Transfusion and Apheresis Science 47 (2012) 269–270

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Transfusion and Apheresis Science

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Case Report

Autoimmune hemolytic anemia due to auto anti-N in a patient with sepsis

Lakhvinder Singh, Sheetal Malhotra, Gagandeep Kaur, Sabita Basu ⇑, Ravneet KaurDepartment of Transfusion Medicine, Government Medical College and Hospital, Chandigarh 160 032, India

a r t i c l e i n f o

Keywords:Autoimmune hemolytic anemia

Alloantibody

1473-0502/$ - see front matter � 2012 Elsevier Ltdhttp://dx.doi.org/10.1016/j.transci.2012.07.022

⇑ Corresponding author. Address: House No. 544,garh 160 012, India.

E-mail address: drsabitabasu@gmail.com (S. Basu

a b s t r a c t

We describe a case of autoimmune hemolytic anemia (AIHA) due to anti-N in a young malepatient with cellulitis. There have been several reports of anti-N in N positive individuals.But in all these reports, auto anti-N was mostly associated with underlying immunologicalconditions. We report here a case of auto anti-N in a patient of bacterial sepsis without anyunderlying immune disorder.

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1. Introduction

Anti-N is a rare antibody that usually occurs as an IgMcold agglutinin, reacting best in vitro at 4 �C, and weaklyor not at all, both at 37 �C and in the anti-globulin phase.There have been several reports of anti-N in N positiveindividuals [1,2]. The IgM type is known to cause autoag-glutination leading to ABO discrepancies and autoimmunehemolytic anemia [3,4]. Anti-N can also present as an IgGtype. There have been reports where IgG type anti-N hascaused hemolytic transfusion reactions, hemolytic diseaseof the newborn and warm autoimmune hemolytic anemia(AIHA) [2,5].

We illustrate the importance of identifying anti-Nantibody if present in any recipient because both IgMand IgG may be present in the same individual. We de-scribe here a case of auto anti-N which is reactive at both22 and 37 �C. It caused both ABO discrepancy and com-patibility problems because of its high thermal amplitudereactivity.

2. Case report

A 26 year old male patient was admitted to the surgeryemergency unit of our hospital with the chief complaints ofswelling, redness and pain in the right leg. On detailed his-

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tory and examination, he was diagnosed as having right legcellulitis with signs suggestive of septic shock.

An intramuscular injection in the right gluteal regionwas found to be the cause of this cellulitis and sepsis. Hewas a known case of left leg poliomyelitis since childhood.The past history of the patient was insignificant. The pa-tient had been in good health with no history suggestiveof chronic anemia. The patient had not been transfusedearlier.

At presentation, he was icteric, hemodynamicallyunstable and was started on crystalloids. A request fortwo units of packed red blood cells was sent to the bloodbank. At this time, the total serum bilirubin (TSB) was19.5 mg/dL, and unconjugated bilirubin 15.5 mg/dL. Hishemoglobin was 5.2 gm/dL with NRBC’s 14/100 RBC’s.The peripheral blood film showed anisocytosis and poikilo-cytosis with microcytes, normocytes, a few macrocytes, afew macro-ovalocytes, fragmented cells, elliptical cellsand bite cells, features consistent with hemolytic anemia.He also had leucocytosis with neutrophila. Blood cultureof the patient was sent and Staphylococcus aureus and Esch-erichia coli organisms were identified. The patient wasstarted on antibiotics and debridement of the inflamed tis-sue was done. The patient received three blood transfu-sions in the hospital and his hemoglobin was 7.5 gm/dl atthe time of discharge.

The antibody was suspected consequent to discrepantresults in ABO cell and serum grouping (Table 1). The pa-tient’s unwashed red cells reacted 4+ with commercialanti-A and 2+ with anti-B monoclonal antisera. The red

Table 1Results of ABO blood grouping with unwashed cells.

Cell grouping Serum grouping

Anti-A Anti-B Anti-D A1 cells B cells O cells Autocontrol4+ 2+ 4+ 2+ 4+ 3+ 4+

Table 2Thermal reactivity of patient’s serum at various temperatures.

�C Auto cells Allo cells (N +ve)

37 3+ 3+22 3+ 3+4 4+ 4+

270 L. Singh et al. / Transfusion and Apheresis Science 47 (2012) 269–270

cells showed spontaneous agglutination with the patient’sown serum, AB serum, six percent albumin and saline.After five cell washings the blood group was confirmedto be group A (RhD +ve). The direct antiglobulin test waspositive with both polyspecific (anti IgG + anti C3d) andmonospecific (anti IgG) anti-human globulin reagents.The indirect antiglobulin test was also positive. An anti-body screen and identification panel was used and theantibody was identified as anti-N. Since the patient wasin urgent need of blood, 10 units of A(RhD) positive redblood cells were phenotyped for the N-antigen. Out of 10units, three units were found to be negative for the N anti-gen and compatible with the patient sera by the gel tech-nique. All the three units were transfused without anyadverse reaction.

Further work up of this case was done to determine theother serological features of this anti-N in the individual.The anti-N was found to react with both patient and donorN positive red cells. Patient serum was tested for thermalreactivity at 37, 22 and 4 �C and it was found to be reactiveat all these temperatures (Table 2). A dosage phenomenonwas not seen on the antibody screen and identificationpanel.

The patient’s red cells were eluted by a gentle heat elu-tion method and then phenotyped. The patient’s red cellswere M+, N+, S+ and s+. Hence, there was heterozygousexpression of the N antigen. The patient’s red cells werepapainized and tested with the serum. Enzyme treatmentof the cells diminished the autocell reactivity with the pa-tient’s serum, which further confirms the specificity andthe auto character of the anti-N. Treatment of the serumwith dithiothreitol (DTT) destroyed the anti-N activity,suggesting that the antibody is an IgM immunoglobulin.Hence, this was a case of auto anti-N with wide thermalreactivity and IgM in nature. Antibody titers were 1:8 bythe tube technique.

3. Discussion

Auto anti-N has been described as a cause of immunehemolytic anemia in various case reports in the past. Ourcase represents a case of immune hemolytic anemia dueto auto anti-N with wide thermal reactivity. There have

been several reports of auto anti-N in healthy blood donors[4], but the antibodies in all these cases were clinicallyinsignificant because they were reactive only at 4 �C.

The frequency of the M+N�, M+N+, M�N+ phenotypesis 38.5%, 36.9% and 24.6% in the Indian population [5].

Anti-N as an alloantibody is very rare. Patients on regu-lar dialysis may develop anti-N, usually active at 20 �C aswell as 4 �C, but never reactive at 37 �C [7]. These antibod-ies can also be found in healthy donors, pregnant females,patients with carcinoma of the bladder, infectious mono-nucleosis and other viral infections [1,6]. Both viral trans-formation and exposure to foreign substances that crossreact with the N substance have been invoked as possibleetiologic factors in the development of anti-N inducedautoimmune hemolytic anemia. Perhaps by the cleavingof sialic acid residues from the red cell membrane, as isknown to occur with neuraminidase producing organisms,precursor N antigen could get exposed and subsequently isnot recognized as self, resulting in the production of anti-N-like antibodies [8,9]. Both S. aureus and E. coli are non-neuraminidase producing organisms; hence an alternativemechanism of production of auto anti-N needs to be con-sidered in the present case. One explanation could behemolysis occurring secondary to sepsis.

Anti-N in our case, if missed, could have resulted in atransfusion reaction because it was reactive at 37 �C. Inthe index case, development of anti-N and hemolytic ane-mia was associated with sepsis. There was a history ofchronic viral infection as poliomyelitis since childhood.However, there is no clear relationship between produc-tion of anti-N and pre-existing disease. There was no his-tory of any previous blood transfusion and drug intake.The presentation of hemolytic anemia was acute in thiscase. Hence, there is high probability that bacterial infec-tion in our patient led to the development of auto-anti N.However, the hypothesis that viruses and possibly otherbacteria can also act as triggering agents for initiating redcell auto-antibodies requires further research.

References

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[2] Cohen DW, Garratty G, Morel P, Petz LD. Autoimmune hemolyticanemia associated with IgG auto anti-N. Transfusion 1979;19:329–31.

[3] Winn LC, Eska PL, Grindon AJ. ABO discrepancy caused by auto anti-N.Transfusion 1975;15:612–3.

[4] Metaxas-Buhler M, Ikin W, Romanski J. Anti-N in the serum of ahealthy blood donor of group MN. Vox Sang 1961;6:574–82.

[5] Thakral B, Saluja K, Sharma RR, Marwaha N. Phenotype frequencies ofblood group systems (Rh, Kell, Kidd, Duffy, MNS, P, Lewis, andLutheran) in north Indian blood donors. Transfus Apher Sci2010;43:17–22.

[6] Telischi M, Behzad O, Issitt PD, Pavone BG. Hemolytic disease of thenewborn due to anti-N. Vox Sang 1976;31:109–16.

[7] Crosson JT, Moulds JJ, Comty CM, Polesky HF. A clinical study of anti-Nin the sera of patients in a large repetitive hemodialysis program.Kidney Int 1978;10:463.

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