Alloimmune Hemolytic Disease Of The Fetus / Newborn: Rh Isoimmunization
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Alloimmune Hemolytic Disease Of The Fetus / Newborn:
Rh Isoimmunization
Professor Hassan A Nasrat
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The Antibodies Arise In The Mother As The Direct Result Of A Blood Group Incompatibility Between The Mother And Fetus. The mother become Isoimmunized.
e.g. When An RhD Negative Mother Carries An RhD Positive Fetus and Develop Isoimmunistion.
In The Fetus: the condition is called: Erythroblastosis Fetalis
In The Newborn: the condition is called: HDN.
Alloimmune Hemolytic Disease Of The Fetus / Newborn:
Definition: The Red Cells Of The Fetus Or Newborn Are Destroyed By Maternally Derived Alloantibodies
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Antibodies That May Be Detected During Pregnancy:
Innocuous Antibodies:
Most Of These Antibody Are IgM Therefore Cannot Cross The Placental Barrier E.G. Those Directed Against Such Specificities As A, P(1), Le(a), M, I, IH And Sd(a).
Antibodies Capable Of Causing Significant Hemolytic Transfusion Reactions:
IgG antibodies, Their Corresponding Antigens Are Not Well Developed At Birth E.g. Lu (b), Yt (a), And VEL —
Antibodies That Are Responsible For HDN : Anti-c, Anti-d, Anti-e, And Anti-k (Kell)
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ISO: is a prefix means similar, equal or uniform.
Isoimmunization: is the process of immunizing a species with antigen derived from the same subject.
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RhD D negativity primarily occurs among Caucasians; the average incidence is 15 percent in this group.
Examples of the blood group distribution in various populations are illustrated below:
• Basques — 30 to 35 percent
Finland — 10 to 12 percent
American blacks — 8 percent
Indo-Eurasians — 2 percent
Native Americans and Inuit Eskimos — 1 to 2 percent.
• Local Studies (population) + 8%
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The RH Antigen – Biochemical and Genetic Aspects
Mechanism of Development of Maternal Rh Isoimmunization
Natural History of Maternal isoimmunization /HD of the Newborn
Pathogenesis of Fetal Erythroblastosis Fetalis
Diagnosis of Rh isoimmunization
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The RH Antigen – Biochemical and Genetic Aspects
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The Rh Antigen- Biochemical Aspects:
The Rh Antigen Is A Complex Lipoprotein.
It Has A Molecular Weight Of Approximately 30,000.
It Is Distributed Throughout The Erythrocyte Membrane In
A Nonrandom Fashion.
The Antigens Can Not Be Seen By Routine
Microscopy, But Can Be Identified By Specific Antisera
Function of the Rh antigen:
Its Precise Function Is Unknown. Rh Null Erythrocytes Have Increased Osmotic Fragility And Abnormal Shapes.
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The RH Antigen- Genetic Aspect
The Rh gene complex is located on the distal end of the short arm of chromosome one.
A given Rh antigen complex is determined by a specific gene sequence inherited in a Mendelian fashion from the parents. one haploid from the mother and one from the father.
Three genetic loci, determine the Rh antigen (i.e. Rh blood group).
Each chromosome will be either D positive or D negative (there is no "d" antigen), C or c positive, and E or e positive.
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DcE
eCd
eCd/EcD
PhenotypeGenotype
D positive
C c
E e
D no small d
Three genetic loci, determine the Rh antigen (i.e. Rh blood group).
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There are Grades Of “Positively” Due To Variation In The Degree of Genetic Expression Of The D Antigen.
Incomplete Expression May Result In A Weakly Positive Patient e.g. Du Variant Of Weakly Rh Positive Patient (They May Even Be Determined As Rh Negative).
A Mother With Du Rh Blood Group (Although Genetically Positive) May Become Sensitized From A D-positive Fetus Or The Other Way Around May Take Place.
Genetic Expression (Rh Surface Protein Antigenicity):
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Incomplete Expression Of The D Antigen Result In A Weakly Positive Patient e.g. Du Variant Of Weakly Rh Positive Patient.
Genetic Expression (Rh Surface Protein Antigenicity):
Du VariantFrank D Positive
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Factors Affect The Expression Of The Rh Antigen
The Number Of Specific Rh-antigen Sites: - The Gene Dose, - The Relative Position Of The Alleles, - The Presence Or Absence Of Regulator Genes.
Interaction Of Other Components Of The Rh Blood Group. Erythrocytes Of Individuals Of Genotype Cde/cde Express Less D Antigen Than Do The Erythrocytes Of Individuals Of Genotype cDE/cde.
The Exposure Of The D Antigen On The Surface Of The Red Cell Membrane.
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DcE
eCd
eCd/EcD
PhenotypePhenotypeGenotypeGenotype
D positive
Antigenicity of the Rh surface protein:
genetic expression of the D allele.
Number of specific Rh antigen sites.
Interaction of components of the Rh gene complex.
Exposure of the D antigen on the surface of the red cell
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Mechanism of Development of Maternal Rh Isoimmunization
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The Mechanism of Development of the Rh Immune Response:
Fetal RBC with Rh +ve antigen
Maternal circulation of an Rh –ve mother
(Primary immune response)(Primary immune response)
The Rh +ve antigen will be cleared by macrophages; processed and transferred to plasma stem cell precursors (Develop an almost
permanent immunologic memory)
With subsequent exposure the plasma cell line proliferate to produce humeral antibodies
(Secondary immune response).(Secondary immune response).
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The Primary Response:
Is a slow response (6 weeks to 6 months). IgM antibodies a molecular weight of 900,000 that does not cross the placenta.
The Secondary Response:
Is a Rapid response IgG antibodies a molecular weight of 160,000 that cross the placenta.
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Exposure to maternal antigen in utero “the grandmother theory”:
Explains the development of fetal isoimmunization in a primigravida, who has no history of exposure to incompatible Rh blood.
Rh negative Fetus and the mother is Rh positive
The Fetus is exposed to the maternal Rh antigen through maternal-fetal transplacental bleed.
The fetus immune system develop a permanent template (memory) for the Rh-positive antigen.
When the fetus becomes a mother herself and exposed to a new load of D antigen
from her fetus (hence the grandmother connection) the immune memory is recalled and a secondary immune response occur.
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Natural History of Rh Isoimmunization
And HD Fetus and Newborn
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The Risk of development of Rh Isoimmunization
The Husband Phenotype And Genotype (40 % Of Rh Positive Men Are Homozygous And 60% Are Heterozygous).
The Antigen Load And Frequency Of Exposure.
ABO Incompatibility
Less than 20% of Rh D incompatible pregnancies actually lead to maternal alloimmunization
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The Amount Of Fetal Cells In Maternal Blood
Why Not All the Fetuses of Isoimmunized Women Develop the Same Degree of Disease?
The Non-responders:
ABO Incompatibility:
Antigenic Expression Of The Rh Antigen:
Classes Of IgG Family
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Pathogenesis of Fetal Erythroblastosis Fetalis
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Complications of Fetal-Neonatal Anemia:
Fetal Hydrops And Stillbirth
Hepatosplenomegaly
Neonatal Jaundice
Compilations Of Neonatal Kernicterus (Lethargy,
Hypertonicity, Hearing Loss, Cerebral Palsy And
Learning Disability)
Neonatal Anemia
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ManagementManagement
- Prevention:
- Treatment of Diagnosed cases of Maternal Isoimmunisation:
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Prevention of Rh Isoimmunization
Screening all women for D Factor and antibodies
Prophylaxis (Anti D Immunoglobulin) only for those who are negative for antibodies
The dose of Immunoglobulin depends the volume of Blood
Anti D should be given: 72 hours after delivery, 28-32 weeks, and any other time if there is risk of Fetomateranl
Bleeding
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10 mcg of anti-D Ig should be administered for every mL of fetal blood in the maternal circulation.
Thus, the 300-mcg dose is more than adequate for a typical feto- maternal hemorrhage and covers hemorrhage volumes up to 30 mL of whole fetal blood.
In the less than 1% of cases where the volume of fetomaternal hemorrhage exceeds 30 mL.
The Kleihauer-Betke test to estimate the volume of fetomaternal hemorrhage and administer the appropriate amount of anti-D Ig (10 mcg/mL fetal blood)
Dose of prophylactic Anti-D Ig:
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The amount of fetal cells in maternal blood:
The Kleihauer-Braun-Betke Test
Acid elution technique of Kleihauer. Fetal RBCs stain with eosin (appear dark), adult RBCs do not stain (appear as ghosts). This maternal blood smear contained 11.2% fetal RBCs, representing a transplacental hemorrhage of about 450 mL of blood.
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Management of cases of Rh isoimmunisation
Diagnosis Of RH Isoimmunisation
Evaluation of Fetal Condition
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Diagnosis of Rh isoimmunization
The diagnose is Based on the presence of anti-Rh (D) antibody in maternal serum.
The Enzymatic Method The Antibody Titer In Saline, In Albumin The Indirect Coombs Tests.
Methods of Detecting Anti D Antibodies in Maternal Serum:
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Antibody Titre in Saline: RhD-positive cells suspended in saline solution are agglutinated by IgM anti-RhD antibody, but not IgG anti-RhD antibody. Thus, this test measure IgM, or recent antibody production.
Antibody Titre in Albumin: Reflects the presence of any anti-RhD IgM or IgG antibody in the maternal serum.
The Indirect Coombs Test: o First Step: RhD-positive RBCs are incubated with maternal serumAny anti-RhD antibody present will adhere to the RBCs.
o Second Step:The RBCs are then washed and suspended in serum containing antihuman globulin (Coombs serum). Red cells coated with maternal anti-RhD will be agglutinated by the antihuman globulin (positive indirect Coombs test).
Diagnosis Maternal Isoimmunization
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Is Done After Birth To Detect The Presence Of Maternal Antibody On The Neonate's RBCs.
The Infant's RBCs Are Placed In Coombs Serum.If The Cells Are Agglutinated This Indicate The Presence Of Maternal Antibody
The Direct Coombs Test
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Fetal Rhesus Determination
RHD Type And Zygosity (If RHD-positive) Of The Father
Amniocentesis To Determine The Fetal Blood Type Using The Polymerase Chain Reaction (PCR)
Detection Of Free Fetal RHD DNA (FDNA) Sequences In Maternal Plasma Or Serum Using PCR
Flow Cytometry Of Maternal Blood For Fetal Cells
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Management of cases of Rh isoimmunisation
Diagnosis Of RH Isoimmunisation
Evaluation of Fetal Condition
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The management should be undertaken in specialized fetal units with facilities for therapeutic intervention. Evaluation of the severity of the disease depends on:
(1) history of preceding disease, (2) maternal antibody titers,(3) Determination of fetal Rh blood group, if the technology available (4) Doppler assessment of the fetal blood middle cerebral artery peak systolic velocity for prediction of fetal anemia (5) amniocentesis for amniotic fluid spectrophotometric measurements, and (6) direct fetal blood sampling(7) Ultrasound evaluation
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Methods of Diagnosis and Evaluation of Fetal Rh Isoimmunization
Measurements Of Antibodies in Maternal Serum
Determination of Fetal Rh Blood Group
Ultrasonography
Amniocentesis
Fetal Blood Sampling
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Interpretation of Maternal Anti-D Titer
Antibody Titer Is A Screening Test.
A Positive Anti-d Titer Means That The Fetus Is At Risk For Hemolytic Disease, Not That It Has Occurred Or Will Develop.
Variation In Titer Results Between Laboratories And Intra Laboratory Is Common.
A Truly Stable Titer Should Not Vary By More Than One Dilution When Repeated In A Given Laboratory.
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Ultrasound Image of Transabdominal Chorion Villus Sampling
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To Establish The Correct Gestational Age.
In Guiding Invasive Procedures And Monitoring Fetal Growth And Well-being.
Ultrasonographic Parameters To Determine Fetal Anemia: o Placental Thickness.o Umbilical Vein Diametero Hepatic Size.o Splenic Size.o Polyhydramnios. o Fetal Hydrops (e.g. Ascites, Pleural Effusions, Skin Edema).
Ultrasonography:
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Anemic Fetus Preserves Oxygen Delivery To The Brain By Increasing Cerebral Flow Of Its Already Low Viscosity Blood.
Doppler Velocimetry Of The Fetal Middle Cerebral Artery (MCA)
To Predict The Timing Of A Second Intrauterine Fetal Transfusion.
For Predicting Fetal Anemia
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Amniocentesis
Normally Bilirubin In Amniotic Fluid Decreases With Advanced Gestation.
It Derives From Fetal Pulmonary And Tracheal Effluents.
Its Level Rises in Correlation With Fetal Hemolysis.
Determination Of Amniotic Fluid Bilirubin:
By The Analysis Of The Change In Optical Density Of Amniotic Fluid At 450 nm On The Spectral Absorption Curve (delta OD450)
Procedures Are Undertaken At 10-15 Days Intervals Until Delivery Data Are Plotted On A Normative Curve Based Upon Gestational Age.
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Ultrasound image of amniocentesis at 16 weeks of gestation
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Interpretation Of Amniotic Fluid Bilirubin:
A Falling Curve: Is Reassuring: i.e. An Unaffected Or RhD-negative Fetus.
A Plateauing Or Rising Curve: Suggests Active Hemolysis (Require Close Monitoring And May Require Fetal Blood Sampling And/Or Early Delivery).
A Curve That Reaches To Or Beyond The 80th Percentile Of Zone II On The Liley Graph Or Enters The “ Intrauterine Transfusion" Zone Of The Queenan Curve:
Necessitates Investigation By Fetal Blood Sampling
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Is the gold standard for detection of fetal anemia.
Reserved for cases with: - With an increased MCA-PSV- Increased ΔOD 450
Complications: Total Risk of Fetal Loss Rate 2.7% (Fetal death is 1.4% before 28 weeks and The perinatal death rate is 1.4% after 28 weeks). Bleeding from the puncture site in 23% to 53% of cases. Bradycardia in 3.1% to 12%. Fetal-maternal hemorrhage: occur in 65.5% if the placenta is anterior and 16.6% if the placenta is posterior. Infection and abruptio placentae are rare complications
Fetal blood sampling:
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Diagram of cordocentesis procedure Cordocentesis
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Cordocentesis
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