Abstract

Background: Iron depletion and deferral for low hemoglobin occur rapidly in some donors while others can repeatedly donate without deferral. This variation may be partly explained by significant genetic differences that affect regulation of dietary iron absorption and/or hemoglobin level following blood donation. Previous studies have found that the C282Y and H63D hemochromatosis mutations do not improve dietary iron absorption following donation. TMPRSS6 is a membrane-associated serine protease that degrades hemojuvelin, thereby decreasing hepcidin production with consequent increase in dietary iron absorption. Its physiological importance is best demonstrated by the development of iron resistant iron deficiency anemia in those with TMPRSS6 mutations. A common TMPRSS6 polymorphism, A736V (rs855791), is associated with lower hemoglobin, MCV and transferrin saturation. Its correlation with iron status and hemoglobin production was examined in blood donors undergoing repeated phlebotomy. Methods: Three sample sets from blood donors with well-characterized demographics, blood donation history, hemoglobin and iron parameters enrolled in the approximately 2-year longitudinal Retrovirus Epidemiology Donor Study-II (REDS-II) Iron Status Evaluation Study (RISE) were selected from the NHLBI BioLINCC repository: (Set 1) a random sample of 200 male and 200 female first-time donors and donors reactivated at study enrollment (no donations for 2 years); (Set 2) 114 first-time females who became “frequent” donors by donating at least 4 times in a 2-year period; and (Set 3) 33 repeat male donors, who were deferred for low hemoglobin during the study. Genetic testing for the TMPRSS6 A736V polymorphism was performed on donors in all 3 sets. Variation of hemoglobin and iron status among genotypes, adjusted for donor age, weight, race/ethnicity, number of donations in the previous 12 months and length of time between donations, was assessed using linear models. Results: Genotypic frequencies in Set 1 were 40% AA, 42% A/V and 18% VV. The prevalence was not statistically different in the other two sets of donors, although there was a trend for higher prevalence of VV (30%) in Set 3. Hemoglobin, log-transformed ferritin and body iron stores (calculated based on ferritin and soluble transferring receptor) varied among TMPRSS6 genotypes in females but not in males. For females, average hemoglobin was 0.75 and 0.53 g/dL higher in AA (p<0.0001) and A/V (p=0.0057) than in VV. Average ferritin was 75% and 53% higher in AA (p=0.0024) and A/V (p=0.022) than in VV. Average body iron stores were 2.2 mg/kg and 1.8 mg/kg higher in AA (p=0.0045) and AV (p=0.022) than in VV. For males, average hemoglobin was only 0.13 g/dL higher in AA (p=0.47) and 0.035 g/dL lower in A/V (p=0.83) than in VV. Average ferritin was only 15% higher in AA (p=0.46) and was 8% lower in A/V (p=0.83) than in VV. Average body iron stores were 0.84 mg/kg higher in AA (p=0.10) and 0.15 mg/kg lower in A/V (p=0.75) than in VV. Due to the differences found in females, the longitudinal models were repeated using Set 2 subjects, which confirmed a significant association of the TMPRSS6 polymorphism with hemoglobin in females; 0.50 and 0.35 g/dL higher in AA (p<0.0037) and A/V (p=0.0255) than in VV. However, trends for effects on ferritin and body iron stores did not reach significance. Plasma hepcidin values were also available for Set 2 subjects, but significant differences among TMPRSS6 genotypes were not found. Conclusions: The A736V TMPRSS6 polymorphism is associated with significant differences in hemoglobin and iron status of first-time female blood donors after undergoing iron loss from repeated phlebotomy. The apparent difference between males and females in variation among genotypes may be the result of greater baseline iron depletion in females rather than a gender difference per se. This is the first demonstration of an association between a common polymorphism of iron metabolism and altered individual responses to blood donation. The findings are consistent with a model in which the TMPRSS6 genotypes differ in proteolytic activity towards hemojuvelin, thereby altering hepcidin production and dietary iron absorption in otherwise healthy, but iron depleted individuals.