In this randomized double-blind controlled trial, a four-week iron treatment of IDWA initiated one week after a blood donation had no beneficial effect on fatigue and consistently did not improve aerobic capacity, despite having a significant impact on hemoglobin and ferritin levels. Furthermore, a blood donation does not induce significant fatigue measured one week after donation. This study was sufficiently powered to exclude a clinically significant effect of iron supplementation on fatigue. Consequently, these data provide important information on the well-being of donors: blind iron-supplementation after donation is not justified even if it has been shown that adverse events related to a blood donation penalize blood supply . Taking these data into consideration, we have decided not to introduce iron replacement for young female donors at our transfusion center, since no clinical benefit has been documented. However, further trials focusing on long-term iron deficiency or chronic fatigue among donors could lead to a change in our policy.
Most participants of this study were made iron deficient by a single blood donation while all previous experimental studies included participants with long-term IDWA induced by a progressive imbalance between intake and loss of iron [2–9]. Indeed, the median pre-donation ferritin level of donors randomized in our study (34 ng/mL) was above the threshold of an overt iron deficiency (12 ng/mL to 15 ng/mL)  and IDWA was induced by acute bleeding. Interestingly, our results suggest a difference in clinical responses to short-term and long-term IDWA. Such a rapid transition to IDWA possibly has no effect on non-erythroid compartments, such as nervous tissue or muscle. In this context, our results do not conflict with data from the recent non-controlled trial that showed numerous clinical benefits of iron treatment after donation, reducing fatigue, prostration, difficulty in concentrating, headache, hair loss and nail breakage . Besides the methodological limits of this study, donors treated with iron already had IDWA before donation since their inclusion criterion was a pre-donation level of ferritin of < 10 ng/mL. These donors were therefore more likely to have iron deficits in non-erythroid compartments before blood donation.
However, comparing only biological changes between groups, significantly more donors in the placebo group had a decreased ferritin (P < 0.001) and hemoglobin (P = 0.002) level during intervention. Consequently, we should not neglect that iron treatment could prevent symptomatic deterioration of iron status related to further donations.
The total quantity of elemental iron (2,200 mg) orally administered to each participant in our study was set according to iron loss from a donation. While this is certainly not sufficient to compensate for all occurrences of IDWA, the main purpose of this study was to investigate the clinical effect of iron deficiency induced by a single blood donation. Overall mean changes of hemoglobin (Δ 11 g/L) and ferritin (Δ 13 ng/mL) levels between baseline and the end of the treatment were consistent with expected values. Such a biological change, induced by a comparable amount of elemental iron, was enough in some previous randomized placebo controlled trials to obtain a favorable impact on fatigue and endurance [4–6, 8, 9]. The degree of hypoferritinemia could also be critical. Indeed Verdon et al. showed that the treatment effect on fatigue depended on baseline ferritin levels and was not quantitatively significant among subjects with a ferritin level above 50 ng/mL . However, even if the cut-off level of ferritin used for inclusion in our study was not severe (< 30 ng/mL), the mean ferritin value before treatment was 15 ng/mL, which is comparable to other randomized placebo controlled trials dealing with IDWA and showing a clinical improvement after iron treatment [3, 5, 6, 9].
Blood donors come spontaneously to the donation center and are then clinically selected by professionals as being adequately healthy and fit to donate. Consequently, symptoms of fatigue should not be very frequent, as notably observed with half of the donors reporting a level of fatigue before donation of three or less on the VAS. Moreover, blood donation did not induce clinically significant fatigue  in our study and the minor difference detected reflected rather a regression toward the mean due to a natural fluctuation of fatigue. Therefore, treatment effect on fatigue was possibly absent merely because no symptom was perceived before intervention. Indeed, our randomized controlled trial had the particularity to exclusively use biological inclusion criteria to evaluate treatment effect on fatigue among subjects with IDWA. Furthermore, a measurement of aerobic capacity, which is probably more appropriate for healthy volunteers, did not show any significant treatment effect either, thus strengthening our result on fatigue.
Our study suggests that oral FeSO4 administered to donors with IDWA improves quality of life. Surprisingly, this isolated effect is exclusively related to physical condition. More precisely, the main item of significance concerned pain. To our knowledge there are no studies reporting a significant effect of iron treatment on pain and no physiological basis can support the link between iron treatment and pain. Consequently, despite its significance, this result has been considered as spurious.
Our study showed significant side effects in the treatment group. This significance resulted mainly from hardening of stools (absolute difference: 13%, P < 0.01), which could be considered rather as a slight discomfort. Moreover, despite this significant side effect, drop-out rate for a side effect, adherence to treatment and correct guessing of treatment group were similar for each group. Interestingly, Bruner et al. showed no difference in side effects, particularly concerning constipation, between treatment and placebo, in spite of a higher daily dose of FeSO4 (260 mg of elemental iron daily) and a proportion of subjects in the iron treatment group that correctly guessed their group assignment (62%) similar to that of our trial . According to other clinical trials lacking in a placebo, constipation related to oral FeSO4 is the most frequent adverse effect, ranging from 11% in a study comparing intravenous versus oral iron among postpartum patients  to 30% of new cases in a study testing an older population (mean age 62 years) . Among donors, this side effect seems to occur less frequently: 3% to 13% [22, 38]. Our significant result on these moderate side effects adds, however, an argument against broad-based supplementation after each donation.
Our study had several limitations. Firstly, outcomes of this study were restricted to fatigue, physical performance, mood disorder and quality of life but did not include other consequences of IDWA that could affect the well-being of donors. Treatment of IDWA has been shown to improve cognitive function in randomized controlled trials [35, 39]. Moreover, in a recently reported prospective clinical trial among blood donors, restless legs syndrome was frequent (18%) and iron treatment after donation was effective . Concerning hair loss, evidence that iron treatment is beneficial is still lacking , but data from Pittori et al. suggest the beneficial impact of oral iron treatment . Secondly, our study revealed that anemia was a highly prevalent form of iron deficiency (44%) after donation, which contrasts with data obtained from a comparable cohort of menstruating women in the general population . Indeed, 121 donors were excluded from randomization because of anemia and received a three-month iron treatment. No follow-up data are available since the aim of our study was exclusively to explore IDWA. Anemia one week after a blood donation is not surprising since our national recommendation for the hemoglobin threshold for such a donation is 120 g/L. Female participants who became anemic one week after donation were also observed in a study by Rosvik et al., even in the iron-treatment group (one week of oral iron, 100 mg/day) and in spite of an older mean age (43.2 years; SD = 12.1) and a higher pre-donation hemoglobin level (137 g/L; SD = 0.7) . According to Fowler's data, around 75% of donors return to their initial hemoglobin level after eight weeks but the other subjects need a longer recovery period of up to 15 weeks . Anemia induced by a blood donation may be causative of disabling symptoms and it would be fair to explore this clinically. Thirdly, we cannot exclude that clinical effect of treatment was not detected because follow-up took place too early. Indeed, Pittori et al. observed a significant decrease in fatigue after six months of follow-up, but not after only two months .