Pulling back to the nature: A condition to fight IBD

Manoj Patidar; Naveen Yadav; Sarat K. Dalai*

Institute of Science, Nirma University, Ahmedabad-382 481, India

*Correspondence to: Sarat K. Dalai, Institute of Science, Nirma University, Ahmedabad-382481,

India

Chapter 1

Inflammatory Bowel Disease

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1. Introduction

Inflammatoryboweldisease(IBD)groupedasautoimmunediseasearisesduetoinflam-mationofsmallandlargeintestine.BasedonthetargetorganIBDisclassifiedunderUlcer-ativecolitis(UC;affectsthecolon)andCrohn’sdisease(CD;affectswholeintestinalwallbutmainlytotheileum)[1].1-1.6millionpeoplearesufferingfromIBDinUnitedStatesandthemaintargetagegroupisbetween15-30yrs.Theprevalencerateof201/106ofCDand238/106 ofUCinadultsattractsattentiontothedisease[2].Abdominalcramping,weightloss,fever,sweats,fatigue,growthretardation,diarrhoea,constipationandabnormalbowelmovementarethemajorsymptomsofIBD.ItisclearlynoticedthatIBDisnotlimitedtojustinflammationofthedigestivetract,butitcanleadtoothercomplicationslikearthritis,thromboembolism,car-diovascular-,pulmonary-&neurologicaldiseaseaffectingthequalityoflifeofanindividual.ItisclearlyseenthatIBDrunsinthefamily,andthefamilymembersofaffectedindividualsareatthemaximumriskofIBD.InlasttwodecadesitisshownthatenvironmentalfactorsinthepathogenesisofIBDareplayingimportantroleinincreasingtheincidencesofIBDcasesinthecountrieswithhistoricallylowratesofIBD.InNorthAmericaandEurope20-foldincreaseinIBDcaseswerefoundsinceWorldWarIIduetochangeindietaryhabitandenvironmentalfactors.TheenvironmentalfactorsalsocorrelateswiththesimilartrendsinIBDduetoglo-balizationandWesternizingEastAsia[3].VariationinmicrofloraanddisregulatedimmuneresponsesarethekeyeffectorsofIBD.InthischapterwehavediscussedthevariousfactorsaffectingthepathogenesisofIBDincludinggenetic,environmentalandimmunefactors.WehavealsodiscussedthatthedelayinIBDdiagnosisandfailtocontrolit,canresultintrans-

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InflammatoryBowelDiseaseDalaiSK

formationintothecancer.WehavealsodiscussedtheroleofTcellsaswellasmicroflorainpathogenesisofIBDandshedlightonitstherapeuticaspects.

2. Disregulated Inflammation Predisposes to IBD

Healthygutcontains10-100foldmoremicrobesthanmammaliancellsandintestinalepithelial layer hides thismicrobial line from intestinal immune systemandmaintains thepeaceornon-inflammatoryenvironment.Asthenamesuggestsinflammationisthekeytoin-ducingIBDandbysuppressinginflammationIBDcanbeameliorated.InthepathogenesisofIBD,cytokinesholdthecentralpositionasdemonstratedinmanygeneticandimmunologicalstudies(Table 1).Basedontheirnatureofevokingorsuppressingtheinflammation,theyaregroupedaspro-andanti-inflammatorycytokines.Boththeseclassesofcytokinesareinten-sivelystudiedforbetterunderstandingtheIBDbiology.Cytokinesmaintainthenormalim-munehomeostasisbytriggeringtheresponsestoinfectionsandrevertingbackinflammationtoabasallevelaftertheinfectionisresolved.Butoftendysfunctionoruncontrolledexpressionof thesecytokines triggers immunologicaldisorders.Forexample,IL-1receptorantagonist(IL-1Ra)controlstheinflammatoryresponseofIL-1cytokines,andinsteady-stateIL-1/IL-1-Rarationisfoundtobeconstant.ButanincrementintheratiowasobservedincaseofIBDduetooversecretionofIL-1bymonocytesandmacrophages.TheinvolvementofIL-1inIBDpathogenesismakesitareliablemarkerfordiagnosisofIBDandalsomakeitasuitabletargetfortherapy[4].Similarly,IL-6,-8,-12,-13,-17,-15&-21triggertheinflammationandblock-ingthesecytokinesisshowntoalleviateIBD[5,6].Anti-inflammatorycytokinesorcytokineneutralizingAbsarefoundefficaciousforIBD(Table 1).Unlikethesepro-inflammatorycy-tokines,anti-inflammatorycytokinesdownregulatetheinflammatoryconditionandhelpincuringtheIBD.ForexamplesIL-10,acrucialanti-inflammatoryplayerinCD,controlsTh17cell development andhas beneficial effects in IBD.Spontaneous development of IBDob-servedinIL-10-/-mice[7]supportthefactthatIL-10iscriticalincontrollingtheIBD.Further,useofIL-10,TGF-βandIL-4hasshowntoreverseIBD(Table1).GenomewideassociationstudiesestablishedtheimportanceofSTAT-1,-3,-4,CCR6,CCL-2,-13,IL-12R,-23R,JAK2,IL-2,-21,-10,-27andIFN-γ[8]ininductionofIBD.

AmongthecytokinesTNF-αisapotentactivatorofinflammatoryresponsesandhencedrawnattentionof researchersworking in the areaof IBD.TNF-α isnotonly involved inpathogenesisofIBDbutalsoplaysdominantroleinotherdisorderslikerheumatoidarthritis,psoriasis,psoriaticarthritis,andankylosingspondylitis[8].OverexpressionofTNF-αleadstoacascadeofinflammatoryeventswhichresultsinauto-immunediseasesincludingCDandUC.Itshowspleiotropiceffectsbyincreasingtheexpressionofadhesionmolecules,stimulat-ingfibroblastproliferation,IL-1β,IL-6,IL-33,ST2expression[Table1:Cytokinesinvolve-mentinIBD8].ElevatedlevelsofTNF-αwerefoundintheserum,mucosaandstoolofIBDpatients.Deletion ofTNF-α synthesis regulatory elements is found to evoke IBD inmice

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models.Interestingly,modelsofinducingIBDthroughchemicalsfailedinTNF-/-mice[9]showingtheimportanceofTNF-αinIBD.TNF-αinducesIBDthroughseveralmechanisms,forexample,recruitmentofneutrophilsoninflammationsite,activationofcoagulationandfibrinolysis,andgranulomaformation.TNF-αhastwoisoformsanditwasdemonstratedthatmembrane-boundTNF-αisdominantinevokingIBDthansolubleform[8].

Sr. No.

Cytokines In Immune Homeostasis In IBD

Pro-inflammatory Cytokines

1IL-1 family: IL-1α, -1β, -18,-33,-36α,β,γ

1. IL-1Ra controls the activity ofIL-12.ConstantIL-1/IL-1Raratio

1.Overproductionbymonocytes&macrophagehenceincreaseIL-1/IL-1Raratio.2.Reliablemarkerofinflammation.3.TargetingIL-1hastherapeuticvalues[4].

2 TNF-αCriticalplayerfordefenceagainstmicroorganisms and immunesystemdevelopment.

1.Majorsourcemacrophages,monocytes,andTh1cells.2.Showspleiotropiceffects,3.Increasetheexpressionofadhesionmolecules,4.Stimulatefibroblastproliferation,5.StimulateIL-1β,IL-6,IL-33,ST2expression[5].6.TNF-αinhibitorsareefficaciousforIBD.

3IL-6 family:IL-6, IL-11, IL-31

1. IL-6 is an immunoregulatorycytokine2. Acts as both anti- and pro-inflammatory3. Signals through IL-6-sIL-6R-receptorgp130complex.4. Immune response duringinfectionandaftertrauma.

1.Over-secretionbymononuclearcells&intestinalepithelialcells.2.IncreasedlevelofsIL-6RwasseeninUCandCDpatients.3.IL-6inducesNF-kBactivation,STAT-3andexpressionoftheintercellularadhesionmolecule1.4.IL-6helpsinTh17differentiation.5.Controlsanti-apoptoticsignalsofCD4+Tcellsatthesiteofinflammation.6.anti-IL-6receptorantibodiesreversetheIBD[10].

4 IL-8Involves in neutrophil activationandmigration

1.HighertissuelevelwasfoundinactiveUC.2.Poormarkerofdiseaseactivity[11].

5IL-12 family:IL-12, IL-23, IL-27andIL-35

1. Role in Th17 celldifferentiation.2. Produce in response to TLRstimulationorendogenoussignals.

1.ElevatedIL-12levelsinthemucosaofUCpatients.2.IL-23oversecretionleadstoactivationofNKcells,NKTcells,CD4+TcellsandCD8+Tcells.3.IL-35involvesinT-cell-dependentcolitis[5].

6 IL-13

1.Allergicinflammation2. Also show anti-inflammatoryactivityw3.InducesIgEsecretion.

1.HigherexpressionwasfoundinUCmucosa.2.OversecretionbyCD161+NKTcells.3.InhibitorsofIL-13signallingpathwaysshowingpromisingresults[5].

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IL-17 family:IL-17A,B,C,D,E(IL-25),F

1.Allergicresponses2.Delayed-typeimmunereactionsby increasing chemokineproduction and recruitingmonocytes and neutrophils to theinflammatorysite.

1.UpregulationinIL-17levelisassociatedwithUC.2.Anti-IL-17antibodies,IL-17receptorblockerandIL-17inhibitorsareusefulintargetingIBD[5].

8 IL-5 EosinophildifferentiationfactorOverexpressionofmucosalIL-5bymononuclearcellsisseeninactiveUC[5].

Table 1:CytokinesinvolvementinIBD

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3. Uncontrolled IBD May Lead To Cancer

Thelinkbetweeninflammationandcanceriswellaccepted[18].ThedirectrelationofIBDwithcancer,particularlyforcolorectalcancer(CRC)wasshownbyBurrillCrohnin1925[19].Duetochronicintestinalinflammation,theIBDpatientsarefoundtohaveanincreasedriskofdevelopingCRCandtheriskincreaseswiththeprogressionofIBD[20].TheriskofCRCwascalculatedforUCanditwasfoundthatpatientswithUCafter10years,20yearsand30yearsofstartingofdiseasehasriskby2%,8%and18%respectively[21].Similarly,therelativeriskwasfoundtobe2.59forCRCand28.4forsmallbowelcarcinomainCDcases[22].

DuringIBD,inflammationisthemajorcauseforthedevelopmentofCRC[23].Asig-nificantincreaseinthelevelofinflammatorymediatorslikecyclooxygenase-2,nitricoxidesynthase-2and interferon-induciblegene 1–8Uwas found in IBDpatients leading toCRC[24].TLR4inducedCox-2andEGFRsignallingarealsoinvolvedinthepromotionofCRC[25].InactiveIBDelevatedlevelofclaudin-1,-2andBeta-cateninwasalsoobservedwhich

9 IL-211.Th1mediatedinflammation2.InducerofIFN-γproduction

1.UncontrolledsecretionofIL-21byCD4+laminaproprialTcellsleadstoIFN-γproductionresultinIBD.2.IL-21inhibitsTregdifferentiationwhichultimatelyresultsininflammatoryresponses[12].

Anti-inflammatory Cytokines

1

IL-10 family:IL-19, IL-20,IL-22, IL-24 andIL-26

1. Regulate diverse host defensemechanisms from epithelial layerduringvariousinfections2.Importantformaintenanceoftheintegrityandhomeostasisoftissueepitheliallayers3. Down-regulate inflammatoryresponses and controlstissue disruptions caused byinflammation.

1.Crucialanti-inflammatoryplayerinCD.2. IL-10 controls Th17 cell development by inducing IL-1secretion.2.BlockingofIL-10leadstoIL-12andIFN-γproduction.3.LowerexpressionofIL-10wasseenininflamedmucosaandgranulomasofCDpatients[13,14].4.IL-22iselevatedinCDmucosaandserum.

2 TGF-β

1.Actsasaninhibitorycytokine2. Regulate the immunologicalhomeostasis and inflammatoryresponses

1.TGF-β induces IL-13 expression andwhichpromotes theexpressionofcellinvasionproteins.BothcanbetargetedforCD.2.Down-regulatedexpressioninCDpatients,butinterestinglyup-regulatedinUC[15].3.Somereportssuggesttheup-regulationofTGF-β1inbothUCandCD,butinhibitedbySmad7.4.InhibitingSmad7restoresTGF-β1activitywhichsuppressesinflammationandhelpsinrecovery[16].

3 IL-41.StimulatorofBandTcells2. Immunosuppressive activity intheintestine

1. Use of anti-IL-4 antibody decreases Th2-type cytokineproductionandincreasesIFN-γproduction,suggestingroleofIL-4inIBDpathogenesis.2.AdministrationofIL-4resultedindown-regulationofVEGFinactiveCDandUCpatients[5,17].

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iscorrelatedwithdiseasetransformation[26].Similarly, inIBDpatients,IL-6,-23,NF-κBandTNF-αarealsoshowntoinvolveinthedevelopmentofCRC[27-31].AlsotheroleofoxidativestressinCRCdevelopmentwasexploredandfoundthatnitricoxidehascontributingroleinthesame[32].ResearchersaremakingattemptstoreducetheriskofCRCinIBDbytheearlydiagnosisofCRC,treatmentbycolonoscopicsurveillanceandbychemoprevention[33]including5-aminosalicylicacid(5-ASA)i.e.,MesalazineandSulfasalazine[34-36]haveshownverypositiveresults.

4. T cells and IBD

Gutistheprimeentrysiteforpathogenscomingthroughfoodandorallyadministeredsubstances.GALT(gutassociatedlymphoidtissue)directstissuerestrictedandlocalizedim-muneresponseto the intestinalantigens.Tcellsprimedagainst invadingpathogenmigratefromGALT to the site of infection andprotect thegut.After resolutionof infection theseantigen-experiencedTcellsmigratetotheLP(laminapropria)andepithelialcompartmentswheretheyresideaslong-livedeffectormemoryTcellsandaccumulateovertime.Uponre-encounterwiththepathogenthesecellsrespondswiftlyandvigorouslytoeliminatethepatho-gen,preventingsystemicinfection[18,19].TheseeffectormemoryTcellsproliferateslowly,producelowamountofinflammatorycytokinesthathelpmaintainorganintegrity[20].LPisalsoenrichedwiththeTregcellsrequiredforpreventingtheexcessiveresponsesagainstthegutmicrobes,therebymaintainingthegutintegrity[21].

IngutthereispresenceofselfspecificTcellsexpressinghighlevelsofbotheffectorandregulatorymolecules.TheyareusuallyinquiescentstageanduponstrongstimulationtheybecomehighlycytolyticlikeconventionalTcells.TheyrestrictmicrobialinvasionbykillinginfectedordamagedIECs[37,38].TCRγδ+Tcellsproducespro-inflammatorycytokinesandantimicrobialfactorswhichsetthebasalmucosalinflammatorytoneandlimitentericbacterialtranslocation[39].Interestingly,TCRγδ +Tcellssecreteepidermalgrowthfactor(EGF)whichalsohelpsrepairtheepitheliuminsults[40].Thoughthesecellsactlikefirst-linedefence,theiractivationisfine-tunedandhighlyregulatedtoavoiduncontrolledimmuneresponsesandtis-suedestruction.

Varietiesofmicrobialspeciescoexistandestablishcommensalrelationshipinthegutofthehumans.Inreturnofthesupplyoffoodfromthehost,microbeshelpinmetabolismanddevelopmentoftheimmunesystem.BreachingofthisrelationleadstotheIBDduetoaber-rantandpersistentimmuneresponses.AsdescribeaboveGALTisinvolvedinneutralizingthepotentiallyharmful pathogens [41].Controlling theoverwhelming immune responses afterpathogenclearanceisrequiredtoavoidresponseagainsttheselforharmlessantigens.Per-turbationinimmuneregulatorymechanismsmaypredisposethehosttothepathogenesisofIBD[42].Differentcelltypesoftheintestinallymphoidtissueareinvolvedinmaintainingthe

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immunetolerancetotheluminalantigensandprotectionagainstthepathogens.Infiltrationofcellsmediatinginnateimmunity(neutrophils,macrophages,anddendriticcells)andadaptiveimmunity(TandBcells)isassociatedwiththeactiveIBD.CD4+Tcells(Th1,Th2,Th17,andTfollicularhelperTFH)playimportantroleindefenceagainstpathogens.RegulatoryTcells(nTreg,iTreg;Tr1andTh3)arerequiredforcontrollingtheeffectorresponses[41].

TcellsaccumulateintheinflamedgutofIBDpatientsasaresultofenhancedrecruit-ment,increasedcellcyclingandresistanttoapoptoticsignals.PresenceofCD4+TcellsintoinflamedmucosasuggeststheinvolvementofthesecellsinthepathogenesisofIBD.ThekeyroleofCD4+TcellsinIBDiscorroboratedbytheeffectivenessofanti-CD4antibodythera-piesandsuppressionofIBDinHIVpatients[41].Intheanimalmodelsofexperimentalcolitis,theadoptivenaiveTcelltransferintoalymphopenichostisshowntoinducecolitis[41].Ontheotherhand,blockingtheCD4+Tcellfunctionisfoundtoimprovetheexperimentalcolitis.RoleofCD8+TcellsinIBDhasbeendemonstratedinexperimentalanimalmodelsandpa-tients.ElevatednumbersofCD8+TcellsisalsoreportedinIBDpatients[37].Recently,itwasfoundthatapoptosismachineryofTcellshasintrinsicdefectsinIBDpatients,thusthedefec-tivepathwayinmitochondriahasimportantimplicationsforthetreatmentofIBD.Theeffec-tivenessofTNFneutralisingdrugsforIBDtreatmenthavecorrelatedwellwiththeircapacitytoinduceTcellapoptosis.AttemptshavebeenmadetousetheinhibitorsofTcellactivation,proliferation,differentiationorhomingtocontrolIBD.

Apartfromtheincreaseinthenumberofcells,defectiveregulatoryTcellresponsesarealsoconsideredasriskfactorsforIBD.NaturalTreg(nTreg)hastheabilitytorecognizebothselfandforeignantigens,andplaysanimportantroleinthemaintenanceofself-toleranceandpreventionofchronicimmunestimulationinIBD.TCRligationwithAginthepresenceofTGF-ßinducesFoxP3onCD4+Tcells,convertsthemintoTregsandsuppressestheimmunesystem[38].HigherfrequenciesofCD25brightandFoxP3+TregscellswereseeninlaminapropriaofactiveIBDpatientsandincreasedasdiseaseprogresses[38].Interestingly,colonicbiopsiesdemonstratedlessnumberofTregsinIBDpatientscomparedtonon-IBDinflammatorydiseas-es[38].CurrentIBDtherapeuticsmayhaveaneffectonthefrequencyandfunctionofTregs.ManipulationofregulatorypathwayswouldhavetherapeuticimpactagainstIBD. ItisfoundthattheadministrationofTregsinIBDpatientshavebeneficialeffects.ApartfromadministrationofexogenousTregs,attemptshavebeenmadetoexpandtheendogenousTregpopulation,convertingnaiveTcellsintoTregsbyinsertionofFoxP3genes,modulatingCD4+ TcellsinthepresenceofTGF-ßandimprovingthesurvivalofTregsviastabilizedß-catenin&upregulationofBcl-xL[38].InducedTreg(iTreg)cellspredominantlysecreteTGF-βandIL-10tomaintainmucosalhomeostasisandprotectfromexperimentalcolitis,respectively[41].Immuno-suppressorswithabilitytoinduceTregareuseful,butincreasingthenumberofmayalsodown-regulatethedesiredimmuneresponse,therefore,criticalanalysisisrequiredbefore

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targetingthepathwayofTregs.

5. Gut microbiota and IBD

Hostmicrobialinteractionsandimportanceofintestinalmicrobiotaformetabolismandhostdefencearewellestablished.Humansevolvewiththesemicrobesanditisknownthataround1,800generawith15,000-36,000bacterialspeciesco-existintheintestine[43].Itisinterestingtoknowthatthebacterialload(i.e.,1014bacteria)inhumangutis100timesmorethanthehumancells(i.e.,1012cells).Mucosalimmunecellscantargetthesemicrobesandevokeimmuneresponses.Theepitheliallayerscoveringthebowelwallcontroltheentryofgutfloraandminimizetheinteractionofmicrobeswiththeimmunecells[44].Alargebodyofstudiesisfocussedonimmuneresponsesaswellasmaintenanceofimmunetolerancetotheintestinalmicrobes.TheimmuneresponsesarelargelyprovokedbyrecognitionofPAMPs(pathogenassociatedmolecularpatterns)bytoll-likereceptors(TLRs).Apartfrommicrobes,themetabolites frommicrobesare shown toplay important role in inducing inflammation.Sometimesmicrobespenetratethebowelwallandprovokeimmuneresponses[45].TherearemanyreportssuggestingtheroleofentericflorainthepathogenesisofUCandCD[46-48].Itisobservedthattheareawithhighestbacterialloadshowshighestdegreeofinflammation.LoweringthebacterialloadbyantibioticshasbeenshowntoabrogatetheprogressionofIBD.ItisworthnotingthatinoculationofbacteriasuccessfullyinducesIBDinexperimentalmice[45].Genome-wideanalysisofhealthyandIBDpatientsprovidestheclueofmicrobialcon-nectionwithIBDanditisfoundthat>160SNPsareidentifiedasmarkersofriskforIBD.Interestingly,manyofthesegenesarecriticallyinvolvedinhostresponsestothemicroflora.MutationsinNOD2,ATG16L1,IRGM,CARD9andIL23RappeartobecriticalininducingIBD[1].

TounderstandtheinvolvementofbacteriainCrohn’sdiseasealargestudyinvolving668childrenwasconducted.Theresultshowsthedecreaseinnumbersofbeneficialbacteriaandincreaseinnumbersofharmfulbacteriainthegutofpatients[49].FirmicutestoBacteroidetesratiowasfoundtobedecreasedinIBD(bothCDandUC)[50,51].Apartfromtheinvolve-mentofabacterialshiftinIBD,roleoffungusisalsoreported.BysuppressingthefungalflorabyprobioticSchreiberetal.successfullytreatedtheUCsuggestingtherequirementofnotonlybacterialdiversitybutalsofungalandbacterialratio.IBDcanalsodevelopduetoinfectionsofMycobacteria, Shigella, Salmonella, Yersinia, Clostridium difficile andBacteroides vulgates [52].ThereductioninBifidobacterialoadandincrementinE. coliandclostridia loadinrectalmucosawasobservedinUCpatientsanda250-foldchangeinthebalancewasobserved[53].Surprisingly,inCDcasestheE. coliloadwasfoundincreasedbutnochangewasobservedinBifidobacteriaload[53].

6. Microflora and T cells shape each other: a check point for IBD

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GutmicrobiotaispostulatedtodirectandmodulatetheTcellactivationanddifferentia-tion.Itisevidentfromthefactthatgermfree(GF)rodentsdonotdevelopintestinalinflam-mationorimmuneactivationandtheyhavereducednumbersofCD4+andCD8+Tcellsthatgetrestoredfollowingrecolonizationwithmicrobiome[54].TcellclonesspecifictoEnter-obacteriaceae, Bacteroides,andBifidobacteriumwereisolatedfromthegut[55].Theimmunesystemistrainedtodevelopresponsesnotonlyagainstthepathogens,butalsotothecom-mensal trying toescape thehomeostasismechanisms.WhenintestinalparasiteToxoplasma gondiiinfectionoccurs,CD4+Tcellsdevelopresponsesagainstthepathogenandalsoagainstthetranslocatingcommensalbreachingthetolerancetocommensal[41].ThesemicrobesalsomodulateTh-17cellresponsesaswellasIL-17secretion;anditwasobservedthatGFmiceormicetreatedwithantibioticstoeliminatethecommensalbacteriahavecomparativelylowernumbersofTh-17cells.SegmentedFilamentousBacteria(SFB),agroupofgutmicrobehavebeenshowntoplayimportantroleinthedifferentiationoftheTh17TcelllineageaswellasIL-22-producingCD4+Tcellsinthesmallintestineandthecolon,andalsomediateprotectionagainstentericinfectionbyCitrobacter rodentium[41].DisproportionateSFBfrequencyhasdirectassociationwith IBD.Commensalbacteriaalsomodulate thebalancebetween regu-latoryandeffectorTcellsviapatternrecognitionreceptors, includingTLRs.DendriticcellstimulationbyTLR5leadstoanincreaseinIL-23,whichfurtherpromotestheTh17effectorcellpathwaytothedetrimentofFoxp3+Treg[41].

ItisestablishedthatregulatoryTcells(Tregs)alsoplayacriticalroleinthemaintenanceofguthomeostasis.InterestinglyitisobservedthatthegenerationandfunctionofTregsareinfluencedbythecolonfloralikeClostridiaandBacteroides fragilis butnotbythemicrobesinsmallintestine.Clostridiaclusters(Clostridium leptum andClostridium coccoides)arefoundtoinduceproliferationanddifferentiationofperipheralTregandhelpaccumulateCD4+CD25+ Foxp3+Tregincolon[41].ByreconstitutingGFmicewithamixof46ClostridialstrainsitwasshownthatasignificantnumberofTregswereaccumulated,inthececumandthecolon,tolevelssimilartothatofSPFmicebutsimilaraccumulationwasnotpossibleby16Bacteroidesspecies,SFB,and3Lactobacillusspecies.TheprotectiveeffectofClostridiaspeciesmaybemediatedtoalargeextentbySCFA(shortchainfattyacids)whichsupporttheexpansionoftheexistingpoolofcolonicTregs[41].

ItisamplyclearfromtheabovefindingsthatmicrofloracriticallyshapestheTcellre-sponses.Ontheotherhand,interestingly,TcellsappeartoshapetheGutMicrobiome.Thereis emerging evidencewhich demonstrates this reciprocal relationship betweenT cells andtheirabilitytoshapethecompositionofthegutmicrobiota.Thedifferenceinmicrobiotadi-versityandloadwasseeninRag1−/−micevs.normalmice.TheimmunodeficientmicehaveincreasedloadofAkkermansiamuciniphilabutdecreasedloadofLactobacillalesandEnter-obacteriales[41]comparedtothatofnormalmicestronglysupportstheideathatTcellplays

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importantroleinshapingtheGutMicrobiome.

7. Targeting IBD

CurrentlyIBDistreatedbythewaitandwatchapproachwiththehelpofdrugswhichalleviatethesymptomsbyregulatingtheinflammation.AminosalicylatesandCorticosteroidsareusedtocontroltheinflammations[56,57].AntibioticslikeMetronidazoleandCiprofloxa-cinarebeingusedtoreducethebacterialload.Apartfromthesetraditionalmethods,manybiologicsarenowbeingtriedtoforIBD.Theantibodiesthatcontroltheloadoffungus,Sac-charomyces cerevisiae [3]are inuse target thepro-inflammatorycytokines, induceanti-in-flammatorycytokinesandalsotoblocktheintravascularadhesionmolecules.TNF-αinhibi-torslikeInfliximab(Remicade),adalimumab(Humira)andantiα4-integrinlikecertolizumabpegol (Cimzia)arealsoplaying important role incontrolling thepathogenesisof IBD[56](Table 2).

Sr. No.

Therapeutics Examples Mechanism of action Side effects

1 AminosalicylatesMesalazine,Sulfasalazine,Olsalazine,Balsalazide

ActivationofPPARγ1.Inhibitionofmacrophage2.chemotaxisInhibitionofNF-κB,STAT&3.AP-1

Diarrhea,nausea,vomiting,headache,abdominalpain,hepaticabnormalities,Arthralgiaandmyalgia

2 Corticosteroids

Budesonide,Dexamethasone,Hydrocortisone,Methylprednisolone,Prednisolone,Prednisone

Down-regulationofNF-κB1.Inhibitionofcytokine2.productionBlockingtherecruitmentof3.immunecellsInhibitionofexpressionof4.adhesionmolecules

Osteoporosis,metabolicsyndrome,cardiovasculardisease,infections,osteonecrosis,andcataract

3 AntibioticsMetronidazole,Ciprofloxacin,

Decreasebacterialload1.Suppresstheintestine'simmune2.system

Numbness,tingling,musclepain,weakness,tendonrupture

4 ThiopurinesAzathioprine,6-mercaptopurine

Immunesystemsuppressors1.Inhibitionofproteinsynthesis2.InhibitionofLymphocyte3.proliferationInduceapoptosisofactivated4.TcellsBlockingofTRAIL,TNFRS75.&α4-integrin

Suppressionofdesiredimmuneresponsesmeanslossofresistancetoinfection,verylesschanceofdevelopinglymphomaandskincancers,adverseeffectsonliverandpancreas,

5Folicacidantagonists

MethotrexateInhibitionoffolatepathwayandpurineandpyrimidinesynthesis

Hepatotoxicity,nausea,fatigue,pneumonia,bonemarrowsuppression,cancer.

Table 2:TherapeuticsforIBD,theirmodeofactionandpossiblesideeffects

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8. Probiotic as therapy

Itisevidentfromvariousstudiesthatmicroflorapayskeyroleininitiationormodula-tionofinflammationinfluencingtheonsetorprogressionofIBD.Andthefactthatdisruptionin themicrobioticbalanceevokes IBDopensnewavenues foruseofbacterial inoculation(probiotic)totargettheIBDbyrestoringthebalance.Probioticsareclaimedtobesafer,easiertoproduceandadminister topatients than thepharmadrugsand immune-modulators.Thesideeffectofbiologicsandtheirabilitytomodulatetheimmunesystemiswellknownthere-forealternativebetterstrategiesshouldbepreferred.ManyexperimentalmodelsandclinicalstudiessuggestthesuitabilityofprobioticsfortargetingIBDandalsotheinfectionrateduetoprobioticsisnegligiblei.e.,between0.05%and0.4%cases(58).TheperfectmechanismbywhichprobioticalleviatesIBDisstillnotclear.MadsenetalfoundthatprobioticVSL#3(Non-pathogenicSalmonellastrains)actonepithelialbarriersandimprovetheresistancetoIBDinduction[59].KailaetalproposedantibodyproductionbyBifidobacterium breve[60],andMajamaaetalproposedenhancedcellmediatedphagocytosisbytheuseofBifidobacte-rium lactisprobiotic[61].ModulationofNF-kBpathway,preventionofepithelialapoptosis,antimicrobialactivityandimmunetolerancewereobservedbyuseofVSL#3,Lactobacillus rhamnosus GG,Enterococcus faeciumandBifidobacterium lactis respectively[58,59,62].EachstrainactsdifferentiallyanditwasfoundfromastudyconductedbyR.N.Fedoraketal[58]thatfivemechanismsmightberesponsiblefortheactionofprobioticsinalleviatingIBD:[1]probioticscompetewithmicrobialpathogensforreceptorspresentonthesurfaceepithe-lium;[2]immunomodulationofgut-associatedlymphoidandepithelialcells;[3]suppressionofpathogengrowththroughreleaseofantimicrobialfactorsbyprobiotics[4]enhancementofmucosalbarrierfunction;and[5]inductionofT-cellapoptosis[58].

ClinicalstudiesusingprobioticstotargetCDdemonstratedhighlyencouragingresults.Guptaetal.usedLactobacillusGGandobservedsignificantimprovementinthePaediatricCDactivityindexwithinoneweek.McCarthyetal.showedthat76%patientsavoidedothertreatmentsafter3monthsof Lactobacillus salivariustherapy[63].Apartfrominvestigatingthecontrolofactivediseasemanyclinicalstudieswereperformedtoevaluatethemaintenance

6 Anti-TNF

Infliximab,Adalimumab,Certolizumabpegol,Golimumab

InhibitionofcytokineTNF-αthusactasanti-inflammatory

Costly,treatmentfailure,infusionreactions,infections,autoimmunity,lymphomas

7 Anti-α4-integrinNatalizumab,Vedolizumab

InhibitionofT-cellmigrationProgressivemultifocalleukoencephalopathy

8 Probiotics

Lactobacillusspecies,Bifidobacteriumspecies,E.ColiNissle,yeastSaccharomycesboulardii

Inhibitionofdiseasecausing1.bacteriafromstickingtotheliningoftheintestinesInhibitinflammation2.Re-maintenanceofnaturalmicro3.flora

Sofar,noserioussideeffectshavebeenreported

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ofmedical-andsurgical- induced remission [58].Thestudiesdemonstrated thatprobioticsnotonlyalleviateCDbutalsobeneficialforUCandremissionmaintenance.Thestudiesper-formedbyRembackenetal[64],Fedoraketal[65],Guslandi etal[66]andBorody etal[67]withE. coli Nissle,VSL#3,Saccharomyces boulardiiandfecalenemas-showverypromis-ingresults.Geneticallymodifiedmicroorganismforprobiotic,Lactococcus lactis-modifiedtosecretandthusaccumulateIL-10atthemucosalsurfacewasfoundtopreventIBD[14].TheoverallfindingssupportthatprobiotictherapyhelpsmaintainthenaturalgutmicrofloraandthusasuitablecandidateforthetreatmentofIBD.

9. Conclusion and future perspective

The emerging cases during last two decades and severity associatedwith IBDneedfocusedeffortstodevelopsuitabletherapeutics.Thegutmicrobialcompositionanditsrela-tionshipwithimmunesystemarethekeyconsiderationsforIBD.IBDistargetedbyvariousmeansincludingantibiotics,drugsandimmunemodulators.Theseagentsshowsideeffects,highlyexpensiveandproductionisnotadequatetoreachtothelargepopulation.AlthoughthetherapeuticagentsactthroughvariousmechanismstotargetIBD,itiscriticaltomaintainthebalanceofmicroflorainthegut.AsthemodulationofTcellsshapethemicroflora,aprerequi-sitetomaintainahealthygut,weshoulduseprobiotictotreatIBD.

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