A case of pediatric hemoglobinopathy diagnosed through the evaluation of unexplained cyanosis

Variant hemoglobin (Hb) arises from amino acid substitutions in globin chains that create qualitative defects. In Japan, the prevalence of variant Hb is estimated at 1 in 3000. Inheritance is autosomal dominant, with over 1200 known variants. Approximately 70% of cases are asymptomatic, and the 30% that are symptomatic often present with hemolytic anemia or cyanosis.1, 2 We describe the case of a boy with unexplained cyanosis whose arterial oxygen measurements were inconsistent with his clinical condition owing to a hemoglobinopathy. A 3-year-old boy with no notable medical history was admitted to our hospital with a cough and severe cyanosis during acute respiratory syncytial virus infection. On admission, physical examination revealed a body temperature of 39°C, with an oxygen saturation on pulse oximetry (SpO2) of 86% breathing room air. He was active with mild pallor, but no jaundice or hepatosplenomegaly. Despite clinical improvement, his SpO2 remained low (88%–91% in room air); however, echocardiography revealed no evidence of cyanotic congenital heart disease. Chest radiography and computed tomography showed mild pulmonary infiltrates, insufficient to explain the low SpO2. While he was receiving supplemental oxygen, a blood gas test revealed a discrepancy between the partial pressure of arterial oxygen (PaO2), which was above the limit of measurement, and the arterial oxygen saturation (SaO2) (77.6%). Moreover, his family history revealed that his mother and maternal grandfather occasionally demonstrated unexplained low SpO2 readings (85%–93%). Accordingly, a hematological cause was suspected, and informed consent was obtained from his parents to conduct further investigations. A complete blood count revealed mild microcytic, hypochromic anemia (red blood cell [RBC] count 4.32 × 106 cells/μL, hemoglobin concentration 11.2 g/dL, mean corpuscular volume 79.6 fL, mean corpuscular hemoglobin concentration 25.9 pg, and red-cell distribution width of 13.6%, low reticulocyte count (1.4%), and a Mentzer index of 18.4), suggesting iron deficiency anemia. Peripheral blood smears revealed numerous RBCs of unequal size, with hypopigmentation, and malformations (Figure 1a). Blood biochemistry revealed a lactate dehydrogenase level of 225 U/L, total bilirubin level of 0.2 mg/dL, normal haptoglobin level, and HbA1c of 5.6%. Tests for autoimmune hemolytic anemia, including direct and indirect Coombs tests and cold agglutinin tests, were negative. Flow cytometry for CD55/CD59 expression excluded paroxysmal nocturnal hemoglobinuria. Screening for thalassemia (GLT50 test, HbF/HbA2 quantification, and absence of HbH inclusion bodies) was also negative. The RBC membrane band 3 levels were within normal limits; however, the isopropanol test results were positive, suggesting the presence of an unstable Hb variant. Isoelectric-focusing electrophoresis did not detect any distinct abnormal bands, and α1-globin and α2-globin gene sequencing did not show any abnormalities. β-globin gene sequencing was subsequently performed because a hemoglobinopathy was suspected. This revealed a heterozygous missense variant, NM_000518.5:c.112A>G, p.(Thr38Ala) (Figure 1b). The patient is currently being followed up with regular outpatient visits. The mother, who shared a similar SpO2 pattern, had not yet undergone genetic testing but has been given information on hemoglobinopathies. This case highlights the difficulty in diagnosing hemoglobinopathies in pediatric patients. The differential diagnosis of cyanotic conditions includes central cyanosis, typically associated with respiratory or cardiac disorders; peripheral cyanosis, primarily due to impaired peripheral circulation; and hematological cyanosis.3 In this case, investigations ruled out central and peripheral cyanosis. The marked discrepancy between the measured PaO2 and SaO2 strongly suggested the presence of a hemoglobinopathy.4 Additionally, findings from a peripheral blood smear and family history supported the diagnosis of hematological cyanosis. The patient carried a heterozygous missense variant in the β-globin gene in which a polar threonine in codon 38 was replaced with a nonpolar alanine. This variant is located outside any known functional domain or mutational hotspot and was absent from population databases, including gnomAD v4.1.0 and the 8.3KJPN Genome Reference Panel, fulfilling the ACMG/AMP PM2 criterion of the evidence being insufficient to classify this variant as either pathogenic or benign. Therefore, it was classified as a variant of uncertain significance. However, hemoglobin Hazebrouck, NM_000518.5:c.113A>C, p.(Thr38Pro), a hemoglobin variant caused by a nucleotide substitution in the same codon, has been reported to induce cyanosis and hemolytic anemia.5 In this case, genetic testing revealed a novel β-globin variant, demonstrating the necessity of considering hemoglobinopathies in children with persistent, unexplained cyanosis. When cyanosis persists beyond the anticipated clinical course, particularly in patients with a family history of low oxygen saturation, clinicians should broaden the differential diagnosis to include abnormal hemoglobin levels and perform targeted hematologic testing. Although hemoglobinopathies are generally incurable, recognition of the condition enables unnecessary investigations and treatments to be avoided. Therefore, establishing a definitive diagnosis is essential, and documentation of additional cases of β-globin gene variants is warranted. T.S. drafted the manuscript. K.K., T.F., and T.S. managed the patient and provided input on the manuscript. All authors have read and approved the final manuscript. We gratefully acknowledge the Department of Clinical Laboratory, Fukuyama Clinical Laboratory Center Co., Ltd. for their technical assistance and support and Editage (www.editage.com) for English language editing. The authors declare no conflicts of interest. Written informed consent was obtained from the child's parents to publish this report. The data that support the findings of this study are available on request from the corresponding author. The data are not publicly available due to privacy or ethical restrictions.