A presentation on Rh Isoimmunization presented by Chukwuma Onyeije, M.D.

1. Chukwuma I. Onyeije, M.D. Atlanta Perinatal Associates Rh (Rhesus) Isoimmunization:Perinatal Implications and Management

2. A copy of this lecture can also be found at:http://onyeije.net/present/rhdis

3. Objectives Review Terminology Review Major Blood Group Antigens Review Minor Blood group antigens Overview of Clinical Management Discuss Prevention Strategies Q&A

4. Rh Disease Alloimmunization Isoimmunization HDFN ErythroblastosisFetalis Confusing Terminology

5. Rh (Rhesus) Isoimmunization:

7. Red blood cell antigens and antibodies. Pathophysiology

9. INTRODUCTION: Four blood types ( A, B, AB, and O) Each blood type is additionally classified according to the presence or absence of the Rh factor

10. Rh Incompatibility Occurs when there is a different Rh blood type between that of the pregnant mother (Rh negative) and that of the fetus (Rh positive)

11. Rh Incompatibility Exposure to fetal ANTIGENS causes the mother to produce ANTIBODIES

12. The placenta usually acts as a barrier to fetal blood entering the maternal circulation.

13. Fetal cells can enter the maternal circulation through a “break” in the “placental barrier”.

14. Maternal production of Rhesus antibodies following introduction of Rhesus positive blood

15. Maternal production of Rhesus antibodies following introduction of Rhesus positive blood

16. Causes of RBC Transfer: “A break in the barrier” Abortion/Ectopic Pregnancy Partial molar pregnancy Blighted ovum Antepartumbleeding Procedures (amniocentesis, cordocentesis, CVS) External version Platelet transfusion Abdominal trauma Inadvertent transfusion Rh+ blood Postpartum (Rh+baby)

17. IMMUNIZED PREGNANCY NON-IMMUNIZED PREGNANCY Rh Incompatibility

18. Pathophysiology After first antigenic exposure, memory B lymphocytes recognize appearance of RBC’s containing the antigen in subsequent pregnancies

19. Pathophysiology- Fetal Events Maternal antibodies cross the placenta & attach to fetal RBC’s- leading to RBC destruction Sequestration by macrophages in fetal spleen (extravascularhemolysis) produces fetal anemia

20. METABOLISM OF BILIRUBIN:

21. JAUNDICE DOES NOT OCCUR BEFORE DELIVERY:

22. * Hemolysis/Anemia is basis of MCA-PSV ** Increased Bilirubuin is basis of Delta OD-450 * *

23. What’s harmless? What’s dangerous? Antigen Nomenclature

24. CDE (Rhesus) System Clinically Important Includes c, C, D, e, E Rh negative status indicates the absence of D antigen 87% of Caucasians carry the D antigen

25. Other Antibodies Antigens such as A, P, Le (a), M, I, IH, and Sd (a) are innocuous Most are IgM Lewis antibodies and cold agglutinins of I are prevalent but not clinically significant “Lewis Lives”

26. Antibodies Associated with HDFN Anti-c, Anti-D, Anti-E, and Anti-Kell

27. Antibodies Associated with HDFN RhoGAM has decreased HDFN caused by anti-D Anti-D antibody is still MOST COMMON CAUSE of red cell isoimmunization

28. Minor RBC Antigens Kell is most common of minor Responsible for 10% of cases of severe antibody-mediated anemia Mechanism of anemia two-fold 1. Hemolysis 2. Suppression of erythropoiesis **Transfuse women with Kell(-) blood**

29. Minor RBC Antigens Duffy antigens Fy(a) and Fy(b) Only anti-Fy(a) antibody associated with HDFN- may range from mild to severe “Duffy Dies” We treat sensitization to minor RBC antigens similar to those with Rhisoimmunization

30. Minor Antigens MNS system = M, N, S, s, U antigens Anti-M and anti-N naturally occurring- no clinical significance Anti-S, anti-s, and anti-U antibodies ~ mild to severe HDFN

31. Clinical Management Routine blood type screen Repeat Ab screen at 24-28 weeks for Rhnegative women PRIOR to receiving RhoGAM If Abscreen is (+), identify antibody and potential for HDFN

32. Clinical Management For Positive Antibody Screen Determine risk factors for isoimmunization Past pregnancies Transfusions Shared needles Determine father’s RBC antigen status and zygosity If paternity unknown or father is (+) for antigen, fetus is at RISK

33. Clinical Management For Positive Antibody Screen Obtain antibody titer Consider invasive testing at titer of 1:32 or greater by indirect Coombs (1:16 most often used)

34. Clinical Management For Positive Antibody Screen If AB titer remains below critical titer… Invasive testing can be deferred Evaluate serial Ab titers Serial titers are NOT necessary before 18-20 weeks If critical titer noted at first visit, amnio for delta OD450 at 22-24 weeks

35. Clinical Management Fetal Testing Obtain amniocytes to determine fetal blood type When father is heterozygous for the antigen responsible for alloimmunization When paternal status is unknown MCA-PSV can be used as early as 18 weeks~ if greater than 1.5 MoM, consider fetal blood sampling

36. Normal and Abnormal MCA Dopplers

37. The Middle Cerebral Artery Should be examined close to its origin in the internal carotid artery. The angle of the ultrasound beam and the direction of blood flow should be zero degrees. The risk of anemia is highest in fetuses with a peak systolic velocity of 1.5 times the median or higher.

39. MCA Doppler and Fetal Anemia Fetuses with anemia show an increased peak velocity of systolic blood flow in the middle cerebral artery (MCA) MCA Doppler is useful in the determination of fetal anemia in Rh-isoimmunizedpregnancies MCA Doppler is also used to follow fetal response to intrauterine transfusion and to assist in timing subsequent transfusions.

40. MCA Doppler and Fetal Anemia Method: MCA closest to the maternal skin should be measured using a minimal angle of insonation The Doppler gate is placed over the vessel as it bifurcates from the carotid siphon. Serial MCA Doppler studies can be used to generate a curve that plots MCA peak systolic velocity as a function of gestational age. After 35 weeks' gestation, accuracy in determining MCA PSV appears to decrease; therefore, at this gestational age amniocentesis for deltaOD450 is indicated.

41. Clinical Management (cont) Perform serial amniocenteses to measure delta OD450 AND Plot values on Liley Curve “Belt and Suspenders” Approach

42. Delta OD450 Spectral analysis of amniotic fluid at 450 nm measures change in OD Measures the level of bilirubin and predicts severity of hemolytic disease after 27 weeks Delivery or intrauterine transfusion if delta OD450 falls into zone III or upper zone II

43. RhIsoimmunized Pregnancy Worksheet

44. Cordocentesis Gold standard for detection of fetal anemia Complications 2.7% total risk of fetal loss Reserved for patients with increased MCA-PSV or delta OD450

45. Advantages of MCA-PSV Non-invasive NO risk for worsening isoimmunization Utility with alloantibodies other than Rh-D, including anti-Kell antibodies

46. Caution Regarding MCA-PSV From: http://youtu.be/FGUFC39Bgu0

47. Review of Management for RhIsoimmunization Monthly indirect coombs titer (in first sensitized pregnancy) If critical titer reached, determine paternal and fetal antigen status Amniocentesis and delta OD450 OR MCA-PSV ** For 2nd or greater sensitized pregnancy, initiate amnio or MCA at 18-20 weeks**

48. Prevention (cont) Give 300 mcg dose within 72 hrs of delivery to unsensitizedRh (-) women (Rh positive infant) ACOG: 300 mcg at 28 weeks UNLESS father known to be Rh(-)

50. Prevention Test for excessive fetal-maternal hemorrhage after blunt trauma, abruption, cordocentesis, and bleeding assoc. with previa KleihauerBetke Give RhoGAM for partial molar pregnancy, SAB, TAB, ectopic, chorionic villus sampling, amniocentesis, external version

51. SUMMARY Remember the instances in which to consider RhoGAM SAB, TAB, threatened AB (controversial), ectopic, previa/bleeding, abruption, partial molar, CVS, blunt trauma, cordocentesis Clinically important antibodies: Anti-c, Anti-D, Anti-E, and Anti-Kell, Rarely Anti-Duffy Fy(a) Usually not associated with severe HDFN: , ABO incompatibilities, Anti-Duffy(Fy-b) antibodies, (Duffy Fy-a causes mild to severe HDFN), Anti-A, Anti-P, Anti-M, Anti-I, Anti-IH, Anti-Sd(a)

52. SUMMARY Anti-D still most common cause of red cell alloimmunization, despite RhoGAM Kell = most common minorantigen Critical titer most often used is 1:16 by indirect Coombs Amnio with delta OD450 & MCA-PSV Antibody screens and indications for RhoGAM

53. References Gabbe Obstetrics – Normal and Problem Pregnancies, 4th edition. Creasy R., Resnik R., Iams J., Maternal Fetal Medicine Principles and Practice, 5th edition. ACOG Compendium 2005 Harkness U., Spinnato J., Prevention and Management of RhDisoimmunization. Clinics in Perinatology, Dec 2004 31:4. Pereira L., Jenkins T., Conventional management of maternal red cell alloimmunization compared with management by Doppler assessment of MCA-PSV. American Journal of Obstetrics and Gynecology, Oct 2003 189:4. Cohen D., Hemolytic disease of the newborn: RBC alloantibodies in pregnancy and associated serologic issues. Up to Date, Oct 2004. Barss V., Moise K., Significance of minor red blood cell antibodies during pregnancy. Up to Date, Apr 2005. Online resources: www.austincc.edu/mlt/bb/bb_HDN.ppt http://www.perinatology.com/Archive/Isoimmunization.htm http://emedicine.medscape.com/article/273995-overview http://www.nlm.nih.gov/medlineplus/ency/article/001600.htm

55. Kleihauer-Betke Test % fetal RBC in maternal circulation Fetal erythrocytes contain Hbg F which is more resistant to acid elution than HbgA so after exposure to acid, only fetal cells remain & can be identified with stain 1/1000 deliveries result in fetal hemorrhage > 30ml Risk factors only identify 50%

57. Kleihauer Calculations Fetal red cells = MBV X maternal Hct X % fetal cells in KB newborn Hct MBV – maternal blood volume (usually 5000ml) Fetal cells X 2 = whole blood