Dietary iprotein icontent iand iin iparticular iamino iacid icomposition iof ithe idiet iare iremarkably iimportant

ito
support ianimal iproduction iand ihealth. iIt iis inow iev-ident ithat iamino iacids iare ione iof ithe imain idrivers
of ifeed iintake i(FI) iand igrowth iperformance iof ibroilers i(Cho, i2012). iAs ia iresult iof iextensive ibreeding
programs, imodern ibroiler ichickens iare inow iapproximately i400% imore iefficient ithan ibreeds igrown iin
i1956(Zuidhof iet ial., i2014). iThis ihas iresulted iin iincreased inutrient ispecifications iwhich iunavoidably

iincreased ithe itotal icrude iprotein iof ithe idiets iin iorder ito iprovide ithe idesired idigestible iamino iacids iin

ibroiler idiets. iIncreased ienvironmental iimpact, iwet ilitter iissues, iand iever iincreasing ireliance ion iquality

iprotein isources iare iamongst ithe imain idrawbacks iof ia ihigh iprotein idiet. iTherefore, iin irecent iyears, ithere

ihas ibeen igreater iinterest iin ifeeding ibirds iwith ireduced iprotein i(RP) idiets isupplemented iwith iamino iacids.

iSupplementation iof iamino iacids ihas ibeen iextensively iresearched iin irelation ito iRP idiets iand ithe iorder iof

ilimiting iamino iacids ihas ibeen iidentified i(Ospina-Rojas iet ial., i2012). iAs ifor ithe iperformance

of ibroiler ichickens, ithis ipractice ihas ioften iresulted iin icontradictory iresults ibut imostly iimpaired
iperformance iand iunbalanced isupply iof iamino iacids. iThe imucosa iof ithe ismall iintestine ihas ifinger-like

iprojections iknown ias ivilli. iA isingle ilayer iof iepithelial icells i(enterocytes) icovers ithese ivilli. iThe iintestinal

iepithelium iis iselectively ipermeable iallowing ithe ipassage iof iwater, inutrients, iand ielectrolytes, iwhile

ipreventing ientry iof iharmful iorganisms, itoxins, iand iantigens i(Groschwitz iand iHogan, i2009). iThis

iselective iabsorption iis imediated ithrough icomplex ipathways iof iparacellular ior itranscellular ipermeability.

iIntestinal ipermeability i(IP) ican ibe iregulated ior ialtered iby iseveral ifactors iincluding, ibut inot ilimited ito idiet,

iintestinal imicroflora, istress, idisease, iinfections, itoxins, iand idrugs i(Bischoff iet ial., i2014). iIncreased iIP iis

ioften iassociated iwith icompromised ihealth, iperformance, ibacterial itranslocation, iimmune iactivation, iand

ilameness i(Gilani iet ial., i2016). iIn ia irecent istudy, iBarekatain iet ial. i(2018) ifound ithat iIP ican ibe imaintained

iin ibirds ifed ian iRP idiet iwhen iessential iamino iacids iwere isupplemented ito imatch ia istandard iprotein

diet. iHowever, iwhen icompared iwith ihigher iconcentrations iof iamino iacids, ia ihigher iIP iwas iobserved
iindicating ia icompromise iin iintestinal ibarrier ifunction iassociated iwith ichanges iin igene iexpression iof itight

ijunction iproteins iand isome inutrient itransporters. iChen iet ial.(2016) ialso ishowed ithat ireducing idietary

iprotein iexacerbated ithe ieffect iof iaflatoxin iin ibroiler iperformance iand inutrient iutilization ialong iwith ia

inumeric itendency ito iincrease iIP. iFor iRP idiets, ispecific iamino iacids ihave ibeen iextensively istudied, iwith

ithe imost ipromising icandidates ibeing iglycine i(Gly) iand iserine i(Ser) ithat ineed ito ibe iclosely iconsidered

iwhen iformulating idiets i(Siegert iet ial., i2016). iDespite iabundant idata ion ithe iperformance iparameters

iassociated iwith iamino iacid isupplementation ito iRP idiets, ifew istudies ihave ibeen iconducted ito iinvestigate

itheir iunderlying iroles ifor iintestinal ibarrier ifunction iand ipermeability. iA istudy ihas ishown ithe ieffect iof iGly

ion iIP iin ipiglets ibut iwithout iconsideration iof idietary iprotein i(Li ietal., i2016). iAdditionally, isome iamino

iacids iincluding iglutamine i(Gln), iarginine i(Arg), iand ithreonine i i(Thr) ihave ibeen idocumented ito ialter

iintestinal iintegrity iin iother ispecies iand imore irecently iin ipoultry

(Bortoluzzi iet ial., i2017).It iis iwell iestablished ithat iGln iplays ia icritical irole iin imaintaining igut iintegrity
ithrough iprovision iof ienergy ifor irapidly iproliferating ienterocytes iand ilymphocytes imediating isuch ieffects

ithrough itight ijunctions, imucosal icell iproliferation, iand idifferentiation(Soares iet ial., i2014). iGln iis iinvolved

iin imucin isynthesis. iN-Acetyl iglucosamine iis ia iglycoprotein iand ia icomponent iof ithe imucin ithat iprotects

imucosal isurfaces iand iits iformation iis ifully idependent ion iGln i(Coster iet ial.,

2004). iGln imay ibe iconsidered ia iconditionally iessential iamino iacid iduring istress, iinjury, ior imalnutrition.
iUnder isuch icircumstances ithe irequirement imay iexceed ithe iendogenous imetabolic icapacity irequired ito

imaintain igut iintegrity iand ireduce iinflammation ias iimmunocytes iincrease itheir iuptake iof iGln i(Coster iet

ial., i2004). iPrevious istudies ihave idemonstrated isome ipositive iresponse ito iGln iin iterms iof igrowth

iperformance iand igut idevelopment iin ibroiler ichickens i(Bartell iand iBatal, i2007; iMurakami iet ial., i2007).
iHowever, iour irecent istudies ihave ifailed ito ishow iany ipositive iresponse ito iGln iin
broiler ichickens iin iterms iof igut ipermeability ior iperformance i(Gilani iet ial., i2018c). iGilani iet ial. i(2018c)
ifound ithat iproviding i1% isupplemental iGln iin idrinking iwater ito inewly ihatched ibroilers idid inot ireduce iIP

iin idelayed ifed ibirds. iIn ianother istudy, iGilani iet ial. i(2018b) isupplemented i10 ig/kg iGln ito ia iwheat-based

idiet iand ifound ino ieffect ion iIP iassessed iby ifluorescein iisothiocyanate idextran i(FITC-d) iuptake ior

ilactulose, imannitol, iand irhamnose isugar iassays. iIt ishould ibe inoted ithat iin iboth

trials i(Gilani iet ial., i2018b, ic), ithe iGln iwas igiven ialongside ia inormal icommercial idiet ias iopposed ito ian iRP
diet. iIt imay ibe ihypothesized ithat ithe iL-Gln icould ibe imore ieffective iwhen ithe idietary iprotein iis ireduced ior
the ianimal iis iunder ia iserious istressor ior idisease ichallenge. iAs ian iessential iamino iacid ifor ichickens,
imounting ievidence iis iemerging ion ithe ifundamental iroles ithat iArg iplays iin ivarious imetabolic ipathways

iand iregulation iof iintestinal ifunction iand iultimately iprotein isynthesis iand iperformance. iThe isynthesis iof

initric ioxide i(NO) iand ipolyamines iis idependent ion iArg i(Wu iandKnabe, i1995). iProduction iof iNO ialong

iwith ienterocyte imigration iis icrucial ifor irestoration iof iepithelial icontinuity i(Jacobi iand iOdle, i2012).

iTherefore, iArg ican imodulate iimmune iresponses, iinflammation, iand ithe iprocess

of irecovery ifrom iinjury ior istress. iRecent idata iin ipoultry isuggest ipotential ifor iincreasing ifeed ispecification
ifor iArg ibeyond icurrent iindustry ipractices ito isupport ioptimal igrowth iand iintestinal ifunction i(Tan iet ial.,

i2014a). iSupplementation iof iArg ican itherefore iimprove igut ibarrier ifunction ias ishown iby ireducing iileal

ipermeability imeasured iin iUsing ichambers iby iZhang iet ial. i(2017).

However, ithe ispecific irole iof iArg iin iRP idiets ifor iintestinal ibarrier ifunction iand ipermeability ihas inot ibeen
investigated. iThus, iit iwas ihypothesized ithat iindividual iamino iacids iincluding iGln, iGly, iand iArg iwould
ipositively iimpact iintestinal ibarrier ifunction iin ibirds ifed iRP idiets. iSpecifically, ithe ipresent istudy

iinvestigated ithe irole iof iincreased iconcentrations iof iL-Gln, iGly, iand iL-Arg

on igrowth iperformance iand iIP, igene iexpression iof iselected itight ijunction iproteins, iand iintestinal
iarchitecture iof ibroiler ichickens iutilizing ia idexamethasone i(DEX)-induced imodel iof ileaky igut.