Apoptosis & Ros Prof Pur

7/25/2019 Apoptosis & Ros Prof Pur

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THE MOLECULAR BASISTHE MOLECULAR BASIS

OF APOPTOSISOF APOPTOSIS

OlehOleh

Prof. dr. Purnomo SuryohudoyoProf. dr. Purnomo Suryohudoyo


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APOPTOSISAPOPTOSIS NECROSISNECROSIS

1.1. OccurrenceOccurrence Physiological ofPhysiological ofPathologicalPathological Always PathologicalAlways Pathological

.. Cell!sCell!s

In"ol"ementIn"ol"ementActi"eActi"e Passi"ePassi"e

#.#. In$ammationIn$ammation %% &&

'.'. (or)hological(or)hologicalfeaturesfeaturesCell mem*raneCell mem*raneCell sha)eCell sha)e

+inal outcome+inal outcome

IntactIntactShrin,ing- *ecomeShrin,ing- *ecomemore rounded-more rounded-

loose contact withloose contact withneig*ouring cellsneig*ouring cells

A)o)totic *odiesA)o)totic *odies

amagedamagedSwollenSwollen

Cell lysasCell lysas

/./. Cell remnantCell remnantremo"alremo"al

Neig*ouring cellsNeig*ouring cellsresident 0Oresident 0O

In$ammatoryIn$ammatory)hagocyt es NO-)hagocyt es NO-

P0NP0NApoptosis can be induced by extracellular as well as intracellular stimultApoptosis can be induced by extracellular as well as intracellular stimult

APOPTOSIS NECROSISAPOPTOSIS NECROSIS


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APOPTOTIC PAT(2A3SAPOPTOTIC PAT(2A3S

4 EXTRINSIC PATHWAYEXTRINSIC PATHWAYINITIATE +RO0 O5TSIE T(E CE66INITIATE +RO0 O5TSIE T(E CE66RE75IRE8RE75IRE8

SI9NA6IN9 0O6EC56E 8SI9NA6IN9 0O6EC56E 8DEATH LIGANDDEATH LIGANDAN A SPECIA6 RECEPTOR 8AN A SPECIA6 RECEPTOR 8

DEATH RECEPTORDEATH RECEPTOR

4 INTRINSIC PATHWAYINTRINSIC PATHWAYINITIATE +RO0 INSIE T(E CE66INITIATE +RO0 INSIE T(E CE66RE75IRE T(E PRESENCE O+ NITOC(ONRIARE75IRE T(E PRESENCE O+ NITOC(ONRIA

THESE 2 PATHWAYS MAYBE INTERCONNECTEDTHESE 2 PATHWAYS MAYBE INTERCONNECTED


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DEATH LIGANDDEATH LIGAND

DEATH RECEPTORDEATH RECEPTOR

TRANSDUCTOR NOLECULESTRANSDUCTOR NOLECULES

INITIATOR CASPASEINITIATOR CASPASE(Caspase 8)(Caspase 8)

(FLICE)(FLICE)

EXCECUTOR CASPASESEXCECUTOR CASPASES

Cleaved protein substrateCleaved protein substrate

APOPTOSISAPOPTOSIS

FLICE : Fas like interleukin converting enzymeFLICE : Fas like interleukin converting enzyme

CASPASE ACTI:ATIONCASPASE ACTI:ATION;1


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DAMAGING STIMULIDAMAGING STIMULIOndantsOndants

BaxBaxCeramideCeramideCa ++Ca ++

HITOCHONDRIAHITOCHONDRIA

OPENING OF PT-POREOPENING OF PT-PORE

Apof-1Apof-1Cytochrome CCytochrome C

d-ATPd-ATP

INITIATOR CASPASE C!"p!"e #$INITIATOR CASPASE C!"p!"e #$

E%CECUTOR CASPASESE%CECUTOR CASPASES

APOPTOSISAPOPTOSIS PT & Perme!'()(ty tr!*"(t(o*PT & Perme!'()(ty tr!*"(t(o*

CASPASE ACTI:ATION;1


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DDDD : DEATH DOMAIN: DEATH DOMAINDEDDED : DEATH EFFECTOR DOMAIN: DEATH EFFECTOR DOMAINTNFTNF : TUMOR NECROSIS FACTOR: TUMOR NECROSIS FACTORTNF-R1TNF-R1 : TNF-RECEPTOR 1: TNF-RECEPTOR 1

FADDFADD : Fas ASSOCIATED DEATH: Fas ASSOCIATED DEATHDOMAIN PROTEIN! DOMAIN PROTEIN!TRADDTRADD : TNF-R ASSOCIATED DEATH: TNF-R ASSOCIATED DEATH

DOMAIN PROTEIN! DOMAIN PROTEIN!F"ICEF"ICE : FADD-"I#E ICE: FADD-"I#E ICEICEICE : INTER"EU#IN 1B: INTER"EU#IN 1B

CON$ERTIN% EN&'ME CON$ERTIN% EN&'ME


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EAT( 6I9ANS- EAT( RECEPTOR EAT( 6I9ANS- EAT( RECEPTOR

TRANS5CTORSTRANS5CTORS

LIGAND (L)LIGAND (L) RECEPTOR (R)RECEPTOR (R) TRANSDUCTORTRANSDUCTOR

1.1. Fas LFas L(CD95L)(CD95L)

2.2. TNFTNF

.. A!" LA!" L

#.#. TRAILTRAIL

FasFas(CD 95)(CD 95)(A!" 1)(A!" 1)(DR 2)(DR 2)

TNFRTNFR

(DR 1)(DR 1)A!" A!" (TRAMP)(TRAMP)(DR )(DR )

TRAIL$R1TRAIL$R1(DR #)(DR #)TRAIL$R2TRAIL$R2

(DR 5)(DR 5)TRAIL$RTRAIL$R(TRID)(TRID)(D% R1)(D% R1)TRAIL$R#TRAIL$R#(TRUNDD)(TRUNDD)(D% R2)(D% R2)

FADDFADD

TRADD& FADDTRADD& FADD

TRADD& FADDTRADD& FADD

''

''

$ (%"*$ (%"*+%!,"+)+%!,"+)

$ (%"*$ (%"*+%!,"+)+%!,"+)

A((re)iati*ns :A((re)iati*ns :Fas : Fi(r*(ast ass*,iated CD : C.ster */Fas : Fi(r*(ast ass*,iated CD : C.ster */di//erentiati*n APO : A0*0t*sis DR : Deatdi//erentiati*n APO : A0*0t*sis DR : Deatre,e0t*r D,R : de,*2 re,e0t*r FADD : Fasre,e0t*r D,R : de,*2 re,e0t*r FADD : Fasass*,tiated deat d*mein TNF : t.m*rass*,tiated deat d*mein TNF : t.m*rne,r*sis /a,t*r TRADD : TNF ass*,iatedne,r*sis /a,t*r TRADD : TNF ass*,iateddeat d*main TRAMP : TNF reateddeat d*main TRAMP : TNF reateda0*0t*sis mediatin3 0r*tein TRAI" : TNFa0*0t*sis mediatin3 0r*tein TRAI" : TNFreated a0*0t*sis ind.,in3 i3and TRID :reated a0*0t*sis ind.,in3 i3and TRID :TRAI" re,e0t*r 4it*.t intra,e.ar d*mainTRAI" re,e0t*r 4it*.t intra,e.ar d*mainTRUNDD : tr.n,ated deat d*mainTRUNDD : tr.n,ated deat d*main


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APOPTOSIS 8 RO6E O+ (ITOC(ONRIAAPOPTOSIS 8 RO6E O+ (ITOC(ONRIA


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CASPASESCASPASES

CASPASESCASPASESPRO$APTOPTOTICPRO$APTOPTOTIC

ANITIATORANITIATOR

CASPASECASPASE 2 (')2 (')99

--

1 (')1 (')

EFFECTOREFFECTOR

CASPASECASPASE 2 (')2 (')

//

00

PROCASPASEPROCASPASE LOW LOW

ACTIITYACTIITY

CASPASECASPASE HIGH ACTIITY HIGH ACTIITY

PROINFLAMMATORYPROINFLAMMATORY

CASPASECASPASE 2 (')2 (')99

--

1 (')1 (')


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CASPASE S5=STRATESCASPASE S5=STRATESSUBSTRATESSUBSTRATES EFFECTSEFFECTS RESULTSRESULTS

1.1. ICAICA

.. Nuclear 6aminsNuclear 6amins

#.#. 9elsolin9elsolin

'.'. +A>+A>

/./. PA>%PA>%

?.?. NA%P>NA%P>

Release of CARelease of CA

e)olymerisatione)olymerisation

Acti"e gelsolinActi"e gelsolin

Clea"ageClea"age

Clea"ageClea"ageClea"ageClea"age

NA Clea"ageNA Clea"age

issa))earance of nuclearissa))earance of nuclearmem*rane- chromatinmem*rane- chromatincondensationcondensation

Actin @lament degradationActin @lament degradation

Inhi*ition of NA re)airInhi*ition of NA re)air

Appro+(m!te)y , prote(*" !re .*o/* "0'"tr!te" of c!"p!"e" ho/e2er theAppro+(m!te)y , prote(*" !re .*o/* "0'"tr!te" of c!"p!"e" ho/e2er the

re)!t(o*"h(p of the(r c)e!2!3e to the "pec(f(c morpho)o3(c ch!*3e" !*d ce)) de!thre)!t(o*"h(p of the(r c)e!2!3e to the "pec(f(c morpho)o3(c ch!*3e" !*d ce)) de!th

rem!(* for mo"t of them 0*.*o/*4rem!(* for mo"t of them 0*.*o/*4

A((re)iati*ns :A((re)iati*ns :CADCAD & C!"p!"e !ct(2!ted deo+y-r(2o*0c)e!re& C!"p!"e !ct(2!ted deo+y-r(2o*0c)e!re

ICADICAD & I*h('(tor of CAD& I*h('(tor of CAD

FA5FA5 & Foc!) !dhe"(o* .(*!"e& Foc!) !dhe"(o* .(*!"e

PA5-6PA5-6 & P61 !ct(2!ted .(*!"e-6& P61 !ct(2!ted .(*!"e-6

DNA-P5DNA-P5 & DNA !ct(2!ted prote(* .(*!"e& DNA !ct(2!ted prote(* .(*!"e


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0O56ATION O+ APOPTOSIS ;1


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0O56ATION O+ APOPTOSIS0O56ATION O+ APOPTOSIS

A33+4a,"6s 7A33+4a,"6s 766 8 eath ligands- R 8 eath rece)tors- cR 8 ecoy rece)tors8 eath ligands- R 8 eath rece)tors- cR 8 ecoy rece)tors

+6IP+6IP 8 +A%li,e ICE inhi*itory )rotein8 +A%li,e ICE inhi*itory )rotein

+6ICE+6ICE 8 +A%li,e interleu,in con"erting enFyme ;+A%li,e ICE


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=IO6O9ICA6 RO6E O+ APOPTOSIS=IO6O9ICA6 RO6E O+ APOPTOSIS

1.1. RE0O:A6 O+ 5NNEEE CE66SRE0O:A6 O+ 5NNEEE CE66S

;e.g. during em*ryonal de"elo)ment


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CASPASECASPASE

INDEPENDENTINDEPENDENT

APOPTOSISAPOPTOSIS


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NMDANMDA 7 N 8 ,:*; 8 D 8 as!a+,a,7 N 8 ,:*; 8 D 8 as!a+,a,

NOSNOS 7 N,+" 6?+"6a;)

PARPPARP 7 P";* (ADP$+3"s) !";*+as7 P";* (ADP$+3"s) !";*+as

PARGPARG 7 P";* (ADP$+3"s)


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APOPTOSISAPOPTOSIS

TOO MUCH :TOO MUCH :55 DE%ENERATI$E DISEASESDE%ENERATI$E DISEASES

E6%6 : A"&HEIMER DISEASESE6%6 : A"&HEIMER DISEASES : C6$6A6 STRO#E!: C6$6A6 STRO#E!

TOO "ITT"E :TOO "ITT"E :

55 CANCERCANCERMUTATION IF E7TENSI$E 5 P89 5 APOPTOSISMUTATION IF E7TENSI$E 5 P89 5 APOPTOSISIF P89 IS INACTI$ATED 5 NO APOPTOSISIF P89 IS INACTI$ATED 5 NO APOPTOSIS55 CANCERCANCER


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R.O.S.R.O.S.

;REACTI:E OH39EN SPECIES


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R.O.S.R.O.S. ;1


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R.O.S.R.O.S. ;


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4 NON$RADICALNON$RADICAL(3RO9EN PEROHIE(3RO9EN PEROHIE ((OO ( O O (( O O (

(3POC(6ORITE ION(3POC(6ORITE ION ClOClOJJ Cl OICl OIKK

SIN96ET OH39ENSIN96ET OH39EN OO O OIO OI

4 FREE RADICALFREE RADICAL(3ROH36 RAICA6(3ROH36 RAICA6 44O(O( ( O( O44

PEROH36 RAICA6PEROH36 RAICA6 44OO(OO( ( O O( O O44

S5PEROHIE IONS5PEROHIE ION 44OO IO OIO O44KK

R.O.S.R.O.S. ;#>O ? OO ? O>>@ATER H@ATER HO!O! H ? O ? HH ? O ? H

H'DRO7'" ION OHH'DRO7'" ION OH!! H ? OIH ? OI


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1.1. T(E RE5CTION O+ OT(E RE5CTION O+ OTO (TO (O IN:O6:ES T(EO IN:O6:ES T(E

TRANS+ER O+ ' ;+O5R< E6ECTRONSTRANS+ER O+ ' ;+O5R< E6ECTRONS

OO & ' e& ' eJ & ' (J & ' (&& L ( L (OO

.. T(ESE TRANS+ERS OCC5R ONE E6ECTRONT(ESE TRANS+ERS OCC5R ONE E6ECTRON

EAC( TI0EEAC( TI0E#.#. A66 R.O.S. EHCEPT ClOA66 R.O.S. EHCEPT ClOJJ 11OO CAN =ECAN =E

CONSIERE AS =EIN9 T(E RES56T O+CONSIERE AS =EIN9 T(E RES56T O+

INCO0P6ETE OH39EN RE5CTIONINCO0P6ETE OH39EN RE5CTION

R.O.S.R.O.S. ;'OOH PERO7'" RADICA"!

66 T@O E"ECTRONS :T@O E"ECTRONS : OO + e + H+ e + H++ 5 H5 HOOH'DRO%EN PERO7IDE!H'DRO%EN PERO7IDE!

9696 THREE E"ECTRONS :THREE E"ECTRONS : OO + 9 e + 9 H+ 9 e + 9 H++ 5 H 5 HO + >OH H'DRO7'" RADICA"!O + >OH H'DRO7'" RADICA"!


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OHIANTS +REE RAICA6SOHIANTS +REE RAICA6SOHIANTS 8OHIANTS 8 ARE E6ECTRON ACCEPTORSARE E6ECTRON ACCEPTORS

i.e. IT ATTRACTS E6ECTRONSi.e. IT ATTRACTS E6ECTRONS+e+e ++++++& eJ& eJ +e+e ++++

+REE RAICA6S 8+REE RAICA6S 8

=ECA5SE O+ T(E PRESENCE O+ AN 5NPAIRE E6ECTRON=ECA5SE O+ T(E PRESENCE O+ AN 5NPAIRE E6ECTRONA6SO S(O2 A TENENC3 TO ATTRACT E6ECTRONS AN CANA6SO S(O2 A TENENC3 TO ATTRACT E6ECTRONS AN CAN

T(5S =E CONSIERE OHIANTST(5S =E CONSIERE OHIANTSa.a. E6ECTRON ONOR ANOT(ER RAICA6E6ECTRON ONOR ANOT(ER RAICA6

RR114 & R4 & R44 RR118R8R ;R ;R11 R R


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OHIANTS +REE RAICA6SOHIANTS +REE RAICA6S

1.1. +REE RAICA6S ARE 0ORE AN9ERO5S T(AN+REE RAICA6S ARE 0ORE AN9ERO5S T(AN

NON%RAICA6S OHIANTS =ECA5SE T(E CANNON%RAICA6S OHIANTS =ECA5SE T(E CANTRI99ER C(AIN REACTIONTRI99ER C(AIN REACTION

.. 2(EN A +REE RAICA6 ATTAC>S A NON%RAICA6- IT 2I66 RES56T2(EN A +REE RAICA6 ATTAC>S A NON%RAICA6- IT 2I66 RES56TIN T(E +OR0ATION O+ A NE2 RAICA6 8IN T(E +OR0ATION O+ A NE2 RAICA6 8

RR114 & R4 & R ( ( RR11 ( & R ( & R44

#.#. T(E NE263 +OR0E RAICA6 CAN A9AIN ATTAC>T(E NE263 +OR0E RAICA6 CAN A9AIN ATTAC>A NON%RAICA6 8A NON%RAICA6 8

RR4 & R4 & R## ( ( RR ( & R ( & R##44

'.'. T(IS PROCESS CAN =E REPEATE A9AIN AN A9AIN RES56TIN9 INT(IS PROCESS CAN =E REPEATE A9AIN AN A9AIN RES56TIN9 INA C(AIN REACTION 8A C(AIN REACTION 8

RR##4 & R4 & R'' ( ( RR## ( & R ( & R''44

RR''4 & R4 & R// ( ( RR'' ( & R ( & R//4 ETC4 ETC

/./. S5C( A C(AIN REACTION 2I66 ON63 STOP 2(EN RAICA6SS5C( A C(AIN REACTION 2I66 ON63 STOP 2(EN RAICA6S0EET 80EET 8

RR114 & R4 & R1144 RR11 R R11

RR114 & R4 & R44 RR11 R R

RR4 & R4 & R44 RR R R ETC ETC


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+OR0ATION O+ R.O.S.+OR0ATION O+ R.O.S.

RE6ATIONS(IP =ET2EEN R.O.S.RE6ATIONS(IP =ET2EEN R.O.S.

;S500AR3


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+OR0ATION O+ OT(ER R.O.S.+OR0ATION O+ OT(ER R.O.S.

44OO(OO(44OOJ & (J & (++ 4OO(4OO(

44O(O(

44OOJ & (J & (OO OO & O(J & 4O(& O(J & 4O(

((OO & +e& +e++++MCuMCu++ +e+e++++++MCuMCu++++ & O(J & 4O(& O(J & 4O(

ClOJClOJ((OO & ClJ& ClJ ((O & ClOJO & ClOJ

OO((OO & ClOJ& ClOJ ((O & ClJ & OO & ClJ & O

M=ELOPERO%IDASE$M=ELOPERO%IDASE$

M=ELOPERO%IDASE$M=ELOPERO%IDASE$

FENTON REACTION$FENTON REACTION$

HA8ER-9EISS REACTIONHA8ER-9EISS REACTIONFeFe> > >> > >

C0C0>>>>$$

11

11


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+OR0ATION O+ PRI0AR3 R.O.S.+OR0ATION O+ PRI0AR3 R.O.S.

44OOJJ

TRANSITION 0ETA6S 8TRANSITION 0ETA6S 8OO & +e& +e++++;Cu;Cu++


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E++ECT O+ R.O.S.E++ECT O+ R.O.S.

1.1. ROS CAN REACT WITH MANYROS CAN REACT WITH MANYSUBSTANCES 7SUBSTANCES 7%% SI0P6E CO0PO5NS 8SI0P6E CO0PO5NS 8

A0INO ACIS- +ATT3 ACIS-A0INO ACIS- +ATT3 ACIS- C(O6ESTERO6 S59ARC(O6ESTERO6 S59AR%% CO0P6EH S5=STANCES 8CO0P6EH S5=STANCES 8

PROTEINS- NAPROTEINS- NA

2.2. THE MOST DAMAGING EFFECT OF ROSTHE MOST DAMAGING EFFECT OF ROS

HOWEER& IS ON IMPORTANTHOWEER& IS ON IMPORTANTCOMPONENTS OF CELLS 7COMPONENTS OF CELLS 7%% 0E0=RANE 6IPIS0E0=RANE 6IPIS

%% PROTEINSPROTEINS

%% NANA


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E++ECT O+ ROS ON 0E0=RANE 6IPISE++ECT O+ ROS ON 0E0=RANE 6IPIS


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E++ECT O+ ROS ON PROTEINSE++ECT O+ ROS ON PROTEINS


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E++ECT O+ ROS ON NAE++ECT O+ ROS ON NA;1


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E++ECT O+ ROS ON NAE++ECT O+ ROS ON NA;


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E++ECT O+ ROS ON NAE++ECT O+ ROS ON NA;#


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ANTI%OHIANTSANTI%OHIANTS;1


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ANTI%OHIANTSANTI%OHIANTS;


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PRE:ENTI:E ANTI%OHIANTSPRE:ENTI:E ANTI%OHIANTS1.1. 0ETA6 =ININ9 PROTEINS0ETA6 =ININ9 PROTEINS

+e+e 8 TRANS+ERIN AN +ERRITIN8 TRANS+ERIN AN +ERRITIN

CuCu 8 CER56OP6AS0IN AN A6=50IN8 CER56OP6AS0IN AN A6=50INPRE:ENTS 8 (A=ER%2EISS REACTION 8PRE:ENTS 8 (A=ER%2EISS REACTION 8

44OOJJ & ( & (OO OO & O(& O(JJ && 44O(O(

+c+c&&&&&&;Cu;Cu&&&&


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C(AIN =REA>IN9 ANTI%OHIANTC(AIN =REA>IN9 ANTI%OHIANT

:ITA0IN E ;TOCOP(ERO6S< 8 Toc( 8:ITA0IN E ;TOCOP(ERO6S< 8 Toc( 81.1.Toc( & 6Toc( & 644 6( & Toc4 6( & Toc4

;TOCOP(ERO6< ;6IPI RAICA6< ;6IPI< ;TOCOP(ER36 RAICA6


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TOCOP(ERO6 ;:ITA0IN E


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ASCOR=IC ACI ITSASCOR=IC ACI ITSERI:ATI:ESERI:ATI:ES

9?P9?P ANTI OHIANTANTI OHIANT


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9?P9?P ANTI%OHIANTANTI%OHIANT

E+ENSE S3STE0E+ENSE S3STE0GLUTATHIONE PLAYS AN IMPORTANT ROLE IN ANTI$OXIDANT DEFENSE&GLUTATHIONE PLAYS AN IMPORTANT ROLE IN ANTI$OXIDANT DEFENSE&

IT CAN REACT DIRECTLY WITHIT CAN REACT DIRECTLY WITH OH (HYDROXYL RADICAL) 7OH (HYDROXYL RADICAL) 7

2 GSH 22 GSH 2 OHOH GSSG 2 HGSSG 2 H22

OR WITH HOR WITH H22OO22(HYDROGEN PEROXIDE) CATALYED BY GLUTATHION(HYDROGEN PEROXIDE) CATALYED BY GLUTATHION

PEROXIDASE (GPPEROXIDASE (GPXX) 7) 7

2 GSH H2 GSH H22OO22 GSSG 2 HGSSG 2 H22OO

THE RATIO 7 (GSH 7 GSSG) IN NORMAL CELLS ARE ALWAYS HIGH& SOTHE RATIO 7 (GSH 7 GSSG) IN NORMAL CELLS ARE ALWAYS HIGH& SO

THAT THERE MUST BE A MECHANISM TO REGERATE GSH FROM GSSG.THAT THERE MUST BE A MECHANISM TO REGERATE GSH FROM GSSG.

THIS IS DONE BY THE ACTION OF 2 ENYMES 7THIS IS DONE BY THE ACTION OF 2 ENYMES 7

GLUTATHIONE REDUCTASE AND GLUCOSE $G$ PHOSPHATEGLUTATHIONE REDUCTASE AND GLUCOSE $G$ PHOSPHATE

DEHYDROGENASE (G/PD)DEHYDROGENASE (G/PD)

GSSG NADPH HGSSG NADPH H 2 GSH NADP2 GSH NADP

GLUCOSE 8 / 8 PHOSPHATE NADPGLUCOSE 8 / 8 PHOSPHATE NADP / 8 PHOSPHOGLUCONATE/ 8 PHOSPHOGLUCONATE

NADPH H NADPH H

GPGP%%$$

GLUTATHIONEGLUTATHIONE

REDUCTASE$REDUCTASE$

G?PD$G?PD$

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Apoptosis & Ros Prof Pur