzmann E Chiela U Matte N Marroni and C AMarroni ldquoN-acetylcysteine improves antitumoural response ofInterferon alpha byNF-kB downregulation in liver cancer cellsrdquoComparative Hepatology vol 11 no 1 p 4 2012

[25] T Zhu D X Wang W Zhang et al ldquoAndrographolide protectsagainst LPS-Induced Acute Lung Injury by Inactivation of NF-120581Brdquo PLoS ONE vol 8 no 2 Article ID e56407 2013

[26] K Modis A Asimakopoulou C Coletta A Papapetropoulosand C Szabo ldquoOxidative stress suppresses the cellular bioener-getic effect of the 3-mercaptopyruvate sulfurtransferasehydro-gen sulfide pathwayrdquo Biochemical and Biophysical ResearchCommunications vol 433 no 4 pp 401ndash407 2013

[27] X-Y Zhu E Daghini A R Chade et al ldquoMyocardialmicrovas-cular function during acute coronary artery stenosis effectof hypertension and hypercholesterolaemiardquo CardiovascularResearch vol 83 no 2 pp 371ndash380 2009

[28] T Wu X Gan S Zhou M Ge Z Zhang and Z Hei ldquoHis-tamine at low concentrations aggravates rat liver BRL-3A cellinjury induced by hypoxiareoxygenation through histamineH2 receptor in vitrordquo Toxicology In Vitro vol 27 no 1 pp 378ndash386 2013

[29] A M Abdelrahman S Al Salam A S Almahruqi I S AlHusseni M A Mansour and B H Ali ldquoN-acetylcysteineimproves renal hemodynamics in rats with cisplatin-inducednephrotoxicityrdquo Journal of Applied Toxicology vol 30 no 1 pp15ndash21 2010

[30] G Inagawa K Sato T Kikuchi et al ldquoChronic ethanol con-sumption does not affect action of propofol on rat hippocampalacetylcholine release in vivordquo British Journal of Anaesthesia vol93 no 5 pp 737ndash739 2004

[31] K-X Liu T RinneWHe FWang and Z Xia ldquoPropofol atten-uates intestinal mucosa injury induced by intestinal ischemia-reperfusion in the ratrdquo Canadian Journal of Anesthesia vol 54no 5 pp 366ndash374 2007

[32] C J Chiu H J Scott and F N Gurd ldquoIntestinal mucosallesion in low-flow states IIThe protective effect of intraluminalglucose as energy substraterdquo Archives of Surgery vol 101 no 4pp 484ndash488 1970

[33] Z Xia D V Godin and D M Ansley ldquoPropofol enhancesischemic tolerance of middle-aged rat hearts effects on 15-F2t-isoprostane formation and tissue antioxidant capacityrdquoCardiovascular Research vol 59 no 1 pp 113ndash121 2003

[34] H Oda M Fujimoto M S Patrick et al ldquoRhoH plays criticalroles in Fc120576RI-dependent signal transduction in mast cellsrdquoJournal of Immunology vol 182 no 2 pp 957ndash962 2009


[35] J E Krawisz P Sharon and W F Stenson ldquoQuantitative assayfor acute intestinal inflammation based on myeloperoxidaseactivity assessment of inflammation in rat and hamster mod-elsrdquo Gastroenterology vol 87 no 6 pp 1344ndash1350 1984

[36] B Xu X Gao J Xu et al ldquoIschemic postconditioning attenuateslung reperfusion injury and reduces systemic proinflammatorycytokine release via heme oxygenase 1rdquo Journal of SurgicalResearch vol 166 no 2 pp e157ndashe164 2011

[37] M Karabeyoglu B Unal B Bozkurt et al ldquoThe effect ofethyl pyruvate on oxidative stress in intestine and bacterialtranslocation after thermal injuryrdquo Journal of Surgical Researchvol 144 no 1 pp 59ndash63 2008

[38] G Pejler E Ronnberg I Waern and S Wernersson ldquoMastcell proteases multifaceted regulators of inflammatory diseaserdquoBlood vol 115 no 24 pp 4981ndash4990 2010

[39] S Basu A Miclescu H Sharma and L Wiklund ldquoPropo-fol mitigates systemic oxidative injury during experimentalcardiopulmonary cerebral resuscitationrdquo Prostaglandins Leuko-trienes and Essential Fatty Acids vol 84 no 5-6 pp 123ndash1302011

[40] F Schiemann E Brandt R Gross et al ldquoThe cathelicidin LL-37activates humanmast cells and is degraded bymast cell tryptasecounter-regulation by CXCL4rdquo Journal of Immunology vol 183no 4 pp 2223ndash2231 2009

[41] K Yoshiya P H Lapchak T-H Thai et al ldquoDepletion of gutcommensal bacteria attenuates intestinal ischemiareperfusioninjuryrdquo American Journal of Physiology Gastrointestinal andLiver Physiology vol 301 no 6 pp G1020ndashG1030 2011

[42] M Ghyczy C Torday J Kaszaki A Szabo M Czobel andM Boros ldquoOral phosphatidylcholine pretreatment decreasesischemia-reperfusion induced methane generation and theinflammatory response in the small intestinerdquo Shock vol 30 no5 pp 596ndash602 2008

[43] A B Penissi M I Rudolph and R S Piezzi ldquoRole of mast cellsin gastrointestinal mucosal defenserdquo Biocell vol 27 no 2 pp163ndash172 2003

[44] N G Frangogiannis C W Smith and M L Entman ldquoTheinflammatory response in myocardial infarctionrdquo Cardiovascu-lar Research vol 53 no 1 pp 31ndash47 2002

[45] L Ramos G Pena B Cai E A Deitch and L Ulloa ldquoMastcell stabilization improves survival by preventing apoptosis insepsisrdquo Journal of Immunology vol 185 no 1 pp 709ndash716 2010

[46] C Wolber F Baur and J R Fozard ldquoInteraction betweenadenosine and allergen or compound 4880 on lung parenchy-mal strips from actively sensitized Brown Norway ratsrdquo Euro-pean Journal of Pharmacology vol 498 no 1ndash3 pp 227ndash2322004

[47] Y Liu S Lei X Gao et al ldquoPKC120573 inhibition with ruboxistaurinreduces oxidative stress and attenuates left ventricular hyper-trophy and dysfuntion in rats with streptozotocin-induceddiabetesrdquo Clinical Science vol 122 no 4 pp 161ndash173 2012

[48] Z Guo Z Xia J Jiang and J H McNeill ldquoDownregulationof NADPH oxidase antioxidant enzymes and inflammatorymarkers in the heart of streptozotocin-induced diabetic ratsby N-acetyl-L-cysteinerdquo The American Journal of PhysiologymdashHeart and Circulatory Physiology vol 292 no 4 pp H1728ndashH1736 2007

[49] S Lei W Su H Liu et al ldquoNitroglycerine-induced nitratetolerance compromises propofol protection of the endothelialcells against TNF-120572 the role of PKC-120573

2and nadph oxidaserdquo

Oxidative Medicine and Cellular Longevity vol 2013 Article ID678484 9 pages 2013

[50] G R Drummond S Selemidis K K Griendling and C GSobey ldquoCombating oxidative stress in vascular disease NADPHoxidases as therapeutic targetsrdquoNature Reviews DrugDiscoveryvol 10 no 6 pp 453ndash471 2011

[51] T Inoue Y Suzuki T Yoshimaru and C Ra ldquoReactive oxygenspecies produced up- or downstream of calcium influx regulateproinflammatory mediator release from mast cells role ofNADPH oxidase and mitochondriardquo Biochimica et BiophysicaActaMolecular Cell Research vol 1783 no 5 pp 789ndash802 2008

[52] YGuan R TWorrell T A Pritts andMHMontrose ldquoIntesti-nal ischemia-reperfusion injury reversible and irreversibledamage imaged in vivordquo American Journal of Physiology-Gastrointestinal and Liver Physiology vol 297 no 1 pp G187ndashG196 2009

[53] M-C Lee K Kobayashi F Yoshino et al ldquoDirect assess-ments of the antioxidant effects of propofol medium chaintriglyceridelong chain triglyceride on the brain of stroke-pronespontaneously hypertensive rats using electron spin resonancespectroscopyrdquoAnesthesiology vol 109 no 3 pp 426ndash435 2008

[54] W Yao G Luo G Zhu et al ldquoPropofol activation of the nrf2pathway is associated with amelioration of acute lung injuryin a rat liver transplantation modelrdquo Oxidative Medicine andCellular Longevity vol 2014 Article ID 258567 9 pages 2014

[55] K Hama-Tomioka H Kinoshita K Nakahata et al ldquoRoles ofneuronal nitric oxide synthase oxidative stress and propofolin N-methyl-D-aspartate-induced dilatation of cerebral arteri-olesrdquoTheBritish Journal of Anaesthesia vol 108 no 1 pp 21ndash292012

[56] K M Ho and J Y Ng ldquoThe use of propofol for mediumand long-term sedation in critically ill adult patients a meta-analysisrdquo Intensive Care Medicine vol 34 no 11 pp 1969ndash19792008

[57] F Spiller M I L Orrico D C Nascimento et al ldquoHydrogensulfide improves neutrophil migration and survival in sepsisvia K+ATP channel activationrdquoAmerican Journal of Respiratoryand Critical Care Medicine vol 182 no 3 pp 360ndash368 2010

[58] S J Compton J A Cairns S T Holgate and A F WallsldquoHuman mast cell tryptase stimulates the release of an IL-8-dependent neutrophil chemotactic activity from human umbil-ical vein endothelial cells (HUVEC)rdquoClinical and ExperimentalImmunology vol 121 no 1 pp 31ndash36 2000

[59] X Zhao M Dib X Wang B Widegren and R AnderssonldquoInfluence ofmast cells on the expression of adhesionmoleculeson circulating and migrating leukocytes in acute pancreatitis-associated lung injuryrdquo Lung vol 183 no 4 pp 253ndash264 2005

[60] C A Efthymiou and W I Weir ldquoSalmonella sepsis simulatinggastrointestinal ischaemia following cardiopulmonary bypassrdquoInteractive Cardiovascular and Thoracic Surgery vol 12 no 2pp 334ndash336 2011

[61] W T Kassahun T Schulz O Richter and J Hauss ldquoUnchangedhighmortality rates fromacute occlusive intestinal ischemia sixyear reviewrdquo Langenbecks Archives of Surgery vol 393 no 2 pp163ndash171 2008

Submit your manuscripts athttpwwwhindawicom

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014


MEDIATORSINFLAMMATION

of


Behavioural Neurology

EndocrinologyInternational Journal of



Disease Markers


BioMed Research International

OncologyJournal of



Oxidative Medicine and Cellular Longevity


PPAR Research

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Immunology ResearchHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Journal of

ObesityJournal of



Computational and Mathematical Methods in Medicine

OphthalmologyJournal of


Diabetes ResearchJournal of



Research and TreatmentAIDS


Gastroenterology Research and Practice


Parkinsonrsquos Disease

Evidence-Based Complementary and Alternative Medicine

Volume 2014Hindawi Publishing Corporationhttpwwwhindawicom


can alleviate IIR injury [14 15] We have previously foundthat ROS production was significantly increased after IIRwhich is parallel with the enhancements inMCdegranulation[16] but it is unknown whether a causal relationship existsbetween increased ROS production and MC degranulationin the setting of IIR in vivo These findings [6 17] togetherwith reports showing that MC can be activated in a ROS-dependent pathway [18] both in vitro [18] and in vivo [16]prompted us to postulate that during IIR increased ROSproduction initiates andor exacerbates IIR injury primarilyvia activating MC and that NADPH oxidase activation isincreased during IIR which may be a major source of ROSoverproduction during IIR

Propofol an intravenous anesthetic with antioxidantproperty that we widely used in intensive care unit andoperation theatre has been shown to dose-dependentlyattenuate myocardial ischemia reperfusion injury in patients[19] Propofol has also been shown to inhibit mast cellexocytosis in a dose-dependent manner in vitro [20] A mostrecent study shows that propofol attenuates brain traumainduced cerebral injury through inhibiting NADPH oxidaseactivation [21] We therefore hypothesized that inhibition ofROS mediated MC activation subsequent to attenuation ofintestinal NADPH oxidase activation may represent a majormechanism by which propofol attenuates IIR injury Thishypothesis was tested in a rat model of mesenteric ischemiareperfusion in vivo and a rat cell line of mast cell exposed toROS in vitro

2 Materials and Methods

21 Cell Culture Ratmast cell line (RBL-2H3) was purchasedfrom the cell bank of the Chinese Academy of Sciences(Shanghai China) and was cultured in Eaglersquos minimumessential medium containing 10 fetal bovine serum sup-plemented with 100UmL penicillin and 100 120583gmL strep-tomycin at 37∘C in a humidified atmosphere of 5 CO

2

as described [22] A rat small intestinal epithelial cell line(IEC-6) was also obtained from the cell bank of the ChineseAcademy of Sciences (Shanghai China) This cell line wascultured in 1640 RPMI medium with 10 fetal bovine serumat 37∘C in a 5 CO

2incubator

22 Measurement of Cell Degranulation Degranulation ofRBL-2H3 cells was measured by determining the activityof released 120573-hexosaminidase in the culture supernatantsand cell lysates [23] The RBL-2H3 cells were incubatedwith different concentrations of NAC [24] or propofol [20]for 12 h after the cells were washed with 1 times PBS for 3times RBL-2H3 cells were incubated in a 24-well plate (1 times106 cellswell) at 37∘C overnightThe above cells were washedwith 1timesPBS and then incubatedwith different concentrationsof lipopolysaccharides (LPS) [25] or H

2O2[26] or Com-

pound 4880 [27] for 1 h or 4 h To measure the amount of120573-hexosaminidase activity released from the cells the cul-tured media were transferred and centrifuged at 4∘C Thesupernatant (25 120583L) was mixed with 50120583L p-NAG (10mM)in 01M sodium citrate buffer (pH 45) into a 96-well plate

and incubated for 1 h at 37∘CThe reaction was terminated bythe addition of stop buffer (01M sodium carbonate bufferpH 100) The 120573-hexosaminidase activity was determinedby measuring the difference of absorbance at wavelength405 nm

23 Measurement of Cell Viability IEC-6 cells were seededinto 96-well flat bottom culture plates at a density of 5 times104 cellswell and incubated for 24 h After exposure totryptase (0ndash1000 ngmL) for 12 h in RPMI 1640 culturemedium the cell viability was measured by modified MTTassay as described [28] The assay detected the reduction ofthe tetrazolium to formazan product The cell viability wasevaluated using the following formula Survival rate () =(119860 sample ndash 119860blank)(119860control ndash 119860blank) times 100

24 Assessment of Cell Cytotoxicity Cytotoxicity (cell necro-sis) was assessed by measuring the release of lactate dehy-drogenase (LDH) After exposure of the cells to various con-centrations of tryptase (0ndash1000 ngmL) for 2 h in RPMI 1640culture medium cell necrosis was assessed by LDH-releaseassay using a commercial kit (Jiancheng China) according tothe manufacturerrsquos instruction

25 Animals Female Sprague-Dawley (SD) rats weighing180ndash200 g purchased from Animal Center of GuangdongProvince (Guangzhou China) were housed individually inwire-bottomed cages and were placed under pathogen freecondition illuminated from 800 AM to 800 PM (1212 hlight-dark cycle) for one week before use The experimentalprotocol and design was approved by the Sun Yat-sen Uni-versity Animal Experimentation Committee and performedaccording to Sun Yat-sen University Guidelines for AnimalExperimentation

26 In Vivo Rat IIR Model and Treatments All the animals were fasted for 16 h (while free access to water wasallowed) before surgery Rats were respectively injectedwithN-acetylcysteine (NAC 05 gkg from Sigma company)propofol (50mgkg commercial product Diprivan fromAstraZeneca) intralipid (50mgkg 20 emulsion fromSigma) or normal saline (05mL100 g) which served as thecontrol group intraperitoneally at 600 PM for 3 successivedays The dosages of NAC were chosen based on the resultsshowing that treatment of rats with ip NAC (500mg kgminus1per day for 9 days) improved the renal hemodynamic changestriggered by cisplatin-mediated nephrotoxicity [29]The doseof propofol was chosen based on the finding that propofol50mgkg given intraperitoneally provided sedative effect butnot anesthetic effect [30] and that propofol when used at thisdosage attenuated IIR injury in rats [31] At the 4th day partsof the rats were sacrificed by overdose of anesthetic chloralhydrate and the intestinal mucous was obtained and scrapedfor further determination and the intestinal morphologicalchanges were assessed



























3 Results


2O2dose-dependently




2O2(Figure 1(e))





0

5

10

15

20120573

-Hex

osam

inid

ase r

eleas

ed (

)

Control LPS10ngmL

LPS100ngmL

LPS1000ngmL

1h4h

(a)

0

20

40

60

80

Compound 4880

120573-H

exos

amin

idas

e rele

ased

()


lowast

lowast

(b)

0

10

20

30

120573-H

exos

amin

idas

e rele

ased

()

Control

H2O2

5120583M50120583M500120583M

lowast

lowast

(c)

0

10

20

30

40(

)

H2O2 500120583MNAC 10mM

NAC 20mM

minus

minus

minus

minus minus

minus minus

+ + +

+

+

lowast

(d)

0

10

20

30

40

()

lowast




+ ++

+

+

+

+

lowast

lowast

lowast$



(e)

0

20

40

60

80

100

Tryptase

Control

10ngmL1ngmL

100ngmL1000ngmL

lowast

lowast

Cel

l via

bilit

y (

)

lowast

(f)

Figure 1 Continued


0

200

400

600

Tryptase

Control

10ngmL1ngmL

100ngmL1000ngmL

lowastlowast lowast

lowast

LDH

(UL

)

(g)














(a) (b)

(c) (d)

00

02

04

NS NAC Pro Lip

Chiu

rsquos sc

ores

(e)





0

200

400

600

800

1000

NS NAC Pro Lip

lowast lowast15

-F2t

-isop

rost

ane (

pgm

g)

(a)

0

50

100

150

SOD

activ

ities

(uni

tsm

g)

NS NAC Pro Lip

lowast

lowast

(b)


p47phox

gp91phox

120572-Tubulin

(c)

00

05

10

15

NS NAC Pro Lip

p47ph

oxex

pres

sions

(au

)

lowast lowast

(d)

00

05

10

15

NS NAC Pro Lip

gp91

phox

expr

essio

ns (a

u)

lowast lowast

(e)

00

05

10

15

Tryptase

Tryp

tase

expr

essio

ns (a

u)

NS NAC Pro Lip

120572-Tubulin

(f)

00

05

10

15

NS NAC Pro Lip

120573-H

exos

amin

idas

e (nM

)

(g)





0 20 40 60 80 100 1200

20

40

60

80

100

IIRIIR + CP

P lt 00001

(min)

Surv

ival

rate

s (

)(a)

0 20 40 60 80 100 120(min)

0

20

40

60

80

100

P lt 00001


IIR + CPSurv

ival

rate

s (

)

(b)

0 20 40 60 80 100 120(min)

0

20

40

60

80

100

P lt 00001

P =

05

005

P =

00

384IIR

Lip + IIR

Lip + IIR + CP

IIR + CPSu

rviv

al ra

tes (

)

(c)

SHAM IIR

IIR + CP

IIR

IIR + CP

IIR

IIR + CP

IIR

IIR + CP


(d)

0

2

4

6

Propofol

Intralipid

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

lowast lowast

Chiu

rsquos sc

ores NAC

lowast$ lowast$

lowast998771lowast

lowastlowast

(e)











4 Discussion





0

1000

2000

3000

4000

NAC Propofol

Intralipid

15-F

2t-is

opro

stan

e (pg

mg)

lowast lowast

$ $

lowast lowast998771

(a)

NAC Propofol

Intralipid

0

50

100

150

200

lowast lowast

lowast

SOD

activ

ities

(uni

tsm

g)

$

$

lowast998771

(b)

NAC Propofol

Intralipid

00

05

10

15

20

25

lowast lowast$ $

p47phox

120572-Tubulin

p47ph

oxex

pres

sions

(au


(c)

NAC Propofol

Intralipid

00

05

10

15

20

25

lowast lowast

$ $

gp91phox

120572-Tubulin

gp91

phox

expr

essio

ns (a


(d)

NAC Propofol

Intralipid

0

5000

10000

15000

lowast lowast

lowast$ lowast$

IL-6

leve

ls (p

gm

g)

lowastlowast998771

(e)

NAC Propofol

Intralipid

0

10

20

30

40

lowast lowast


MPO

activ

ities

(uni

tsg)

lowast lowast998771

(f)


Intralipid

00

05

10

15

20

25

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

120572-Tubulin

ICAM-1

ICA

M-1

expr

essio

ns (a

u)

lowast lowast998771

(g)

$$

lowastlowast

lowast

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

NAC Propofol

Intralipid

00

05

10

15

20

25

120572-Tubulin

P-selectin

P-se

lect

in ex

pres

sions

(au

)

lowast998771

(h)








NAC Propofol

Intralipidlowast

lowast998771

120572-Tubulin

lowast lowast

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

3

2

1

0

Tryptase120573

-Hex

osam

inid

ase (

nM)

lowast$$

(a)

NAC Propofol

Intralipid

lowast lowast998771

lowastlowast

3

2

1

0

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

Tryp

tase

expr

essio

ns (a

u)

$$

(b)


















Acknowledgments


References








































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of










































3 Results


2O2dose-dependently




2O2(Figure 1(e))





0

5

10

15

20120573

-Hex

osam

inid

ase r

eleas

ed (

)

Control LPS10ngmL

LPS100ngmL

LPS1000ngmL

1h4h

(a)

0

20

40

60

80

Compound 4880

120573-H

exos

amin

idas

e rele

ased

()


lowast

lowast

(b)

0

10

20

30

120573-H

exos

amin

idas

e rele

ased

()

Control

H2O2

5120583M50120583M500120583M

lowast

lowast

(c)

0

10

20

30

40(

)

H2O2 500120583MNAC 10mM

NAC 20mM

minus

minus

minus

minus minus

minus minus

+ + +

+

+

lowast

(d)

0

10

20

30

40

()

lowast




+ ++

+

+

+

+

lowast

lowast

lowast$



(e)

0

20

40

60

80

100

Tryptase

Control

10ngmL1ngmL

100ngmL1000ngmL

lowast

lowast

Cel

l via

bilit

y (

)

lowast

(f)

Figure 1 Continued


0

200

400

600

Tryptase

Control

10ngmL1ngmL

100ngmL1000ngmL

lowastlowast lowast

lowast

LDH

(UL

)

(g)














(a) (b)

(c) (d)

00

02

04

NS NAC Pro Lip

Chiu

rsquos sc

ores

(e)





0

200

400

600

800

1000

NS NAC Pro Lip

lowast lowast15

-F2t

-isop

rost

ane (

pgm

g)

(a)

0

50

100

150

SOD

activ

ities

(uni

tsm

g)

NS NAC Pro Lip

lowast

lowast

(b)


p47phox

gp91phox

120572-Tubulin

(c)

00

05

10

15

NS NAC Pro Lip

p47ph

oxex

pres

sions

(au

)

lowast lowast

(d)

00

05

10

15

NS NAC Pro Lip

gp91

phox

expr

essio

ns (a

u)

lowast lowast

(e)

00

05

10

15

Tryptase

Tryp

tase

expr

essio

ns (a

u)

NS NAC Pro Lip

120572-Tubulin

(f)

00

05

10

15

NS NAC Pro Lip

120573-H

exos

amin

idas

e (nM

)

(g)





0 20 40 60 80 100 1200

20

40

60

80

100

IIRIIR + CP

P lt 00001

(min)

Surv

ival

rate

s (

)(a)

0 20 40 60 80 100 120(min)

0

20

40

60

80

100

P lt 00001


IIR + CPSurv

ival

rate

s (

)

(b)

0 20 40 60 80 100 120(min)

0

20

40

60

80

100

P lt 00001

P =

05

005

P =

00

384IIR

Lip + IIR

Lip + IIR + CP

IIR + CPSu

rviv

al ra

tes (

)

(c)

SHAM IIR

IIR + CP

IIR

IIR + CP

IIR

IIR + CP

IIR

IIR + CP


(d)

0

2

4

6

Propofol

Intralipid

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

lowast lowast

Chiu

rsquos sc

ores NAC

lowast$ lowast$

lowast998771lowast

lowastlowast

(e)











4 Discussion





0

1000

2000

3000

4000

NAC Propofol

Intralipid

15-F

2t-is

opro

stan

e (pg

mg)

lowast lowast

$ $

lowast lowast998771

(a)

NAC Propofol

Intralipid

0

50

100

150

200

lowast lowast

lowast

SOD

activ

ities

(uni

tsm

g)

$

$

lowast998771

(b)

NAC Propofol

Intralipid

00

05

10

15

20

25

lowast lowast$ $

p47phox

120572-Tubulin

p47ph

oxex

pres

sions

(au


(c)

NAC Propofol

Intralipid

00

05

10

15

20

25

lowast lowast

$ $

gp91phox

120572-Tubulin

gp91

phox

expr

essio

ns (a


(d)

NAC Propofol

Intralipid

0

5000

10000

15000

lowast lowast

lowast$ lowast$

IL-6

leve

ls (p

gm

g)

lowastlowast998771

(e)

NAC Propofol

Intralipid

0

10

20

30

40

lowast lowast


MPO

activ

ities

(uni

tsg)

lowast lowast998771

(f)


Intralipid

00

05

10

15

20

25

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

120572-Tubulin

ICAM-1

ICA

M-1

expr

essio

ns (a

u)

lowast lowast998771

(g)

$$

lowastlowast

lowast

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

NAC Propofol

Intralipid

00

05

10

15

20

25

120572-Tubulin

P-selectin

P-se

lect

in ex

pres

sions

(au

)

lowast998771

(h)








NAC Propofol

Intralipidlowast

lowast998771

120572-Tubulin

lowast lowast

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

3

2

1

0

Tryptase120573

-Hex

osam

inid

ase (

nM)

lowast$$

(a)

NAC Propofol

Intralipid

lowast lowast998771

lowastlowast

3

2

1

0

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

Tryp

tase

expr

essio

ns (a

u)

$$

(b)


















Acknowledgments


References








































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of

















0

5

10

15

20120573

-Hex

osam

inid

ase r

eleas

ed (

)

Control LPS10ngmL

LPS100ngmL

LPS1000ngmL

1h4h

(a)

0

20

40

60

80

Compound 4880

120573-H

exos

amin

idas

e rele

ased

()


lowast

lowast

(b)

0

10

20

30

120573-H

exos

amin

idas

e rele

ased

()

Control

H2O2

5120583M50120583M500120583M

lowast

lowast

(c)

0

10

20

30

40(

)

H2O2 500120583MNAC 10mM

NAC 20mM

minus

minus

minus

minus minus

minus minus

+ + +

+

+

lowast

(d)

0

10

20

30

40

()

lowast




+ ++

+

+

+

+

lowast

lowast

lowast$



(e)

0

20

40

60

80

100

Tryptase

Control

10ngmL1ngmL

100ngmL1000ngmL

lowast

lowast

Cel

l via

bilit

y (

)

lowast

(f)

Figure 1 Continued


0

200

400

600

Tryptase

Control

10ngmL1ngmL

100ngmL1000ngmL

lowastlowast lowast

lowast

LDH

(UL

)

(g)














(a) (b)

(c) (d)

00

02

04

NS NAC Pro Lip

Chiu

rsquos sc

ores

(e)





0

200

400

600

800

1000

NS NAC Pro Lip

lowast lowast15

-F2t

-isop

rost

ane (

pgm

g)

(a)

0

50

100

150

SOD

activ

ities

(uni

tsm

g)

NS NAC Pro Lip

lowast

lowast

(b)


p47phox

gp91phox

120572-Tubulin

(c)

00

05

10

15

NS NAC Pro Lip

p47ph

oxex

pres

sions

(au

)

lowast lowast

(d)

00

05

10

15

NS NAC Pro Lip

gp91

phox

expr

essio

ns (a

u)

lowast lowast

(e)

00

05

10

15

Tryptase

Tryp

tase

expr

essio

ns (a

u)

NS NAC Pro Lip

120572-Tubulin

(f)

00

05

10

15

NS NAC Pro Lip

120573-H

exos

amin

idas

e (nM

)

(g)





0 20 40 60 80 100 1200

20

40

60

80

100

IIRIIR + CP

P lt 00001

(min)

Surv

ival

rate

s (

)(a)

0 20 40 60 80 100 120(min)

0

20

40

60

80

100

P lt 00001


IIR + CPSurv

ival

rate

s (

)

(b)

0 20 40 60 80 100 120(min)

0

20

40

60

80

100

P lt 00001

P =

05

005

P =

00

384IIR

Lip + IIR

Lip + IIR + CP

IIR + CPSu

rviv

al ra

tes (

)

(c)

SHAM IIR

IIR + CP

IIR

IIR + CP

IIR

IIR + CP

IIR

IIR + CP


(d)

0

2

4

6

Propofol

Intralipid

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

lowast lowast

Chiu

rsquos sc

ores NAC

lowast$ lowast$

lowast998771lowast

lowastlowast

(e)











4 Discussion





0

1000

2000

3000

4000

NAC Propofol

Intralipid

15-F

2t-is

opro

stan

e (pg

mg)

lowast lowast

$ $

lowast lowast998771

(a)

NAC Propofol

Intralipid

0

50

100

150

200

lowast lowast

lowast

SOD

activ

ities

(uni

tsm

g)

$

$

lowast998771

(b)

NAC Propofol

Intralipid

00

05

10

15

20

25

lowast lowast$ $

p47phox

120572-Tubulin

p47ph

oxex

pres

sions

(au


(c)

NAC Propofol

Intralipid

00

05

10

15

20

25

lowast lowast

$ $

gp91phox

120572-Tubulin

gp91

phox

expr

essio

ns (a


(d)

NAC Propofol

Intralipid

0

5000

10000

15000

lowast lowast

lowast$ lowast$

IL-6

leve

ls (p

gm

g)

lowastlowast998771

(e)

NAC Propofol

Intralipid

0

10

20

30

40

lowast lowast


MPO

activ

ities

(uni

tsg)

lowast lowast998771

(f)


Intralipid

00

05

10

15

20

25

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

120572-Tubulin

ICAM-1

ICA

M-1

expr

essio

ns (a

u)

lowast lowast998771

(g)

$$

lowastlowast

lowast

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

NAC Propofol

Intralipid

00

05

10

15

20

25

120572-Tubulin

P-selectin

P-se

lect

in ex

pres

sions

(au

)

lowast998771

(h)








NAC Propofol

Intralipidlowast

lowast998771

120572-Tubulin

lowast lowast

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

3

2

1

0

Tryptase120573

-Hex

osam

inid

ase (

nM)

lowast$$

(a)

NAC Propofol

Intralipid

lowast lowast998771

lowastlowast

3

2

1

0

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

Tryp

tase

expr

essio

ns (a

u)

$$

(b)


















Acknowledgments


References








































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of

















0

200

400

600

Tryptase

Control

10ngmL1ngmL

100ngmL1000ngmL

lowastlowast lowast

lowast

LDH

(UL

)

(g)














(a) (b)

(c) (d)

00

02

04

NS NAC Pro Lip

Chiu

rsquos sc

ores

(e)





0

200

400

600

800

1000

NS NAC Pro Lip

lowast lowast15

-F2t

-isop

rost

ane (

pgm

g)

(a)

0

50

100

150

SOD

activ

ities

(uni

tsm

g)

NS NAC Pro Lip

lowast

lowast

(b)


p47phox

gp91phox

120572-Tubulin

(c)

00

05

10

15

NS NAC Pro Lip

p47ph

oxex

pres

sions

(au

)

lowast lowast

(d)

00

05

10

15

NS NAC Pro Lip

gp91

phox

expr

essio

ns (a

u)

lowast lowast

(e)

00

05

10

15

Tryptase

Tryp

tase

expr

essio

ns (a

u)

NS NAC Pro Lip

120572-Tubulin

(f)

00

05

10

15

NS NAC Pro Lip

120573-H

exos

amin

idas

e (nM

)

(g)





0 20 40 60 80 100 1200

20

40

60

80

100

IIRIIR + CP

P lt 00001

(min)

Surv

ival

rate

s (

)(a)

0 20 40 60 80 100 120(min)

0

20

40

60

80

100

P lt 00001


IIR + CPSurv

ival

rate

s (

)

(b)

0 20 40 60 80 100 120(min)

0

20

40

60

80

100

P lt 00001

P =

05

005

P =

00

384IIR

Lip + IIR

Lip + IIR + CP

IIR + CPSu

rviv

al ra

tes (

)

(c)

SHAM IIR

IIR + CP

IIR

IIR + CP

IIR

IIR + CP

IIR

IIR + CP


(d)

0

2

4

6

Propofol

Intralipid

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

lowast lowast

Chiu

rsquos sc

ores NAC

lowast$ lowast$

lowast998771lowast

lowastlowast

(e)











4 Discussion





0

1000

2000

3000

4000

NAC Propofol

Intralipid

15-F

2t-is

opro

stan

e (pg

mg)

lowast lowast

$ $

lowast lowast998771

(a)

NAC Propofol

Intralipid

0

50

100

150

200

lowast lowast

lowast

SOD

activ

ities

(uni

tsm

g)

$

$

lowast998771

(b)

NAC Propofol

Intralipid

00

05

10

15

20

25

lowast lowast$ $

p47phox

120572-Tubulin

p47ph

oxex

pres

sions

(au


(c)

NAC Propofol

Intralipid

00

05

10

15

20

25

lowast lowast

$ $

gp91phox

120572-Tubulin

gp91

phox

expr

essio

ns (a


(d)

NAC Propofol

Intralipid

0

5000

10000

15000

lowast lowast

lowast$ lowast$

IL-6

leve

ls (p

gm

g)

lowastlowast998771

(e)

NAC Propofol

Intralipid

0

10

20

30

40

lowast lowast


MPO

activ

ities

(uni

tsg)

lowast lowast998771

(f)


Intralipid

00

05

10

15

20

25

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

120572-Tubulin

ICAM-1

ICA

M-1

expr

essio

ns (a

u)

lowast lowast998771

(g)

$$

lowastlowast

lowast

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

NAC Propofol

Intralipid

00

05

10

15

20

25

120572-Tubulin

P-selectin

P-se

lect

in ex

pres

sions

(au

)

lowast998771

(h)








NAC Propofol

Intralipidlowast

lowast998771

120572-Tubulin

lowast lowast

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

3

2

1

0

Tryptase120573

-Hex

osam

inid

ase (

nM)

lowast$$

(a)

NAC Propofol

Intralipid

lowast lowast998771

lowastlowast

3

2

1

0

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

Tryp

tase

expr

essio

ns (a

u)

$$

(b)


















Acknowledgments


References








































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of

















(a) (b)

(c) (d)

00

02

04

NS NAC Pro Lip

Chiu

rsquos sc

ores

(e)





0

200

400

600

800

1000

NS NAC Pro Lip

lowast lowast15

-F2t

-isop

rost

ane (

pgm

g)

(a)

0

50

100

150

SOD

activ

ities

(uni

tsm

g)

NS NAC Pro Lip

lowast

lowast

(b)


p47phox

gp91phox

120572-Tubulin

(c)

00

05

10

15

NS NAC Pro Lip

p47ph

oxex

pres

sions

(au

)

lowast lowast

(d)

00

05

10

15

NS NAC Pro Lip

gp91

phox

expr

essio

ns (a

u)

lowast lowast

(e)

00

05

10

15

Tryptase

Tryp

tase

expr

essio

ns (a

u)

NS NAC Pro Lip

120572-Tubulin

(f)

00

05

10

15

NS NAC Pro Lip

120573-H

exos

amin

idas

e (nM

)

(g)





0 20 40 60 80 100 1200

20

40

60

80

100

IIRIIR + CP

P lt 00001

(min)

Surv

ival

rate

s (

)(a)

0 20 40 60 80 100 120(min)

0

20

40

60

80

100

P lt 00001


IIR + CPSurv

ival

rate

s (

)

(b)

0 20 40 60 80 100 120(min)

0

20

40

60

80

100

P lt 00001

P =

05

005

P =

00

384IIR

Lip + IIR

Lip + IIR + CP

IIR + CPSu

rviv

al ra

tes (

)

(c)

SHAM IIR

IIR + CP

IIR

IIR + CP

IIR

IIR + CP

IIR

IIR + CP


(d)

0

2

4

6

Propofol

Intralipid

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

lowast lowast

Chiu

rsquos sc

ores NAC

lowast$ lowast$

lowast998771lowast

lowastlowast

(e)











4 Discussion





0

1000

2000

3000

4000

NAC Propofol

Intralipid

15-F

2t-is

opro

stan

e (pg

mg)

lowast lowast

$ $

lowast lowast998771

(a)

NAC Propofol

Intralipid

0

50

100

150

200

lowast lowast

lowast

SOD

activ

ities

(uni

tsm

g)

$

$

lowast998771

(b)

NAC Propofol

Intralipid

00

05

10

15

20

25

lowast lowast$ $

p47phox

120572-Tubulin

p47ph

oxex

pres

sions

(au


(c)

NAC Propofol

Intralipid

00

05

10

15

20

25

lowast lowast

$ $

gp91phox

120572-Tubulin

gp91

phox

expr

essio

ns (a


(d)

NAC Propofol

Intralipid

0

5000

10000

15000

lowast lowast

lowast$ lowast$

IL-6

leve

ls (p

gm

g)

lowastlowast998771

(e)

NAC Propofol

Intralipid

0

10

20

30

40

lowast lowast


MPO

activ

ities

(uni

tsg)

lowast lowast998771

(f)


Intralipid

00

05

10

15

20

25

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

120572-Tubulin

ICAM-1

ICA

M-1

expr

essio

ns (a

u)

lowast lowast998771

(g)

$$

lowastlowast

lowast

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

NAC Propofol

Intralipid

00

05

10

15

20

25

120572-Tubulin

P-selectin

P-se

lect

in ex

pres

sions

(au

)

lowast998771

(h)








NAC Propofol

Intralipidlowast

lowast998771

120572-Tubulin

lowast lowast

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

3

2

1

0

Tryptase120573

-Hex

osam

inid

ase (

nM)

lowast$$

(a)

NAC Propofol

Intralipid

lowast lowast998771

lowastlowast

3

2

1

0

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

Tryp

tase

expr

essio

ns (a

u)

$$

(b)


















Acknowledgments


References








































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of

















0

200

400

600

800

1000

NS NAC Pro Lip

lowast lowast15

-F2t

-isop

rost

ane (

pgm

g)

(a)

0

50

100

150

SOD

activ

ities

(uni

tsm

g)

NS NAC Pro Lip

lowast

lowast

(b)


p47phox

gp91phox

120572-Tubulin

(c)

00

05

10

15

NS NAC Pro Lip

p47ph

oxex

pres

sions

(au

)

lowast lowast

(d)

00

05

10

15

NS NAC Pro Lip

gp91

phox

expr

essio

ns (a

u)

lowast lowast

(e)

00

05

10

15

Tryptase

Tryp

tase

expr

essio

ns (a

u)

NS NAC Pro Lip

120572-Tubulin

(f)

00

05

10

15

NS NAC Pro Lip

120573-H

exos

amin

idas

e (nM

)

(g)





0 20 40 60 80 100 1200

20

40

60

80

100

IIRIIR + CP

P lt 00001

(min)

Surv

ival

rate

s (

)(a)

0 20 40 60 80 100 120(min)

0

20

40

60

80

100

P lt 00001


IIR + CPSurv

ival

rate

s (

)

(b)

0 20 40 60 80 100 120(min)

0

20

40

60

80

100

P lt 00001

P =

05

005

P =

00

384IIR

Lip + IIR

Lip + IIR + CP

IIR + CPSu

rviv

al ra

tes (

)

(c)

SHAM IIR

IIR + CP

IIR

IIR + CP

IIR

IIR + CP

IIR

IIR + CP


(d)

0

2

4

6

Propofol

Intralipid

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

lowast lowast

Chiu

rsquos sc

ores NAC

lowast$ lowast$

lowast998771lowast

lowastlowast

(e)











4 Discussion





0

1000

2000

3000

4000

NAC Propofol

Intralipid

15-F

2t-is

opro

stan

e (pg

mg)

lowast lowast

$ $

lowast lowast998771

(a)

NAC Propofol

Intralipid

0

50

100

150

200

lowast lowast

lowast

SOD

activ

ities

(uni

tsm

g)

$

$

lowast998771

(b)

NAC Propofol

Intralipid

00

05

10

15

20

25

lowast lowast$ $

p47phox

120572-Tubulin

p47ph

oxex

pres

sions

(au


(c)

NAC Propofol

Intralipid

00

05

10

15

20

25

lowast lowast

$ $

gp91phox

120572-Tubulin

gp91

phox

expr

essio

ns (a


(d)

NAC Propofol

Intralipid

0

5000

10000

15000

lowast lowast

lowast$ lowast$

IL-6

leve

ls (p

gm

g)

lowastlowast998771

(e)

NAC Propofol

Intralipid

0

10

20

30

40

lowast lowast


MPO

activ

ities

(uni

tsg)

lowast lowast998771

(f)


Intralipid

00

05

10

15

20

25

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

120572-Tubulin

ICAM-1

ICA

M-1

expr

essio

ns (a

u)

lowast lowast998771

(g)

$$

lowastlowast

lowast

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

NAC Propofol

Intralipid

00

05

10

15

20

25

120572-Tubulin

P-selectin

P-se

lect

in ex

pres

sions

(au

)

lowast998771

(h)








NAC Propofol

Intralipidlowast

lowast998771

120572-Tubulin

lowast lowast

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

3

2

1

0

Tryptase120573

-Hex

osam

inid

ase (

nM)

lowast$$

(a)

NAC Propofol

Intralipid

lowast lowast998771

lowastlowast

3

2

1

0

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

Tryp

tase

expr

essio

ns (a

u)

$$

(b)


















Acknowledgments


References








































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of

















0 20 40 60 80 100 1200

20

40

60

80

100

IIRIIR + CP

P lt 00001

(min)

Surv

ival

rate

s (

)(a)

0 20 40 60 80 100 120(min)

0

20

40

60

80

100

P lt 00001


IIR + CPSurv

ival

rate

s (

)

(b)

0 20 40 60 80 100 120(min)

0

20

40

60

80

100

P lt 00001

P =

05

005

P =

00

384IIR

Lip + IIR

Lip + IIR + CP

IIR + CPSu

rviv

al ra

tes (

)

(c)

SHAM IIR

IIR + CP

IIR

IIR + CP

IIR

IIR + CP

IIR

IIR + CP


(d)

0

2

4

6

Propofol

Intralipid

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

lowast lowast

Chiu

rsquos sc

ores NAC

lowast$ lowast$

lowast998771lowast

lowastlowast

(e)











4 Discussion





0

1000

2000

3000

4000

NAC Propofol

Intralipid

15-F

2t-is

opro

stan

e (pg

mg)

lowast lowast

$ $

lowast lowast998771

(a)

NAC Propofol

Intralipid

0

50

100

150

200

lowast lowast

lowast

SOD

activ

ities

(uni

tsm

g)

$

$

lowast998771

(b)

NAC Propofol

Intralipid

00

05

10

15

20

25

lowast lowast$ $

p47phox

120572-Tubulin

p47ph

oxex

pres

sions

(au


(c)

NAC Propofol

Intralipid

00

05

10

15

20

25

lowast lowast

$ $

gp91phox

120572-Tubulin

gp91

phox

expr

essio

ns (a


(d)

NAC Propofol

Intralipid

0

5000

10000

15000

lowast lowast

lowast$ lowast$

IL-6

leve

ls (p

gm

g)

lowastlowast998771

(e)

NAC Propofol

Intralipid

0

10

20

30

40

lowast lowast


MPO

activ

ities

(uni

tsg)

lowast lowast998771

(f)


Intralipid

00

05

10

15

20

25

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

120572-Tubulin

ICAM-1

ICA

M-1

expr

essio

ns (a

u)

lowast lowast998771

(g)

$$

lowastlowast

lowast

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

NAC Propofol

Intralipid

00

05

10

15

20

25

120572-Tubulin

P-selectin

P-se

lect

in ex

pres

sions

(au

)

lowast998771

(h)








NAC Propofol

Intralipidlowast

lowast998771

120572-Tubulin

lowast lowast

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

3

2

1

0

Tryptase120573

-Hex

osam

inid

ase (

nM)

lowast$$

(a)

NAC Propofol

Intralipid

lowast lowast998771

lowastlowast

3

2

1

0

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

Tryp

tase

expr

essio

ns (a

u)

$$

(b)


















Acknowledgments


References








































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of

























4 Discussion





0

1000

2000

3000

4000

NAC Propofol

Intralipid

15-F

2t-is

opro

stan

e (pg

mg)

lowast lowast

$ $

lowast lowast998771

(a)

NAC Propofol

Intralipid

0

50

100

150

200

lowast lowast

lowast

SOD

activ

ities

(uni

tsm

g)

$

$

lowast998771

(b)

NAC Propofol

Intralipid

00

05

10

15

20

25

lowast lowast$ $

p47phox

120572-Tubulin

p47ph

oxex

pres

sions

(au


(c)

NAC Propofol

Intralipid

00

05

10

15

20

25

lowast lowast

$ $

gp91phox

120572-Tubulin

gp91

phox

expr

essio

ns (a


(d)

NAC Propofol

Intralipid

0

5000

10000

15000

lowast lowast

lowast$ lowast$

IL-6

leve

ls (p

gm

g)

lowastlowast998771

(e)

NAC Propofol

Intralipid

0

10

20

30

40

lowast lowast


MPO

activ

ities

(uni

tsg)

lowast lowast998771

(f)


Intralipid

00

05

10

15

20

25

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

120572-Tubulin

ICAM-1

ICA

M-1

expr

essio

ns (a

u)

lowast lowast998771

(g)

$$

lowastlowast

lowast

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

NAC Propofol

Intralipid

00

05

10

15

20

25

120572-Tubulin

P-selectin

P-se

lect

in ex

pres

sions

(au

)

lowast998771

(h)








NAC Propofol

Intralipidlowast

lowast998771

120572-Tubulin

lowast lowast

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

3

2

1

0

Tryptase120573

-Hex

osam

inid

ase (

nM)

lowast$$

(a)

NAC Propofol

Intralipid

lowast lowast998771

lowastlowast

3

2

1

0

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

Tryp

tase

expr

essio

ns (a

u)

$$

(b)


















Acknowledgments


References








































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of

















0

1000

2000

3000

4000

NAC Propofol

Intralipid

15-F

2t-is

opro

stan

e (pg

mg)

lowast lowast

$ $

lowast lowast998771

(a)

NAC Propofol

Intralipid

0

50

100

150

200

lowast lowast

lowast

SOD

activ

ities

(uni

tsm

g)

$

$

lowast998771

(b)

NAC Propofol

Intralipid

00

05

10

15

20

25

lowast lowast$ $

p47phox

120572-Tubulin

p47ph

oxex

pres

sions

(au


(c)

NAC Propofol

Intralipid

00

05

10

15

20

25

lowast lowast

$ $

gp91phox

120572-Tubulin

gp91

phox

expr

essio

ns (a


(d)

NAC Propofol

Intralipid

0

5000

10000

15000

lowast lowast

lowast$ lowast$

IL-6

leve

ls (p

gm

g)

lowastlowast998771

(e)

NAC Propofol

Intralipid

0

10

20

30

40

lowast lowast


MPO

activ

ities

(uni

tsg)

lowast lowast998771

(f)


Intralipid

00

05

10

15

20

25

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

120572-Tubulin

ICAM-1

ICA

M-1

expr

essio

ns (a

u)

lowast lowast998771

(g)

$$

lowastlowast

lowast

SHA

M IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

IIR

IIR +

CP

NAC Propofol

Intralipid

00

05

10

15

20

25

120572-Tubulin

P-selectin

P-se

lect

in ex

pres

sions

(au

)

lowast998771

(h)








NAC Propofol

Intralipidlowast

lowast998771

120572-Tubulin

lowast lowast

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

3

2

1

0

Tryptase120573

-Hex

osam

inid

ase (

nM)

lowast$$

(a)

NAC Propofol

Intralipid

lowast lowast998771

lowastlowast

3

2

1

0

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

Tryp

tase

expr

essio

ns (a

u)

$$

(b)


















Acknowledgments


References








































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of

















NAC Propofol

Intralipidlowast

lowast998771

120572-Tubulin

lowast lowast

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

3

2

1

0

Tryptase120573

-Hex

osam

inid

ase (

nM)

lowast$$

(a)

NAC Propofol

Intralipid

lowast lowast998771

lowastlowast

3

2

1

0

SHA

M IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP IIR

IIR+

CP

Tryp

tase

expr

essio

ns (a

u)

$$

(b)


















Acknowledgments


References








































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of



























Acknowledgments


References








































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of



















































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of
















research article propofol attenuates small intestinal...

Documents