Hemoglobinopathies and thalassemia's

Question 1

How do you screen for Hb S: Sickle Solubility test

Answer 1

screen for hgb S
-based on principle that hgb s is less soluble than hb A when it gets deoxygenated and precipitates causing turbidity
-blood is added to buffer salt solution with a reducing agent like (NA dithionite, Na hydrosulfite - reduces ferric iron so it cant bind o2 = deO2) and detergent lysing agent like Saponin (hgb gets released from RBC by dissolving membrane lipids)
-DeO2 Hgb S polymerizes in solution creating turbidity. Solutions WITHOUT Hgb S, remain clear
-look at solution against lined paper

Question 2

What gives you A FALSE NEG when screening for Hb S: Sickle Solubility test

Answer 2

-Infants <6 months:
-mutant beta chains did not replace normal gamma chains. Thus low levels of Hgb S present, means no polymerization occurs

-low hematocrit

Question 3

What gives you A FALSE POS when screening for Hb S: Sickle Solubility test

Answer 3

-hyperlipidemia

-Hemoglobinopathies (ex. Hgb C Harlem)
-Has 2 substitutions – one is the one seen in Hgb S (valine replacement)

too much blood added

-hgb like harlem which have two aa substitutions so the HbS and then one other one.

Question 4

Screening Tests for Hb S: Sickling Test

what is usually pos or neg

Answer 4

-Induces sickling on a glass slide

-one drop of whole blood mixed with 2 drops of Na Metabisulphite (reducing agent) on slide
-seal under cover slip to prevent O2 from entering - inducing deO2 hgb S- polymerization

Hb C Harlem is negative when using this test however newborns that contain < 10% Hb S are weakly positive and easily missed (mostly Hb F)

Question 5

What are some Screening Tests for Sickle Cell Anemia

Answer 5

Sickling and Solubility tests are used for screening

Detects presence of Hgb S but does not distinguish (AS) and disease SS

Question 6

Alkaline Hemoglobin Electrophoresis:
What is it used of for

Answer 6

-confirms hemoglobinopathies
-separates hgb molecules in electric field based on differences in total molecular charge
-charged molecules move through pH 8.4 (alkaline) producing mobility patterns to detect variant and normal hgb (hgb assumes a neg charge and moves toward the anode (positive)
-some hgbs have the same charge so they move with similar mobility patterns so you need to run in acidic environment to ID the hgb present

Question 7

Alkaline Electrophoresis vs acidic

Answer 7

Hgb molecules have negative charge and migrate toward the positive pole (anode) in the gel, others will take on positive charge (go to cathode) - like Hb S migrates with D and G on alkaline eletrophoresis but separates
l from D and G on aide.
-likewise C migrates with E and O on alkaline but separates on acid

Alkaline must be done first and is enough to diagnose SS, AS and Thalassemia but acidic is needed for confimation

Question 8

What is HPLC

Answer 8

-Confirmed Hgb variants with electrophoresis best used for B thalassemia because it can quantify A, A2, and F
-Molecular methods are best for determining definitively

Process:
Patient’s RBC lysate is injected into a cation exchange column.
Both normal and variant Hgbs will bind to column
Elution buffer is injected and forms a gradient of varying ionic strength
Various Hgb types will be differentially eluted from the column at their own retention time
A detect will measure the absorbance of the fraction at 415 nm
The area under the peak is used to quantify Hgb fraction – reported as % of total Hgb

Question 9

Thalassemia

Quantitative Hemoglobin Mutations

Answer 9

Reduced production of globin chain
-mutations in Alpha or beta chain are most sign because they are the major type of hemoglobin
-Disrupts regular alpha-beta ratio, resulting in premature destruction of RBCs
-divided into alpha and beta thalassemia

-thalassemia belt coincides with Malarial prevalence

possibility because a reduction in parasite invasion and growth in thalassemia cells
Reduced attachment of infected thalassemia cells to endothelial cells and greater binding to anti-malarial antibodies to infected cells
-Therefore, increased phagocytosis and immune clearance of parasites

Question 10

process of normal HGB

Answer 10

-switch from embryonic to fetal Hb occurs between 6 and 10 weeks of gestation
-fetal to adult Hb occurs at approximately the time of birth.

New born has Hb A (20-30%), Hb F (65-90), hbA2 (<1)

Adult Hb A (95-100), hb F 0-2, hb a 0-3.5

Question 11

Normal Individual Genotype:

Answer 11

one cluster of B globin genes are inherited on chromosome 11
normal B chain synthesis = BB
2 copies of a-globin gene are inherited on chromosome 16 aa/aa

Question 12

clinical manifestations of thalassemia result from:

Answer 12

-Reduced or absent transcription of messenger RNA
-mRNA processing Errors because of mutations that add or remove splice sites resulting in nog globin chains or altered chain production
-Translation errors due to mutations that change codon reading frame - add a stop codon, substitute an incorrect on

Unequal production of the α- or β-globin chains causing an imbalance in alpha beta ratio > Reduced RBC precursors ** more important

PBF:
-Hypo/micro RBCs

Question 13

Mechanisms in β - Thalassemia

unpaired excessive chains

Answer 13

-Unpaired, excess alpha chains precipitate and form inclusion bodies
-Inclusion bodies cause oxidative stress and damage to cellular membranes.
-Apoptosis occurs

Question 14

Mechanisms in β - Thalassemia
Iron Overload

Answer 14

Children (Iron causes growth retardation and absence of sexual maturity),
Adults (Cardiomyopathy, Fibrosis, Cirrhosis, Exocrine dysfunction)
-Mostly due to transfusion required by beta thalassemia major

Consistently elevated EPO = erythropoietin
-Promotes erythroferrone from erythroblasts which suppresses hepcidin
-Low hepcidin levels allows more iron absorption by the intestines
-in ineffective erythropoiesis , this hyperabsorption of iron is not needed because iron is recycled back into plasma

Question 15

Mechanisms in β - Thalassemia
Ineffective Erythropoiesis

Answer 15

ineffective erythropoiesis and increased destruction
-bone marrow cannot release sufficiently viable cells into circulation
-cells that have been released have inclusions so they are quickly destroyed by splenic macrophages (EXTRAvas)

Question 16

Mechanisms in β - Thalassemia
Ineffective Erythropoiesis

Answer 16

enlargened spleen and liver
Worsens anemia and cause neutropenia and thrombocytopenia
The released Hgb leads to increased bilirubin and jaundice

Question 17

when do symptoms of thalassemia occur

Answer 17

-asypm during fetal life and then symps start showing up between 6-24 months

-decreasing of alpha chains causes increase of gamma chains in fetus/newborn and then B chains after birth
-delta and beta chains can form a tetramer
-these tetramers precpitate in mature RBCs forming inclusion bodies which are removed by splenic macrophages = anemia

Question 18

α – Thalassemia – Pathological Hemoglobins

Answer 18

Unlike β thal, α thal affects fetus & newborns

• a decrease in alpha causes excess γ chains form Hb Bart’s - γ4. Stable and dont precipitate but form tetramers. After 6 months they will switch to Beta and a decrease in alpha chains will now be an increase in Beta chains

Very high oxygen affinity
Poor/no release of oxygen to tissue

excess β chains form Hb H - β4
beginning few weeks after birth and throughout life
Vulnerable to oxidation
Precipitates out forming many bodies of denatured Hgb

Question 19

β-thalassemia is divided into four categories:

Answer 19

-β-thalassemia silent carrier (heterozygous state) – no anemia, no clinical symptoms

-β-thalassemia minor (heterozygous state – mild hemolytic anemia, no clinical symptoms

-β-thalassemia major (homozygous or compound heterozygous state) – severe anemia, severe clinical symptoms, needs transfusion

-β-thalassemia intermedia –mild/moderate anemia, moderate clinical symptoms, doesn’t need transfusion

Question 20

Beta(silent)Beta – Silent carrier state
what does it result in

Answer 20

nearly normal alpha, beta chain ratios AND no hematologic abnormalities

Question 21

β-thalassemia minor trait what does it result in

Answer 21

Micro/hypo; target cells, elliptocytes, teardrop cells and basophilic stippling
-low mcv and mchc

when 1 beta globin gene is affected by a mutation which decreases or abolishes its expression AND the other gene is normal
-mild, asymptomatic anemia
-Hepatomegaly, splenomegaly

Question 22

β Thalassemia Major

Answer 22

characterized by severe anemia needing regular transfusions
-diagnosed between 6months - 2 years old (after gamma to beta switch)

HAP: reduced or absent
LD: markedly elevated

NO Hb A or decreased Hb A

marked erythroid hyperplasia
low retics occurs when erythroid precursors undergo apoptosis due to ineffective erythropoiesis

Question 23

β Thalassemia Major
PBS

Answer 23

marked hypo/micro - because of decreased hgb production

poly - ineffective but increased EPO production

Poik - increased dectruction in BM and Spleen Targets, tears, ellip, frags and all inclusions

Question 24

β Thalassemia Major Treatment
Transfusion & Iron chelation

Answer 24

-major therapeutic option
-use donor cells that are max 10 days old
-regular transfusions are used to suppress anemia and marked erythropoiesis.
-if there is a decrease in erythropoiesis there is decreased iron absorption in the intestines
-children in this treatment will not get hepatosplenomegaly

-however , chronic transfusions can lead to iron overload because there is no way for iron to leave the body so the iron in transfused RBCs accumulate in organs outside the bone marrow such as the heart liver and pancreas

Question 25

β Thalassemia Major Treatment Hematopoietic stem cell transplant (HSCT)

Answer 25

Only curative therapy. Teens with HLA donor match and no iron overload have good outcomes allogenic cells (cord blood unit)

Question 26

β Thalassemia Major Treatment Hydroxyurea (Hb F induction agents)

Answer 26

Switches on the gamma gene to produce more gamma chains. Gamma chains will combine with excess alpha chains to form Hgb F. Thus partially correcting alpha:beta ratio imbalance

Question 27

β Thalassemia Intermedia

Answer 27

-symptoms fall between b-thalassemia minor and b-thalassemia major, but without the need for regular transfusion therapy to maintain the hemoglobin level and quality of life -Patient may develop iron overload 🡪 must monitor for iron overload after age 10 even without regular transfusions - ineffective erythropoiesis which suppresses hepcidin production by the liver caused increased iron absorption by the intestines chelation therapy Patients have higher risk of thrombosis due to iron overload and precipitation of RBC chains

Question 28

What is B^0 b^+ b^s

Answer 28

B^0 - no B chain production b^+ - decreased chain production b^s -mildly decreased always look at family history

Question 29

how to differentiate between IDA and thal

Answer 29

thall - marked hypo micro, targets tears, increased poly , iron and Fe increased and TIBC decreased IDA - moderate to marked hypo/micro, pencils, tears, iron and ferritin decreased but TIBC increased Hgb A2 Levels - IDA should be ruled out FIRST, before doing this **bc we might see FALSELY reduced Hgb A2 levels if Beta thalassemia minor patient has low iron High RBC count, Low MCV is more common to Beta Thalassemia Minor

Question 30

Clinical Syndromes of α Thalassemia

Answer 30

Silent carrier state α-thalassemia minor Hb H disease Hb Bart hydrops fetalis syndrome determined by number of genes affected and the amount of α chains produced.

Question 31

Genetic Designation in α Thalassemia

Answer 31

αα/αα Normal α genes -α/αα One gene deletion – silent carrier - -/αα Heterozygous deletion – α thal minor -α/-α Homozygous deletion – α thal minor - -/-α Hb H disease - α thal intermedia - -/- - Hydrops fetalis - α thal major

Question 32

Silent Carrier of α Thalassemia

Answer 32

Deletion of one α-globin gene, leaving three functional α-globin genes -α/β chain ratio is nearly normal -one α-globin gene is absent, there is a slight decrease in α chain production with a slight increase in gamma chain during birth forming the hgb Bart No hematologic abnormalities are present -rare to have inclusions

Question 33

α Thalassemia Minor (Trait) what do you expect to see

Answer 33

-deletion of two α-globin genes It exists in two forms: homozygous α+ (– α/– α) 1 alpha from each parent missing heterozygous α0 (– –/αα) 2 alpha genes missing from one parent -few inclusion symptomatic and characterized by a mild microcytic anemia with MCV < 80fL and MCH < 27pg

Question 34

Hemoglobin H Disease Clinical Syndromes of α Thalassemia

Answer 34

Deletion of three α-globin genes is the cause of Hb H disease -accumulation of excess unpaired β chains that form tetramers of Hgb H in adults and hgb bart in newborn. -Supravital Stain: Hb H bodies (“golf-ball” like appearance) Hgb H is vulnerable to oxidation and gradually precipitates in circulating RBCs to form inclusions - pitted golf balls -Inclusions alter RBC shape and vasoelasticity of RBCs -Therefore, decreased RBC survival due to splenic macrophages DIFFERENT FROM HEINZ BODIES (these are larger and fewer in number, attached to inner membrane of RBC)

Question 35

Hemoglobin H Disease Treatment

Answer 35

-no treatment required, can have hemolytic episodes and get transfusions -treat infections promptly and avoid oxidant drugs -can develop iron overload - iron status should be monitored at 10-15 years old

Question 36

Alpha Null Thalassemia Major or Hgb Bart Hydrops Fetalis Syndrome:

Answer 36

Absence of ALL alpha chain production -Hb Bart (γ4) is the predominant hemoglobin🡪 has very high oxygen affinity and will not release oxygen to tissues -Hgb Portland – Helps fetus survive up to third trimester -fetus is delivered stillborn with failure and severe edema (hydrops fetalis) Treatment: -Intrauterine transfusion. Lifelong transfusions will be required if it survives

Question 37

Thalassemia Associated with Structural Hemoglobin Variants Hemoglobin S – Thalassemia (Sickle- Thalassemia): ****** check slide

Answer 37

-Heterozygous condition which results in inheritance of beta thalassemia gene from 1 parent and Hgb S gene from the other -severity depends on how much hb A is produced -differentiate from SS by seeing microcytes and increased levels of A2 Hgb S -B0 can be worse off than SCT and on HPLC you can see increased HGB S vs Hgb A

Question 38

What is the hemoglobin change made up of

Answer 38

4 Heme molecules & 4 globin chains Globin chains – Consists of 2 identical pairs of unlike polypeptide chains

Question 39

What hemoglobin genetics like

Answer 39

6 functional human globin genes located on Chromosome 16 and Chromosome 11 Chromosome 16 – Alpha and Zeta (these are considered as Alpha-like genes) Chromosome 11 – Beta, Gamma, Delta, Epsilon (referred to as Beta-like genes)

Question 40

Hemoglobin Assembly

Answer 40

-Globin chain production occurs in erythroid precursors from Pronormoblast to Polychromatic erythrocyte -nOT IN MATURE RBCs bc these don’t have ribosomes and mitochondria to produce Hgb -β-globin mRNA is translated more efficiently than α meaning equal amounts of alpha and beta chains -transcription from hgb to messenger RNA happens in the nucleus and translation from mrna to globin peptide chain happens in the cytoplasm -Globin chains come off a ribosome (ex. Alpha and non-alpha) will bind one heme and pair to form a Heterodimer -Two heterodimers will bind to form a Tetramer alpha has positive charge and has a high affinity for beta which is neg charged, then gamma then delta

Question 41

how does hgb composition differ

Answer 41

depending on prenatal gestation time and post natal age – Globin genes activate and inactivate during development -changes in non a-globin chains is similar to change into location of hematopoiesis First 3 months: -1 alpha-like gene (Zeta) and 1 beta-like gene (Epsilon) are activated. -Hemoglobin Gower 1 is formed – 2 Zeta, 2 Epsilon After 3 months: -Alpha and Gamma chain synthesis occurs -Hemoglobin Gower 2 is formed – 2 Alpha, 2 Epsilon -Hemoglobin Portland is formed – 2 Zeta, 2 Gamma 10 months of Gestation: Zeta and Epsilon synthesis ceases Alpha and Gamma upregulated -Hgb F is formed – 2 Alpha, 2 Gamma (also known as Fetal Hemoglobin) Birth Occurs: -Gamma to Beta Switch: Gamma chain decreases, Beta chain production is increased -Hemoglobin A is formed – 2 Alpha, 2 Beta (also known as Adult Hemoglobin) -Delta chain is being produced at low levels -Hemoglobin A2 is formed – 2 Alpha, 2 Delta

Question 42

In a healthy adult, typically we see

Answer 42

>95% Hgb A <3.5% Hgb A2 1-2% Hgb F The higher Hgb F, the less clinical complication of SSD

Question 43

What are Hemoglobinopathies: which can affect globin sysnthesis in what ways

Answer 43

Gene mutations in the Globin genes. Qualitatively – Structural changes, ex. Sickle cell anemias- hgb production at normal rate Quantitatively – Reduced production, ex. Thalassemias but dont affect aa sequence

Question 44

Genetic Mutations include: Point mutation

Answer 44

Most common in Hemoglobinopathies 1 nucleotide change Change to protein coding or regulatory sequence

Question 45

Genetic Mutations include: Deletion or insertion (1 or more nucleotides)

Answer 45

loss or addition of nucleotide causes a frameshift Affects structure and function of Hgb molecule

Question 46

Genetic Mutations include: Chain extensions

Answer 46

Mutation in stop codon, translation continues resulting in longer globin chains AAs are added until another STOP codon is added Affects structure and function of Hgb molecule

Question 47

Genetic Mutations include: Gene fusions

Answer 47

Two normal genes break between nucleotides, switch positions and anneal to the opposite gene Can form hybrid globin chains – Affects Hgb function

Question 48

Zygosity

Answer 48

Link between to Number of gene mutations and the severity of the genetic defect -normal adult globin there are 4 alpha and delta genes and 2 of beta and delta genes which means 4 level of severity for alpha/gamma mutations and 2 levels of severity for beta and delta mutations Homozygous (disease) genes from both parents are affected Heterozygous (trait) only one gene affected, one normal gene (trait) OR One mutation on one gene, a different mutation on the other beta mutations impact hgb function more than alpha for the SAME # of mutations 1 beta gene mutated - 1 gene compensates 1 alpha then 3 others have to compensate why there are more B gene variants

Question 49

Impact of mutation on globin function depends on

Answer 49

1. Chemical nature of substituted AA >> Charge and size of AA -Charge – affects how substituted AA sits next to neighbor AA -Size – affects shape of globin chain by adding or subtracting 2. Location of the mutation on globin chain 3. Number of genes mutated (also known as Zygosity) -Homozygous mutation – likely severe both beta genes are mutated and the variant becomes the dominant -Heterozygous mutation – not as severe one Beta is mutated and the other is normal = 50/50 distribution of hemoglobin

Question 50

Hemoglobin S what is it

Answer 50

point mutation Glutamic Acid is replaced by Valine bc change to 6th AA in Beta-globin chain most common in africa parallels maralial incidences because SS gene given protection against cerebral falicparum malaria -malaria uses O2 in cells but sickling is caused by reduced O2. Injured rbcs get stuck in BVs, spleen increasing the chances of phagocytosis therefore less malaria

Question 51

Hemoglobin S Inheritance Pattern

Answer 51

homozygous SS = Sickle cell disease - produce no Hgb A Heterozygous for Hb S AS = Sickle cell trait B hgb variants in one gene from parents autosomal codominants so person with SSD has inherited the trait from both parents someone with SCT has inherited a trait from one parent - cant do high level athletics The higher the hgb F the less clinical complications

Question 52

how does Amino acid substitution affect the way hemoglobin molecules interact:

Answer 52

-Valine is hydrophobic in comparison to Glutamic acid which is hydrophilic -GA is on the surface of the hgb and binds water for solubility, valine also extends but tries to hide in hydrophobic niche -but when Hgb S is oxygenated there is no hydrophobic socket so its soluble but when deO2 there will be a pocket so the hgb is not soluble in the cytosol = crystals making the deO2 the more favored state

Question 53

What happens when the cells start becoming sickled

Answer 53

-blood starts getting thick -hypoxic environment , pH reduced ;acidic -flow of blood is slowed -when O2 saturation is less than 85% in SS or <40% in AS -increase in 2,3 bpg Low pH + High 2,3-BPG reduces hgb affinity for O2 -further deO2 the environment Occlusion of capillaries and arterioles by sickled RBCs infarction of surrounding tissue -Infarction = Dead tissue

Question 54

What are the 2 forms of sickle cells

Answer 54

Reversible sickle cells (RBC with Hgb S) -start normal as environment is anoxic , sickle. They are able to travel to the microvasculature with normal shape but then distort in response to O2 tension and polymerize. otherwise circulate normally Irreversible Sickle Cells Do not change shape regardless of oxygen tension. Seen on PBF as elongated sickle cell. These are removed from circulation by spleen. THEY DO NOT CAUSE VASOOCCULSION intracellular hydration can also cause sickling efflux of water leading to increased Hb S concentration inside the cell which increased polymerization

Question 55

When will symptoms arise -

Answer 55

Symptom variety is due to: Intracellular ratio to Hgb S to Hgb F pts Asymptomatic up until 6 months of age due to protective Hgb F >6 months – Progressive anemia -Mutated beta chains replace normal gamma chains -Hgb S levels rise while Hgb F levels decrease

Question 56

What is a hallmark of SCD

Answer 56

vasoocclusive crises (VOC) -event that causes reversible sickle cells to change shape -triggered by acidosis, hypoxia, dehydration, infection and fever, exposure to extreme cold -Painful episodes manifest in the bones, lungs, liver, spleen, penis, eyes, CNS, urinary tract -Occurs in capillaries and postcapillary venules Possible Risk Factors to VOC: -Polymerization, Decrease deformability, Sickle cell endothelial cell adherence (results in slow blood flow), Endothelial cell activation, WBC and PLT activation, Homeostatic activation, Altered vascular tone Example: Clinical dehydration: -Increased VWF – Triggers RBC adherence to endothelium and can precipitate into a VOC

Question 57

Hemoglobin S Clinical Features BIG CHART

Answer 57

Spleen sequestration characterized by trapping of blood in spleen that causes decrease in hgb gradual loss of spleen is known as autosplectomy - hjb and pph -loss of spleen function leads to increased risk of bacterial infection -Acute chest syndrome or ACS is repeat infractions that cause decreased alveolar oxygen tension that induces hgb s formation and sickle formation -blood gas to monitor lung function -O2 is needed to maintain saturation at 95% -SS pt have increased susceptibility to Staph A, Strep Pneu and Haem influ Retinopathy - ppl can lose visual acuity so at 10 kids with SCD should have dilated exams to detect early retinal injury to help resolve vitreous hemorrhage

Question 58

Treatment of SCD (Supportive Care)

Answer 58

Hydration – For good blood flow, reduces VOC -Avoid low oxygen environments: Strenuous exercise High altitudes Small planes Surgery with anesthesia -Treatments to increase Hb F (Hydroxyurea) (THE MORE SEVERE THE LESS HGB F) -Prophylactic Oral Penicillin V children <5 --Analgesics and opioid's are used to manage pain Blood Exchange Transfusion BET -severe VOC hgb S RBC replaced with donor - possible iron overload with storage in heart, liver, pancreas may need iron chelation therapy Chronic Transfusions 8 or more a year Hydroxyurea Therapy – Increases amount of Hgb F in RBCs in SCD patients -Hgb F doesn’t copolymerize with Hgb S, this will help avoid SCD severity Bone Marrow/Stem Cell Transplantation – Only cure usually pts are younger than 17 -Not performed often due to lack of HLA matched donors. -can use cord blood but the sample is too little for an adult , or genetic alteration of fetal hematopoietic cells -Gene therapy with CRISPR - increases hgb F

Question 59

What does the name sickle cell trait mean

Answer 59

-benign condition that is a heterozygous state (Hb AS). -asypm unless extreme hypoxic conditions causing systemic sickling, VOC in spleen and brain SCT will have kidney impairment due to iminished perfusion to kidneys

Question 60

What is Hemoglobin C

Answer 60

point mutation in the β - globin gene Result – change in 6th amino acid in β - globin chain Lysine replaces Glutamic Acid -change is the charge of molecule leaving thick Hb C crystals even in an oxygenated state - intracellular polymerization -minimal shape disruption therefore less hemolysis and splenic sequestration -if homozygous hb CC then mild disease while hbAC is asymp

Question 61

Hemoglobin E

Answer 61

point mutation in the β - globin gene DIFFERENT AA position than Hgb S/ Hgb C - 26th position -Lysine replaces Glutamic Acid -changes charge of molecule -Leads to abnormal alternative splicing and decreased transcription of mRNA for Hgb E chain -Therefore, reduction of Hgb E production -polymerization doesn’t occur Hgb EE homozygote – Mild anemia with microcytes and targets Hgb E heterozygote – asymptomatic no treatment genetic counselling

Question 62

Hemoglobin C-Harlem

Answer 62

Double substitution on the β chain valine for glutamic acid and aspartic acid for asparagine asymptomatic.

Question 63

Hemoglobin SC

Answer 63

compound heterozygosity glutamic acid is replaced by valine (Hb S) on one β-globin chain and by lysine (Hb C) on the other β-globin chain -inherited Hb S from one parent and Hb C from the other parent. dont produce any Hb A -MILDER than SCD, but severe in some cases: Hgb SC doesn’t present significant symptoms until teen years -VOC can occur but not alot of damage -Retinopathy and Respiratory tract infections are more common here than SCD

Question 64

Unstable Hemoglobin why it is unstable

Answer 64

-genetic mutations in globin genes that create Hgb products that precipitate in vivo making Heinz bodies and causing HA -substituting a charge AA for UNcharged in the molecule -Substituting polar for non polar AA in the hydrophobic heme pocket -AA substitution at intersubunit contact point -replace AA with valine in alpha helix part of chain -deleting or elongating the chain

Question 65

Unstable Hemoglobin Clinical Findings - causes?

Answer 65

-detected in childhood with HA, jaundice and splenomegaly -made worse with fever or eating oxidant - Hgb precipitates from RBCs as Heinz Bodies -Precipitates in response to factors that don’t affect normal Hgb like drug ingestion, exposure to heat/cold -Precipitated hemoglobin causing clustering of RBC antigens causing the attachment of immunoglobulins and macrophage activation. -Heinz bodies can be trapped in the spleen which will shorten RBC survival Heinz bodies seen on supravital stain Isopropanol precipitation test 🡪 shows flocculent precipitation if unstable hemoglobin is present

Question 66

Sickle Cell Disease

Answer 66

-Polychromasia seen – due to bone marrow hyperplasia -NRBCs seen -Elevated Bilirubin – Jaundice can be seen -marked sickle cells and target cells is the hallmark of SCD.

Question 67

Hemoglobin C what do you see on a pbs

Answer 67

n/n anemia -marked targets -poly -mild retic -bar shaped crystals INTRA and EXTRAcellular with no cell membrane -Folded cells 🡪 ‘envelope’ forms

Question 68

Hemoglobin e what will you see on a pbs

Answer 68

Few to many target cells normal retic

Question 69

Hemoglobin SC what do you see on pbf

Answer 69

Increased polychromasia Few sickle cells Targets, folded cells Intraerythrocytic blunt ended crystals 🡪 ‘mitten cells’ or ‘hand in glove These crystals protrude from the membrane, they’re often branched

Question 70

What you should see in gels for each Beta Thalassemia Minor – Beta thalassemia Major Sickle Cell Trait – Sickle Cell Disease –

Answer 70

Always check the age of a patient An older baby’s distribution of hemoglobin and a patient with β thal intermedia’s distribution can look very similar. Beta Thalassemia Minor – Increased Hgb A2 is hallmark Beta thalassemia Major – Increased Hgb F, Increased Hgb A2. Decreased Hgb A Sickle Cell Trait – Hgb A is reduced, Hgb S is formed Sickle Cell Disease – No Hgb A, increased Hgb F and increased Hgb S