Hb Santa Juana (β 108(G10) Asn > Ser): a low oxygen affinity hemoglobin variant in a family of Bosnian background

ABSTRACT We present a family that carries the β-hemoglobin variant Hb Santa Juana (HBB:c.326A>G, β 108(G10) Asn>Ser), also known as Hb Serres, in three generations. All affected family members had an anomal hemoglobin fraction as detected by HPLC but normal blood count without evidence of anemia or hemolysis. Oxygen affinity (p50 (O2) = 31.9–40.4 mmHg) was decreased in all probands, compared to 24.9–28.1 mmHg in unaffected individuals. Clinical symptoms likely related to the hemoglobin variant were cyanosis during anaesthesia, while other complaints such as shortness of breath or dizziness were less clearly linked with the hemoglobin variant.

oxygen dissociation curve is right-shifted and characterized by a higher oxygen tension required to reach 50% saturation (p 50 ).However, additional factors contribute to oxygen release into the tissue, such as the concentration of 2,3-bisphosphoglycerate, pH, and CO 2 tension.Weak oxygen binding to hemoglobin favors oxygen release into the tissue, suppressing erythropoietin secretion and causing normocytic anemia.Although some patients expressing hemoglobin variants with decreased oxygen affinity present with cyanosis, most are asymptomatic.A low oxygen affinity hemoglobin variant may be suggested by otherwise unexplained reduced pulse oximetry measurements and normocytic anemia.
Here, we report the clinical presentation and laboratory parameters of four heterozygous carriers of the rare hemoglobin variant Hb Santa Juana inherited over three generations of a family of Bosnian origin (Figure 1, Table 1).At the time when the family presented in our clinic for counseling, a literature search had revealed one published case that described falsely elevated HbA1c measurements in a carrier of Hb Santa Juana [3] and one entry on the variant in the HbVar database that did not provide clinical details [4].
The 60-year-old grandfather II-2 used to play competitive sports (soccer and boxing) at a younger age, but reports to have experienced shortness of breath and a loss of strength during maximum exertion.On the occasion of a medical check-up, he was found to have a reduced pulse oximetry reading 90%.Intraoperative drops in oxygen saturation were not described.He had always been very pale, especially in the face, and frequently developed lip cyanosis.He is regularly taking pantoprazole for symptomatic gastroesophageal reflux.
His 30-year-old daughter III-2 reported to have experienced slow recovery after trivial infections.After a Covid-19 infection in March 2021, chronic fatigue and exhaustion had occurred.Gastroesophageal reflux is known.During surgical procedures, oxygen saturation drops (down to sO 2 = 70%) were repeatedly observed.She had two pregnancies, one with our seven-year-old patient IV-1, the other ending in a late spontaneous abortion at 18 + 3 weeks of pregnancy.During pregnancy, she had developed gestational hypertension which was resistant to therapy with α-methyldopa.With persistent hypertension after pregnancy, she is now taking ramipril.As she is wishing further children, she is currently undergoing evaluation by a fertility center.
Another 13-year-old daughter III-4 also reports drops of oxygen saturation (down to sO 2 = 86%) that was noticed during an esophagogastroduodenoscopy and resulted in the premature termination of the procedure.Like her older sister, III-4 was reported to  slowly after trivial infections and to suffer from gastroesophageal reflux.In addition, she complains of chronic fatigue and exhaustion with accompanying headaches, which is why school attendance is currently incomplete.She is unable to participate in school sports because she quickly develops shortness of breath and dizziness during exercise.For suspected iron deficiency, she had been prescribed an oral iron supplement prior to the presentation in our clinic.
Our youngest patient, the 7-year-old girl IV-1, was born by secondary caesarian section in the 32nd week of gestation with chronic placental insufficiency.She had been diagnosed with latent hypothyroidism and a routine endocrinology examination had revealed an elevated HbA1c (7.5%) that prompted, in combination with the family history, the evaluation for Hb Santa Juana.Further development was unremarkable.Decreased oxygen saturation or signs of anemia were not observed.
We have incomplete information on further family members.The grandfather's 50-year-old sister, II-6, was reported to carry Hb Santa Juana that was diagnosed in her home country, Bosnia.She had experienced decreases of oxygen saturation in the course of two pregnancies.
No other family members have been diagnosed to carry Hb Santa Juana.The family believes the trait has been inherited from the great-grandfather I-1, who had always been pale.He died of cancer at the age of 65 years.
None of the affected individuals showed any abnormalities of blood count, parameters of hemolysis, iron metabolism or erythropoietin levels.The hemoglobin analysis showed a Hb variant with the chromatographic properties of HbE.Molecular genetics [5] confirmed the heterozygous genotype for the hemoglobinopathy Hb Santa Juana (β 108(G10) Asn > Ser; HBB: c.326A > G) in all affected family members.
Because of the history of decreased oxygen saturation during anesthesia and the low-normal hemoglobin levels we hypothesized that Hb Santa Juana, similarly to Hb Schlierbach, Hb Yoshizuka and Hb Presbyterian, which are characterized by point mutations at the same position (supplemental table), may be associated with a decreased oxygen affinity.Because the methodology for directly determining the oxygen saturation curve was not available we validated this hypothesis with a well-established and readily available alternative method that calculates p 50 (O 2 ) using results from venous and capillary blood gas analyses [6].This analysis revealed decreased oxygen affinity in all family members, with p 50 (O 2 ) ranging from 31.9 to 37.5 mmHg (calculated from venous blood samples) and 37.1 to 40.4 mmHg (capillary samples).We compared these results with p 50 (O 2 ) from eight patients not carrying a hemoglobin variant which resulted in significantly lower readings (24.9-28.1 mmHg, p < 0.01), consistent with the normal values reported in the literature (26 ± 1.3 mmHg, [6]).
Decreased oxygen affinity, characterized by a rightward shift in the oxygen binding curve, may be clinically associated with cyanosis, hypoxemia and normocytic anemia.The latter is thought to be a compensatory mechanism resulting from increased oxygen delivery to tissues and consecutive low erythropoietin levels.Because the lower oxygen affinity allows the loaded oxygen of the arterial blood to be exploited very effectively, hemoglobin variants with reduced oxygen affinity typically do not impair physical performance [1].The most apparent symptom cyanosis, either episodically or constantly, appears to be directly linked to oxygen affinity.Variants with an p 50 (O 2 ) that is higher than twice the normal, such as Hb Kansas, Beth Israel, St. Mandé or Bassett, are associated with cyanosis (see Supplemental Table, [7][8][9][10][11][12][13][14][15][16][17][18][19]).In contrast, variants with a p 50 (O 2 ) below 50 mmHg, such as Hb Chico, Sunshine Seth, St. George or Santa Juana, do not or only under certain conditions present with cyanosis but may be detected by low pulse oximetry readings.Mild normocytic anemia has been observed in many of these variants but is typically asymptomatic.The complications and symptoms that had been attributed to Hb Santa Juana in the family we report here, most importantly miscarriage, gastroesophageal reflux disease and severe fatigue, are likely not causally related to the hemoglobin variant.
Another aspect related to Hb Santa Juana is the chromatographic mobility that mimics HbA1c and prompted evaluation of our youngest patient.Aberrantly high HbA1c readings have been described for the hemoglobin variant Santa Juana and need to be taken into account in the differential diagnosis of diabetes mellitus in these patients [3].
In summary, single blood gas analysis can be used to calculate the p 50 (O 2 ) and indicate an abnormal oxygen affinity.Similar to other hemoglobin variants of moderately reduced oxygen affinity, Hb Santa Juana presents with a propensity for low pulsoxymetry readings and cyanosis, especially during anesthesia, but without anemia.

Figure 1 .
Figure 1.Pedigree.Filled shapes indicate family members known to be heterozygous for Hb Santa Juana, while open shapes indicate individuals not carrying Hb Santa Juana.Shapes with questions marks indicate individuals with an unknown carrier status.

Table 1 .
[6]oratory parameters in the four carriers of Hb Santa Juana, reference range in parentheses.p50(O 2 ) was calculated according to Lichtman et al.[6].
*Patient III-4 had received iron supplementation prior to the analysis.