Hemoglobinmetabolismanditsclinicalapplications
Dr.Rohini CSane
FunctionsofHemoglobinmoleculeinoxygentransport
FunctionsofHemoglobinmoleculeincarbondioxidetransport
DegradationofHemetobilepigments
Macrophagesofreticuloendothelial (RE)inspleen/liver/bonemarrow-Hb
RBCà 12Odays
Nonprotein‘Heme’Globin
6gm /dayheme brokendown/resynthesisà Bilirubin(300mg/day)=250mg/dayHb+50mg/daymyoglobin
Fateglobin(reutilization)
• FormationofHb(resynthesis)• Degradedtoaminoacids–metabolismincludingformationofHb
SourcesofHeme
Heme
20%immatureRBC/Myoglobin/globin/cytochrome/peroxidase/catalaseTrppyrrolase80%RBCHb
(GREEN–excretedbybirds&hibians)→
Transferrin
←αGlobinchains
←βGlobinchains
RBC(Agederythrocytes)withHbphagocytosisbymacrophages→
specificityforα methylenebridge←
Presenceinmammals←
Yellowwith36H←
→Bilirubin- AlbumincomplexinBlood
+
Hemeoxygenase
1. Microsomalenzymes2. NADPH,O₂,Methylenebridgesbetweentwopyrroleringsà
biliverdin3. Fe²⁺à Fe³⁺
Heme oxygenaseHemeBiliverdin+Fe³⁺+CO• Biliverdinà excretedbybirds,amphibians&mammals
DegradationofHemetobilepigments
BiliverdinreductaseBiliverdin(green) Bilirubin(nonfunctionalthereforeexcreted)1gmhemoglobin*à 35mgBilirubinBiliverdin+ Bilirubinà Hemederivatives• (*sources- 80%RBC+10%ineffectiveerythropoiesis+10%Myoglobin)• 6gm /dayheme brokendown/resynthesisà Bilirubin(300mg/day)=250mg/dayHb+50mg/daymyoglobin
TransportofBilirubintoLiver
• Bilirubin- (lipophilic–insolubleinwater)• Bilirubin+Albuminà Transportedinplasma• Per100mlplasmaà 25mgBilirubinboundtighttoAlbumin• BilirubinhaslowaffinityforAlbuminà easilydetached&entertissue
Drugs :sulphonomides /salicylates/PenicillindisplaceBilirubinfromAlbuminbindingsites
BilirubinentersCNS
Damagetoneuronsà Kernicterus
DegradationofHemetobilepigments
Conjugation0fBilirubinLiverAlbumin–Bilirubincomplex
IUptake carriermediatedactivetransportSinusoidalsurfaceofhepatocytes
Bilirubin+protein(Ligandin )2UDPGlucoronate
IIConjugation UDPEndoplasmic-reticulum(hepaticmicroosomes ) BilirubinGlucuronyl transferase(chromosome2)
Bilirubin+2moleculesofGlucuronate80%BilirubinDiglucuronide+20%Bilirubinmonoglucuronide
(MOAT–multiplespecificorganiciontransport)-presentincanalculi
IIIsecretionBilirubinOligonucleotide
Bilesalts
FateofBilirubininhumanbody
FactorsaffectingconjugationofBilirubininliver
1. DrugslikePrimaquine/Novobiocin/Chloramphenicol,/Androgen/Pregnanediolinterferewithconjugationprocessmaycausejaundice.
2. DecreaseconcentrationofUDP-Glucuronyl Transferase3. PhenobarbitalinducesUDP-Glucuronyl Transferase
Production&excretionofBilirubinRetiiculoendothelialsystem(RES)
Hb↓
Biliverdin(38H)↓
Bilirubin(36H)
LiverBilirubin
Diglucuronide
BilirubinDiglucuronide↓DeconjugationFreeBilirubin(36H)
↓reductionUrobilinogen (44H)byEcoli
(colorless)
Reductionofvinylgroupofstercobilinogen (48H)
↓stercoblin(46H)
DARKBROWN /200MG/DAY
BLOODUROBILINOGEN
KIDNEYUROBILINOGEN
UROBILLINOGEN(44H)↓
UROBILININURINE(42H)(<4GM/DAY)
BILEDUCT→←PORTALCIRCULATION
EHC↓
←SMALLPORTION
EHC–Entero hepaticcirculation
ExcretionofBilirubinintobile
ConjugatedBilirubinActivetransport*(againstconcentrationgradient)Ratelimitingstep
Bile>98%Bilirubinentersbileinconjugatedform
*InducedbyPhenobarbitone
Production&excretionofBilirubin
• ExcretionofBiliverdinà greencolorstool(children&prolongedantibioticstherapy)• Blackcolorstoolduringconstipation• Schlesingertest:Stercobilin (SB)&Urobilin (UB)+Zn²⁺à greenfluroscent
Fouchet’s DiagnostictestforBilirubininurine
Bilirubin–yellow,Biliverdin–green ,Bilinofuschin –Red, Bilicyanin-violet
←URINE→
Gmelin’s Test
←CONCHNO₃→
Bilirubin–yellow,Biliverdin–green ,Bilifuschin–Red, Bilicyanin-violet
FateofBilirubinBilirubinGlucuronides
Hydrolysis↓bacterialenzyme(β Glucuronidase )Bilirubin↓smallportionreabsorbed
Urobilinogen (colorless)à SBG(Erhlich +ve )↙↘
Stercobilin Urobilin(excretedinstool-)(excretedinurinebykidney)↓↓BrowncolorofstoolpaleyellowcolorofurineSerumBilirubinà VanDerBergTest,UrineBilirubinà Fouchet’s Test,Gmelin Test
Conjugatedandunconjugatedbilirubin(Directandindirectbilirubin)
Stercobilin andUrobilin
FormationofconjugatedbilirubininHepatocytes
TypesofBilirubinFreebilirubin Conjugatedbilirubin
1 InH₂O insoluble soluble2 Inalcohol soluble soluble3 Normalplasmalevels 0.2mg/dl 0.2-0.8mg/dl4 Inbile Absent present5 Inurine Alwaysabsent Normallyabsent(presentin
hemolytic&Obstructivejaundice)6 AbsorptionfromGIT Absorbed NotAbsorbed7 Diffusionintissue Diffusestocauseyellow Dosenotdiffuse
8 VanderBerghTest Indirectpositive directpositive
Bilirubin&itsreductionproductsnumberofHatoms color
Bilirubin(BR) 36 RedyellowMeso bilirubin(MB) 40 yellowUrobillinogen (UBG) 44 colorlessStercobilinogen (SBG) 48 colorlessUrobilin (UB) 42 OrangebrownStercobillin (SB) 46 Darkbrown
Jaundice- HyperbilirubinemiaPhysiological– TotalBilirubinconcentrationinserum=(0.2- 0.8mg/dl)
0.2-0.6mg/dl(unconjugated)0- 0.2mg/dl(conjugated)Yellowcolorsclera&skin(depositionofbilirubin)-conc ofserumbilirubin>2mg/dl
q ConjugatedHyperbilirubinemiaq UnconjugatedHyperbilirubinemiavSymptoms1. Nausea2. Vomiting3. AppetiteØ LatentJaundice(1-2mg/dl)
*Diagnosticlaboratorytosetupownqualitycontrolsforreferenceranges
Prehepatic,Hepaticandposthepaticjaundice– aclassificationbasedonsite(causeofofHyperbilirubinemia)
ClassificationofJaundicebasedontypeofBilirubin
Intrahepaticjaundice(Genetic/inheritedcauses)
Cause• HemolyticJaundice
• Causes– HemolysisMalariabloodtransfusionsicklecellanemia
• LiverfailstoconjugateexcessofBilirubin
• Therefore↑unconjugatedbilirubin↑Urobilinogen↑stercobilinogen(browncolorstool)
• SGPT/ALP-Normal
• HepaticJaundice• Causes–dysfunctionofLiverHepatitisalinfectionpoisons/toxinscirrhosis/CCF
• ↑unconjugatedbilirubin↑conjugatedbilirubin↑Urobilinogen↑stercobilinogen(browncolorstool)↑SGPT/ALP
• ObstructiveJaundice• Gallstonestoolcontainsfatunavailabilityofbilesalts
• ↑conjugatedbilirubin↑Urobilinogen↓ stercobilinogen (palecolorstool)↑SGPT/ALP
ComparisonofHemolytic/hepaticandobstructiveJaundice
DifferentialdiagnosisJaundice/Icterus
HemolyticDiseaseofAdultsà unconjugatedHyperbilirubinemia
1. increaseinunconjugatedbilirubininblood2. Absenceofbilirubininurine3. Excretionofurobilinogenà (EhrlichTestpositive)4. Excretionofstercobilinogen infaecesvDiseasesassociatedwitha) congenitalspherocytosisb) G6PDdeficiencyc) Autoimmunehemolyticanemiad) Toxincarbontetrachloride
HemolyticDiseaseofAdultsà unconjugatedHyperbilirubinemiaIncompatibilitybetweenmaternal&fetalbloodgroupsAntiantibodiesABOIgM typecannotbetransferredtoplacentavRh incompatibilityFetusmotherRh(+ve )Rh(-ve )RBCRBC elicitimmuneresponseAnti-D(IgG)
Destruction Anti-D (IgG )
Of RBC←Placenta
Secondpregnancy(beforebirth–destructionofRBC)à CHILDbornwithseverehemolyticdiseaseàErythroblastosis fetalis
Erythroblastosis Fetalis• SerumBilirubinà >20mg/dlà nomoreboundtoAlbumin
• BilirubinBrain(Kernicterus-depositionofbilirubininbrain)• Basalgangliaàmentalretardation• ↓ATPasemitochondriaà fits,spasticity,toxicencephalitis• Treatment:(1)phototherapybeforeage<1yearà
isomerizationZZà ZE(2)Bloodtransfusion
HepatocellularJaundice
1. ViralHepatitis(virusesA,B,C,DorG)à purehepatocellulardiseases
2. ↓conjugatedbilirubintherefore↑freeBilirubin3. Inflammatoryodemaofcellsà intracellularcanalculicompressed
à obstruction(atsiteofbileformation)4. Increaseobstructionà↑obstructionà↑conjugatedBilirubin
thereforeBiphasic5. Bilirubinuria6. UrobilinogenàNORMAL ordecreasedinhepatocellularJaundice
ObstructiveJaundiceà conjugatedHyperbilirubinemia1. ↑conjugatedbilirubin2. Bilirubininurine3. ↓urobilinogen (nilifobstructiveiscomplete)4. Faeces claycolor(↓stercobilinogen )5. Absenceofbilesaltsà Stetorrhoeamayresult6. CausesIIntrahepaticCholestasisa) Activehepatitisb) Biliarycirrhosisc) Lymphomasd) Hepatomae) Viralhepatitis
ObstructiveJaundiceà conjugatedHyperbilirubinemia
• CausesIIExtrahepaticCholestasis1. Stonesinbiliarytract2. Stonesingallbladder3. Biliaryatresia4. Carcinomaheadofpancreasà lymphglandsenlarged5. Porta hepatitis
Acquiredhyperbilinogen(unconjugatedhyperbilinogen)
vPhysiologicalJaundiceofnewborn(neonates)1. Afterseconddayoflife,transienthyperbilinogen –Jaundice2. ↑rateofdestructionofRBC(transient)3. à immaturehepaticsystemofconjugation4. à Bilirubindonot exceed5mg/dl5. à 2weeksphototherapy+barbitonetoinduceconjugation(ZZ
bilirubinà ZE+EEà excretedwithoutconjugationinurine)6. Undueprolongationà crenism7. Breastmilkjaundice(estrogenà↓enzymeGlucuronyl transferase)
Bilepigment DiagnosticTestBilirubin VanDerBergh(serum),Fouchet’s Test&Gmelin ‘stest
inUrineUrobilinogen (UBG) Ehrlich’sTestUrobilin Schlesinger’stest
DIRECTBILIRUBIIN- Azo pigment(pH5)àPURPLECOLOR-↓
INDIRECTREACTION–FREEBILIRUBIN(H2Oinsoluble,alcoholsoluble)
*Diagnosticlaboratorytosetupownqualitycontrolsforreferenceranges
EnzymeestimationshelpindifferentialdiagnosisofJaundice
EnzymeestimationshelpindifferentialdiagnosisofJaundice
*Diagnosticlaboratorytosetupownqualitycontrolsforreferenceranges
EnzymesindicatingHepatocellulardamage
v↑ALT↑ASTViralHepatitis
v↑IHDHepatocellularnecrosis
v ALT/ALT>1AlcoholliverdiseaseObstructiveliverdisease:cholestasis/hepaticcarcinoma/parenchymalcell
damage
Parameter HemolyticJaundice HepaticJaundice
ObstructiveJaundice
1 Bloodfreebilirubin ↑ ↑ normal2 Conjugatedbilirubin normal ↑ ↑
3 ALP normal ↑ ↑↑
4 Bilesalts Nil Nil PRESENT5 UrineBilirubin Nil Nil PRESENT6 Urineurobilinogen ↑ Nil Nil7 Stool Darkbrown Claycolor
DifferentialDiagnosisofJaundice
FateofBilirubin
CongenitalHyperbilirubinemiavAbnormaluptake,conjugationorexcretionofbilirubinduetoinheriteddefects• 1.GilbertDisease–defectinuptake• 2.CriglerNajjar syndrome–defectinconjugation
CongenitalHyperbilirubinemiav(1) Gilbert’sDiseasea) Autosomaldominanttraitb) DefectuptakeofBilirubinc) Asymptomaticd) Presenceofjaundice(Bilirubinà 3mg/dl)
CongenitalHyperbilirubinemia(2)CriglerNajjar syndrome:DefectinconjugationTypeIa) CongenitalnonhemolyticJaundiceb) DeficiencyofUDP–Glucuronyl transferasec) Fatal(deathbeforetwoyearsofage)d) Jaundiceappearswithin24hrsoflifee) Unconjugatedbilirubin(>20mg/dl)f) Kernicterusg) Barbituratesnoeffect(enzymenodefect)
CongenitalHyperbilirubinemia(2)CriglerNajjar syndrome:DefectinconjugationTypeIIa) CongenitalnonhemolyticJaundiceb) DeficiencyofUDP–Glucuronyl transferasec) Diseaserunsmorebenigncoursed) Bilirubin–mononucleotidepresente) Totalbilirubinseldome rises:Unconjugatedbilirubin(>15mg/dl)f) NoKernicterusg) Barbituratesuseintreatmentà Jaundiceimproves
ManagementofCrigler Najjar Syndrome
CongenitalHyperbilirubinemiavDubin Johnson’ssyndromea) Autosomalrecessivetraitb) ↑conjugatedbilirubininbloodc) DefectinexcretionofconjugatedBilirubin(ATPase dependent
organicanionTransporterprotein-MOATmutationd) DefectATPdependentorganicaniontransportinbilecanalculie) BilirubingetsdepositedinliverthereforeBilirubingetsdepositedin
liver(BlackLiverJaundice)f) DiagnosticTest:BromosulphthaleinTest
CongenitalHyperbilirubinemiavDubin Johnson’ssyndromeDiagnosticTest:BromosulphthaleinTest250mgBromosulphthalein(intravenous)
Normal(dyeremainingplasma) Dubin Johnson’ssyndrome(dyeremainingplasma)
45min <5% >2hrslevels(<2%)2hrs <2% <45minlevels(>5%)
(2hrlevelmorethan45min)
At45minBSPtakenupbyhepatocytes/thereforedecreaseinblood levels
EXCRETORYDEFECT:BSPregurgates inblood
CongenitalHyperbilirubinemiavRotorsyndromea) Causenotknown(autosomalrecessive)b) Bilirubinexcretiondefectivec) Nodepositionofpigmentinliver
?
ComparisonofDubin Johnson’sandRotor‘ssyndrome
ComparisonofGilbert‘sandCrigler- Najjar syndrome
BiosynthesisofHeme(Porphyrinring)• Siteà Liver/RBCproducingcellsofbonemarrow(erythroid cells)/othertissue• ExcretiontoruleàmatureRBClackingmitochondria• IFormationofδ aminoLevulinate (mitochondria)Glycine+succinylCoA
* Pyridoxalphosphatedependentδ aminoLevulinate synthtase
δ aminoLevulinate (ALA)*ratecontrollingstep
BiosynthesisofHeme(Porphyrinring)IISynthesisofPorphobilinogen2. δ aminoLevulinate (ALA)
ALADehydratase *
Porphobilinogen(PBG)• ActivitydecreasesbyPb /Hg&Zinccontainingenzyme
BiosynthesisofHeme(Porphyrinring)1. SuccinylCoA+Glycine→ALA2. ALA+ALAà Porphobilinogen(PBG)+2H₂O3. 4X(PBG)4. Uroporbilinogen III(UBG)5. Coporphyrinogen III(CPGIII)+CO₂6. Protoporphyrinogen III(PPGIII)7. HEME
BiosynthesisofHeme(Porphyrinring)
SuccinylCoA+Glycine→ALA
ALA+ALAà Porphobilinogen(PBG)+2H₂O4
4X(PBG)Uroporphyrinogen
Coporphyrinogen III(CPGIII))
Protoporphyrinogen III(PPGIII)
HEME
HEMGLOBIN1. ALA SYNTHTASE 2.ALA DEHYDRATASE 3.PBG DEAMINASE&UPGIIICOSYNTHTASE 4.UROPORPHYRIN
DECARBOXYLASE 5COPORPHYRINOGENOXIDASE 6.Hemesynthtase
→4CO₂
NADP+O₂→ →NADPH+H+2CO₂
ACETYLàMETHYL
PROPINYLàVINYL
ACETYLàMETHYL,4CO₂
PROPINYLà VINYL+2CO₂
BiosynthesisofHeme
Metallo- porphyrin
1. UroporphyrinI&III2. Coporphyrin I&III3. PROTOPORPHYRINIX&HEMEvHemecontainingproteins- hemoglobin,cytochromes,catalase,tryptophanpyrrolase
Hemesynthesis
• Site :mitochondriaofallmammalian tissueexceptRBCpredominantlyinLiver&bonemarrow
vStepI :*ALAsynthasePLP CO₂CoASH
SuccinylCoA+Glycineδ ALA• δ ALA:DeltaaminoLevullinic acid• *Regulatoryenzyme=ratelimitingstepqINHtreatmentà PLP↓à ALASYNTHESIS↓
Hemesynthesis
vStepII :Site–MITOCHONDRIA
*ALADEHYDRATASEPLP 2H₂O2MoleculesofALAPorphobilinogen(PBG )*activitydecreasedbyheavymetals
Hemesynthesis
vStepIII : Site–MITOCHONDRIA
*PBGDEAMINASE4NH₃4MoleculesofPBGUroporphyrinogen(UBG )
• *UroporphyrinogenIsynthtase&UroporphyrinogenIIIsynthtase• *deficiencyofUroporphyrinogenIIIsynthtaseleadstoformationofTypeIUBGà Porphyria
Hemesynthesis
vStepIV :Site–MITOCHONDRIA
*Uroporphyrinogendecarboxylase4CO₂• Uroporphyrinogen(UBG )Type III CoproporphyrinogenIII
• *AcetylàMethyl• *deficiencyofà Porphyria
Hemesynthesis
vStepV : Site-CYTOSOLIC
*Coproporphyrinogenoxidase2CO₂Coproporphyrinogen IIIProtoporphyrinogenIII
O₂• *Methylà Vinyl• *deficiencyofCoproporphyrinogenoxidaseà Porphyria
Hemesynthesis
vStepVI :Site-CYTOSOLIC
*Protoporphyrinogenoxidase4HProtoporphyrinogenIIIProtoporphyrin III (IX)
Methylene(CH2)àMethenyl (CH- )• *deficiencyofProtoporphyrinogen oxidaseà Porphyria
Hemesynthesis
vStepVII :Site-CYTOSOLIC
*HemesynthtaseProtoporphyrinIII (IX)Heme
Fe²⁺• Ferrochelatase• Heme+ Protein= HEMOGLOBIN
1.SuccinylCoA+Glycineà
ALA
ALA
2.ALAà PBG
3.PBGà UBGIII 4.UBGà CPGIII
CPGIII
CPGIII
5.CPGIIIàPPGIII 6.PPGà PP 7.PPà HEME
CYTOSOLIC-2,3,4
MITOCHONDRIAL
HEMESYNTHESIS
RegulationofHemesynthesis1.↑Glucoseà↑catabolismofCRP(repressorprotein)à decreaseALA(inductionprevented,Glucoseadministeredfortreatmentofporphyria
2.↓ALAsynthasebyHematin,ExcessoffreeHeme(Fe²⁺à Fe³⁺)
3.Compartmentizationofenzymesà easierforregulation,ratelimitingstepsinmitochondria
4. Hemesynthesiscontrolledbya)globinsynthesisb)Druglikebarbiturates(requirecytochrome450)
5.lead/Mercury↓ALASynthase↓Ferrochelatase
RegulationofHemesynthesis
ThreemechanismscontrollingactivityofALAsynthasebyHeme/HematinFe³⁺
a.Feedbackinhibition(Heme/HematinFe³⁺)
b.RepressionofALAsynthase
C.InhibitionofALAsynthasetransportfromcytosoltomitochondria(thesiteofaction)
Hemesynthesissite:liver(ALAsynthaseà regulatoryenzyme)
EffectofdrugsonALAsynthaseactivity1. *Phenobarbital2. *Insecticides ↑activityofALAsynthase3. *Carcinogen* MetabolizedbyHEMEcontainingproteincytochrome450
↓IncreasedincorporationofHemeincytochrome450
↓↓cellularlevelofHeme
↓↑ALAsynthase(derepression)tomeetcellulardemands
RegulationintheErythroidcells
vALAsynthasedosenotregulateHemesynthesisinErythroidcellsvUroporphyrinogensynthase&Ferrochelatase regulateHemesynthesis
vCellularuptakeofironregulateHemesynthesisvHemestimulateglobinsynthesis
RegulationofHemesynthesis
Porphyrin&BilirubinMetabolism
• Porphyrin :Groupofcompoundsof4substitutedpyrroleringslinkedbyMethanebridges•à formationofcomplexeswithmetalionsboundtoN₂atomofpyrrolering
IRON
Heme
Hb
MAGNASIUM
Chlorophyll
Photosynthetic pigment
Porphyrin&BilirubinMetabolism
vIronPorphyrins :1. Hemoglobin2. Myooglobin3. Cytochrome4. Catalase&peroxidase5. Tryptophanpyrrolase
PorphyrinMetabolism• Porphyria's:GroupofinbornerrorsofmetabolismassociatedwiththebiosynthesisofHeme• Characteristics of Porphyrias :increaseinproduction&excretionofporphyrin&theirporphyrinprecursorsALA&PBG• AutosomaldominantorAutosomalrecessive• Diagnosis ofPorphyria's:• 1.urine(exoposure toUVà red fluorescence )• 2.urine+CHCl₃à extraction• Aqueouslayer(PBG)+Ehrlichreagentà pink color
Porphyria's:
• TypesPorphyria's:1.inheriteda) Erythropoietin :enzymesdeficiencyoccursinerythrocytesb) Hepatic : enzymesdeficiencyliesintheliver2.Acquired💟
Autosomalrecessive
Autosomaldominant
Erythrodontia
RBC-FLUORESCENCE
Liver,skinfluorescence
AcuteintermittentPorphyrias• DeficiencyofenzymesUroporphyrinogensynthaseI
SuccinylCoA+Glycine
δ aminolevilinate(ALA)↑
Porphobilinogen ↑
UroporphyrinogenIII
δ aminolevilinatesynthase
ALADehydratase
Uroporphyrinogen synthase
Uroporphyrinogen I
↑ALA↑PORPHYRINOGEN
1.AcuteintermittentPorphyria• Age :afterpuberty(Artist–Vincet ,KingGeorgeIII–MAD)• Deficient enzyme :UroporphyrinogensynthaseI(↑ALA–nofeedbackmechanism)• Noporphyria,nophotosensitivity• Excretion inurine:δ aminolevilinate(ALA)&porphobilinogen (PBG)• Exposureofporphobilinogen (PBG)toairdarkens(formationofPorphobilin )• Diagnosis:urinaryconc ofPBG(darkenson exposuretoair)• Symptoms:Abdominalpain,Vomiting,cardiovascularabnormalities,neuropsychiatricdisturbances(therefore↓Trppyrrolase ,↑Trp,5ʹOHTyramine• Treatment:Hematinà↓ALA, ↓PBG• Adverse effects :Barbiturate(ppt ofattack)à↑ALAsynthase(ascytochrome450)àPBG↑,increasewithcarbohydratediet&menopause
2.Congenital erythropoietin PorphyriavCongenitalvDeficient enzyme :↓UroporphyrinogencosynthtaseIII• Excretioninurine:UroporphyrinogenI&Coporphyrinogen Ià
↓oxidation↓UroporphyrinI&Coporphyrin I
Symptoms :1.↑Hemolysis2.Erythrodontia(teeth),portwinecolorurine3.ExposureOFSKINà UVsensitizationofporphyrinà absorbed&emitredfluorescentlight4.Dermatitis,scarringburning&itchingofskin- ear&nose(Leprosylike),(asROS↑à accumulationofPorphyrin)5.portwinecolorurine(UroporphyrinI&Coporphyrin I)
Congenital erythropoietin Porphyria
PhotosensitivityErythrodontia (teeth )
Congenital erythropoietin Porphyria
Dermatitis,scarringburning&itchingofskin- ear&nose(Leprosylike,(asROS↑à accumulationofPorphyrins )
Dermatitis,scarringburning&itchingofskin- ear&nose(Leprosylike)(asROS↑à accumulationofPorphyrins
Congenital erythropoietin Porphyria
3.Porphyria Cutanea Tarda• Cutaneoushepaticporphyria• Deficient enzyme :↓Uroporphyrinogendecarboxylase• Excretioninurine:Porphobilinogen&Uroporphyrin(I&III)rarely• Symptoms :1. Cutaneous2. Photosensitivity3. LiverexhibitsfluorescenceTreatment :Hematin(↓ALAsynthase,↓accumulationvariousintermediate)
4.Hereditary Coporphyria• Deficient enzyme :↓Coporphyrinogenoxidase• Excretioninurine:UroporphyrinogenI&Coporphyrinogen Ià
↓oxidation↓UroporphyrinI&Coporphyrin I
• Urinedarkcolor:presenceof UroporphyrinI ,Coporphyrin I,ALA,PBG• Treatment :Hematin(↓ALAsynthase,↓accumulationvariousintermediate)•
5.Variegate porphyria• Deficient enzyme :Protoporphyrinogen oxidase• Accumulationinplasma&excretioninurineof:PorphobilinogenUroporphyrinogen,Uroporphyrin,Coproporphyrinogen,Coproporphyrin,Protoporphyrin(↓Hemesynthesis)• Photosensitivity exhibited• Plasmaexhibitsredfluorescence(duepresenceofCoproporphyrinogen• Neuropsychiatric manifestation
6.Herediatory Protophyria (Erythropoietic protoporphyria )
• Deficient enzyme:Ferrochelatase• Accumulationinplasma(increaseinconcentration) ,excretioninurine (increaseinconcentration)& faeces (increaseinconcentration)of:PROTOPORPHYRINIX• RBC,skinexhibitredfluorescence•
7.Acquired (toxic )Porphyrias• Exposureofbodytotoxiccompounds(toys/Xeroxink)Examples:Heavymetals(Lead)↓ALADehydratase
Hexchlorobenzene ↓UroporphyrinsynthaseIDrugs(Griseoflavin )↓Ferrochelatase
Therefore↓Heme↑ALAsynthaseAquried porphyrias associatedwithanamia
ReactionsofHeme synthesis
PrenataldiagnosisofPorphyriasEnzymeestimations/PCR
Anemia• 75%Indianpopulationsufferfromanemia• Hbconcentrationdecreasesinanemia<10gm%(normalconcentrationofbloodHb– 14-16mg%-male,13-15mg%-female)vIIronDeficiencyanemia–mostcommona) Nutritionalb) LackofabsorptionofIron(Gastrectomy&Achlorhydria )c) Hookworminfection(0.3ml/day/hookworm)d) Repeatedpregnancy(↓hemoglobin-1gmperdelivery)e) Nephrosisà Glomerularfiltration→proteinuriaf) LossofHaptoglobin/Hemopexin /Transferring) Heavymetalpoisoningh) Lossofblood(menustrual cycle,piles,pepticulcers,Uterinehemorrhages)
AnemiaII-ImpairedproductionofRBC
DefectinHemesynthesis
• Nutritional• Deficiencyofiron,Copper,VitaminB12,VitaminC
• Leadpoisoning
DeficiencyofERYTHROPOITIN
• Physiological–kidneycellssynthesizeERYTHROPOIETIN
↓• ↑RBCsynthesis• Chronicrenalfailureàno ERYTHROPOIETIN
Decreaseinstemcells
• APLASTICANEMIA
• MALIGNANTINFITRATION
• INFLECTION
Hemin crystals
• Fe²⁺↔Fe³⁺à Fe³⁺+Cl⁻à HematinchlorideorHEMINvMedicallegalcases• Blood+Bloodstains+Nippe ’sfluid(1%KCL+KBr +KI+Glacialaceticacidà darkbrownRHOMBICcrystalàmicroscopicinspectionHemepartofBloodà Testpositive
Hemin crystals
HemoglobinestimationvDrabkin’s TestforHemoglobinestimation:Cyanmeth HemoglobinMethod• Blood(Hb)+Drabkin’s reagentàCyanmeth Hemoglobin(Absorbance-540nm)à colorimetricestimation• Hb+potassiumferricyanideàMeth- Hb
Sulph Hemoglobinemia• Oxy–Hb+H₂Sà Sulph –Hb(absorptionpeakat620nm)
vFormationofSulph –Hbbysulphonamide ,Phenacetin ,Dapsone ,AcetonevBasophilicstripingofRBC
←Irreversible
AbnormalHborHbvariants1.Sicklesyndromea) 1. sicklecelltrait(AS)b) 2.sicklecelldiseasewithSS,SC,SD,SO,Sβ Thalassemia2.UnstableHaemoglobinsCongenitalHeinzbodyanemia–Hb Zurich3.HbwithabnormaloxygenaffinityA.Highaffinityà Polycythemia(familial)à HbChesapeake,OlympiaB.Low affinityà Cyanosis(familial)à HbM,Hb–Kansas,Hb- Hoppe4. StructuralvariationleadingtoThalassemia'sphenotypeA. AlphaThalassemiaà HbConstantspring,DeltabetaThalassemia,Hb–
LeporeB. BetaThalassemia:HbQuongC. 5.Hemoglobinthatdonot produceanyclinicalsymptoms:HbP HbQ ,HbJ
SICKLE CELL DISEASEvCharacteristics of Sickle cell disease a) Glutamicacidà Valine (6th positiononbetachain)b) Hydrophilicà Hydrophobic(stickinessonsurfaceofmolecule)c) PolymerizationofHb inRBCàdistortionofRBCintosickleshapedd) DeoxyHbS hasprotrusionononeside&cavityonothersideà
manymoleculesadheretogether
SICKLE CELL DISEASE
SignsandsymptomsofThalassemiamajor
InheritanceofThalassemia
AbnormalHborHbvariantsIIIHemolyticanaemias duetointra-corpusculardefecta) Hemoglobinopathies :HbS ,Hbc ,HbMb) Thalessemias :major&minorc) Abnormalshapespherocytosis&elliptocytosisd) Enzymedeficiencyà Glucose6PhosphodehydrogenaseIVHemolyticanemiadueextracorpuscularcause/defectsa) Infection–malarialparasites,streptococcusb) AutoimmuneHemolysis–RBCmembranecomponent,Syphilis,
Lympholenticular neoplasiac) Isoimmune hemolysisà RhincompatibilityinNewbornd) Hemolysisduetodrugsensitization–Dopa quinone (fixedonRBC)à
Abnormalantibodies againstalteredmembrane
AbnormalHborHbvariantsVhemorrhages
a) Hematuriab) Hematomesisc) Hemoptysisd) Pepticulcerse) Hemorrhoidesf) Thrombocytopeniag) Menorrhagiah) Bleedingtendencies