metabolism of hemoglobinporphyrins & porphyrias

Metabolism of HemoglobinPorphyrins & Porphyrias

11Dr Gihan GawishDr Gihan Gawish

Hematology Block- Medical Biochemistry and Molecular Genetics

file://upload.wikimedia.org/wikipedia/commons/3/3d/1GZX_Haemoglobin.png

Case 1A 10-years-old boy with a family history of prophyrias has complained of fatigue with exposure to sun. Lab investigations have reported PBG, uroprophryin, and 5-ALA accumulated in the plasma and excreted in urine. Which of the following porphyria explain his findings?Congenital erythropoetic porphyriaErythropoetic protoporphyriaAcute intermittent porphyria Variegate porphyria


A 46-years-old worker male has had worsening headaches, poor appetite, abdominal pain with vomiting and constipation. A CBC shows: Hgb 10.8 g/dL, Hct 33.1%, MCV 104 fL, platelet count 239,000/microliter, and WBC count 7720/microliter. The peripheral blood smear shows normal WBC morphology and RBCs. His serum vitamin B6 is in normal level. Porphobilinogen, uroprophryin and d-aminolevulinic acid (ALA) were detected and elevated in urinalysis. Which of the following underlying conditions explain his findings?Chronic lymphocytic leukemia Hereditary spherocytosisLead poisoningPeptic ulcer disease

Dr Gihan GawishDr Gihan Gawish 33

Case 2

Case 3A 12-years-old girl with a family history of prophyrias has complained of fatigue with exposure to sun. The case was recorded as photosensitivity with fragility of exposed skin. Lab investigations have reported increased urinary uroporphyrin and stool isocoproporphyrin. Which of the following porphyria explain her findings?Congenital erythropoetic porphyriaErythropoetic protoporphyriaPorphyria cutanea tardaVariegate porphyria


Case 4Mrs. Maha is a 36 year old who presents with yellow discoloration of her eyes which she noticed two days ago while washing her face. At first she thought the color was due to the lighting in her bathroom, but this morning, she thought her hands "looked yellow." The examination shows the abdomen is round and slightly tympanitic. The lab data have represented that AST was 150 U/L, ALT60 U/L, total Bilirubin 22 mg/dL, Alkaline phosphatase 400 U/L and Prothrombin time 13.2 second. During, hospitalization WBC rose to 42,000/mm3; total bilirubin rose to 32 mg/dL. Which of the following is the main clinical problem?JaundiceProphyriaWilson’s diseaseHypoglycemia


Case 5An 80-year-old Saudi female with a past medical history of osteoarthritis was admitted to the hospital with a chief complaint of jaundice for 2 months. The patient did not notice the jaundice or felt any differently but her physician was worried and she was admitted to a different hospital 2 months ago. The liver ultrasound (U/S) showed gallstones and she had a laparoscopic cholecystectomy 2 months ago. Which of the following lab results represented her case?


(a) (b) (c)

Describe the structure, synthesis, degradation and types of haemoglobin.

Specific Objectives: Describe the pathway and rate-limiting steps by which heme is synthesized.

List the normal hemoglobins found in fetal and adult blood.

Describe the pathway for the degradation of heme.

List the clinical conditions associated with abnormal metabolism of heme.


Metabolism of Hemoglobinporphyrins & porphyrias


HaemoglobinHaemoglobins is globular composed of haem and globin.

Globin is a simple protein “globin” in hemoglobin refers to the individual protein subunits

Haem is composed of Fe++ & Porphyrins


Typical Hemoglobin Values

Hemoglobin levels are measured by the amount of hemoglobin in grams (gm) per deciliter (dl) of blood. The normal ranges for hemoglobin values are dependent on the age and sex. Normal ranges are:

Newborns: 17-22 gm/dlOne (1) week of age: 15-20 gm/dlOne (1) month of age: 11-15gm/dlChildren: 11-13 gm/dlAdult women: 12-16 gm/dlAdult males: 14-18 gm/dlWomen after middle age: 11.7-13.8 gm/dl Men after middle age: 12.4-14.9 gm/dl


Hemoglobin

Haem synthesis occurs largely in the mitochondria.

Globin synthesis occurs in the polyribosomes.

65% of the Hb is synthesized in the erythroblasts, and 35% at the reticulocyte.


Haemoglobin Each molecule of normal adult haemoglobin (Hb-A) consists of four polypeptide chains a2b2, each with its own haem group.


Globin synthesis polypeptide subunits: two alpha chains and two beta chains, each with 141 and 146 amino acids respectively.

The synthesis of these globins is under genetic control.

Humans normally carry functional globin chains, arranged in two, duplicated gene clusters:

-like cluster (, , and globin genes) on the short arm of chromosome 11 and

-like cluster ( and globin genes) on the short arm of chromosome 16. 1313Dr Gihan GawishDr Gihan Gawish

α Alpha β Beta γ Gamma δ Delta ε Epsilon ζ Zeta and η Eta 1414Dr Gihan GawishDr Gihan Gawish

Hb in adult bloodHb A Hb A2 Hb F

Structure a2b2 a2d2 a2g2

Normal % 96-98 % 1.5-3.2 % 0.5-0.8 %

The major switch from fetal to adult haemoglobin occurs 3-6 months after birth.


STRUCTURE OF HEME

Ferrous iron (Fe2+) Protoporphyrin IX: comprised of 4 pyrrole rings linked together by methenyl bridges


Iron MetabolismGu

tBlood

Cells

• Low pH of stomach solubilizes Fe-containing ionic compounds.• Fe transporters facilitate absorption into blood stream• Fe3+ ions are bound and chelated by Transferrin (Tf).• Transferrin transports Fe to tissues

•Maintains solubility•Keeps Fe ions unreactive

• Transferrin endocytosis is receptor-mediated (TfR)• Endocytosis results in Fe3+ release• Fe is distributed to distinct regions of the cell via Fe transporter and/or by the help of channels• Function of iron in the cell: Protein components (Heme)• Storage: Ferritin (Fe2+)• Toxicity


Iron transport to the Cell

Transferrin Receptors (TfR)

Fe(III)2-Tf Tf

Proteins: Catalysis Electron, oxygen transport Structural stabilization Formation of protein-bound radicals

Storage and Sequestration: FerritinFerritin

Toxicity: Oxidative stress

[Fe]

[Fe]

[Fe]1818Dr Gihan GawishDr Gihan Gawish

Heme

8

8

Succinyl CoA

Glycine**

Heme Synthesis85% of total heme synthesis occurs in red

blood cells (RBC)

** Amino acid (building blocks of protein) synthesized in your body2020Dr Gihan GawishDr Gihan Gawish

Heme synthesis


Heme Synthesis


SummaryIRON

AMINO CI

D

Pyrrole

ring2323Dr Gihan GawishDr Gihan Gawish

Regulation Of Heme And Globin Synthesis:

Heme:

Inhibition of the synthase and stimulation of globin synthesis are the most important aspects in balancing hemoglobin production.-ALA synthase Requires pyridoxal phosphate (Vitamin B6) as a coenzyme

diminishes the transport of -ALA synthase from cytoplasm to mitochondria. Represses the production of -ALA synthase by regulating gene transcription.

Inhibits activity of pre-existing -ALA synthase

Stimulates globin synthesis to ensure that levels of free heme remain low in concentration.


Disorders of Heme Synthesis

Acquired: Lead poisoning

Congenital: Porphyrias

Deficiency of heme has serious effects in relation to hemoglobin and cytochromes.


LEAD TOXICITYSymptoms

Irritibility Poor appetite Lethargy Abdominal pain (with

or without vomiting) Sleeplessness Headaches ConstipationLEAD

Binds to any compound with a sulfhydryl groupInhibits multiple enzyme reactions including those involved in heme biosynthesis (PBG synthase &ferrochelatase)


lead

Vitamin B6

Lead Toxicity


PORPHYRIAS A group of rare disorders caused by deficiencies of enzymes of the heme biosynthetic pathway

The majority of the porphyrias are inherited in an autosomal dominant fashion - thus, affected individuals have 50% normal levels of the enzymes, and can still synthesize some heme

Affected individuals have an accumulation of heme precursors (porphyrins), which are toxic at high concentrations

Attacks of the disease are triggered by certain drugs, chemicals, and foods, and also by exposure to sun


PORPHYRIASGLYCINE + SuccinylCoA

-aminolevulinic acid(ALA)

Porphobilinogen(PBG)

hydroxymethylbilane

uroporphyrinogen III

coprophyrinogene III

Protoporphyrinogene IX

protoporphyrin IX

Heme

ALA synthase

ALA dehydratase

PBG deaminase

Uroporphyrinogen IIIcosynthase

Uroporphyrinogendecarboxylase

Coproporphyrinogenoxidase

Protoporphyrinogenoxidase

Ferrochelatase

ALA-dehydrataseDeficiency porphyria

Acute intermittentporphyria

Congenital erythropoieticporphyria

Prophyria cutanea tarda

Herediatarycoproporphyria

Variegateporphyria

Erythropoieticprotoporphyria

Mitochondria

Agent Orange


Acute Intermittent Porphyria Autosomal dominant

Caused by a deficiency in porphobilinogen deaminase, which is involved in the conversion of porphobilinogen (PBG) to uroporphyrinogen III

PBG, uroprophryin, and 5-ALA accumulate in the plasmaand the excreted in urine

Patients have neuropyschiatric symptoms and abdominal pain.


Porphyria Cutanea TardaMost common porphyria

Autosomal dominant

Disease is caused by a deficiency in uroporphyrinogen decarboxylase, which is involved in the conversion of uroporphyrinogen III to coproporphyrinogen III

Uroporphyrinogen accumulates inplasma and appear in urine Patients are photosensitive (cutaneous photosensitivity)

Accumulation of porphyrinogens results in their conversion to porphyrins by light

Porphyrins react with molecular oxygen to form oxygen radicalsOxygen radicals can cause severe damage to the skin 3131Dr Gihan GawishDr Gihan Gawish

Heme Catabolism


Destruction of Normal Red Cell

Red cell destruction usually occurs after a mean life span of 120 days.

The cells are removed extravascularly by macrophages of the reticuloendothelial system (RES), specially in the bone marrow but also in the liver and spleen.


Normal Red Cell Destruction

The breakdown of red cells liberates

1- Iron for recirculation via plasma transferrin to marrow erythroblasts

2- Protoporphyrin which is broken down to bilirubin.

3- Globins which are converted to amino acids.


The bilirubin circulates to the liver where it is conjugated to glucuronides which are excreted into the gut via bile and converted to stercobilinogen and stercobilin (excreted in faeces).

Stercobilinogen and stercobilin are partly reabsorbed and excreted in urine as urobilinogen and urobilin.

A small fraction of protoporphyrin is converted to carbon monoxide (CO) and excreted via the lungs.

Bilirubin


Normal red cell breakdownhaemoglobin

haem

protoporphyriniron

Bilirubin(free)

COExpired airtransferrin

erythroblastBilirubin glucuronides

Stercobilin(ogen)Urobilin(ogen)

Urine

Liverconjugation

faeces

globin

Amino acids


DEGRADATION OF HEME TO BILIRUBIN

P450 cytochrome

75% is derived from RBCs

In normal adults this results in a daily load of 250-300 mg of bilirubin

Normal plasma concentrations are less then 1 mg/dL

Hydrophobic – transported by albumin to the liver for further metabolism prior to its excretion

“unconjugated” bilirubin

39Dr Gihan Gawish

BILIRUBIN PRODUCTION

Heme(250 to 400 mg/day)

Heme oxygenase

Biliverdin reductase

Hemoglobin(70 to 80%)

Erythroid cellsHeme proteinsmyoglobin, cytochromes

(20 to 25%)

Biliverdin

Bilirubin

NADPH + H+

NADP+

3 [O]Fe3+ + CO

apoferritinferritin

indirectunconjugatedpre-hepatic

albumin

DrGihanGawish40Dr Gihan Gawish

BILIRUBIN PROCESSINGalbumin-Bilirubin

ligandin

Bilirubin diglucuronide

ER

hepatocyte

UDP-Glucuronyl transferase

albumin

ligandin-Bilirubin

bile (gall bladder)

directconjugatedpost-hepatic

2 UDP-glucuronate

2 UDP

41Dr Gihan Gawish

BILIRUBIN EXCRETION

Bilirubin diglucuronide

Intrahepatic urobilinogen

cycle

StercobilinogenBacterial enzymes

BilirubinBacterial enzyme2 glucuronate

Bacterial enzyme

Urobilinogen

8H

liver

Urobilinkidneys urine

Stercobilin feces

kidneysintestines

42Dr Gihan Gawish

HYPERBILIRUBINEMIA

Increased plasma concentrations of bilirubin (> 3 mg/dL) occurs

when there is an imbalance between its production and excretion

Recognized clinically as jaundice

DrGihanGawish DrGihanGawish 43Dr Gihan Gawish

Diseases of Haem catabolism

1.Jaundice2.Haemoglobinopathies3.Anaemia


A. Hemolytic anemia

excess hemolysis

unconjugated bilirubin (in blood) conjugated bilirubin (released to bile duct)

B. Hepatitis

unconjugated bilirubin (in blood) conjugated bilirubin (in blood)

C. Biliary duct stone

unconjugated bilirubin (in blood) conjugated bilirubin (in blood)

Hyperbilirubinemia


Causes of Jaundice1- Increased production of bilirubin by hemolysis or blood disease:

•Increase in blood indirect bilirubin•Called pre-hepatic jaundice•Stool color remains normal.

2- Abnormal uptake or conjugation of bilirubin: •Leads to non-hemolytic unconjugated hyperbilirubinemia•Increased indirect bilirubin.•Stool color turns gray.•Caused by liver damage or disease. 4646Dr Gihan GawishDr Gihan Gawish

3- Cholestasis: Problems with bile flow.1. Intrahepatic cholestasis: hyper conjugated bilirubinemia

•Increase in blood indirect and direct bilirubin•Caused by liver damage or disease: eg cirrhosis, hepatitis•Can also occur in pregnancy:

2. Extrahepatic cholestasis: •Blockage of bilirubin transport in the bilary tract.•Increased direct bilirubin.•Stool color turns gray.•Caused by: Tumors or gall stones.


Diagnoses of JaundiceDiagnoses of Jaundice


Haemoglobinopathies There are mainly two types normalities:

Quantitative abnormalities: where there is reduction in the production of certain types of globins e.g. A. thalassaemia & B thalassaemiaQualitative abnormalities: where there is production of abnormal haemoglobin e.g. sickle cell anaemia.


Iron Deficiency Anemia The characteristic sequence of events ensues when the total body iron level begins to fall:

1. decreases the iron stores in the macrophages of the liver, spleen and bone marrow

2. increases the amount of free erythrocyte protoporphiryn (FEP)

3. begins the production of microcytic erythrocytes

4. decreases the blood haemoglobin concentration


Factors leading to Iron deficiency anemia

The end result of a long period of negative iron balanceDecreased iron intake

Inadequate diet, impaired absorption, gastric surgery, celiac disease

Increased iron lossGastrointestinal bleeding (haemorrhoids, salicylate ingestion, peptic ulcer, neoplasm, ulcerative colitis)

Excessive menstrual flow, blood donation, disorders of hemostasis

Increased physiologic requirements for ironInfancy, pregnancy, lactation


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metabolism of hemoglobinporphyrins & porphyrias

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