Early Human Development (2008) 84, 515523

a v a i l a b l e a t w w w. s c i e n c e d i r e c t . c o m

w w w. e l s e v i e r. c o m / l o c a t e / e a r l h u m d e v

BEST PRACTICE GUIDELINE

The changing face of haemolytic disease of the newborn

Irene A.G. Roberts
Department of Haematology, 4th Floor Commonwealth Building, Hammersmith Hospital, Du Cane Road, London W12 0NN, United Kingdom

KEYWORDS
Neonate; Hyperbilirubinaemia; Immune; Haemolytic; Transfusion; Immunoglobulin; Metalloporphyrin

Abstract The diagnosis, acute management and follow-up of neonates with haemolytic disease of the newborn (HDN) still represents a significant area of activity for maternity/neonatal services. ABO incompatability is now the single largest cause of HDN in the western world. However, with increasing knowledge at the molecular level, and closer liaison between neonatal paediatricians and haematologists, the diagnosis of non-immune causes of HDN is increasing. As these conditions have an inherited basis and therefore have implications for other family members (or future children), it remains a high priority for all neonatal paediatricians to achieve an accurate diagnosis in all cases of HDN. As the efficacy of phototherapy increases the role of exchange transfusion in acute management is rapidly decreasing. This makes gauging the appropriate time to intervene with exchange transfusion a difficult clinical decision, and guidelines appropriate to the spectrum of contemporary disease are required. In the future intravenous immunoglobulin and/or intramuscular metalloporphyrins may further reduce the need for exchange transfusion and continue to change the spectrum of HDN faced by neonatal paediatricians. 2008 Published by Elsevier Ireland Ltd.

Contents 1. 2. 3. 4. 5. 6. 7. 8. Clinical presentation of HDN . . . . . . . . . . . . . . . . . . . . . . Severe unconjugated hyperbilirubinaemia. . . . . . . . . . . . . . . ABO incompatibility . . . . . . . . . . . . . . . . . . . . . . . . . . . Positive direct anti-globulin test . . . . . . . . . . . . . . . . . . . . Haemolysis in neonates with a known positive maternal antibody test Screening for maternal red cell allo-antibodies . . . . . . . . . . . . . Clinical significance of maternal red cell allo-antibodies . . . . . . . Anti-Kell antibodies . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 516 516 516 517 517 517 518 518

Tel.: +44 208 383 2163; fax: +44 208 742 9335. E-mail address: irene.roberts@imperial.ac.uk.

0378-3782/$ - see front matter 2008 Published by Elsevier Ireland Ltd. doi:10.1016/j.earlhumdev.2008.06.005

516 9. Management of HDN due to red cell allo-antibodies . . . . . . . . . 10.Intrauterine transfusion . . . . . . . . . . . . . . . . . . . . . . . . . 11.Haemolysis detected on the blood film . . . . . . . . . . . . . . . . 12.Prolonged hyperbilirubinaemia . . . . . . . . . . . . . . . . . . . . . 13.How to distinguish HDN from other causes of neonatal haemolysis 13.1. Red blood cell membrane disorders [23,24] . . . . . . . . . 13.2. Red blood cell enzyme defects [2531] . . . . . . . . . . . 13.3. Neonatal haemolysis due to haemoglobinopathies . . . . . 13.4. Alpha thalassaemia major . . . . . . . . . . . . . . . . . . . 14. Developments in therapy for neonatal hyperbilirubinaemia. . . . . 14.1. Improved phototherapy . . . . . . . . . . . . . . . . . . . . 14.2. High dose intravenous immunoglobulin . . . . . . . . . . . 14.3. Metalloporphyrins . . . . . . . . . . . . . . . . . . . . . . . 15. Practice points . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16. Research directions . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

I.A.G. Roberts . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 518 519 519 519 519 519 519 519 520 520 520 520 521 521 521 522

In the space of most paediatricians working lifetime the spectrum of haemolytic disease of the newborn (HDN) has changed beyond recognition. Thirty years ago HDN was almost synonymous with rhesus (Rh) D allo-immunisation and was a common neonatal problem. It was usually unmodified by antenatal therapy and caused overt fetal or neonatal haemolysis leading to marked neonatal hyperbilirubinaemia and attendant anaemia. Affected neonates were commonly sick and unstable at birth, required multiple exchange transfusions and suffered considerable neonatal morbidity and mortality. However, the introduction of routine post-natal prophylactic anti-D immunoglobulin in the 1970s dramatically reduced this form of HDN making this one of the success stories of modern perinatal care. This review aims to discuss HDN in the context of contemporary neonatal services and review its varying presentations. The role of established therapies, particularly improved phototherapy, and potential new therapies, including intravenous immunoglobulin (IVIG), and intramuscular metalloporphyrins will also be assessed.

neonate. If jaundice is clinically obvious within the first 2448 h of life then the neonate may still be hospital-based with their mother. However, with increasing early maternal discharge from maternity services many affected neonates will be re-admitted from the community. In such cases the plasma bilirubin at presentation is often greater than 300 mmol/L. The commonest cause of HDN presenting in this way is that due to ABO incompatibility (see below). However, other clinically significant causes of neonatal haemolysis may also present in this way (Table 1). Therefore all neonates with early-onset, severe and/or prolonged hyperbilirubinaemia should be thoroughly investigated to exclude the possibility of neonatal haemolysis. This is especially pertinent if common causes of HDN (e.g. ABO incompatibility) are excluded, but the degree of hyperbilirubinaemia cannot be satisfactorily explained by the neonate's general course since birth (e.g. state of hydration/ weight loss, establishment of feeding, concurrent neonatal complications i.e. infection).

1. Clinical presentation of HDN

Although the clinical burden imposed on neonatal services by HDN has greatly reduced, it has not disappeared. Neonatal paediatricians continue to recognise a number of different presentations of neonatal haemolysis, many of which are more subtle than the traditional neonatal emergency presentation of severe RhD disease. HDN should be considered where there is one or more of the following: rapidly developing or severe unconjugated hyperbilirubinaemia; a positive direct antiglobulin test (DAT); positive maternal antenatal antibody screening and/or a severely anaemic or hydropic fetus; haemolysis detected on blood film examination; prolonged hyperbilirubinaemia.

3. ABO incompatibility
ABO HDN occurs almost exclusively in the offspring of women of blood group O (although reports exist of occasional cases in group A mothers with high titre anti-group B IgG [1]. In general 15 to 25% of all maternal/fetal pairs are ABO incompatible but ABO HDN is confined to the 1% of group O women that have high-titre IgG antibodies [2]. Haemolysis due to anti-A is more common (1 in 150 births) than anti-B. Affected neonates will usually, but not always, be DAT positive. The proportion of cases of ABO HDN with a positive DAT varies according to the laboratory reagents used; in a recent report 40% of babies requiring phototherapy due to ABO incompatibility had a negative DAT [3]. In contrast to the clinical picture with anti-Rh antibodies, both anti-A and antiB HDN usually result predominately in hyperbilirubinaemia without significant neonatal anaemia. This is mainly because of the relatively low numbers of group A or B sites on neonatal red cells, allowing the antibody-coated cells to remain in the circulation for a longer period than in Rh-D disease [4]. As a reflection of this the blood film in ABO HDN characteristically shows large numbers of spherocytes with little or no increase in nucleated red cells (nRBCs), whereas in

2. Severe unconjugated hyperbilirubinaemia

In contemporary neonatal practice HDN presenting in this way is typically seen without predictive antenatal factors in the early neonatal course of an otherwise well term

The changing face of haemolytic disease of the newborn of black African origin. In such cases severe anaemia as well as hyperbilirubinaemia [5] can occur and antenatal hydrops fetalis has been described [6]. The severity of hyperbilirubinaemia is increased in neonates who also have G6PD deficiency and in those with Gilbert's syndrome [7].

517

Table 1 Causes of unexpected haemolytic disease presenting as early onset or rapidly progressive hyperbilirubinaemia Causes and conditions Comments

ABO incompatability Usually maternal Usually DAT group O, babypositive. Can be group Asevere in cases of maternal group O, baby group B (where mother is of African origin) UndiagnosedAnti-rhesusDAT positive. maternal allo-(D, E, c, C)Following antibodiessensitizing event in pregnancy Anti-KellDAT positive. May present with marked anaemia but mild hyperbilirubinaemia Red blood cellHereditaryDAT negative. membrane defects spherocytosisVariable clinical Hereditarycourse. eliptocytosisCharacteristic red Hereditarycell morphology on pyropoikilocytosis blood film Red blood cellG6PD deficiency DAT negative. May enzyme defectspresent with marked hyperbilirubinaemia without anaemia or blood film signs of haemolysis Pyruvate kinase DAT negative. deficiencyVariable clinical course but severe hyperbilirubinaemia possible Heamoglobinopathies Alpha-DAT negative. May thalassemiararely present with majorhydrops, severe anaemia and hyperbilirubinaemia in liveborn preterm neonates

4. Positive direct anti-globulin test

The direct anti-globulin test (DAT) is a screening test for nonagglutinating antibodies on red blood cells. If maternal serum contains an IgG class immunoglobulin directed against a fetal red blood cell antigen transplacental passage of this antibody will result in red blood cell antibody coating and a positive fetal and neonatal DAT. As a screening test on its own the DAT has a poor positive predictive value for identifying neonates who require treatment of HDN by phototherapy. Dinesh [3] recently reported routine DAT screening of 1724 cord blood samples collected over the first 6 months of 2001 in Wellington, New Zealand. Of these 94 samples (5.5%) showed a positive DAT. However, only 23% of neonates found to have a positive DAT on neonatal screening went on to develop hyperbilirubinaemia requiring phototherapy and it was only when the DATwas strongly positive (4+) that 100% of such neonates required phototherapy. This study confirmed ABO incompatibility as the main cause of HDN in modern neonatal practice as 76/94 cases of DAT positivity occurred in ABO incompatible mother/neonate pairs. Only 6 cases of DAT positivity occurred in neonates where there was antenatal detection of maternal antibodies to red cell antigens with the potential to cause HDN. It should also be appreciated that in a unit where RhD negative women receive routine antenatal anti-D immunoglobulin in the third trimester up to 15% of the resulting neonates will have a positive DAT simply from passive transfer of prophylactic anti-D administered to the mother during pregnancy [8] These antibodies do not cause significant fetal or neonatal haemolysis and so no special postnatal investigation or monitoring of such neonates is required. If there is doubt about the cause of the positive DAT in such neonates blood film examination will point to the correct diagnosis as neither spherocytes nor nRBCs will be present in unusual numbers.

5. Haemolysis in neonates with a known positive maternal antibody test

Although most acute presentations of neonatal haemolysis now occur without warning, a significant number of cases of HDN are predicted by the antenatal detection of known maternal red cell allo-antibodies.

RhD HDN there are few spherocytes and large numbers of circulating nRBCs. Blood film examination is therefore usually diagnostic in DAT negative cases of suspected ABO HDN. Management of ABO HDN is usually successful with phototherapy alone provided by modern equipment (see below). Recently the use of high dose intravenous immunoglobulin (IVIG) has been assessed in ABO HDN and this therapy may be beneficial in selected cases (see below). Even with these therapies close monitoring of the affected neonate is essential as exchange transfusion is occasionally required. This is particularly the case in ABO HDN due to antiB antibodies where racial differences in disease severity exist - severe cases being prevalent in mothers and neonates

6. Screening for maternal red cell allo-antibodies

Recent large population screening studies demonstrate the prevalence of anti-RhD antibody has greatly reduced reflecting the reduction in RhD alloimmunisation achieved by the use of prophylactic anti-D immunoglobulin. Geifman-Holtzman et al. [9] reported antibody screening in 37 506 female serum samples, the majority of which were from women of reproductive age. Of the total, 452 (1.2%) showed positive antibody screens. In these positive women the frequencies of

518 specific antibodies were: anti-D, 18.4%; anti-E, 14%; anti-c 5.8%; anti-C, 4.7%; anti-Kell group, 22%; anti-MNS, 4.7%; antiFya (Duffy), 5.4%; and anti-Jka, (Kidd) 1.5%. This gives an overall rate of anti-D positivity of 2.6 per 1000 samples analysed, compared with 43.3 per 1000 samples reported in a study by Polsky et al. in 1967. In a further study assessing antibody screening in 21 730 pregnant women in Yugoslavia, 630 samples gave positive screens: 254 being anti-D and 376 other antibodies. However, only 522 samples (2.4% overall) were felt to contain clinically significant antibodies.

I.A.G. Roberts proliferation of these cells may be responsible for the thrombocytopenia and leukopenia sometimes seen in newborns with HDN due to anti-Kell [12]. In anti-Kell disease the lack of a consistent correlation between the degree of fetal anaemia and the standard methods of monitoring pregnancies affected by HDN (maternal antibody titres and amniotic fluid OD 450) has previously made the assessment of such pregnancies problematic. This has lead to recommendations that such pregnancies should be monitored closely with the use of repeated estimation of fetal haemoglobin concentration by cordocentesis. However, non-invasive methods to assess the risk of fetal anaemia are now being developed and validated to reduce the need for repeated invasive procedures in such pregnancies (see below).

7. Clinical significance of maternal red cell allo-antibodies

Despite this reduction in the prevalence of anti-D antibodies, anti-D continues to be the commonest antibody leading to fetal and neonatal morbidity and mortality. Howard et al. [10] report a study of antenatal antibody screening in 22 264 women, including the fetal and neonatal outcomes in women with positive antibody screens to antigens known to cause HDN. Clinically important antibodies were detected in 244 (1%) of women, of which 100 were anti-D, 73 were directed against other Rh antigens, and 71 were non-rhesus antibodies (anti-Kell being most common). In these antibody positive pregnancies there were 3 intrauterine deaths and 3 fetuses were given intrauterine transfusions. Exchange or top-up transfusions were required in 10 neonates and 27 other neonates had phototherapy. However, the most significant clinical outcomes including: fetal loss prior to 24 weeks gestation, intrauterine death, and the requirement for intrauterine red cell transfusion and/or post-natal exchange transfusion were all more frequent in the fetuses and neonates of mothers with anti-D antibodies. In this study the other maternal antibodies most commonly associated with adverse fetal and neonatal outcomes were: anti-c, anti-E, and anti-Kell (although numbers of affected fetuses/ neonates are too small for meaningful comparisons of severity of outcome related to maternal antibody type).

9. Management of HDN due to red cell allo-antibodies

Although detailed descriptions are beyond the scope of this review, significant advances have been made in the monitoring of fetuses where maternal red cell allo-antibodies put the fetus at risk of haemolysis. Antenatal blood group genotyping by polymerase chain reaction (PCR) from fetal cells obtained by amniocentesis [13] or, more recently, from maternal blood samples [1416] helps to differentiate susceptible form non-susceptible fetuses. In addition, assessing the degree of fetal anaemia non-invasively by middle cerebral artery Doppler studies reduces the need for repeated invasive procedures [17]. For a more detailed overview of these advances the reader is referred to recent reviews [18,19]. The management of affected neonates requires close cooperation between obstetric, neonatal and haematology teams. All neonates at risk should have cord blood taken for measurement of haemoglobin, DAT and bilirubin (to confirm the diagnosis and assess the need for postnatal therapy). All potentially affected neonates should remain in hospital until hyperbilirubinaemia and/or anaemia have been properly assessed and managed and appropriate follow-up arranged. Trial evidence is lacking to support the use of prophylactic phototherapy in affected neonates [20], but in busy post-natal wards this approach at least ensures that those who require effective phototherapy receive it in a timely manner and this may avoid exchange transfusion in a small number of neonates. Appropriate guidelines for exchange transfusion in neonates with HDN are more controversial. Established guidelines suggest exchange transfusion in HDN is required for:

8. Anti-Kell antibodies
HDN due to anti-Kell antibodies appears to have distinct features compared to other commonly occurring maternal antibodies. The degree of fetal and neonatal anaemia in Kell alloimmunised pregnancies can be severe and may be the main clinical feature of the disease without significant hyperbilirubinaemia. Neonatal nucleated red blood cell counts (reflecting fetal erythropoietic activity) may also be inappropriately low. In addition, the degree of fetal anaemia is not well reflected by maternal anti-Kell antibodies titres or by amniotic fluid OD 450. All of these features point to a degree of fetal erythropoietic suppression as part of the clinical picture in anti-Kell alloimmunised pregnancies. This feature of the disease was confirmed by Vaughan et al. [11] who found that using an in-vitro culture system the growth of Kell positive neonatal erythroid progenitor cells was inhibited by the addition of anti-Kell antibodies, suggesting this was, at least in part, the mechanism responsible for the anaemia seen in anti-Kell HDN. It has subsequently been demonstrated that Kell antigens are present on other myeloid progenitor cells and that inhibition of the fetal

severe anaemia: haemoglobin b 10 g/dL at birth (with the possible exception of anti-K which as outlined above can cause significant anaemia without marked hyperbilirubinaemia) and/or severe hyperbilirubinaemia (total serum bilirubin N350 N 10 mmol/L per hour).

However, these values are derived from clinical experience in preventing kerniterus in sick neonates with untreated RhD HDN (an increasingly uncommon presentation). As outlined previously the typical presentation of HDN is now the well term neonate with ABO incompatibility who presents with a total serum bilirubin already in excess of 300 mmol/L (often as

The changing face of haemolytic disease of the newborn a readmission from home), without significant anaemia. Intensive phototherapy (during the 46 h it normally takes to obtain appropriate blood product and organise exchange transfusion) almost always results in a significant drop in the total serum bilirubin. Such neonates may also be candidates for therapy with IVIG (see below) as this will almost certainly be available more quickly than blood for exchange transfusion. This causes a practical clinical dilemma as the blood product(s) used for exchange transfusion in the UK are in short supply and potential wastage has to be avoided. Therefore, whilst these guidelines for exchange transfusion remain pertinent for untreated HDN associated with defined antenatally-detected maternal antibodies, the majority of cases must now be assessed individually depending upon the general condition of the neonate (state of hydration, gestational and post-natal age), family history, an accurate clinical/laboratory diagnosis and the initial response to modern intensive phototherapy (see below). Following this approach the majority of large maternity/neonatal services now perform only a handful of exchange transfusions per year without apparent detriment to the long-term health of neonates affected by HDN [21].

519

12. Prolonged hyperbilirubinaemia

All neonates with prolonged hyperbilirubinaemia should be thoroughly investigated to exclude the possibility of HDN (Table 1). Persistent jaundice is a common presentation of HDN and virtually all the conditions mentioned in this review may present with prolonged hyperbilirubinaemia. Achieving the correct diagnosis may well have implications not only for the jaundiced neonate but also for other family members.

13. How to distinguish HDN from other causes of neonatal haemolysis

13.1. Red blood cell membrane disorders [23,24]
The main clues that a neonate with early-onset or rapidly progressive hyperbilirubinaemia has a red cell membrane disorder are a family history, negative DAT and an abnormal blood film. The most common red cell membrane disorder to present with neonatal hyperbilirubinaemia is hereditary spherocytosis. Most affected neonates are not anaemic but a small number require transfusion. The diagnosis is usually made by the combination of the characteristic red cell spherocytosis on the blood film and family history but more specialised tests such as red cell dye-binding studies or membrane electrophoresis may be helpful in difficult cases. The other red cell membrane disorders that may present with neonatal hyperbilirubinaemia are hereditary elliptocytosis and hereditary pyropoikilocytosis (HPP), both of which are identified by examination of the blood film.

10. Intrauterine transfusion

Neonates who have received repeated intrauterine transfusions (IUT) for treatment of haemolysis have an altered neonatal course. The effect of the first intrauterine transfusion on fetal haemoglobin is short-lived due the presence of residual antibody-coated fetal RBC. However, with subsequent IUT fetal haemopoiesis is suppressed so that newborns who have been transfused until near term, neonatal jaundice is usually mild, they may require no or minimal phototherapy, and neonatal anaemia is absent [22]. By contrast late anaemia in such neonates is common as the haemopoietic suppression induced by IUT may last for many weeks following delivery. Management of such neonates is expectant, initially ensuring adequate nutritional support (including appropriate iron and folic acid supplementation). Following discharge monitoring of haemoglobin concentration, blood film morphology, reticulocyte count and bilirubin is necessary to gauge when haemolysis has stopped and endogenous red cell production has re-commenced. The appropriate interval for this must be tailored to the individual neonate taking into consideration the extent of the underlying haemolysis and, most importantly, the general condition and growth and development of the affected neonate (simply bleeding such neonates weekly often only raises maternal anxiety without necessarily adding useful clinical information). Red cell transfusions are only required in those with symptomatic anaemia (notoriously difficult to evaluate clinically) with most neonates requiring no RBC top-ups, although in some clinically well neonates the haemoglobin level may fall as low as 5 g/dL before rising spontaneously (personal experience).

13.2. Red blood cell enzyme defects [2531]

Inherited red cell enzyme deficiencies, especially glucose-6phosphate dehydrogenase (G6PD) deficiency, are a relatively common cause of neonatal hyperbilirubinaemia. G6PD deficiency presents with early-onset unconjugated hyperbilirubinaemia, which is often severe; significant anaemia is uncommon (refs). It is seen in all ethnic groups with a high prevalence in individuals from central Africa (20%) and the Mediterranean (10%). It is X-linked and so mainly affects males although female heterozygotes occasionally develop neonatal jaundice. The main clues to this diagnosis are the negative DAT, ethnic origin and gender, early onset and usually normal blood film; the diagnosis is made by assaying G6PD in peripheral blood cells. Pyruvate kinase (PK) deficiency is the second most common red cell enzymopathy in neonates. It is autosomal recessive and clinically heterogeneous with presentation varying from hydrops fetalis through severe early onset neonatal hyperbilirubinaemia requiring exchange transfusion to a mild unconjugated hyperbilirubinaemia. The diagnosis is made by measuring pre-transfusion red cell PK activity; the blood film is sometimes distinctive but more often shows nonspecific changes of non-spherocytic haemolysis.

11. Haemolysis detected on the blood film

In modern neonatal practice haemolysis may be detected during routine blood film examination (e.g. during review for evidence of neonatal sepsis). This emphasises the importance of requesting blood film examination on all neonates with non-physiological jaundice (Table 1).

13.3. Neonatal haemolysis due to haemoglobinopathies

The haemoglobinopathies, with the exception of alpha-thalassaemia major, do not usually present in the neonatal period.

520

I.A.G. Roberts are employed correctly (i.e. ensuring correct distance between device and patient, proper maintenance and servicing of phototherapy units). Phototherapy units are now smaller, easier to use around the cot, more efficient particularly high-intensity gallium nitride light-emitting diodes (LEDs) [38,39], and more powerful- the total irradiance that can be applied to an individual neonate has vastly increased. In short phototherapy is now a viable alternative to the planned use of exchange transfusion in the therapy of even moderate to severe HDN [40], and as devices continue to develop and improve phototherapy is likely to play an even greater role in the therapy of HDN. For a fuller description of developments in neonatal phototherapy since its first use the reader is referred to recent reviews [36,41].

13.4. Alpha thalassaemia major

Alpha-thalassaemia major occurs when all 4 alpha-globin genes on chromosome 16 are deleted [32]. It predominantly affects families of south east Asian origin and presents with mid-trimester fetal anaemia or hydrops fetalis which is fatal within hours of delivery (occasional babies have lived a few days). It can occasionally cause diagnostic confusion if a fetus developing the marked anaemia and hydrops characteristic of the condition is delivered preterm and rapidly then develops hyperbilirubinaemia. Therefore, the diagnosis of alpha-thalassaemia major should be suspected in any case of severe fetal anaemia presenting in the second trimester and any case of hydrops fetalis with severe anaemia in which the parents come from south east Asia (it is also seen occasionally in families who originate from India, the Middle East or the Mediterranean). Checking the blood counts of the parents will immediately identify whether they are at risk of having a child with alpha-thalassaemia major- both parents will be carriers of a chromosome 16 in which both of the 2 alpha-globin genes are deleted and so they will have hypochromic, microcytic red cell indices (MCV usually b 74 fl and MCH usually b24 pg). The diagnosis of alpha-thalassaemia major is confirmed by Hb electrophoresis or HPLC (which shows only Hb Barts and Hb Portland; HbF and HbA are absent). The blood film shows hypochromic, microcytic red cells with vast numbers of circulating NRBCs. Neonatal management of alpha-thalassaemia major has no impact on survival unless the baby has received a planned programme of intrauterine transfusions. For fetuses/neonates treated in this way post-natal management is the same as for beta-thalassaemia, i.e. life-long red cell transfusions or bone marrow transplantation after the age of 2 years [33,34].

14.2. High dose intravenous immunoglobulin

In the last 1015 years a number of studies of high dose intravenous immunoglobulin (IVIG) as adjuvant treatment for HDN have been published [4247], and two systematic reviews have been carried out [48,49]. In 2004 Miqdad et al. [42] reported the use of IVIG in a study of 112 well term neonates with hyperbilirubinaemia resulting from DATpositive ABO HDN. In addition to phototherapy the intervention group (n = 56) received 500 mg/kg IVIG over 4 h if the serum bilirubin was rising by 8.5 mmol/L per hour or greater. Exchange transfusion was carried out in all neonates if the serum bilirubin exceeded 340 mmol/L, or was rising by greater than 8.5 mmol/L per hour in the phototherapy only group. In the phototherapy only group 16 neonates were treated with exchange transfusion whereas only 4 neonates in the IVIG group required exchange transfusion. The duration of phototherapy was also reduced in the IVIG group. No side-effects of IVIG were seen. Similar results were seen in a study reported by Alpay et al. in 1999 [44]. They studied 116 neonates with hyperbilirubinaemia resulting from DAT-positive ABO or Rh HDN of whom 58 received IVIG 1 g/kg over 4 h when the serum bilirubin exceeded 204 mmol/L. Exchange transfusion was performed if the serum bilirubin exceeded 290 mmol/L or was rising by more than 17 mmol/L per hour. In the phototherapy only group 22 neonates were treated with exchange transfusion whereas only 8 neonates in the IVIG group required exchange transfusion. Again the duration of phototherapy and hospital stay were significantly reduced in the IVIG group. No adverse effects of IVIG were reported. Similar results have been found in previous smaller studies assessing the use of IVIG in the treatment of HDN. Despite the positive benefits of IVIG suggested by these studies there are methodological difficulties and questions about the safety of IVIG that potentially limit the size of the role IVIG may have in the treatment of HDN. The preponderance of ABO HDN in the larger studies suggests that the neonates assessed are relatively well and the vast majority would be expected to respond to intensive phototherapy alone unless low thresholds for exchange transfusion (bilirubin 290340 mmol/L) are employed. There is also variation in the timing of administration and dose of IVIG between studies. Late anaemia may be more prevalent in those treated with IVIG, presumably because fewer neonates have exchange transfusion and therefore removal of maternal antibody. No major side effects have been reported in the neonates treated with

14. Developments in therapy for neonatal hyperbilirubinaemia

14.1. Improved phototherapy
The changing clinical practice surrounding HDN is, in no small way, the result of improvements in both the understanding and delivery of phototherapy. Phototherapy was first introduced for the treatment of neonatal hyperbilirubinaemia in the late 1950s [35], when white light was the mainstay of treatment. Since then considerable advances have been made and it is now appreciated more fully that the efficacy of phototherapy in reducing neonatal hyperbilirubinaemia is dependant on a number of factors [36]:

the spectral qualities of the delivered light (optimal wavelength range 400520 nm, with peak emissions of 460 nm) irradiance (intensity of light) [37] body surface area receiving phototherapy skin pigmentation total serum bilirubin concentration at commencement of phototherapy duration of exposure.

Modern phototherapy devices are designed to maximise the efficacy of phototherapy to the neonate and clinicians are more appreciative of the importance of ensuring such devices

The changing face of haemolytic disease of the newborn IVIG but since IVIG is a pooled blood product the potential for transmission of blood borne infections remains [50,51]. Given these facts how should neonatal paediatricians approach the use of IVIG in patients with HDN. The current trial evidence clearly points to positive benefits, particularly the reduction in the need for exchange transfusion. Paediatricians are less experienced with this technique due to the reduction of RhD disease and so morbidity associated with this procedure may increase in the future. Therefore the use of a more straightforward but effective therapy should be considered in the limited number of patients where the likelihood of exchange transfusion is greatest. These would include neonates with red cell alloimmunisation unmodified by antenatal therapy or neonates with potential ABO HDN where a previous sibling has suffered from severe disease requiring exchange transfusion. Also the neonate with severe DAT-positive hyperbilirubinaemia readmitted from the community, where the serum bilirubin already exceeds local guidelines for exchange transfusion, but where initial therapy with IVIG is liable to be available more quickly than exchange transfusion. In these relatively rare circumstances adjuvant therapy with IVIG seems justified. A single dose of IVIG of 500 mg/kg appears to be as effective as any other regimen. haemolytic disease) with moderate hyperbilirubinaemia (plasma bilirubin 256308 mmol/L) developing between 48 96 h of age. Despite being a population of relatively uncomplicated neonates a significant number of these would be expected to go on to be treated with phototherapy, often causing maternal anxiety and lengthening hospital stay. The study enrolled a total of 84 neonates, 40 of who received a single intramuscular dose of SnMp at 6 mg/kg body weight. In the control neonates 12 (27%) required phototherapy at a predetermined level of 333 mmol/L, whereas none of the SnMp treated neonates required phototherapy. SnMp treated neonates also required a shorter period of plasma bilirubin monitoring and a reduced number of plasma bilirubin measurements. No adverse effects of SnMp use were observed. Positive effects of SnMP in reducing peak plasma bilirubin concentrations have also been observed in neonates with glucose-6-phosphate dehydrogenase deficiency [55]. In addition neonates of Jehovah's Witness parents have been given SnMP to reduce the likelihood of jaundiced neonates requiring therapy with exchange transfusion [56]. Given these data there is good evidence to suggest that a single dose of SnMP in uncomplicated neonates reduces peak plasma bilirubin concentrations and reduces the need for phototherapy. As these are outcomes that themselves are not likely to result in harm it can be argued that SnMP therapy presents an unknown risk as the long-term consequences of such therapy are not yet fully known. However, phototherapy in relatively well neonates often provokes a high degree of maternal concern and prolongs hospital stay, both of which are unwanted outcomes in modern hospital-based medical practice. Further studies are underway to more fully assess the efficacy and safety of SnMP but the use of metalloporphyrins to reduce the medical burden of neonatal hyperbilirubinaemia may well find a role in the future as models of health care become increasingly community centred.

521

14.3. Metalloporphyrins
Metalloporphyrins are heme analogs that competitively inhibit the activity of heme oxygenase, the rate-limiting enzyme in heme catabolism. This action reduces the formation of bilirubin and makes them potential agents for both the prophylactic and therapeutic reduction of hyperbilirubinaemia in the newborn. Tin (Sn) mesoporphyrins are the most fully studied compounds in this context. In 1988 Kappas et al. [52] reported the prophylactic use of Sn-Protoporphyrin (SnPP) in 122 term infants with DAT-positive ABO incompatability. At doses up to 2.25 mg/kg body weight, administered by 2 or 3 intramuscular injections, they demonstrated a significant reduction in the rate of rise of plasma bilirubin levels beginning at 48 h post SnPP administration that continued until 96 h. The only reported side effect in SnPP treated neonates was transient erythema during the concurrent use of phototherapy in two neonates. In 1994 Valaes et al. [53]. reported the results of 5 sequential studies of the prophylactic use of Sn-Mesoporphyrin (SnMp) in preterm neonates between 30 and 36 weeks gestational age. SnMp was administered at doses up to 6 mg/ kg body weight by intramuscular injection beginning within the first 24 h of life. 517 neonates were studied over 4 years between 1988 and 1992. As the study population were preterm newborns prophylactic phototherapy was commenced at predetermined low levels and the main outcome of the study was a reduction in the requirement for phototherapy in SnMp treated neonates. This was most marked in those neonates receiving the highest dose of SnMp (6 mg/kg) where mean peak incremental plasma bilirubin concentration was reduced by 41% and phototherapy requirements by 76%, compared to control neonates. Transient erythema was again noted in conjunction with phototherapy in SnMp treated neonates but no other adverse effects were noted during the study or at follow-up at 3 and 18 months. More recently Martinez et al. [54] have looked at the therapeutic effect of SnMp in healthy term neonates (without

15. Practice points

The diagnosis, acute management and follow-up of neonates with HDN still represent a significant area of activity for maternity/neonatal services. ABO incompatability is now the single largest cause of HDN in the western world. As the efficacy of phototherapy increases the role of exchange transfusion in the acute management of HDN is rapidly decreasing. With increasing knowledge at the molecular level, and closer liaison between neonatal paediatricians and haematologists, the diagnosis of non-immune causes of HDN is increasing. As these conditions have an inherited basis and therefore have implications for other family members (or future children), it is a high priority for all neonatal paediatricians to achieve an accurate diagnosis in all cases of HDN. All neonates with persistent haemolysis should always be fully investigated and followed up and appropriately.

16. Research directions

Although difficult to achieve, evidence-based guidelines for indications for exchange transfusion in the post-RhD HDN era are required.

522 Maternal sensitization to red cell allo-antigens remains a significant cause of HDN. Further studies aimed at investigating the causes of, and ways to reduce, such sensitization are necessary. The ease of delivery of phototherapy will always mean that further research to advance it's effectiveness in the treatment of HDN is worthwhile. IVIG and IM metalloporphyrins appear to reduce the need for exchange transfusion in neonates with hyperbilirubinaemia. Both are readily available and easy to administer (as apposed to blood for, and the process of, exchange transfusion). Further trials to define optimal patient populations and doses are required.

I.A.G. Roberts
[16] Bianchi DW, Avent ND, Costa JM, van der Schoot CE, et al. Noninvasive prenatal diagnosis of fetal rhesus D: ready for prime(r) time. Obstet Gynecol 2005;106:8414. [17] K.J. Moise Jr. The usefulness of middle cerebral artery Doppler assessment in the treatment of the fetus at risk for anemia. Am J Obstet Gynecol. submitted for publication;198(2):161.e14. [18] Urbaniak SJ, Greiss MA. RhD haemolytic disease of the fetus and the newborn. Blood Rev 2000;14:4461. [19] Harkness UF, Spinnato JA. Prevention and management of RhD isoimmunization. Clin Perinatol 2004;31:72142. [20] Yaseen H, Khalaf M, Rashid N, Darwich M. Does prophylactic phototherapy prevent hyperbilirubinemia in neonates with ABO incompatibility and positive Coombs' test. J Perinatol 2005;25:5904. [21] Newman TB, Liljestrand P, Jeremy RJ, Ferriero DM, Wu YW, Hudes ES, et al. Outcomes among newborns with total serum bilirubin levels of 25 mg per deciliter or more. N Engl J Med 2006;4 (354):1889900. [22] van Kamp IL, Klumper FJ, Meerman RH, Oepkes D, Scherjon SA, Kanhai HH. Treatment of fetal anemia due to red-cell alloimmunization with intrauterine transfusions in the Netherlands, 19881999. Acta Obstet Gynecol Scand 2004;83:7317. [23] Delaunay J. Molecular basis of red cell membrane disorders. Acta Haematol 2002;108:2108. [24] Tse WT, Lux SE. Red blood cell membrane disorders. Br J Haematol 1999;104:213. [25] Luzzatto L. Glucose-6-phosphate dehydrogenase deficiency. In: Saunders WB, Ed Nathan A, Oski FA, editors. Hematology of infancy and childhood, 4th edition. 1993. p. 67495. [26] Kaplan M. Genetic interactions in the pathogenesis of neonatal hyperbilirubinemia: Gilbert's Syndrome and glucose-6-phosphate dehydrogenase deficiency. J Perinatol 2001;21(Suppl 1): S3539. [27] Kaplan M, Hammerman C. Glucose-6-phosphate dehydrogenase deficiency: a potential source of severe neonatal hyperbilirubinaemia and kernicterus. Semin Neonatol 2002;7:1218. [28] www.rialto.com/favism/english/index.mv. [29] Gilsanz F, Vega MA, Gomez-Castillo E, Ruiz-Balda JA, Omeaca F. Fetal anaemia due to pyruvate kinase deficiency. Arch Dis Child 1993;69:5234. [30] Zanella A, Bianchi P. Red cell pyruvate kinase deficiency: from genetics to clinical manifestations. Bailliere's Best Pract Res Clin Haematol 2000;13:5781. [31] Schneider AS. Triosephosphate isomerase deficiency: historical perspectives and molecular aspects. Bailliere's Best Pract Res Clin Haematol 2000;13:11940. [32] Higgs DR. Alpha-thalassaemia. Bailliere's Clin Haematol 1993;6:11750. [33] Chik KW, Shing MM, Li CK, Leung TF, Tsang KS, Yuen HL, et al. Treatment of hemoglobin Bart's hydrops with bone marrow transplantation. J Pediatr 1998;132:103942. [34] Sohan K, Billington M, Pamphilon D, Goulden N, Kyle P. Normal growth and development following in utero diagnosis and treatment of homozygous alpha-thalassaemia. BJOG 2002;109:130810. [35] Cremer RJ, Perryman PW, Richards DH. Influence of light on the hyperbilirubinaemia of infants. Lancet 1958 24;1(7030):10947. [36] Vreman HJ, Wong RJ, Stevenson DK. Phototherapy: current methods and future directions. Semin Perinatol 2004;28:32633. [37] Hart G, Cameron R. The importance of irradiance and area in neonatal phototherapy. Arch Dis Child Fetal Neonatal Ed 2005;90:F437440. [38] Vreman HJ, Wong RJ, Stevenson DK, Route RK, Reader SD, Fejer MM, et al. Light-emitting diodes: a novel light source for phototherapy. Pediatr Res 1998;44:8049. [39] Seidman DS, Moise J, Ergaz Z, Laor A, Vreman HJ, Stevenson DK, et al. A prospective randomized controlled study of

References
[1] Wang M, Hays T, Ambruso DR, Silliman CC, Dickey WC. Hemolytic disease of the newborn caused by a high titer antigroup B IgG from a group A mother. Pediatr Blood Cancer 2005;45:8612. [2] Chen JY, Ling UP. Prediction of the development of neonatal hyperbilirubinemia in ABO incompatibility. Zhonghua Yi Xue Za Zhi (Taipei) 1994;53:138. [3] Dinesh D. Review of positive direct antiglobulin tests found on cord blood sampling. J Paediatr Child Health 2005;41:5047. [4] Oski FA. The erythrocyte and its disorders. In: Saunders WB, Ed Nathan A, Oski FA, editors. Hematology of Infancy and Childhood; 1993. p. 1843. [5] Waldron P, de Alarcon P. ABO hemolytic disease of the newborn: a unique constellation of findings in siblings and review of protective mechanisms in the fetal-maternal system. Am J Perinatol 1999;16:3918. [6] Ziprin JH, Payne E, Hamidi L, Roberts I, Regan F. ABO incompatibility due to immunoglobulin G anti-B antibodies presenting with severe fetal anaemia. Transfus Med 2005;15:5760. [7] Kaplan M. Genetic interactions in the pathogenesis of neonatal hyperbilirubinemia: Gilbert's Syndrome and glucose-6-phosphate dehydrogenase deficiency. J Perinatol 2001;21(Suppl 1):S3539. [8] Cortey A, Brossard Y. [Adverse effects and patient information]. J Gynecol Obstet Biol Reprod (Paris) 2006;35(1 Suppl):1S1128. [9] O. Geifman-Holtzman, M. Wojtowycz, E. Kosmas, R. Artal. Female alloimmunization with antibodies known to cause haemolytic disease. Obstet Gynecol. submitted for publication;89(2):272275. [10] Howard H, Martlew V, McFadyen I, Clarke C, Duguid J, Bromilow I, et al. Consequences for fetus and neonate of maternal red cell allo-immunisation. Arch Dis Child Fetal Neonatal Ed 1998 Jan;78(1):F6266. [11] Vaughan JI, Manning M, Warwick RM, Letsky EA, Murray NA, Roberts IAG. Inhibition of erythroid progenitor cells by anti-Kell antibodies in fetal alloimmune anemia. N Engl J Med Mar 19 1998;338(12):798803. [12] T. Wagner, G. Lanzer, K. Geissler. Kell expression on myeloid progenitor cells. Leuk Lymphoma. submitted for publication;43 (3):479485. [13] Bennett PR, Le Van Kim C, Colin Y, Warwick RM, Chrif-Zahar B, Fisk NM, et al. Prenatal determination of fetal RhD type by DNA amplification. N Engl J Med 1993 26;329: 60710. [14] Birch L, English CA, O'Donoghue K, Barigye O, Fisk NM, Keer JT. Accurate and robust quantification of circulating fetal and total DNA in maternal plasma from 5 to 41 weeks of gestation. Clin Chem 2005 Feb;51(2):31220. [15] Al-Mufti R, Howard C, Overton T, Holzgreve W, Gaenshirt D, Fisk NM, et al. Detection of fetal messenger ribonucleic acid in maternal blood to determine fetal RhD status as a strategy for noninvasive prenatal diagnosis. Am J Obstet Gynecol 1998;179: 2104.

The changing face of haemolytic disease of the newborn

phototherapy using blue and blue-green light-emitting devices, and conventional halogen-quartz phototherapy. J Perinatol 2003;23:1237. Thaithumyanon P, Visutiratmanee C. Double phototherapy in jaundiced term infants with hemolysis. J Med Assoc Thai 2002;85:117681. McDonagh AF. Phototherapy: from ancient Egypt to the new millennium. J Perinatol 2001;1(;21Suppl):S7S12. Miqdad AM, Abdelbasit OB, Shaheed MM, Seidahmed MZ, Abomelha AM, Arcala OP. Intravenous immunoglobulin G (IVIG) therapy for significant hyperbilirubinemia in ABO hemolytic disease of the newborn. J Matern Fetal Neonatal Med 2004;16:1636. Mukhopadhyay K, Murki S, Narang A, Dutta S. Intravenous immunoglobulins in rhesus hemolytic disease. Indian J Pediatr 2003;70:6979. Alpay F, Sarici SU, Okutan V, Erdem G, Ozcan O, Gkc E.Say High-dose intravenous immunoglobulin therapy in neonatal immune haemolytic jaundice. Acta Paediatr 1999;88:2169. Dagoglu T, Ovali F, Samanci N, Bengisu E. High-dose intravenous immunoglobulin therapy for rhesus haemolytic disease. J Int Med Res 1995;23:26471. Voto LS, Sexer H, Ferreiro G, Tavosnanska J, Orti J, Mathet ER, et al. Neonatal administration of high-dose intravenous immunoglobulin in rhesus hemolytic disease. J Perinat Med 1995;23:44351. Rubo J, Albrecht K, Lasch P, Laufktter E, Leititis J, Marsan D, et al. High-dose intravenous immune globulin therapy for hyperbilirubinemia caused by Rh hemolytic disease. J Pediatr 1992;121:937. Alcock GS, Liley H. Immunoglobulin infusion for isoimmune haemolytic jaundice in neonates. Cochrane Database Syst Rev 2002(3):CD003313. [49] Gottstein R, Cooke RW. Systematic review of intravenous immunoglobulin in haemolytic disease of the newborn. Arch Dis Child Fetal Neonatal Ed 2003;88:F6F10. [50] Hayakawa F, Imada K, Towatari M, Saito H. Life-threatening human parvovirus B19 infection transmitted by intravenous immune globulin. Br J Haematol 2002;118:11879. [51] Quinti I, Pierdominici M, Marziali M, Giovannetti A, Donnanno S, Chapel H, et al. European surveillance of immunoglobulin safety results of initial survey of 1243 patients with primary immunodeficiencies in 16 countries. Clin Immunol 2002;104:2316. [52] Kappas A, Drummond GS, Manola T, Petmezaki S, Valaes T. Snprotoporphyrin use in the management of hyperbilirubinemia in term newborns with direct Coombs-positive ABO incompatibility. Pediatrics 1988;81:48597. [53] Valaes T, Petmezaki S, Henschke C, Drummond GS, Kappas A. Control of jaundice in preterm newborns by an inhibitor of bilirubin production: studies with tin-mesoporphyrin. Pediatrics 1994;93:111. [54] Martinez JC, Garcia HO, Otheguy LE, Drummond GS, Kappas A. Control of severe hyperbilirubinemia in full-term newborns with the inhibitor of bilirubin production Sn-Mesoporphyrin. Pediatrics 1999;103:15. [55] Valaes T, Drummond GS, Kappas A. Control of hyperbilirubinemia in glucose-6-phosphate dehydrogenase-deficient newborns using an inhibitor of bilirubin production, Sn-Mesoporphyrin. Pediatrics 1998;101:E1. [56] Kappas A, Drummond GS, Munson DP, Marshall JR. SnMesoporphyrin interdiction of severe hyperbilirubinemia in Jehovah's Witness newborns as an alternative to exchange transfusion. Pediatrics 2001;108:13747.

523

[40]

[41] [42]

[43]

[44]

[45]

[46]

[47]

[48]