ResearchArticle Portulaca Extract Attenuates...

Research ArticlePortulaca Extract Attenuates Development of Dextran SulfateSodium Induced Colitis in Mice through Activation of PPAR120574

Rui Kong 12 Hui Luo1 NanWang 1 Jingjing Li 1 Shizan Xu13 Kan Chen 1

Jiao Feng1 Liwei Wu1 Sainan Li1 Tong Liu1 Xiya Lu1 Yujing Xia 1 Yanhong Shi1

Yingqun Zhou 1 Weigang He 4 Qi Dai 5 Yuejuan Zheng 4 and Jie Lu 1

1Department of Gastroenterology Shanghai Tenth Peoplersquos Hospital Tongji University School of Medicine Shanghai 200072 China2The School of Medicine Soochow University Suzhou 215006 China3Department of Gastroenterology Shanghai Tenth Hospital School of Clinical Medicine Nanjing Medical UniversityShanghai 200072 China4Department of Immunology and Microbiology Shanghai University of Traditional Chinese Medicine Shanghai 201203 China5The Eye Hospital Wenzhou Medical University Wenzhou City Zhejiang 325027 China

Correspondence should be addressed to Qi Dai dqmaileyeaccn Yuejuan Zheng 13641776412163comand Jie Lu kennisrenhotmailcom

Received 27 July 2017 Revised 14 September 2017 Accepted 28 November 2017 Published 1 February 2018

Academic Editor Yuewen Gong

Copyright copy 2018 Rui Kong et alThis is an open access article distributed under the Creative Commons Attribution License whichpermits unrestricted use distribution and reproduction in any medium provided the original work is properly cited

Portulaca oleracea L is a traditional Chinese medicine which has been used as adjuvant therapy for inflammatory bowel disease(IBD) However the mechanism of its activity in IBD still remains unclear Since previous studies have documented the anti-inflammatory effect of peroxisome proliferator activated receptors-120574 (PPAR-120574) Portulaca regulation of PPAR-120574 in inflammationwas examined in current study Ulcerative colitis (UC) was generated by 5 dextran sulfate sodium (DSS) in mice and four groupswere established as normal control DSS alone DSS plusmesalamine andDSS plus Portulaca Severity of UCwas evaluated by bodyweight stool blood form and length of colorectum Inflammation was examined by determination of inflammatory cytokines(TNF-a IL-6 and IL-1a) Portulaca extract was able to attenuate development of UC in DSS model similar to the treatment ofmesalazine Moreover Portulaca extract inhibited proinflammatory cytokines release and reduced the level of DSS-induced NF-120581Bphosphorylation Furthermore Portulaca extract restored PPAR-120574 level which was reduced by DSS In addition Portulaca extractprotected DSS induced apoptosis inmice In conclusion Portulaca extract can alleviate colitis inmice through regulation of inflam-matory reaction apoptosis and PPAR-120574 level therefore Portulaca extract can be a potential candidate for the treatment of IBD

1 Introduction

Inflammatory bowel disease (IBD) consists of Crohnrsquos diseaseand ulcerative colitis and has been considered as a globalhealth threat to children and adults [1] The distributionof IBD varies in geographical regions with high incidencerate in North American and Northern Europe however themorbidity remains stable in these regions In the developingregions in the world such as Asia the incidence and mor-bidity are increased every year [2] Enormous efforts havebeen made to find an effective therapy for IBD althoughcurrent drugs including sulfasalazine mesalamine and cor-ticosteroids taken alone or in combination contribute to

decelerating disease progression [3] However with currenttreatments and medications some patients suffer from sideeffects and complications of these drugs such as weakenedimmunity infectious diseases or increased malignant poten-tialityTherefore a promisingmedicationwith no ormild sideeffects is needed for IBD treatment

Portulaca oleracea L (Portulaca) is a traditional Chi-nese herb praised with rich multiminerals proteins 120572-amyrin 120573-carotene terpenoids vitamins and fatty acids[4] The medicinal properties and usage of Portulaca havebeen recorded in many ancient Chinese books [5] More-over current researches reveal that Portulaca has several

HindawiPPAR ResearchVolume 2018 Article ID 6079101 11 pageshttpsdoiorg10115520186079101

2 PPAR Research

biological functions such as antibacteria neuroprotectionanti-inflammation antiatherosclerosis and antitumor [6ndash10] Furthermore decoction of Portulaca has been usedas a remedy for inflammatory bowel disease to alleviatesymptoms of bloody stools diarrhea and abdominal pain inclinic Several clinical trials have been conducted on the safetyand effectiveness of Portulaca which showed no adverseeffects of Portulaca Reports from animal studies also indicatelow toxicity of Portulaca with LD

50value of 1853mgkg and

high therapeutic index [11]Peroxisome proliferator-activated receptor-120574 (PPAR-120574) is

an important member of the nuclear receptor superfamilyThe essential role of PPAR-120574 in acute inflammation [12]adipogenesis [13] and several carcinomas has been iden-tified [14ndash16] Moreover PPAR-120574 activation was found invarious types of immune cells including primary peritonealmacrophages dendritic cells and T cells [17] Several studieshave shown that activation of PPAR-120574 could downregulatesome vital proinflammatory cytokines such as IL-1120573 IL-2IL-6 and TNF-120572 [18] as well as inhibition of NF-120581B phos-phorylation in monocyte and other types of cells Thereforeactivation of PPAR-120574 exerts a direct anti-inflammatory effectin IBD while the mechanism of PPAR-120574 activation remainsto be further explored [19]

In the current study the anti-inflammatory activity andmechanism of Portulaca in dextran sulfate sodium (DSS)induced UC animal model were explored

2 Materials and Methods

21 Plant Material and Preparation of the Extracts ThePortulaca material was obtained from the Department ofTraditional Chinese Medicine Pharmacy of the ShanghaiTenth Peoplersquos Hospital The herb was triturated into pow-der and water-soluble substance was extracted as describedpreviously [20] Briefly the prepared powder was boiled indistilled water (80∘C 25 g120ml water wv) for 60min andthen smashed via ultrasonic concussion (3 cycles of 15 sec)Themixture of water and concussion was filtered using 2mmpores strainer and the resultant material was lyophilizedunder vacuum

22 Animals and Acute Colitis Induction Forty adult femalemice weighting 18ndash22 g were purchased from the Shang-hai Laboratory Animal Co Ltd (Shanghai China) andmaintained in an environment of 25∘C with controlled 12 hlightdark cycle Mice were permitted free access to waterand standardmouse chowThe animal protocol was approvedby the Ethics Committee of the Tongji University whichmeets the recommendations in the Guide for Care and Useof Laboratory Animals of the National Institutes of HealthThe dextran sulfate sodium reagent (molecular weight36000ndash50000 fromMP Biochemicals Solon OH USA) wasdiluted with drinking water to make final concentration of3

The mice were divided into 4 groups at random

(1) Normal control given normal food and water for 7days

Table 1 DAI assessment (disease activity index)

Weightloss Shape of stool FOBTlowast

blood stool Score

0 Normal Normal 01ndash5 Shapeless stool FOBT positive 16ndash10 211ndash15 Loose stool Gross blood stool 3gt16 4lowastFOBT fecal occult blood test

(2) DSS group 3 DSS + saline by gavage for 7 days(3) Mesalamine positive control group 3 DSS +

74mgkg mesalamine daily by gavage for 7 days(4) Portulaca group 3 DSS + 100mgkg daily by gavage

for 7 days

During the period of modeling mice weight stool formand stool occult blood were recorded every day to assess thedisease activity index (DAI) of acute colitis the evaluativecriteria were clarified in Table 1

23 SerumCytokine Analysis The serum levels of TNF-120572 IL-1120573 and IL-6 concentrations were determined by using ELISAKits (Elabscience Wuhan China) respectively according tothe manufacturerrsquos instructionThe OD values of absorbanceat 450 nm were examined byThermoMax microplate reader

24 Determination of MPO (Myeloperoxidase) Activity inSerum Sample Myeloperoxidase is found in activated neu-trophils and macrophages which plays a defensive role dur-ing inflammatory response via catalyzing hydrogen peroxideinto hypochlorous acid The serum samples reacted withthe mixed solution including 331015840-dimethoxybiphenyl-441015840-diamine and hydrogen peroxide pH 60 according to theMPO assay kit (Nanjing Jiancheng Bioengineering InstituteA044 China)The absorbance of control well and testing wellat 460 nm was determined by microplate reader The activityof MPO was calculated as the following formula

MPO performance (UL)

=(testing OD values minus control OD values)

113

times sample volume (L)

(1)

25 Fecal Occult Blood Testing The form of stool wasrecorded and the severity of stool occult blood was analyzedby urine fecal occult blood test kit (Nanjing JianchengBioengineering Institute C027 China) A small amount offeces samples was picked and smeared on the slides Addorthotolidine and hydrogen peroxide reagent on the surfaceof stool samples The results were analyzed according to theindications negative (minus) the samples do not show colorwithin 3 minutes weakly positive (+) the samples show bluewithin 30 to 60 seconds positive (++) the samples appear

PPAR Research 3

Table 2 The sequence of primers used in experiments

Gene Primer sequence (51015840-31015840)Annealingtemperature

(∘C)GADPH

F GGACCTCATGGCCTACATGG 576R TAGGGCCTCTCTTGCTCAGT

IL-6F GGCGGATCGGATGTTGTGAT 573R GGACCCCAGACAATCGGTTG

IL-1120573F CTTTGAAGTTGACGGACCC 531R TGAGTGATACTGCCTGCCTG

TNF-120572F CACCACCATCAAGGACTCAA 545R AGGCAACCTGACCACTCTCC

PPAR-120574F CCCAATGGTTGCTGATTACA 544R GGACGCAGGCTCTACTTTGA

NF120581Bp65F TGCGATTCCGCTATAAATGCG 602R ACAAGTTCATGTGGATGAGGC

BaxF AGACAGGGGCCTTTTTGCTAC 573R AATTCGCCGGAGACACTCG

Bcl-2F GAGCCTGTGAGAGACGTGG 573R CGAGTCTGTGTATAGCAATCCCA

bluish green strongly positive (+++) samples showmazarineimmediately

26 Gene Expression Analysis by Real-Time Reverse Transcrip-tase Polymerase Chain Reaction Total RNA was extractedfrom fresh colorectum tissues by the Trizol reagent from theThermo Fisher Scientific (Waltham MA USA) and washedby 75 ethanol One microgram total RNA was transcribedinto cDNA using reverse transcription kit (Takara Biotech-nology Dalian China) The first transcribed cDNA wasdiluted ten times and applied to real-time polymerase chainreaction (SYBR Premix EX Taq Takara Biotechnology) withthe following cycle of denaturing at 94∘C for 30 secondsannealing at different temperature according to the primersfor 30 seconds and elongation at 72∘C for 45 seconds usinga 7900HT Fast Real-Time PCR System (Thermo FisherScientific) All primers used in experiments were listed inTable 2

27 Hematoxylin and Eosin Staining Fresh colorectum tis-sues were fixed in formalin and embedded in paraffin Thetissue blot was cut into 4 120583m thick section for histologicalexamination using hematoxylin and eosin (HampE)The injuryindex of histology was evaluated according to three aspects

the destructive degree of the intestine epithelium the pres-ence of the immune cell infiltration in intestine mucosa andthe infiltration degree of immune cell in submucosa Thehistology score was depicted in Table 3 Two experiencedpathologists made the assessment independently

28 Western Blotting Analysis Total protein was extractedfrom fresh colorectum tissue by cell lysis buffer with proteaseand phosphatase inhibitor mixture (New Cell amp MolecularBiotech Co Ltd China) The protein quantification wasdetermined by BCA methods (Beyotime Shanghai China)Equal amount of protein (25 ug) fromeach samplewas loadedon to 12 SDS-PAGEAfter separation through electrophore-sis the proteins were transferred onto PVDF membranes(Millipore Corp Billerica MA USA) The membranes werethen incubated with 5 nonfat milk in tris-buffered saline(TBS) to block nonspecific binding with antibody Themembranes were then incubated with primary antibodies at4∘C overnight The primary antibodies used in experimentswere as follows 120573-actin PPAR-120574 NF-120581B pNF-120581B (all fromCell Signaling Technology Danvers MA USA) Bcl-2 Bax(all from Proteintech Group Inc Chicago IL USA) andcaspase 3 (Cell Signaling Technology Danvers MA USA)with dilution of 1 1000 1 1000 1 1000 1 500 1 500 and1 1000 respectively After incubation with primary antibodythe membranes were washed three times with TBST andthen incubated with the appropriate horseradish peroxidase-conjugated secondary antibodies The signals were detectedand the intensity of bands was analyzed by Odyssey

29 Immunohistochemical Staining Tissue specimen col-lected from mice was fixed in 4 paraformaldehydeovernight and embedded in paraffin Sections (4 120583m) weredewaxed and hydrated through serial graded xylene ethanolAfter being washed with phosphate-buffered saline (PBS)solution three times antigen retrieval was performed usinga microwave with heating to 95∘C and cooling to roomtemperature (four times) Primary antibody PPAR-120574 (CellSignaling Technology Danvers MA USA) was diluted intoworking concentration and then incubated with the slices at4∘C overnight Following incubation of secondary antibodyDAB substrate was used as the chromogen Positive expres-sion areas were observed under microscope

210 Statistical Analysis The statistical analysis was per-formed using the SPSS 200 software package (IBMCorporation Armonk NY USA) All values wereexpressed with the mean plusmn standard deviation using theStudentndashNewmanndashKeuls test or one-way analysis of varianceA 119901 value lt 005 was considered statistically significant Thepositively stained areas of IHC were evaluated and analyzedusing Image-Pro Plus 60 software (Media Cybernetics SilverSpring MD USA)

3 Results

31 Portulaca Extract Decreases the Disease Activity Indexthe Length of Colorectum and Myeloperoxidase Activity ofDextran Sulfate Sodium Induced Colitis Since severity of

4 PPAR Research

Table 3 Histology scoring

Lesions of colonic epithelium Granulocyte infiltrationof intestine mucosa

Granulocyte infiltrationof submucosa Score

Normal Normal Normal 0Cellular proliferationanomalous cryptabsence of goblet cell Mild Mild to moderate 1Absence of intestinal crypt (10ndash50) Moderate Severe 2Absence of intestinal crypt (50ndash90) Severe 3Entire absence of intestinal crypt with integral epithelia 4Mild to moderate ulceration epithelia 5Severe ulceration 6

colitis is evaluated by the DAI MPO and the length ofcolorectum the effects of Portulaca extract on these param-eters were examined As shown in Figure 1(b) mice in DSSgroup have significant higher DAI than those in all othergroups starting from day 3 (119901 lt 005) Mice in Portulacaextract and mesalamine groups showed higher DAI startingfrom day 3 and reaching to the peak at day 5 comparedto those in normal control After day 5 DAI was graduallydecreased although they were still higher than that in normalcontrol Since DAI consists of three aspects including weightloss diarrhea and the bloody stool scores The weight lossand bloody stool scores were further evaluated and shownin Figures 1(c) and 1(d) DSS induced ulcerative colitis andsignificantly reduced body weight of the mice in DSS group(119901 lt 001) However the treatment of Portulaca extract andmesalamine significantly restored the body weights as shownin the Portulaca and mesalamine groups In addition therewas a higher bloody stool score in DSS group than all othergroups Treatment of Portulaca and mesalamine reduced thescore (Figure 1(d))

The length of colorectum was an evaluation content ofcolitis which means the total length from cecum to anusof mice Compared with that of control group there was asignificant decrease in the length of colorectum in DSS group(119901 lt 00001) But the length of colorectum in mesalamineand Portulaca group was longer than that in DSS group(Figure 1(e))Moreover myeloperoxidase (MPO) serves as anindependent predictor for oxidative stress and inflammatoryreactionsThe average MPO activity in mice of control groupwas 056 plusmn 005 times 10minus3Uml while DSS exposure increasedtheMPO activity to 219 plusmn 082 times 10minus3UmlThe treatment ofmesalamine and Portulaca group reduced the MPO activityto 084 plusmn 011 times 10minus3Uml and 071 plusmn 004 times 10minus3Umlrespectively (Figure 1(f))

32 Effect of Portulaca Extract on Proinflammatory CytokinesExpression and Colorectum Injury Since DSS induced ulcer-ative colitis the expressions of three proinflammatorycytokines (TNF-120572 IL-6 and IL-1120573) were examined in theserum and colorectum tissue As shown in Figures 2(a) and2(b) there were significant increases in all three proinflam-matory cytokines at both mRNA and protein levels (119901 lt005) Moreover treatment of Portulaca and mesalaminesignificantly reduced the DSS-induced cytokine expression

at both mRNA and protein levels (119901 lt 005) Two inde-pendent experienced pathologists examined the tissue injuryof colorectum The injury score resulted from the evaluationof enterocyte lessons and inflammatory cell infiltration inmucosa and submucosa regions Figure 2(c) showed HEstaining of colorectum tissues from mice of four differentgroups with magnification of 200x in the upper row anddifferentmagnification of colorectum inDSSmiceThere wasa significant increase in tissue injury score in DSS mice com-pared to all other mice (119901 lt 00001) Treatment of Portulacaextract and mesalamine resulted in lesser enterocyte lesionsand inflammatory cell infiltration in mucosa and submucosa(Figures 2(c) and 2(d))

33 Regulation of NF-120581B and PPAR-120574 by Portulaca Extracts inColorectum BothNF-120581B and PPAR-120574 play an important rolein inflammation activation of NF-120581B is the critical step toincrease the expression of different inflammatory cytokinessuch as TNF-120572 Moreover previous studies documented thatPPAR-120574 could antagonize inflammatory responses throughinhibition of transcriptional activation of inflammatoryresponse genes such as transcriptional factor NF-120581B There-fore the expressions of NF-120581B and PPAR-120574 were examinedin the study Figure 3(a) showed the staining of PPAR-120574in colorectum tissues from mice of four different groupsSignificant reduction of PPAR-120574 staining was observed in thecolorectum of mice in DSS groups while treatment of eitherPortulaca extract or mesalamine restored back the stainingof PPAR-120574 in the colorectum Figure 3(b) displayed both NF-kBp65 and PPAR-120574 proteins by western blot and Figure 3(c)showed the mRNA levels of both NF-kB and PPAR-120574 Therewas significant decrease in PPAR-120574 expression at bothmRNAand protein levels in DSS group compared to that in normalcontrol group (119901 lt 005) Both Portulaca extract andmesalamine restored PPAR-120574 levels back to that in normalcontrol group DSS treatment significantly induced NF-kBexpression at bothmRNA and protein levels (119901 lt 005) whiletreatments of either Portulaca extract or mesalamine broughtNF-kB levels back to that in normal control group

34 Regulation of Apoptosis Protein in Colorectum by Por-tulaca Extract Inflammatory reaction usually results in celldeath however whether it is through apoptosis or necrosisremains to be explored In the current study three apoptoticproteinswere examined including proapoptotic proteins (Bax

PPAR Research 5

(a)D

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alam

ine(

74)NC

Seru

m M

PO (times

10minus3５

ml)

lowastlowast

(f)

Figure 1 Effect of Portulaca treatment on disease severity of DSS-induced UCmodel Panel (a) shows the appearance of representative micefrom four experiment groups from left to right control group DSS-induced groupmesalamine group and Portulaca group Panel (b) displaysthe graph of disease activity index (DAI) of four groups at different days of the experiment Panel (c) represents the weight of mice daily indifferent group Panel (d) shows the image of fecal occult blood testing at 8th day with the darker color and more severe bloody stool andshows the score of bloody stool at different days of each group Panel (e) represents the length of colorectum from four groups at 8th day afterDSS administration Panel (f) indicates the MPO activity in serum (Data were presented as mean plusmn SD from six rats lowast indicates comparisonbetween DSS and NC indicates comparison between mesalamine and DSS + indicates the comparison between Portulaca and DSS Thenumber of symbols indicates significance of difference for example four symbols indicate 119901 lt 00001 three symbols indicate 119901 lt 0001 twosymbols indicate 119901 lt 001 and one symbol indicates 119901 lt 005)

and cleaved caspase 3) and antiapoptotic protein (Bcl-2) Asshown in Figure 4 abundance of Bcl-2 Bax and caspase 3wasdocumented by western blot analyses There was significantdecrease in Bcl-2 level (119901 lt 0001) but significant increasein Bax level in colorectum of mice in DSS group comparedto all other groups (119901 lt 00001) Although whole caspase 3

showed no change in the colorectum of mice in DSS groupthe cleaved form of caspase 3 was significantly increased(119901 lt 001) All these changes were corrected after treatmentof either Portulaca extract or mesalazine Moreover mRNAlevels of Bcl-2 and Bax showed the same changes as theirproteins in the four different groups of mice

6 PPAR Research

0

2

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Relat

ive m

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of T

NF-

lowastlowastD

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NF-

а (pg

ml)

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(pg

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200x M200x DSS 200x P

200x DSS100x DSS 400x DSS

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(c)

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DSSNC


Portulaca(10

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alam

ine(

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(d)

Figure 2 Effect of Portulaca extract on serum cytokines and tissue injury scores of DSS-induced acute colitis Panel (a) indicates that themRNA expressions of TNF-120572 IL-6 and IL-1120573 were evaluated in each group with qRT- PCR Panel (b) shows that the levels of TNF-120572 IL-6and IL-1120573 in serumwere detected via ELISA assay Panel (c) displays the HampE stained colon images (NC refers to negative control DSS refersto DSS alone group M refers to mesalamine group P refers to Portulaca group) The dotted potion indicated the hyperplasia of lymphoidfollicle red arrow pointed the infiltration of inflammatory cells Panel (d) indicates the histological scores of HampE stained colorectum images(Data were presented as mean plusmn SD from six rats lowast indicates comparison between DSS and NC indicates comparison betweenmesalamineand DSS + indicates the comparison between Portulaca and DSS The number of symbols indicates significance of difference for examplefour symbols indicate 119901 lt 00001 three symbols indicate 119901 lt 0001 two symbols indicate 119901 lt 001 and one symbol indicates 119901 lt 005)

PPAR Research 7

200x P200x M200x DSS200x NC(a)

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pNF-B

PPAR-

-Actin

(b)

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+

000

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in le

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(d)

Figure 3 Effect of Portulaca extracts on NF-120581B and PPAR-120574 expression in colonic tissue Panel (a) represents the immunohistochemistrystaining of PPAR-120574 (NC refers to negative control DSS refers to DSS alone group M refers to mesalamine group P refers to Portulaca grouporiginal magnification times200) Panel (b) shows the expression of PPAR-120574 NF-120581B and pNF-120581B in colorectum tissues from western blot Panel(c) shows the histograms of grey intensity for PPAR-120574 and NF-120581B relative to 120573-actin Panel (d) shows the relative mRNA expression of PPAR-120574 and NF-120581B from quantitative real-time PCR (Data are presented as mean plusmn SD from six experiments lowast indicates comparison betweenDSS and NC indicates comparison between mesalamine and DSS + indicates the comparison between Portulaca and DSSThe number ofsymbols indicates significance of difference for example four symbols indicate 119901 lt 00001 three symbols indicate 119901 lt 0001 two symbolsindicate 119901 lt 001 and one symbol indicates 119901 lt 005)

4 Discussion

Inflammatory bowel disease commonly presents with intes-tine and extraintestine manifestations [21]The typical symp-tom of IBD clinically is the blood and mucus mixed withstool Other symptoms may include fevers weight lossarthritis mucocutaneous lesions and extraintestine presen-tations which are useful for diagnosis evaluating patientrsquoscondition and drug selection [22] In the current studyweight loss and feces changes accompanied by blood and

mucus mixed with stool were recorded daily to estimatethe DAI (disease activity index) of the colitis Significanthigh index of DAI in DSS-induced UC mice in the currentstudy indicates successful establishment of UC model inmice Moreover with this model therapeutic activity ofPortulaca extract was also demonstrated showing that micewith treatment of Portulaca extract had significant lower DAIindex than that without Portulaca treatment Furthermorewith this successful model of UC the effect of Portulacaextract on the length of colorectum was also documented

8 PPAR Research

NC DSS M P

Bcl-2

Bax

Caspase 3Cleaved

caspase 3

-Actin

(a)

Rela

tive p

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005

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DSSNC

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lowastlowastlowastlowastlowastlowast

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ax

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alam

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74)

Portulaca(10

0)

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alam

ine(

74)

(c)

Figure 4 Effects of Portulaca extract on apoptosis in mice on inflammatory bowel disease Panel (a) shows the protein level of Bcl-2 Baxcaspase 3 and cleaved caspase 3 in each group Panel (b) represents the histogram of density for Bcl-2 Bax and cleaved caspase 3 Panel (c)represents the relative mRNA level of Bcl-2 and Bax from quantitative real-time PCR respectively (Data are presented as mean plusmn SD from sixexperimentslowast indicates comparison betweenDSS andNC indicates comparison betweenmesalamine andDSS + indicates the comparisonbetween Portulaca and DSSThe number of symbols indicates significance of difference for example four symbols indicate 119901 lt 00001 threesymbols indicate 119901 lt 0001 two symbols indicate 119901 lt 001 and one symbol indicates 119901 lt 005)

Mice with DSS-induced UC showed shorter colorectum thannormal mice which is consistent with the report of UC[23] Treatment of Portulaca extract could also increase thelength of colorectum in rats with DSS-induced colitis Thefindings indicate that Portulaca extract could ameliorategeneral symptoms of IBD

Currently myeloperoxidase activity is considered as anindependent indicator of inflammation and oxidative stressThis enzyme in leukocyte usually has the capacity to cat-alyze the hydrogen peroxide [24] Clinicallymyeloperoxidaseactivity has been used to predict inflammation and oxida-tive stress in renal failure [25] myocardial infarction [26]

inflammatory vascular disease [27] and so on Consistentwith these reports higher level ofMPOwas observed inmiceofDSS treatment which induces inflammation and ulcerativecolitis in the intestine [28] This elevation of MPO in serumis also consistent with intestine injury score observed in thecolorectum tissues especially the infiltration of inflammatorycells in the mucosa and submucosa These findings suggestthatMPOcould be an indicator for eitherDSS-induced colitisor clinical presentation of IBD

Several immune cells and immune-regulatory proteinsparticipated in the disturbance of intestine immunesystem which account for activation and augmentation of

PPAR Research 9

inflammation cascade in IBD The intestinal epithelial cellsusually act as the first barrier for any intestinal disorders[29] Once the epithelial cells are injured by external stressintestine pathogens may enter the intestine which triggersantigen-presenting cells and transform naive T cells intoeffector T cells such as Th1 Th17 Th2 and natural killer TcellsThese cells are responsible for generating multiple typesof proinflammatory cytokines [30] Moreover these cellscould secret interferon 120574 interleukin 1120573 and other cytokinesto activate macrophages which in turn could secrete largeamounts of cytokines including tumor necrosis factor 120572interleukin 1 and interleukin 6 Furthermore several celltypes in the process such as dendritic cells mast cells andleucocytes can also promote the dysfunction of innate andadaptive immune response of inflammatory disease [31]In current study we found application of Portulaca couldsignificantly reduce the expression levels of these cytokinesincluding TNF-120572 IL-6 and IL-1120573 These findings suggestedthat Portulaca may affect T cell activation and participate ininflammation to some extent

In present study three proinflammatory cytokines (TNF-120572 IL-6 and IL-1120573) were examined These cytokines aremainly released by macrophages and differentiated T cells[32] In addition TNF-120572 could also exert its inflammationresponse through upregulation of IL-1120573 and IL-6 and inducetissue damage through necrosis and apoptosis [33] andactivate NF-120581B [34] Moreover all three cytokines are ableto exert other functions by interacting with other moleculesthrough different pathway The findings of TNF-120572 IL-1120573and IL-6 in the current study are in line with the reports inpatients with either UC or CD [35 36]

Except cytokines transcriptional factors are also impor-tant in regulation of inflammationThe transcriptional factorNF-120581B is known to regulate the level of TNF-120572 IL-6 and IL-1120573during the progression of inflammatory bowel disease [37]Moreover NF-120581B can facilitate cell apoptosis through theupregulation of c-FLIP level while lowering the expressionof antiapoptotic proteins such as Bcl-2 Bcl-xl c-IAP12and x-IAP [38] In the present study overexpressed TNF-120572 IL-6 IL-1120573 and pNF-120581B were observed in DSS treatedmice which indicate an enhanced inflammatory process andaggravated tissue damage On the contrary the addition ofPortulaca extract reduced inflammation response and tissuedamage indicating a role of Portulaca extract in preventionand treatment of inflammatory bowel disease

PPAR-120574 is another transcription factor which plays animportant role in adipogenesis glucose metabolism [39]and anti-inflammatory reaction Several reports suggest thatPPAR-120574 agonists may have beneficial effects for the inflam-matory diseases including mastitis IBD and arthritis [40ndash42] Jiang et al observed that PPAR agonist (15d-PGJ

2) could

abrogate the production of proinflammatory cytokines TNF-120572 and interleukin-6 [43] Moreover the NF-120581B activationis inhibited after PPAR-120574 agonist due to changes of confor-mation in NF-kB subunits The findings from the currentstudy also observed that PPAR-120574 level was increased inDSS-induced group following Portulaca extract interventiontogether with suppressed cytokines and NF-120581B

The mechanism of apoptosis in IBD remains contro-versial Some studies observed an increased apoptosis inlamina propria (LP) and epithelium which contributed to theseverity of UC [44] Moreover activation of PPAR-120574 couldattenuate the cascade of apoptosis [45] In the current studyantiapoptosis protein (Bcl-2) was decreased and proapoptosisproteins (Bax and cleaved caspase 3) were increased inUC mice However treatment of Portulaca extract reversesapoptosis in the colorectum This finding suggests furtherinvestigation of apoptosis and Portulaca in inflammatorybowel disease

5 Conclusion

With the successful model of inflammatory bowel diseasePortulaca extract was demonstrated to reduce severity of thedisease The mechanism of Portulaca extract in alleviationof inflammatory bowel disease could be related to its regu-lation of inflammatory cytokines (TNF-120572 IL-6 and IL-1120573)transcription factors (NF-kB and PPAR-120574) and apoptosisTherefore Portulaca extractmight be a potential agent for thetreatment of patients with inflammatory bowel disease

Conflicts of Interest

The authors declare that there are no conflicts of interestregarding the publication of this paper

Authorsrsquo Contributions

Rui Kong makes main contribution to this work

Acknowledgments

Theauthors would like to sincerely thankDr Jie Lu ProfessorYuejuan Zheng and Dr Qi Dai and the staff of the CentralLaboratory of Shanghai Tenth Peoplersquos Hospital for theirtechnical assistanceThis studywas supported by theNationalNature Science Foundation of China (81300340 81471537)

References

[1] MD Kappelman S L Rifas-Shiman C Q Porter et al ldquoDirecthealth care costs of Crohnrsquos disease and ulcerative colitis in USchildren and adultsrdquo Gastroenterology vol 135 no 6 pp 1907ndash1913 2008

[2] S C Ng C N Bernstein M H Vatn et al ldquoGeographicalvariability and environmental risk factors in inflammatorybowel diseaserdquo Gut vol 62 no 4 pp 630ndash649 2013

[3] B A Hendrickson R Gokhale and J H Cho ldquoClinical aspectsand pathophysiology of inflammatory bowel diseaserdquo ClinicalMicrobiology Reviews vol 15 no 1 pp 79ndash94 2002

[4] M K Uddin A S Juraimi M E Ali and M R IsmailldquoEvaluation of antioxidant properties and mineral compositionof purslane (Portulaca oleracea L) at different growth stagesrdquoInternational Journal of Molecular Sciences vol 13 no 8 pp10257ndash10267 2012

10 PPAR Research

[5] C-J ChenW-YWang X-LWang et al ldquoAnti-hypoxic activityof the ethanol extract from Portulaca oleracea in micerdquo Journalof Ethnopharmacology vol 124 no 2 pp 246ndash250 2009

[6] L F Hu X Y Xu and B Q Wang ldquoResearch and utilizationsituation of PortulacaOleracea L inChinardquo Journal of PharmacyPractice and Community Medicine vol 20 pp 315-316 2003

[7] L Xiang D Xing W Wang R Wang Y Ding and L DuldquoAlkaloids from Portulaca oleracea Lrdquo Phytochemistry vol 66no 21 pp 2595ndash2601 2005

[8] A N Rashed F U Afifi and A M Disi ldquoSimple evaluationof the wound healing activity of a crude extract of Portulacaoleracea L (growing in Jordan) inMusmusculus JVI-1rdquo Journalof Ethnopharmacology vol 88 no 2-3 pp 131ndash136 2003

[9] X J Zhang Y B Ji QuZh Y and et al ldquoExperimental studieson antibiotic functions of Portulacaoleracea L in vitrordquoChineseJournal of Microbiology and Immunology vol 14 pp 277ndash2802002

[10] W Wang L Gu L Dong X Wang C Ling and M LildquoProtective effect of Portulacaoleracea extracts on hypoxicnerve tissue and its mechanismrdquo Asia Pacific Journal of ClinicalNutrition pp 227ndash233 2007

[11] K Y Musa A Ahmed G Ibrahim et al ldquoToxicity studieson the methanolic extract of Portulaca oleracea L (FamPortulacaceae)rdquo Journal of Biological Sciences vol 7 no 7 pp1293ndash1295 2007

[12] A T Reddy S P Lakshmi andRC Reddy ldquoPPAR 120574 in BacterialInfections A Friend or Foerdquo PPAR Research vol 2016 ArticleID 7963540 2016

[13] T DHinds K John LMcBeth C J Trabbic and E R SanchezldquoTimcodar (VX-853) Is a Non-FKBP12 Binding MacrolideDerivativeThat Inhibits PPAR 120574 and Suppresses AdipogenesisrdquoPPAR Research vol 2016 Article ID 6218637 2016

[14] G D Demetri C D Fletcher E Mueller et al ldquoInductionof solid tumor differentiation by the peroxisome proliferator-activated receptor-120574 ligand troglitazone in patients with liposar-comardquo Proceedings of the National Acadamy of Sciences of theUnited States of America vol 96 no 7 pp 3951ndash3956 1999

[15] T Kubota K Koshizuka E A Williamson and et al ldquoLigandfor peroxisome proliferator-activated receptor gamma (trogli-tazone) has potent antitumor effect against human prostatecancer both in vitro and in vivordquo Cancer Research vol 58 no15 pp 3344ndash3352 1998

[16] S P Lakshmi A T Reddy A Banno and R C Reddy ldquoPPARAgonists for the Prevention and Treatment of Lung CancerrdquoPPAR Research vol 2017 pp 1ndash8 2017

[17] G Chinetti S Lestavel V Bocher and et al ldquoPPAR-alphaand PPAR-gamma activators induce cholesterol removal fromhuman macrophage foam cells through stimulation of theABCA1 pathwayrdquoNature Medicine vol 7 no 1 pp 53ndash58 2001

[18] L F da Rocha Junior A T Dantas A L B P Duarte M JB de Melo Rego I D R Pitta and M G D R Pitta ldquoPPAR120574agonists in adaptive immunity what do immune disorders andtheir models have to tell usrdquo PPAR Research vol 2013 ArticleID 519724 9 pages 2013

[19] M Ricote A C Li T M Willson C J Kelly and C K GlassldquoThe peroxisome proliferator-activated receptor-gamma is anegative regulator of macrophage activationrdquo Nature vol 391no 6662 pp 79ndash82 1998

[20] W Hozayen M Bastawy and H Elshafeey ldquoEffects of AqueousPurslane (PortulacaOleracea) Extractand Fish Oil on Gentam-icin Nephrotoxicity in Albino RatsrdquoNatural Sciences vol 9 pp47ndash62 2011

[21] A N Ananthakrishnan E L McGinley and D G BinionldquoDoes it matter where you are hospitalized for inflammatorybowel disease A nationwide analysis of hospital volumerdquoAmerican Journal of Gastroenterology vol 103 no 11 pp 2789ndash2798 2008

[22] D Bojic Z Radojicic M Nedeljkovic-Protic M Al-Ali D PJewell and S P L Travis ldquoLong-term outcome after admissionfor acute severe ulcerative colitis in Oxford The 1992-1993cohortrdquo Inflammatory Bowel Diseases vol 15 no 6 pp 823ndash828 2009

[23] S Kang Y Jeon K Moon et al Journal of Medicinal Food vol20 no 7 pp 667ndash675 2017

[24] D Ristovski-Kornic A Stefanovic J Kotur-Stevuljevic et alldquoAssociation of Myeloperoxidase and the Atherogenic Index ofPlasma in Children with End-Stage Renal Diseaserdquo Journal ofMedical Biochemistry vol 36 no 1 pp 23ndash31 2017

[25] A Ece F Gurkan M Kervancıoglu et al ldquoOxidative stressinflammation and early cardiovascular damage in children withchronic renal failurerdquo Pediatric Nephrology vol 21 no 4 pp545ndash552 2006

[26] E Cavusoglu C Ruwende C Eng et al ldquoUsefulness of baselineplasma myeloperoxidase levels as an independent predictor ofmyocardial infarction at two years in patients presenting withacute coronary syndromerdquoAmerican Journal of Cardiology vol99 no 10 pp 1364ndash1368 2007

[27] D Lau and S Baldus ldquoMyeloperoxidase and its contributoryrole in inflammatory vascular diseaserdquo Pharmacology amp Ther-apeutics vol 111 no 1 pp 16ndash26 2006

[28] L Zhang Y ZhangW Zhong C Di X Lin and Z Xia ldquoHemeoxygenase-1 ameliorates dextran sulfate sodium-induced acutemurine colitis by regulatingTh17Treg cell balancerdquoThe Journalof Biological Chemistry vol 289 no 39 pp 26847ndash26858 2014

[29] D C Baumgart and A U Dignass ldquoIntestinal barrier functionrdquoCurrent Opinion in Clinical Nutrition amp Metabolic Care vol 5no 6 pp 685ndash694 2002

[30] M A Degli-Esposti and M J Smyth ldquoClose encounters ofdifferent kinds dendritic cells and NK cells take centre stagerdquoNature Reviews Immunology vol 5 no 2 pp 112ndash124 2005

[31] D Gomez N Munoz R Guerrero O Acosta and C AGuerrero ldquoPPAR 120574 agonists as an anti-inflammatory treatmentinhibiting rotavirus infection of small intestinal villirdquo PPARResearch vol 2016 Article ID 4049373 2016

[32] H Baumann and J Gauldie ldquoThe acute phase responserdquo Trendsin Immunology vol 15 no 2 pp 74ndash80 1994

[33] B Begue H Wajant J-C Bambou et al ldquoImplication of TNF-related apoptosis-inducing ligand in inflammatory intestinalepithelial lesionsrdquo Gastroenterology vol 130 no 7 pp 1962ndash1974 2006

[34] M N Ince and D E Elliott ldquoImmunologic and MolecularMechanisms in Inflammatory Bowel Diseaserdquo Surgical Clinicsof North America vol 87 no 3 pp 681ndash696 2007

[35] J Bauditz S Wedel and H Lochs ldquoThalidomide reducestumour necrosis factor-120572 and interleukin 12 production inpatients with chronic active Crohnrsquos diseaserdquo Gut vol 50 no2 pp 196ndash200 2002

[36] S C Ng S Plamondon M A Kamm et al ldquoImmunosup-pressive effects via human intestinal dendritic cells of probioticbacteria and steroids in the treatment of acute ulcerative colitisrdquoInflammatory Bowel Diseases vol 16 no 8 pp 1286ndash1298 2010

[37] Y Liu J Peng T Sun and et al ldquoEpithelial EZH2 serves asan epigenetic determinant in experimental colitis by inhibiting

PPAR Research 11

TNFalpha-mediated inflammation and apoptosisrdquo in Proceed-ings of the National Academy of Sciences pp E3796ndashE3805 2017

[38] R Ravi G C Bedi L W Engstrom et al ldquoRegulation of deathreceptor expression and TRAILApo2L-induced apoptosis byNF-120581Brdquo Nature Cell Biology vol 3 no 4 pp 409ndash416 2001

[39] Y Chen HMa D Zhu et al ldquoDiscovery of Novel Insulin Sensi-tizers Promising Approaches and Targetsrdquo PPAR Research vol2017 pp 1ndash13 2017

[40] F Rosa J S Osorio E Trevisi F Yanqui-Rivera C T Estilland M Bionaz ldquo24-Thiazolidinedione Treatment Improvesthe Innate Immune Response in Dairy Goats with InducedSubclinical Mastitisrdquo PPAR Research vol 2017 pp 1ndash22 2017

[41] L Dubuquoy C Bourdon M Peuchmaur and et al ldquoPerox-isome proliferator-activated receptor (PPAR) gamma a newtarget for the treatment of inflammatory bowel diseaserdquo Gas-troenterologie Clinique Et Biologique vol 24 no 8-9 pp 719ndash724 2000

[42] Y Kawahito M Kondo Y Tsubouchi et al ldquo15-deoxy-Δ1214-PGJ2 induces synoviocyte apoptosis and suppresses adjuvant-induced arthritis in ratsrdquo The Journal of Clinical Investigationvol 106 no 2 pp 189ndash197 2000

[43] C Jiang A T Ting and B Seed ldquoPPAR-120574 agonists inhibitproduction of monocyte inflammatory cytokinesrdquo Nature vol391 no 6662 pp 82ndash86 1998

[44] H S P Souza C J A Tortori M T L Castelo-Brancoet al ldquoApoptosis in the intestinal mucosa of patients withinflammatory bowel disease evidence of altered expression ofFasL and perforin cytotoxic pathwaysrdquo International Journal ofColorectal Disease vol 20 no 3 pp 277ndash286 2005

[45] R Sreedhar S Arumugam V Karuppagounder et al ldquoJumi-haidokuto effectively inhibits colon inflammation and apoptosisin mice with acute colitisrdquo International Immunopharmacologyvol 29 no 2 pp 957ndash963 2015

Stem Cells International

Hindawiwwwhindawicom Volume 2018


MEDIATORSINFLAMMATION

of

EndocrinologyInternational Journal of



Disease Markers


BioMed Research International

OncologyJournal of



Oxidative Medicine and Cellular Longevity


PPAR Research

Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom

The Scientific World Journal

Volume 2018

Immunology ResearchHindawiwwwhindawicom Volume 2018

Journal of

ObesityJournal of



Computational and Mathematical Methods in Medicine


Behavioural Neurology

OphthalmologyJournal of


Diabetes ResearchJournal of



Research and TreatmentAIDS


Gastroenterology Research and Practice


Parkinsonrsquos Disease

Evidence-Based Complementary andAlternative Medicine

Volume 2018Hindawiwwwhindawicom

Submit your manuscripts atwwwhindawicom

2 PPAR Research

biological functions such as antibacteria neuroprotectionanti-inflammation antiatherosclerosis and antitumor [6ndash10] Furthermore decoction of Portulaca has been usedas a remedy for inflammatory bowel disease to alleviatesymptoms of bloody stools diarrhea and abdominal pain inclinic Several clinical trials have been conducted on the safetyand effectiveness of Portulaca which showed no adverseeffects of Portulaca Reports from animal studies also indicatelow toxicity of Portulaca with LD

50value of 1853mgkg and

high therapeutic index [11]Peroxisome proliferator-activated receptor-120574 (PPAR-120574) is

an important member of the nuclear receptor superfamilyThe essential role of PPAR-120574 in acute inflammation [12]adipogenesis [13] and several carcinomas has been iden-tified [14ndash16] Moreover PPAR-120574 activation was found invarious types of immune cells including primary peritonealmacrophages dendritic cells and T cells [17] Several studieshave shown that activation of PPAR-120574 could downregulatesome vital proinflammatory cytokines such as IL-1120573 IL-2IL-6 and TNF-120572 [18] as well as inhibition of NF-120581B phos-phorylation in monocyte and other types of cells Thereforeactivation of PPAR-120574 exerts a direct anti-inflammatory effectin IBD while the mechanism of PPAR-120574 activation remainsto be further explored [19]

In the current study the anti-inflammatory activity andmechanism of Portulaca in dextran sulfate sodium (DSS)induced UC animal model were explored

2 Materials and Methods

21 Plant Material and Preparation of the Extracts ThePortulaca material was obtained from the Department ofTraditional Chinese Medicine Pharmacy of the ShanghaiTenth Peoplersquos Hospital The herb was triturated into pow-der and water-soluble substance was extracted as describedpreviously [20] Briefly the prepared powder was boiled indistilled water (80∘C 25 g120ml water wv) for 60min andthen smashed via ultrasonic concussion (3 cycles of 15 sec)Themixture of water and concussion was filtered using 2mmpores strainer and the resultant material was lyophilizedunder vacuum

22 Animals and Acute Colitis Induction Forty adult femalemice weighting 18ndash22 g were purchased from the Shang-hai Laboratory Animal Co Ltd (Shanghai China) andmaintained in an environment of 25∘C with controlled 12 hlightdark cycle Mice were permitted free access to waterand standardmouse chowThe animal protocol was approvedby the Ethics Committee of the Tongji University whichmeets the recommendations in the Guide for Care and Useof Laboratory Animals of the National Institutes of HealthThe dextran sulfate sodium reagent (molecular weight36000ndash50000 fromMP Biochemicals Solon OH USA) wasdiluted with drinking water to make final concentration of3

The mice were divided into 4 groups at random

(1) Normal control given normal food and water for 7days

Table 1 DAI assessment (disease activity index)

Weightloss Shape of stool FOBTlowast

blood stool Score

0 Normal Normal 01ndash5 Shapeless stool FOBT positive 16ndash10 211ndash15 Loose stool Gross blood stool 3gt16 4lowastFOBT fecal occult blood test

(2) DSS group 3 DSS + saline by gavage for 7 days(3) Mesalamine positive control group 3 DSS +

74mgkg mesalamine daily by gavage for 7 days(4) Portulaca group 3 DSS + 100mgkg daily by gavage

for 7 days

During the period of modeling mice weight stool formand stool occult blood were recorded every day to assess thedisease activity index (DAI) of acute colitis the evaluativecriteria were clarified in Table 1

23 SerumCytokine Analysis The serum levels of TNF-120572 IL-1120573 and IL-6 concentrations were determined by using ELISAKits (Elabscience Wuhan China) respectively according tothe manufacturerrsquos instructionThe OD values of absorbanceat 450 nm were examined byThermoMax microplate reader

24 Determination of MPO (Myeloperoxidase) Activity inSerum Sample Myeloperoxidase is found in activated neu-trophils and macrophages which plays a defensive role dur-ing inflammatory response via catalyzing hydrogen peroxideinto hypochlorous acid The serum samples reacted withthe mixed solution including 331015840-dimethoxybiphenyl-441015840-diamine and hydrogen peroxide pH 60 according to theMPO assay kit (Nanjing Jiancheng Bioengineering InstituteA044 China)The absorbance of control well and testing wellat 460 nm was determined by microplate reader The activityof MPO was calculated as the following formula

MPO performance (UL)

=(testing OD values minus control OD values)

113

times sample volume (L)

(1)

25 Fecal Occult Blood Testing The form of stool wasrecorded and the severity of stool occult blood was analyzedby urine fecal occult blood test kit (Nanjing JianchengBioengineering Institute C027 China) A small amount offeces samples was picked and smeared on the slides Addorthotolidine and hydrogen peroxide reagent on the surfaceof stool samples The results were analyzed according to theindications negative (minus) the samples do not show colorwithin 3 minutes weakly positive (+) the samples show bluewithin 30 to 60 seconds positive (++) the samples appear

PPAR Research 3



(∘C)GADPH
















3 Results


4 PPAR Research











PPAR Research 5

(a)D

AI i

ndex

0

1

2

3

4

NCDSS

MesalaminePortulaca

day

7

day

6

day

5

day

4

day

3

day

2

day

1

lowast

+

(b)NCDSS

Wei

ght (

g)

10

15

20

25

lowastlowast

+

MesalaminePortulaca

day

7

day

6

day

5

day

4

day

3

day

2

day

1

(c)

Bloo

dy sc

ore

minus1

0

1

2

3

4

5

+

lowast

NCDSS

MesalaminePortulaca

day

7

day

6

day

5

day

4

day

3

day

2

day

1

(d)

Leng

th o

f col

orec

tum

(cm

)

0

5

10

15

++

Portulaca(10

0)

DSS

Mes

alam

ine(

74)NC


(e)

0

1

2

3

4

++Portulaca(10

0)

DSS

Mes

alam

ine(

74)NC

Seru

m M

PO (times

10minus3５

ml)

lowastlowast

(f)




6 PPAR Research

0

2

4

6

8

10

+

Relat

ive m

RNA

of T

NF-

lowastlowastD

SSNC

Relat

ive m

RNA

of I

L-6

0

5

10

15

+

lowastlowast

DSSNC

0

5

10

15

20

25

+Relat

ive m

RNA

of I

L-1

lowast

DSSNC

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(a)

Seru

m T

NF-

а (pg

ml)

0

5

10

15

20

++

lowastlowast

DSSNC

0

50

100

Seru

m IL

-6 (p

gm

l)

+++

lowastlowastlowast

DSSNC

0

5

10

15

20

25

+

Seru

m IL

-1

(pg

ml) lowastlowast

DSSNC

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(b)

200x M200x DSS 200x P


200x NC

(c)

Hist

olog

ical

scor

e

0

5

10

15

++++

DSSNC


Portulaca(10

0)

Mes

alam

ine(

74)

(d)


PPAR Research 7


NC DSS M P

NF-B

pNF-B

PPAR-

-Actin

(b)

000

001

002

003

004

+

000

001

002

003

004

++

Relat

ive p

rote

in le

vel o

fPP

AR

Relat

ive p

rote

in le

vel

of N

F

DSSNC

DSSNC

lowast

lowastlowast

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(c)

00

05

10

15

20

++D

SSNC

DSSNC

lowast


Relat

ive m

RNA

of N

FB

0

1

2

3

4

5

+ +

Relat

ive m

RNA

of P

PAR-

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(d)


4 Discussion



8 PPAR Research

NC DSS M P

Bcl-2

Bax

Caspase 3Cleaved

caspase 3

-Actin

(a)

Rela

tive p

rote

inle

vel o

f Bax

000

005

010

015

+

Rela

tive p

rote

inle

vel o

f Bcl-

2

000

002

004

006

008+

Relat

ive p

rote

in le

vel o

fcle

aved

Cas

pase

3

00

01

02

03

04

++++

DSSNC

DSSNC

DSSNC


lowastlowastlowast

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(b)

Relat

ive m

RNA

of B

ax

00

05

10

15

20

25

+++

Relat

ive m

RNA

of B

cl-2

0

2

4

6

++


DSSNC

DSSNC

lowast

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(c)






PPAR Research 9





2) could



5 Conclusion






Acknowledgments


References





10 PPAR Research


































PPAR Research 11














of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of



















PPAR Research 3



(∘C)GADPH
















3 Results


4 PPAR Research











PPAR Research 5

(a)D

AI i

ndex

0

1

2

3

4

NCDSS

MesalaminePortulaca

day

7

day

6

day

5

day

4

day

3

day

2

day

1

lowast

+

(b)NCDSS

Wei

ght (

g)

10

15

20

25

lowastlowast

+

MesalaminePortulaca

day

7

day

6

day

5

day

4

day

3

day

2

day

1

(c)

Bloo

dy sc

ore

minus1

0

1

2

3

4

5

+

lowast

NCDSS

MesalaminePortulaca

day

7

day

6

day

5

day

4

day

3

day

2

day

1

(d)

Leng

th o

f col

orec

tum

(cm

)

0

5

10

15

++

Portulaca(10

0)

DSS

Mes

alam

ine(

74)NC


(e)

0

1

2

3

4

++Portulaca(10

0)

DSS

Mes

alam

ine(

74)NC

Seru

m M

PO (times

10minus3５

ml)

lowastlowast

(f)




6 PPAR Research

0

2

4

6

8

10

+

Relat

ive m

RNA

of T

NF-

lowastlowastD

SSNC

Relat

ive m

RNA

of I

L-6

0

5

10

15

+

lowastlowast

DSSNC

0

5

10

15

20

25

+Relat

ive m

RNA

of I

L-1

lowast

DSSNC

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(a)

Seru

m T

NF-

а (pg

ml)

0

5

10

15

20

++

lowastlowast

DSSNC

0

50

100

Seru

m IL

-6 (p

gm

l)

+++

lowastlowastlowast

DSSNC

0

5

10

15

20

25

+

Seru

m IL

-1

(pg

ml) lowastlowast

DSSNC

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(b)

200x M200x DSS 200x P


200x NC

(c)

Hist

olog

ical

scor

e

0

5

10

15

++++

DSSNC


Portulaca(10

0)

Mes

alam

ine(

74)

(d)


PPAR Research 7


NC DSS M P

NF-B

pNF-B

PPAR-

-Actin

(b)

000

001

002

003

004

+

000

001

002

003

004

++

Relat

ive p

rote

in le

vel o

fPP

AR

Relat

ive p

rote

in le

vel

of N

F

DSSNC

DSSNC

lowast

lowastlowast

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(c)

00

05

10

15

20

++D

SSNC

DSSNC

lowast


Relat

ive m

RNA

of N

FB

0

1

2

3

4

5

+ +

Relat

ive m

RNA

of P

PAR-

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(d)


4 Discussion



8 PPAR Research

NC DSS M P

Bcl-2

Bax

Caspase 3Cleaved

caspase 3

-Actin

(a)

Rela

tive p

rote

inle

vel o

f Bax

000

005

010

015

+

Rela

tive p

rote

inle

vel o

f Bcl-

2

000

002

004

006

008+

Relat

ive p

rote

in le

vel o

fcle

aved

Cas

pase

3

00

01

02

03

04

++++

DSSNC

DSSNC

DSSNC


lowastlowastlowast

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(b)

Relat

ive m

RNA

of B

ax

00

05

10

15

20

25

+++

Relat

ive m

RNA

of B

cl-2

0

2

4

6

++


DSSNC

DSSNC

lowast

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(c)






PPAR Research 9





2) could



5 Conclusion






Acknowledgments


References





10 PPAR Research


































PPAR Research 11














of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of



















4 PPAR Research











PPAR Research 5

(a)D

AI i

ndex

0

1

2

3

4

NCDSS

MesalaminePortulaca

day

7

day

6

day

5

day

4

day

3

day

2

day

1

lowast

+

(b)NCDSS

Wei

ght (

g)

10

15

20

25

lowastlowast

+

MesalaminePortulaca

day

7

day

6

day

5

day

4

day

3

day

2

day

1

(c)

Bloo

dy sc

ore

minus1

0

1

2

3

4

5

+

lowast

NCDSS

MesalaminePortulaca

day

7

day

6

day

5

day

4

day

3

day

2

day

1

(d)

Leng

th o

f col

orec

tum

(cm

)

0

5

10

15

++

Portulaca(10

0)

DSS

Mes

alam

ine(

74)NC


(e)

0

1

2

3

4

++Portulaca(10

0)

DSS

Mes

alam

ine(

74)NC

Seru

m M

PO (times

10minus3５

ml)

lowastlowast

(f)




6 PPAR Research

0

2

4

6

8

10

+

Relat

ive m

RNA

of T

NF-

lowastlowastD

SSNC

Relat

ive m

RNA

of I

L-6

0

5

10

15

+

lowastlowast

DSSNC

0

5

10

15

20

25

+Relat

ive m

RNA

of I

L-1

lowast

DSSNC

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(a)

Seru

m T

NF-

а (pg

ml)

0

5

10

15

20

++

lowastlowast

DSSNC

0

50

100

Seru

m IL

-6 (p

gm

l)

+++

lowastlowastlowast

DSSNC

0

5

10

15

20

25

+

Seru

m IL

-1

(pg

ml) lowastlowast

DSSNC

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(b)

200x M200x DSS 200x P


200x NC

(c)

Hist

olog

ical

scor

e

0

5

10

15

++++

DSSNC


Portulaca(10

0)

Mes

alam

ine(

74)

(d)


PPAR Research 7


NC DSS M P

NF-B

pNF-B

PPAR-

-Actin

(b)

000

001

002

003

004

+

000

001

002

003

004

++

Relat

ive p

rote

in le

vel o

fPP

AR

Relat

ive p

rote

in le

vel

of N

F

DSSNC

DSSNC

lowast

lowastlowast

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(c)

00

05

10

15

20

++D

SSNC

DSSNC

lowast


Relat

ive m

RNA

of N

FB

0

1

2

3

4

5

+ +

Relat

ive m

RNA

of P

PAR-

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(d)


4 Discussion



8 PPAR Research

NC DSS M P

Bcl-2

Bax

Caspase 3Cleaved

caspase 3

-Actin

(a)

Rela

tive p

rote

inle

vel o

f Bax

000

005

010

015

+

Rela

tive p

rote

inle

vel o

f Bcl-

2

000

002

004

006

008+

Relat

ive p

rote

in le

vel o

fcle

aved

Cas

pase

3

00

01

02

03

04

++++

DSSNC

DSSNC

DSSNC


lowastlowastlowast

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(b)

Relat

ive m

RNA

of B

ax

00

05

10

15

20

25

+++

Relat

ive m

RNA

of B

cl-2

0

2

4

6

++


DSSNC

DSSNC

lowast

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(c)






PPAR Research 9





2) could



5 Conclusion






Acknowledgments


References





10 PPAR Research


































PPAR Research 11














of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of



















PPAR Research 5

(a)D

AI i

ndex

0

1

2

3

4

NCDSS

MesalaminePortulaca

day

7

day

6

day

5

day

4

day

3

day

2

day

1

lowast

+

(b)NCDSS

Wei

ght (

g)

10

15

20

25

lowastlowast

+

MesalaminePortulaca

day

7

day

6

day

5

day

4

day

3

day

2

day

1

(c)

Bloo

dy sc

ore

minus1

0

1

2

3

4

5

+

lowast

NCDSS

MesalaminePortulaca

day

7

day

6

day

5

day

4

day

3

day

2

day

1

(d)

Leng

th o

f col

orec

tum

(cm

)

0

5

10

15

++

Portulaca(10

0)

DSS

Mes

alam

ine(

74)NC


(e)

0

1

2

3

4

++Portulaca(10

0)

DSS

Mes

alam

ine(

74)NC

Seru

m M

PO (times

10minus3５

ml)

lowastlowast

(f)




6 PPAR Research

0

2

4

6

8

10

+

Relat

ive m

RNA

of T

NF-

lowastlowastD

SSNC

Relat

ive m

RNA

of I

L-6

0

5

10

15

+

lowastlowast

DSSNC

0

5

10

15

20

25

+Relat

ive m

RNA

of I

L-1

lowast

DSSNC

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(a)

Seru

m T

NF-

а (pg

ml)

0

5

10

15

20

++

lowastlowast

DSSNC

0

50

100

Seru

m IL

-6 (p

gm

l)

+++

lowastlowastlowast

DSSNC

0

5

10

15

20

25

+

Seru

m IL

-1

(pg

ml) lowastlowast

DSSNC

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(b)

200x M200x DSS 200x P


200x NC

(c)

Hist

olog

ical

scor

e

0

5

10

15

++++

DSSNC


Portulaca(10

0)

Mes

alam

ine(

74)

(d)


PPAR Research 7


NC DSS M P

NF-B

pNF-B

PPAR-

-Actin

(b)

000

001

002

003

004

+

000

001

002

003

004

++

Relat

ive p

rote

in le

vel o

fPP

AR

Relat

ive p

rote

in le

vel

of N

F

DSSNC

DSSNC

lowast

lowastlowast

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(c)

00

05

10

15

20

++D

SSNC

DSSNC

lowast


Relat

ive m

RNA

of N

FB

0

1

2

3

4

5

+ +

Relat

ive m

RNA

of P

PAR-

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(d)


4 Discussion



8 PPAR Research

NC DSS M P

Bcl-2

Bax

Caspase 3Cleaved

caspase 3

-Actin

(a)

Rela

tive p

rote

inle

vel o

f Bax

000

005

010

015

+

Rela

tive p

rote

inle

vel o

f Bcl-

2

000

002

004

006

008+

Relat

ive p

rote

in le

vel o

fcle

aved

Cas

pase

3

00

01

02

03

04

++++

DSSNC

DSSNC

DSSNC


lowastlowastlowast

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(b)

Relat

ive m

RNA

of B

ax

00

05

10

15

20

25

+++

Relat

ive m

RNA

of B

cl-2

0

2

4

6

++


DSSNC

DSSNC

lowast

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(c)






PPAR Research 9





2) could



5 Conclusion






Acknowledgments


References





10 PPAR Research


































PPAR Research 11














of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of



















6 PPAR Research

0

2

4

6

8

10

+

Relat

ive m

RNA

of T

NF-

lowastlowastD

SSNC

Relat

ive m

RNA

of I

L-6

0

5

10

15

+

lowastlowast

DSSNC

0

5

10

15

20

25

+Relat

ive m

RNA

of I

L-1

lowast

DSSNC

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(a)

Seru

m T

NF-

а (pg

ml)

0

5

10

15

20

++

lowastlowast

DSSNC

0

50

100

Seru

m IL

-6 (p

gm

l)

+++

lowastlowastlowast

DSSNC

0

5

10

15

20

25

+

Seru

m IL

-1

(pg

ml) lowastlowast

DSSNC

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(b)

200x M200x DSS 200x P


200x NC

(c)

Hist

olog

ical

scor

e

0

5

10

15

++++

DSSNC


Portulaca(10

0)

Mes

alam

ine(

74)

(d)


PPAR Research 7


NC DSS M P

NF-B

pNF-B

PPAR-

-Actin

(b)

000

001

002

003

004

+

000

001

002

003

004

++

Relat

ive p

rote

in le

vel o

fPP

AR

Relat

ive p

rote

in le

vel

of N

F

DSSNC

DSSNC

lowast

lowastlowast

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(c)

00

05

10

15

20

++D

SSNC

DSSNC

lowast


Relat

ive m

RNA

of N

FB

0

1

2

3

4

5

+ +

Relat

ive m

RNA

of P

PAR-

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(d)


4 Discussion



8 PPAR Research

NC DSS M P

Bcl-2

Bax

Caspase 3Cleaved

caspase 3

-Actin

(a)

Rela

tive p

rote

inle

vel o

f Bax

000

005

010

015

+

Rela

tive p

rote

inle

vel o

f Bcl-

2

000

002

004

006

008+

Relat

ive p

rote

in le

vel o

fcle

aved

Cas

pase

3

00

01

02

03

04

++++

DSSNC

DSSNC

DSSNC


lowastlowastlowast

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(b)

Relat

ive m

RNA

of B

ax

00

05

10

15

20

25

+++

Relat

ive m

RNA

of B

cl-2

0

2

4

6

++


DSSNC

DSSNC

lowast

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(c)






PPAR Research 9





2) could



5 Conclusion






Acknowledgments


References





10 PPAR Research


































PPAR Research 11














of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of



















PPAR Research 7


NC DSS M P

NF-B

pNF-B

PPAR-

-Actin

(b)

000

001

002

003

004

+

000

001

002

003

004

++

Relat

ive p

rote

in le

vel o

fPP

AR

Relat

ive p

rote

in le

vel

of N

F

DSSNC

DSSNC

lowast

lowastlowast

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(c)

00

05

10

15

20

++D

SSNC

DSSNC

lowast


Relat

ive m

RNA

of N

FB

0

1

2

3

4

5

+ +

Relat

ive m

RNA

of P

PAR-

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(d)


4 Discussion



8 PPAR Research

NC DSS M P

Bcl-2

Bax

Caspase 3Cleaved

caspase 3

-Actin

(a)

Rela

tive p

rote

inle

vel o

f Bax

000

005

010

015

+

Rela

tive p

rote

inle

vel o

f Bcl-

2

000

002

004

006

008+

Relat

ive p

rote

in le

vel o

fcle

aved

Cas

pase

3

00

01

02

03

04

++++

DSSNC

DSSNC

DSSNC


lowastlowastlowast

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(b)

Relat

ive m

RNA

of B

ax

00

05

10

15

20

25

+++

Relat

ive m

RNA

of B

cl-2

0

2

4

6

++


DSSNC

DSSNC

lowast

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(c)






PPAR Research 9





2) could



5 Conclusion






Acknowledgments


References





10 PPAR Research


































PPAR Research 11














of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of



















8 PPAR Research

NC DSS M P

Bcl-2

Bax

Caspase 3Cleaved

caspase 3

-Actin

(a)

Rela

tive p

rote

inle

vel o

f Bax

000

005

010

015

+

Rela

tive p

rote

inle

vel o

f Bcl-

2

000

002

004

006

008+

Relat

ive p

rote

in le

vel o

fcle

aved

Cas

pase

3

00

01

02

03

04

++++

DSSNC

DSSNC

DSSNC


lowastlowastlowast

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(b)

Relat

ive m

RNA

of B

ax

00

05

10

15

20

25

+++

Relat

ive m

RNA

of B

cl-2

0

2

4

6

++


DSSNC

DSSNC

lowast

Portulaca(10

0)

Mes

alam

ine(

74)

Portulaca(10

0)

Mes

alam

ine(

74)

(c)






PPAR Research 9





2) could



5 Conclusion






Acknowledgments


References





10 PPAR Research


































PPAR Research 11














of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of



















PPAR Research 9





2) could



5 Conclusion






Acknowledgments


References





10 PPAR Research


































PPAR Research 11














of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of



















10 PPAR Research


































PPAR Research 11














of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of



















PPAR Research 11














of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of























of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of



















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