Hb, haemoglobin; MCV, mean corpuscular volume; MCHC, mean corpuscular haemoglobin concentration; RDW, red blood cell distribution width; TIBC, total iron binding capacity.
The homozygous mutation G75R in the human SLC11A2 gene leads to microcytic anaemia and iron overload
Version of Record online: 8 FEB 2012
© 2012 Blackwell Publishing Ltd
British Journal of Haematology
Volume 157, Issue 4, pages 514–516, May 2012
How to Cite
Barrios, M., Moreno-Carralero, M.-I., Cuadrado-Grande, N., Baro, M., Vivanco, J.-L. and Morán-Jiménez, M.-J. (2012), The homozygous mutation G75R in the human SLC11A2 gene leads to microcytic anaemia and iron overload. British Journal of Haematology, 157: 514–516. doi: 10.1111/j.1365-2141.2012.09043.x
- Issue online: 18 APR 2012
- Version of Record online: 8 FEB 2012
- Fondo de Investigaciones Sanitarias. Grant Number: 10/196
- Fundación Mutua Madrileña de Investigación Biomédica. Grant Number: 2010-008
- Instituto de Investigación del Hospital 12 de Octubre
- microcytic anaemia;
- liver iron overload
Microcytic hypochromic anaemia is the most common form of anaemia in children. It can be caused by different abnormalities in the synthesis of globin or haem, or in the availability or acquisition of iron. Solute carrier family 11 (proton-coupled divalent metal ion transporters), member 2 [SLC11A2; also known as (DMT1)] is essential for iron utilization by most types of cells. Mutations in the SLC11A2 gene cause microcytic hypochromic anaemia due to decreased erythroid iron utilization (Iolascon & De Falco, 2009; Iolascon et al, 2009). To date, five patients of Czech, Italian, French and Ecuadorian origin, have been reported with microcytic hypochromic anaemia due to mutations in the SLC11A2 gene. These patients presented anaemia without sideroblasts from birth, reduced mean corpuscular volume, high transferrin saturation and low or moderately high serum ferritin; all patients had hepatic iron overload except the one of Ecuadorian origin. Mutations reported in the SLC11A2 gene are: c.1197G>C (p.E339D) in homozygous state, c.310-3_5delCTT and c.1246C>T (p.R416C) in compound heterozygous state, c.340_342delGTG and c.635G>T (p.G212V) in compound heterozygous state, c.223G>A (p.G75R) in homozygous state, c.1472A>G (p.N491S) and c.635G>T (p.G212V) in compound heterozygous state (Mims et al, 2005; Beaumont et al, 2006; Iolascon et al, 2006; Blanco et al, 2009; Bardou-Jacquet et al, 2011).
We report a 10-year-old boy with non-consanguineous Ecuadorian parents. He was born by vaginal breech delivery after 37 weeks controlled gestation, birth weight of 2·500 kg. He suffered from intrauterine growth retardation; hypertrophic cardiomyopathy resolved after 7 months of age, and he was surgically treated for intestinal atresia in the neonatal period, and was diagnosed with Noonan syndrome. He was under study for chronic hypochromic microcytic anaemia, which was present at birth, when he required transfusion of packed red blood cells (10 ml/kg) (Table 1). Anaemia due to connatal infection, maternal and fetal transfusion, and haemolysis were ruled out. The child was discharged aged 1 month with oral ferrous sulfate supplements.
|Haematological parameters||At birth, 2001||2002||2004||2009||2011||Normal laboratory range|
|Red cells (×1012/l)||4·3||4·2||5·9||3·9||5·6||[3·9–5·2]|
|Reticulocytes cells (×109/l)||87||244||99||194||24||[20–100]|
|Reticulocytes (% RBCs)||2||5·8||1·8||4·9||0·46|
|Serum iron (μmol/l)||40||37||1||8||[9–27]|
|Transferrin saturation (%)||94||96||15||66|
|Serum ferritin (μg/l)||5·5||94||47||164||[20–200]|
In May 2002, the patient, aged 13 months, was referred to the Paediatric Haemato-Oncology Unit in the Hospital Universitario 12 de Octubre of Madrid, for follow-up study. At this time, he had discontinued iron supplements and his haematological parameters were as shown in Table 1: reactive thrombocytosis, normal coagulation and a peripheral blood smear showed hypocromia, microcytosis and anysopoikilocytosis. The white cell series showed no alterations. Oral ferrous sulfate treatment (4–6 mg/kg/d) was restarted, but the patient was refractory despite adequate compliance. Intravenous iron therapy was commenced [50 mg of iron hydroxide per week for 8 weeks, total dose 400 mg (33 mg/kg)], producing a partial response (Hb increased from 56 to 74 g/l). Bone marrow investigations showed hyperplasia of the red cell series with decreased iron deposits and without ringed sideroblasts. Erythrocyte pathology (haemoglobin electrophoresis, enzymopathy and membrane defects) were normal. Oral treatment was suspended in February 2004, after 20 months, when his Hb was 82 g/l (Table 1).
From 2006 to 2009 the patient did not return to consultation. In 2009, aged 8 years, he was referred again with severe microcytic hypochromic anaemia (Table 1), confirmed by preoperative analysis performed before surgery for hypospadias and cryptorchidism, which required packed red blood cell transfusion twice (approximately 525 mg Fe). Once red cell series abnormalities and lead poisoning had been excluded, we resumed the study of microcytic anaemia.
After 5 months of oral iron therapy, the bone marrow study showed iron depletion, and dyserythropoiesis with mild dysplasia, discarding sideroblastic anaemia. Oral iron treatment was suspended in August 2010 after he had maintained a Hb of 80 g/l for 12 months. In October 2010, when an abdominal magnetic resonance imaging (MRI) was performed, the values were 75–140 μmol Fe/g dry weight liver, corresponding to an estimated deposit amount of 90±30 μmol Fe/g, and indicating mild haemosiderosis in accordance with the protocol of iron detection deposits at the University of Rennes.
The patients’ parents gave their informed consent for genetic study following local guidelines. Polymerase chain reaction (PCR) was performed on patient DNA using specific oligonucleotides to amplify the entire coding regions, splice junctions and 5′ and 3′ untranslated regions of the SLC11A2 gene. PCR products were sequenced bidirectionally using the ABI Prism 3130x1 DNA sequencer and compared with the reference sequence (National Center for Biotechnology Information: NC_000012.11) by ClustalW2 software (http://www.ebi.ac.uk/Tools/msa/clustalw2/). The prediction of functional effect was analysed using polyphen software (http://genetics.bwh.harvard.edu/pph2/) and the P49281 protein sequence from UniProtKB/Swiss-Prot database. The study protocol was in accordance with the ethical guidelines of the Declaration of Helsinki.
The patient was found to be homozygous for the mutation c.223G>A in exon 4 of the SLC11A2 gene, which leads to the substitution p.G75R of SLC11A2 protein in the first transmembrane domain. The Polyphen program predicted that this mutation in a highly conserved amino acid is probably damaging. The child in this study had microcytic anaemia since birth with a mild liver iron concentration at 9 years of age; although serum ferritin was not significantly increased. This would be supported by the hypothesis that the defective function of the endosomal SLC11A2 in hepatocytes impairs the iron efflux to the cytosol, leading to iron accumulation in a compartment that does not trigger ferritin synthesis (Priwitzerova et al, 2004; Beaumont et al, 2006). Moreover, the intestinal absorption of haem iron compensates for deficient ferrous iron uptake (Priwitzerova et al, 2004; Beaumont et al, 2006; Iolascon et al, 2006). The volume of transfusions in this patient (approximately 1 g of iron) should not justify the moderate iron deposit in the liver.
This mutation was previously described in a patient, also of Ecuadorian origin, but without deposits of iron in the liver at that time, though authors did not exclude it due to the progressive increase in transferrin saturation (Blanco et al, 2009). The microcytic anaemia caused by the mutation p.G75R in SLC11A2 in these two patients suggests that there may be a common founder origin. A functional study would be necessary to demonstrate whether the mutation p.G75R affects the protein trafficking and/or the iron transport function.
Therefore, it is important to evaluate the liver iron concentration in patients with hypochromic microcytic anaemia with mutations in the SLC11A2 gene, even when serum ferritin levels are normal or moderately elevated.
This work was supported by grants from the “Fondo de Investigaciones Sanitarias” (FIS 10/196) and the “Fundación Mutua Madrileña de Investigación Biomédica” (FMM 2010-008). The authors are grateful for the technical support of the Genomics Facility of “Instituto de Investigación del Hospital 12 de Octubre”.
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