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Contributor:Dr.med.vet.J.Troillet;St.Petersburg24.02.12

VictoriaMaxwell,DVM,MBA2018PotomacRegionalVeterinaryConference

HyattRegencyInnerHarborBaltimore,Maryland

RegenerativeMedicineIRAPandPRPintheEquineAthlete

EquineRegenerativeMedicine•  provideascaffoldforrepair•  providecellsforthehealingprocess•  providegrowthfactorstostimulatethehealingprocess•  Someregenerativetherapiescurrentlyusedinhorses:

–  StemCells–  Platelet-RichPlasma(PRP)–  AutologousConditionedSerum(irap)

StemCells•  Cellsthathavetheabilitytodivideanddevelopinto

manydifferentcelltypesinthebodyduringearlylifeandgrowth.

•  Mayalsohelprepairthebodybydividingtoreplenishcellsthataredamagedbydisease,injury,ornormalwear.–  Whenastemcelldivides,eachnewcellhasthepotentialtoeither:

•  remainastemcell•  becomeanothertypeofcellwithamorespecializedfunction,suchasanervecell,askincell,oraredbloodcell.

StemCells•  3typesofstemcellsidentifiedinhumanandveterinaryresearch:

–  Adultstemcells•  Twoofthemostcommonsourcesofequineadultstemcellsarebonemarrowandadipose(fat)-derivedmesesenchymalstemcells(MCS)

–  Humanembryonicstemcells(hESC)•  Comefromembryosthatdevelopfromeggsthathavebeenfertilizedinvitro(inatesttubeoranotherartificialenvironment).

–  Inducedpluripotentstemcells(iPS)•  Generatedbyreprogrammingadultcellsthathavealreadydifferentiatedintoaspecificcell,suchasaskincell.

•  FDAhasnotapprovedanystemcell-basedproductsforuse,exceptcordblood-derivedhematopoieticprogenitorcells(bloodformingstemcells)forcertainhumanindicationsincludingcertainbloodcancersandsomeinheritedmetabolicandimmunesystemdisorders.

http://www.vet-stem.com/science.php

VeterinaryMedicalDevices

•  Further,adevice"doesnotachieveanyofitsprincipalintendedpurposesthroughchemicalactionwithinoronthebodyofmanorotheranimals,andisnotdependentuponbeingmetabolizedfortheachievementofanyofitsprincipalintendedpurposes."Examplesofdevicesincludesuchthingsasneedles,syringes,surgicalinstruments,prostheticdevices,X-rayequipment,certaindiagnostictestkits,anddentalappliances.

•  Pre-marketApprovalisNotRequired:TheFDAdoesnotrequiresubmissionofa510(k)orformalpre-marketapprovalfordevicesusedinveterinary

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Orthokinevetirap60® Orthokinevetirap10®

MedicalDeviceforPreparationofAutologousConditionedSerum

irapTherapy

•  Whatisirap?–  Interleukin-1ReceptorAntagonistProtein(irap)–  iraptherapyutilizesthepatient’sownbloodandnaturalhealingmechanisms

– ThepatentedirapsyringeproducesAutologousConditionedSerum(ACS),whichcontainsbeneficialanti-inflammatoryandregenerativecytokines.

Cytokine=immunoregulatoryproteins(asinterleukinorinterferon)secretedbycellsespeciallyoftheimmunesystem

AutologousConditionedSerum

Autologous:100%body’sowncomponentsConditioned:AchievedthroughphysicalprocessesSerum:Cell-freesupernatantofcoagulatedwholeblood

•  Howdoesirapwork?–  theirapsyringecontainsmedicalgradeglassbeads

– stimulatethewhitebloodcellstoproducetheanti-inflammatoryandregenerativecytokines

– wholebloodisincubatedandthencentrifugedtocollectserum

–  theserum,(ACS),isthenre-injectedintothepatient.

irapTherapy

GlassBeads

MedicalGrade“A”GlassBeadsTheyareNOTcoatedwithanything

•  Theconditioningofthebloodisachievedthroughexposureofthewholebloodtoaspecificsurfaceofborosilicateglassspheres

•  Certaincellsaccumulatearoundthesesurfacesandbegintoproducecytokinesthatarebeneficialforregenerativeprocess

AutologousProteinProductioninWholeBloodModel:MonocyteAdherencetosurfaces

activemolecules

binding

e.g.glassspheres

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Monocyte

Macrophage

Lymphocyte

Neutrophil granulocyte

Immunologic cell

IL-1Ra Leukocytes(WBCs)

?

Dr.med.vet.J.Troillet;St.Petersburg24.02.12

HowdoesACSwork?

BiomechanicalFactorsActivationofcytokinenetworkcascade

Imbalanceinmatrixmetabolism

ACScanhelptoinhbitthedestructionofcartilageandbreakthisviciouscycle!

Pro-inflammatoryanddestructivecytokines

Destructionofhyalinecartilageandit‘smatrix

InflammatoryMediatorsOneResponsibleCytokine

• Pro-Inflammatorycytokine• Stimulationofcollagenaseandmetallo-proteinase• Destructionofhyalinecartilageanditsmatrix

Interleukin-1Receptorantagonist

• Anti-inflammatoryandchondroprotective

Interleukin-1

• TypeIandIIIL-1receptors

IL- 1

•  Pro-inflammatory •  Syngergy with TNF-α •  Promotes destruction of

chondrocytes abd cartilage matrix

•  IL-RA •  Competitive inhibition of

IL-1 receptors •  Anti-inflammatory and

chondroprotective

ACSModeofAction

IL-1TNFα

IL-1RAIL-10

JointhomeostasisAnabolismandCatabolismareinbalance

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DiseaseModeofAction

IL-1TNFα

IL-1RAIL-10

JointDiseaseAnabolismandCatabolismareinoutofbalance

DZ

ChronicDegeneration

•  Suppressionandsynthesis– Proteoglycans– TypeIICollagenFibers

•  IncreasedDeNovo– MMP’sandAggrecanase–  IL-1onchondrocytes

Dr.med.vet.J.Troillet;St.Petersburg24.02.12

InsideaPathologicJoint

•  Irapisusedtotreatavarietyofinflammatoryconditionsinhorses:–  Jointlameness– Tendonandligamentinjuries– Muscleinjuries

irapTherapy

irapTherapy•  ControlledstudywithOAmodelinhorsesatCSU

showedpositiveeffects•  IL-1Raisgood•  IL-1isbad

•  Results*:↑ IL-1Ra(140fold)↑ IL-4(2.1fold) Anti-inflammatory↑ IL-10(2.2fold)NochangeIL-1beta NochangeTNFalpha Pro-inflammatory*FrisbeeD,KawcakC,WerpyN,ParkR,McIlwraithW,Clinical,biochemicalandhistologicaleffectsof

intra-articularadministrationofautologousconditionedseruminhorseswithexperimentallyinducedosteoarthritis.AmerJourVetResearch,Vol.68,No.3,2007.

irapTherapy-Equine•  Hasirapbeentested?

–  Thepatentediraptherapyhasbeenusedinthousandsofhorsesaroundtheworld.*

–  ControlledstudyatColoradoStateUniversity*

•  resultsindicatedthattherewassignificantclinicalandhistologicalimprovementinosteoarthritis-affectedjointsofhorsesfollowingtreatmentwithACS,comparedwithplacebotreatment.

•  HorsestreatedwithACShadsignificantclinicalimprovementinlameness,unliketheplacebogroup.

*PerOrthogenVeterinary,Duesseldorf,Germany**FrisbeeD,KawcakC,WerpyN,ParkR,McIlwraithW,Clinical,biochemicalandhistologicaleffectsofintra-articularadministrationofautologousconditionedseruminhorseswithexperimentallyinducedosteoarthritis.AmerJourVetResearch,Vol.68,No.3,2007.

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irapTherapy-Equine

•  16horseswithexperimentallyinducedOA–  8horsestreatedw/6mlof

ACSon14,21,28&35–  8horsestreatedw/saline

•  Clinicalexaminations•  Synovialfluidanalysis•  Grossexamination•  Histology•  Biochemical&biomarker

Clinical,biochemical,andhistologiceffectsofintra-articularadministrationofAutologousConditionedSeruminhorseswithexperimentallyinducedosteoarthritis

(Frisbieetal.2007)

Clinical,biochemical,andhistologiceffectsofintra-articularadministrationofAutologousConditionedSeruminhorseswithexperimentallyinducedosteoarthritis

(Frisbieetal.2007)

irapTherapy-Equine

•  Noadversereaction•  Improvedlameness•  Decreasedsynovialmembranehyperplasia•  Lessgrosscartilagefibrillation•  Lesssynovialmembranehemorrhage

PlateletRichPlasma

OsteokinePRPWhatisPRP?•  PRPisapreparationofautologousconcentratedplatelets(thrombocytes)•  PRPproducedbyOsteokinecontainstherapeuticallysignificantconcentrationsof

platelets

Howdoesitwork?•  Wholebloodistakenfromthehorseandplacedintothered-cappedOsteokine

bag•  AfterprocessinginaspecialcentrifugeapelletofPRPisgeneratedwhichcanbe

re-suspendedtotreatthehorsefromwhichitwastaken

WhatcanPRPbeusedfor?•  PRPhasbeenshowntoacceleratehealingininjuriesinvolvingfibroustissues

(tendons,ligaments,skin)andhasalsobeenusedinbonehealing.

Thrombocytes/Platelets•  Plateletsdomorethancoagulation•  α-granulescontainGFindispensablefornormalwoundhealing

•  1stphase:uponinjurytissuethromboplastininducesplateletcoagulationandreducesbleeding

•  2ndphase:plasmafibrinogenbreakstofibrin,furthercoagulationstopsbleeding.AggregationofplateletsstartsoffGFsecretion.GFinducecellproliferation,stemcelldifferentiationandmigration.Macrophagesmigratetothesiteandtakeovertheplaceofplateletsassourceofcytokines.

Platelets

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Osteokine•  Concentratedplateletsarearichsourceof(autologous)growth

factors.

•  Ifplateletsarefreshandfullyfunctional->optimalsequentialgrowthfactoroutput

•  TotaloperatingtimetoretrievePRPisabout40min

•  Averageplateletconcentrationof1,300,000µL±250,000/µlfrom50mLinafinalvolumeof5-10mL

•  Concentrationfactorofplateletsroutinely3-6fold

Average platelet count in native blood: 275,000 / µl

Average platelet count in 8 mL of PRP yielded by Osteokine:

1,300,000 / µl

Characteristics of the Osteokine® PRP system

FactorspresentinOsteokine®PRP•  PDGF•  IGF•  FGF•  TGF•  HGF•  VEGF

•  EGF•  Pluscytokines•  suchasIL-1Ra,IL-6

BenefitsofOsteokine®PRP•  Acceleratedwoundhealinginmultipletissuesincludingskin,tendon,ligament

•  Highercollagencontentinhealinginjuriesresultinginlowerscarformation

•  Reducedperioperativetraumaticpain•  Lowerinfectionrates•  Injectimmediatelyintocorelesions/superficiallesion

•  Improvedtissueformation(e.g.morecollagen)•  improvedbonematuration

TwooptionstoLamenessOsteokine(PRP)•  Useinsofttissue(fibrous)

injuries–  Tendons–  Ligaments–  Skin,muscle&bone

•  Easilyproducedin-clinicwithin30-40minutes

•  Treatinjuredhorse1time•  HighPRPpotency

irap•  Useintra-articular(IA)totreat

osteo-arthritis•  Processedin-clinicforeaseofuse•  Usedinthousandsofhorses

aroundtheworldsince2004•  ProveneffectivebyCSU•  3initialtreatmentsthen

maintenance(horsedictatesneed)

•  Highpotencyandyield•  Proven&trusted!

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Conclusion

•  irapIAworkswithgreatsuccessagainstlamenesscausedbymildtomoderateOA

•  OsteokineisafantasticPRPsystemtotreatsofttissueinjuries

•  Onlyiraphasbeentestedincontrolledstudiesandinrealworldsituations.

Discussion