Unit 11 Heme Derivatives Flashcards

Protoporphyrin Ring + Fe

Heme + 4 globin chains

there are many possible chains, but the common ones are alpha, beta, delta, and gamma]

it is toxic to human tissues

it is the storage form of iron and made up of the Ferric (3+) form, which is stored in the liver

any of a class of pigments (including heme and chlorophyl) whose molecules contain a flat ring of four linked heterocyclic groups, sometimes with a central metal atom.

It is named from the ancient Greek word porphura, meaning purple. These diseases are caused by enzyme deficiencies in the heme production pathway. People generally look purple when they have this disease.

- a black inorganic compound which forms from heme when RBCs are lysed and exposed to air, Can be used as treatment for some porphyrias

Deficiency of Uroporphyrinogen Decarboxylase defined by blistering (most common)

serum porphyrin is the best test to order

Deficiency of PBG Deaminase, defined by blistering and nerve impairments (2nd most common)

urine porphobilinogen is the best test to order

it must show an increase in total erythrocyte protoporphyrin

- Deficiency of ferrochelatase defined by blistering

the Delta-aminolevulinic acid assay

it is important because it’s the rate-limiting step in porphyrin synthesis

Acute Intermittent Porphyria (AIP) is the second most common porphyria. Patients suffer acute neurological disease and urine turns brown with exposure to light.

inhibition of most of the enzymes in the synthetic pathway

elevated levels of succinylacetone which inhibits Aminolevulinic Acid Dehydratase (ALAD) resulting in elevated levels of d-Aminolevulinic Acid

leads to a problem synthesizing heme

leads to an increase in heme production, therefore more precursor substances are needed, potentially leading to a porphyria. Erythrocyte protoporphyrin is increased in these patients.

Iron: Low

Transferrin: High

Transferrin Saturation: Low

Total Iron Binding Capacity: High

Ferritin: Low

Watson-Schwartz - 4-dimethylaminobenzaldehyde (Ehrlich’s reagent) produces a red color in the presence of porphobilinogen which is detected at 555 nm spectrophotometrically.

HPLC with fluorescent detection

note: We typically assess plasma, urine, and stool for porphyrins, using chromatographic analysis (with HPLC)

All of the samples MUST be protected from light

to confirm Acute Intermittent Porphyria (AIP) or to evaluate disease risk in patients with direct family members with AIP

porphobilinogen deaminase

MEMORY TRICK: Think “AIPBG”, the disease together with the diagnostic test

screening test for Lead poisoning

around 120 days

note: Heme is either reused/recycled OR metabolized into bilirubin and excreted

Indirect Bilirubin

Direct bilirubin (hydrophilic)

Increases in unconjugated bilirubin occur with hemolysis or inherited liver disease

Increases in conjugated bilirubin occur with biliary obstruction (tumor) or inherited liver disease

Conjugated bilirubin covalently bound to albumin

Conjugated + Unconjugated + Delta Bilirubin

Indirect bilirubin

1.Dubin-Johnson Syndrome

2.Crigler-Najjar Syndrome

1.Rotor Syndrome

2.Gilbert Syndrome

Dubin-Johnson Syndrome

Rotor Syndrome

Crigler-Najjar Syndrome

Gilbert Syndrome

Indirect bilirubin high

Direct bilirubin high

Creates a red-purple chromogen with maximal absorption at 560 nm

Creates a blue chromogen with maximal absorption at 600 nm

A fasting sample is preferred to reduce alcohol interference

Lipemia will falsely increase the measured bilirubin

Hemolyzed samples will decrease the reaction with the diazo reagent

Bilirubin is very light sensitive and may be 50% destroyed in one hour of exposure

when the serum or plasma is separated from the cells, stored in the dark and for up to 2 days at RT, 1 week at 4˚C, and indefinitely at -20˚C.

First performed on cellulose acetate using an alkaline buffer (pH 8)

Secondarily performed on an acid buffer (pH 6) to distinguish hemoglobins

note: The order of hemoglobins from slowest to fastest is C, S, F, and A

to detect abnormal hemoglobin's

to differentiate abnormal hemoglobins that migrate together on alkaline hemoglobin electrophoresis

The side chains of the abnormally substituted amino acids in the hemoglobin react similarly at a pH of 8.4. The same goes for the group of hemoglobins C, A2, E, and O.

Hemoglobin A2 is normal but Hemoglobin E has a mutated beta-chain. To distinguish, perform capillary electrophoresis

glycated hemoglobin, used for monitoring glucose control in diabetic patients

- carbon monoxide binds to hemoglobin and displaces oxygen and binds with greater affinity, seen in house fires, enclosed areas near an exhaust source, etc.

Alkaline media or oxidizing agents transform normal Fe2+ into Fe3+, conferring a inability to reversibly bind oxygen

- Sulfation of hemoglobin causes an inability to bind oxygen

1.Release iron from transferrin with addition of acid

2.Reduction of Fe3+ to Fe2+ with ascorbic acid

3.React Fe2+ with a chromogen (e.g. ferrozine)

⅔, or 66%

↑TIBC

↓Fe

↓TIBC

↓Fe

↓ TIBC

↑Fe

Inadequate alpha globulin production

Produces Beta Hemoglobin tetramers

2 alpha and 2 beta

Think of Protoporphyrin IX as being Tony
Stark, he’s gotta become Ironman to carry that heavy oxygen around.

Glucose and Heme

Acute Intermittent Porphyria (AIP) is the 2nd most common porphyria. Patients suffer acute neurological disease and urine turns brown with exposure to light.

We call these thalassemias

β thalassemia minor (clinically silent)

β thalassemia intermedia (moderate)

β thalassemia major (severe)

Point mutation in the β globin gene

Her child would have a 1 in 4 chance of having

The correct answer is A, HbA2/S. This is correct because he would have received one normal gene producing HbA2 and one abnormal gene producing HbS. This genotype is considered a “carrier” of the mutation, which may or may not significantly affect their lives. B. HbC/S disease can be quite detrimental to the carriers of both HbC and HbS genes. The red cells in these patients still undergo sickling under hypoxic stress just like in sickle cell disease. HbF/S would be a normal finding in sickle cell disease because the production of HbF is increased in order for the body to be able to carry some more
oxygen. HbS/S is a genotype not a phenotype. A genotype is the genetic instructions, and a phenotype is the physical characteristics that result from your body copying the genetic instructions.

The correct answer is D) 4β chains. Very few α chains are produced in α -thalassemia with 3 out of 4 α -globin chains defective (HbH disease) and no α chains are produced when all 4 chains are defective, and is incompatible with life (hydrops fetalis). Because there are not enough α chains to bind, the β chains just go ahead and form tetramers which are called HbH. A) is the normal variant HbA2 , B) is the normal HbA, and C) is HbF.

The correct answer is B) Decreased HbA and Increased HbF. Let’s dive into why. Normally, we have a lot of HbA, a little HbA2 and adults have very little HbF. These hemoglobins are made up of different hemoglobin chains. HbA is made up of 2α chains and 2β chains and HbF is made up of 2α chains and 2γ chains. In β-thalassemia, the β
globin gene is defective, so all that can be produced are adequate levels of α, γ, δ, and other globin chains. Therefore, because HbA cannot be made in adequate amounts to carry all the oxygen that the body needs, other globin chains are made to attempt to increase the oxygen carrying capacity. These other chains may be present in higher amounts to make up for the lack of the β globins and to create more hemoglobin. This is why the HbF concentration increases in β-thalassemia, because the β-gene cannot be made in adequate amounts, the γ chains are produced in an attempt to make up the difference and bring the patient to a normal oxygen-carrying capacity.. This explains the increase in HbF and the decrease in HbA.
Check the electrophoretic patterns below for a visual representation

heme synthesis

HbA, HbA₂ ,HbS, and HbF. Remember that HbA₂ crawls with HbC.

ALA Dehydratase
Deficiency

Congenital Erythropoietic Porphyria

Porphyria Cutanea Tarda

Erythropoietic porphyria