A Neonate with Coombs-Negative Hemolytic Jaundice With

Journal of Perinatology (2013) 33, 404–406 r 2013 Nature America, Inc. All rights reserved. 0743-8346/13 www.nature.com/jp PERINATAL/NEONATAL CASE PRESENTATION A neonate with Coombs-negative hemolytic jaundice with spherocytes but normal erythrocyte indices: a rare case of autosomal-recessive hereditary spherocytosis due to alpha-spectrin deficiency

HM Yaish1, RD Christensen2 and A Agarwal3 1Division of Hematology/Oncology, Department of Pediatrics, University of Utah School of Medicine, Salt Lake City, UT, USA; 2Department of Women and Newborns, Intermountain Healthcare, McKay-Dee Hospital Center, Salt Lake City, UT, USA and 3Department of Pathology, University of Utah School of Medicine and ARUP Laboratories, Special Genetics Division, Salt Lake City, UT, USA.

<90 fL.2–4 Most neonates with HS, perhaps 70%, inherit the The diagnosis of hereditary spherocytosis (HS) in a newborn infant is condition in an autosomal-dominant manner, with the remainder generally made on the basis of a positive family history, spherocytes on due to either a de novo mutation or rarely inherited as an blood film and Coombs-negative hemolytic jaundice of variable severity autosomal-recessive disorder. with an elevated mean corpuscular hemoglobin concentration (MCHC) and Neonates with Coombs-negative spherocytic hemolytic jaundice, a low mean corpuscular volume (MCV). In general, sodium dodecyl sulfate where the mother or father are known to have HS, generally do not polyacrylamide gel electrophoresis (SDS-PAGE) quantification of erythrocyte need confirmation of the diagnosis using sodium dodecyl sulfate membrane proteins is not needed to make the clinical diagnosis of HS. polyacrylamide gel electrophoresis (SDS-PAGE) of erythrocyte However, we observed that a neonate with no family history of HS, but with membrane proteins.2 However, we cared for a neonate with abundant spherocytosis on repeated blood films, Coombs-negative hemolytic significant Coombs-negative hemolytic jaundice and anemia who jaundice and normal MCHC and MCV measurements, where SDS-PAGE had no family history of HS, but had spherocytes on blood smear revealed alpha-spectrin deficiency, a rare autosomal-recessive variety of HS yet normal red blood cell (RBC) indices. Erythrocyte membrane that generally has a severe clinical phenotype. protein SDS-PAGE revealed an alpha-spectrin deficiency, indicating Journal of Perinatology (2013) 33, 404–406; doi:10.1038/jp.2012.67 an autosomal-recessive variety comprising <5% of HS cases. Keywords: hereditary spherocytosis; neonatal presentation; autosomal recessive; a-spectrin Case Introduction This male was delivered vaginally at 38 weeks gestation after a Neonates with hereditary spherocytosis (HS) can present with early normal pregnancy and labor to non-consanguineous Caucasian and severe hemolytic jaundice and anemia, requiring intensive parents. No family members had anemia or jaundice. He weighed phototherapy and sometimes early erythrocyte transfusions.1–4 3820 g, Apgar scores were 8 and 9 and he appeared healthy with no dysmorphic features. At 24 h, he was noted to be jaundiced with a Because HS is the most common congenital hemolytic anemia of À1 Caucasians, neonatologists often consider HS in the differential total serum bilirubin of 15.2 mg dl (13.9 unconjugated and 1.3 diagnosis of a patient with unexplained Coombs-negative hemolytic conjugated), and phototherapy was begun. Mother’s and baby’s jaundice. Newborn infants with HS typically have spherocytes on blood types were both O ( þ ) and the direct antiglobulin test (Coombs) was negative. After 3 h, the total serum bilirubin was blood smear and distinctive erythrocyte indices with an elevated À1 mean corpuscular hemoglobin concentration (MCHC), generally 13.8 mg dl , hematocrit 34.1%, and three more phototherapy light >36gdlÀ1, and a low mean corpuscular volume (MCV) generally units were added. At 72 h, the total serum bilirubin was 11.0 mg dlÀ1, reticulocytes 13.6% and hematocrit 37%. At about Correspondence: Dr RD Christensen, Department of Women and Newborns, Intermountain 100 h, he was discharged home with a home-phototherapy unit Healthcare, Intermountain Healthcare, McKay-Dee Hospital Center, NICU 4th floor, 4401 and a total serum bilirubin of 14.1 mg dlÀ1 (13.7 unconjugated Harrison Building, Ogden, Salt Lake City, UT 84403, USA. E-mail: [email protected] and 0.7 conjugated). He was bottle-feeding well and other than Received 6 February 2012; revised 9 April 2012; accepted 18 April 2012 jaundice, he appeared healthy. Autosomal recessive hereditary spherocytosis HM Yaish et al 405

Table 1 Characteristics are shown of the main erythrocyte membrane proteins involved in hereditary spherocytosis (HS)

Protein Gene Chromosomal location Percent of HS cases Severity Inheritance

Ankyrin-1 ANK1 8p11.2 40–50% Mild to moderate Autosomal dominant Band 3 SLC4A1 17q21 20–35% Mild to moderate Autosomal dominant b-Spectrin SPTB 14q23–24.1 15–30% Mild to moderate Autosomal dominant a-Spectrin SPTA1 1q22–23 <5% Severe Autosomal recessive Protein 4.2 EPB42 15q15–21 <5% Mild to moderate Autosomal recessive

The genes encoding these proteins, chromosomal locations of these genes, proportion of HS cases involving each protein, typical clinical severity of the condition and the typical pattern of inheritance are shown.

After 1 week, at the first outpatient visit to the pediatrician, he spherocytes on blood film and a high MCHC and a low MCV was reported to be feeding well and had regained birth weight but suggest the diagnosis of HS.1–4 Indeed, among 34 children with HS was obviously pale with a hemoglobin of 5.8 g dlÀ1. That morning followed in our clinic where we have CBCs during their neonatal he was taken to the Primary Children’s Hospital Hematology period, 33 had an MCHC >36 g lÀ1, whereas only this one patient Clinic for further evaluation and treatment. He was judged to be had a normal MCHC (33.5 g lÀ1) (at term the 5th to 95th adequately hydrated and alert but had marked pallor and scleral/ percentile reference range is 32.8 to 35.1 fL 8). Also, all of our other cutaneous icterus. His heart rate was 170 per min and respiratory neonates with HS had a low MCV, generally <90 fL (the 5th to rate 38 per min. Laboratory values included hematocrit 18.8%, 95th % reference range is 100 to 110 fL at term 8), but in this hemoglobin 6.3 g dlÀ1, red cell distribution width 18.4, MCHC patient, the MCV was normal. Besides the normal erythrocyte 33.5 g lÀ1, MCH 31.8 pg, MCV 94.9 fL, reticulocyte count 8%, platelet indices, this patient differs markedly in transfusion requirement count 357 000 per ml, mean platelet volume 11.0 fL and white blood from our other 33. The others have required at most six erythrocyte cell 14 000 per ml with a normal leukocyte differential cell count transfusions during their first 2 years,whereas this one received 26. but with 3.9 nucleated RBC per 100 white blood cell. Hemoglobin Spherocytes, obvious and abundant on our patient’s blood film, electrophoresis on the state metabolic screen was normal with only are not diagnostic of HS. Spherocytes can occur in neonatal immune hemoglobins F and A reported. The blood film examination revealed hemolytic anemia, particularly ABO hemolytic disease, in sepsis, many spherocytes, polychromasia, nucleated RBC and rare echinocytes. microangiopathic hemolytic anemias and hemoglobin H disease. The RBCs were otherwise normochromic and normocytic, and the Osmotic fragility tests can be abnormal in all conditions where leukocytes and the platelets appeared normal in numbers and spherocytes are prominent, yet can be equivocal in neonates with HS. morphology. Pyruvate kinase activity was 17.3 U gÀ1 Hgb (normal Therefore, osmotic fragility testing, like the appearance of spherocytes 9.0 to 22.0) and G6PD activity was 12.9 U gÀ1 Hgb (normal 7.0 to on blood film, is not entirely diagnostic of HS in a neonate. 20.5). His father and mother had normal blood counts and normal Alpha-spectrin, deficient in our patient, is an integral part of the cellular morphology on blood film examinations. Erythrocyte erythrocyte cytoskeleton, linked, along with b-spectrin, to the membrane protein analysis was performed on the neonate by intracellular face of the plasma membrane through ankyrin and SDS-PAGE with densitometric quantitation5,6 and showed normal band 3 (Table 1). The gene encoding alpha-spectrin (SPTA1)is proportions corresponding with ankyrin, band 3, protein 4.2 and located at 1q22-23. Alpha-spectrin is produced in developing b-spectrin, but deficient a-spectrin.7 erythrocytes in quantities exceeding, by three- to four-fold, the During the next several months, RBC transfusions were quantity of b-spectrin. These excess a-chains are degraded administered almost monthly to keep his hemoglobin intracellularly.2,9 Individuals who are heterozygous for an concentration X7.5 to 8.0 g dlÀ1. Between age 12 months and 24 a-spectrin deficiency will still produce a sufficient quantity months his RBC transfusion rate was about 1 every 3 weeks. At 24 of a-spectrin to balance the b-spectrin and therefore their months, his height, weight and head circumference are all normal erythrocytes are normal. Thus, the parents of our patient, obligate (70 to 85 percentile). He has no evidence of bilirubin neurotoxicity. heterozygotes, have normal blood cell counts and morphology. Hearing and speech appear normal and he is meeting However, neonates that are either homozygous for a SPTAI developmental milestones, but is notably pale prior to each mutation or have compound heterozygous mutations in SPTAI are transfusion and is generally mildly icteric. left with little or no erythrocyte a-spectrin, resulting in the rare and clinically severe autosomal-recessive form of HS we speculate in our patient.14 Discussion Guideline recommendations suggest that SDS-PAGE analysis is When evaluating a neonate with Coombs-negative hemolytic not needed to confirm most cases of HS and that this should be jaundice and a non-informative family history, the presence of reserved for selected atypical cases.2 Our neonate was indeed

Journal of Perinatology Autosomal recessive hereditary spherocytosis HM Yaish et al 406 atypical in that, although spherocytes were prominent, the MCHC 6 Coetzer TL, Sahr K, Prchal JT, Blacklock H, Peterson L, Koler R et al. Four different and MCV were normal. We speculate that in other neonates with mutations in codon 28 of alpha spectrin are associated with structurally and Coombs-negative spherocytic hemolytic jaundice, if the erythrocyte functionally abnormal spectrin alpha 1/74 in hereditary elliptocytosis. J Clin Invest indices are normal, SDS-PAGE analysis can indeed be worthwhile. 1991; 88: 743–749. 7 Wichterle H, Hanspal M, Palek J, Jarolim P. Combination of two mutant alpha spectrin Identifying the underlying cause of the erythrocyte disorder will alleles underlies a severe spherocytic hemolytic anemia. J Clin Invest 1996; 15:98 facilitate genetic counseling and permit anticipatory guidance (10): 2300–2307. regarding the predicted severity of clinical phenotype. 8 Christensen RD, Jopling J, Henry E, Wiedmeier SE. The erythrocyte indices of neonates, defined using data from over 12000 patients in a multihospital health care system. J Perinatol 2008; 28: 24–28. Conflict of interest 9 Dhermy D, Steen-Johnsen J, Bournier O, Hetet G, Cynober T, Tchernia G et al. The authors declare no conflict of interest. Coinheritance of two alpha-spectrin gene defects in a recessive spherocytosis family. Clin Lab Haematol 2000; 22(6): 329–336. References 10 Delaunay J. Molecular basis of red cell membrane disorders. Acta Haematol 2002; 108(4): 210–218. 1 Perrotta S, Gallagher PG, Mohandas N. Hereditary spherocytosis. Lancet 2008; 18: 372 11 Tolpinrud W, Maksimova YD, Forget BG, Gallagher PG. Nonsense mutations of the alpha- (9647): 1411–1426. spectrin gene in hereditary pyropoikilocytosis. Haematologica 2008; 93:1752–1754. 2 Gallagher PG, Glader B. Hereditary spherocytosis, hereditary elliptocytosis, and other 12 Wilmotte R, Mare´chal J, Morle´ L, Baklouti F, Philippe N, Kastally R et al. Low disorders associated with abnormalities of the erythrocyte membrane In: Greer JP, expression allele a´-LELY of red cell spectrin is associated with mutations in exon 40 Foerster J, Rodgers GM, Paraskevas F, Glader B, Arber DA, Means RT Jr (eds). (a V/41 polymorphism) and intron 45 and with partial skipping of exon 46. J Clin Wintrobe’s Clinical Hematology. Wolters Kluwer/Lippincott/Williams & Wilkins, Invest 1993; 91: 2091–2096. Philadelphia, 12th edn 2009 pp 913–920. 13 Agarwal N, Swierczek S, Lorenzo FR, Prchal JT. Novel exon 2 alpha spectrin mutation 3 Christensen RD, Henry E. Hereditary spherocytosis in neonates with hyperbilirubine- in a Caucasian family with pyropoikilocytosis, elliptocytosis, and normal morphology: mia. Pediatrics 2010; 125: 120–125. Three phenotypes due to decreased spectrin mRNA and intragenic crossover. Blood 4 Sheffield MJ, Christensen RD. Evaluating neonatal hyperbilirubinemia in late preterm 2008; 3840. Hispanic twins led to the diagnosis of hereditary spherocytosis in them, and in their 14 Mariani M, Barcellini W, Vercellati C, Marcello AP, Fermo E, Pedotti P et al. Clinical sibling, and in their mother. J Perinatol 2011; 31: 625–627. and hematologic features of 300 patients affected by hereditary spherocytosis grouped 5 Coetzer T, Layler J, Prchal JT, Palek J. Molecular determinants of clinical expression of according to the type of the membrane protein defect. Haematologica 2008; 93: hereditary elliptocytosis and pyropoikilocytosis. Blood 1987; 70: 766–772. 1310–1317.

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