**8. Hemodialysis-associated hemosiderosis in the era of erythropoiesisstimulating agents**

Recent studies using SQUID and quantitative MRI to estimate liver iron stores in hemodialysis patients suggest a strong link between the infused iron dose and the risk of iron overload. They also strongly challenge the assumed safety of IV iron products, the reliability of current iron biomarker cutoffs, and current monitoring of iron stores in dialysis patients.

Two recent studies have focused on iron overload in hemodialysis patients with serum ferritin levels well above 500 µg/L: Ferrari et al measured hepatic iron content by magnetic resonance relaxometry R2 in 15 Australian patients with a median ferritin of 782 µg/L and found iron overload in 60% of them [48], whereas Ghoti et al [49] more recently analyzed LIC by T2\*MRI, along with spleen, pancreas and heart iron deposits, in 21 iron-overloaded hemodialysis patients with serum ferritin levels above 1000 µg/L; they found hepatic siderosis in 19 patients (90%)(mild in 8, moderate in 5 and severe in 6), and spleen involvement in every case (21/21); pancreas involvement was sought in only 8 patients (because of poor compliance with the exam) and was found in 3 patients (37%); none of the patients had an abnormal cardiac R2\*[49].

Two modern studies have analyzed hepatic iron stores by SQUID, one in 2004 [50] and the other in 2012 with quantitative MRI based on the Rennes University protocol [6], in cohorts of hemodialysis patients treated according to KDOQI and EDTA-ERBP guidelines with ferritin levels within the target range. Canavese et al used the SQUID technique to study 40 Italian patients and found normal LIC in 30% of them (median ferritin 245 µg/L), mild iron overload in 32.5% (median ferritin 329 µg/L) and moderate iron overload in 37.5% (median ferritin 482 µg/L) [50]. It was subsequently claimed that these findings could not be extrapolated to the general hemodialysis population, owing to possible biased selection of an iron-overloaded population [51].

We recently showed that 84% of a cohort of 119 fit hemodialysis patients treated according to contemporary guidelines had hepatic iron overload on MRI (≥ 51 µmol/g dry weight); mild iron overload was seen in 42 patients (35.3%) and moderate iron overload in 22 patients (18.5%), while 36 of these 119 patients (30%) had severe iron overload (≥ 201 µmol/g dry weight) at levels usually seen in genetic hemochromatosis; MRI also revealed spleen anomalies (a sign of secondary hemosiderosis) in several patients [6].

(according Rostoker G, Griuncelli M, Loridon C et al. Hemodialysis-associated hemosiderosis in the era of erythropoie‐ sis-stimulating agents: a MRI study. Am J Med 2012; 125: 991-999)

#### **Figure 2.** Results of a cross-sectional study of 119 hemodialysis patients

In our cross-sectional study, infused iron, hepcidin and C-reactive protein values correlated with hepatic iron stores in both univariate analysis (p<0.05, Spearman test) and binary logistic regression (p<0.05). We found no relationship between the LIC of hemodialysis patients and alcohol consumption (assessed by the AUDIT score) or the major HFE mutation C282Y [6]. Like Canavese et al [50], we found an increased relative risk of iron overload in female patients (relative risk for females: 3.36 (95% CI: 1.03-10.9))[6]. In 11 patients who were monitored closely during parenteral iron therapy, the iron dose infused per month correlated strongly with both the overall increase and the monthly increase in the liver iron concentration (respectively rho=0.66, p=0.0306 and rho=0.85, p=0.0015, Spearman test)(figures 3 and 4)[6].

hemodialysis patients treated according to KDOQI and EDTA-ERBP guidelines with ferritin levels within the target range. Canavese et al used the SQUID technique to study 40 Italian patients and found normal LIC in 30% of them (median ferritin 245 µg/L), mild iron overload in 32.5% (median ferritin 329 µg/L) and moderate iron overload in 37.5% (median ferritin 482 µg/L) [50]. It was subsequently claimed that these findings could not be extrapolated to the general hemodialysis population, owing to possible biased selection of an iron-overloaded

We recently showed that 84% of a cohort of 119 fit hemodialysis patients treated according to contemporary guidelines had hepatic iron overload on MRI (≥ 51 µmol/g dry weight); mild iron overload was seen in 42 patients (35.3%) and moderate iron overload in 22 patients (18.5%), while 36 of these 119 patients (30%) had severe iron overload (≥ 201 µmol/g dry weight) at levels usually seen in genetic hemochromatosis; MRI also revealed spleen anomalies (a sign

(according Rostoker G, Griuncelli M, Loridon C et al. Hemodialysis-associated hemosiderosis in the era of erythropoie‐

In our cross-sectional study, infused iron, hepcidin and C-reactive protein values correlated with hepatic iron stores in both univariate analysis (p<0.05, Spearman test) and binary logistic regression (p<0.05). We found no relationship between the LIC of hemodialysis patients and alcohol consumption (assessed by the AUDIT score) or the major HFE mutation C282Y [6]. Like Canavese et al [50], we found an increased relative risk of iron overload in female patients (relative risk for females: 3.36 (95% CI: 1.03-10.9))[6]. In 11 patients who were monitored closely during parenteral iron therapy, the iron dose infused per month correlated strongly with both the overall increase and the monthly increase in the liver iron concentration (respectively

rho=0.66, p=0.0306 and rho=0.85, p=0.0015, Spearman test)(figures 3 and 4)[6].

population [51].

66 Updates in Hemodialysis

of secondary hemosiderosis) in several patients [6].

sis-stimulating agents: a MRI study. Am J Med 2012; 125: 991-999)

**Figure 2.** Results of a cross-sectional study of 119 hemodialysis patients

**Figure 3.** Correlation between the infused iron dose and iron stores in 11 hemodialysis patients. Relationship between the monthly infused dose of iron and the monthly increase in iron stores evaluated by magnetic resonance imaging (MRI) in 11 hemodialysis patients. The relationship was studied with the Spearman test, which showed a very strong correlation (rho=0.854; *P*=0.0015).

**Figure 4.** Time course of hepatic iron stores studied by magnetic resonance imaging in hemodialysis patients. (A) Ini‐ tial and final hepatic iron concentrations on MRI in 11 patients during iron therapy. (B) Initial and final hepatic iron concentrations on MRI in 33 patients with hepatic iron overload after iron withdrawal (n=19) or after a major iron dose reduction (n=14) (according Rostoker G, Griuncelli M, Loridon C et al. Hemodialysis-associated hemosiderosis in the era of erythropoiesis-stimulating agents: a MRI study. Am J Med 2012; 125: 991-999).

In the 33 patients with iron overload, iron stores fell significantly after iron withdrawal or after a major reduction in the iron dose (first MRI: 220 µmol/g (CI: 60-340); last MRI: 50 µmol/g (CI: 5-210); p< 0.0001, Wilcoxon's paired test)(figure 4)[6]. The slope of the decline in hepatic iron was not significantly different after iron withdrawal (17.9 µmol/g dry weight/month), iron dose reduction (12.8 µmol/g dry weight/month), and renal transplantation (11.9 µmol/g dry weight/month)(p>0.05, Kruskal-Wallis test) [6]. Thus, the frequency of iron overload appears to be markedly underestimated in hemodialysis patients receiving both erythropoeisisstimulating agents and parenteral iron [6,7]. We concluded that most hemodialysis patients receiving ESA and intravenous iron supplementation likely have hepatic iron overload on MRI and called for a revision of guidelines on iron therapy in this setting, especially regarding the amount of iron infused and the use of non invasive methods for monitoring iron stores [6,7].

## **9. Detrimental effects of iron overload in dialysis patients**

The classical (although rare) clinical picture of hemodialysis-associated hemosiderosis in the pre-ESA era (pigmented skin, cirrhosis and cardiac failure associated with multiple endocrine disorders) has totally disappeared from dialysis centers for at least 3 decades [5]. It is also noteworthy that genetic hemochromatosis and secondary hemosiderosis related to hemato‐ logical disorders are now diagnosed very early, long before any organ dysfunction is detected [37,40]. Therefore, iron overload in dialysis patients in the ESA era is more likely to silently increase the burden of complications of dialysed CKD than to have obvious clinical effects.

Three recent epidemiological studies convergently show that excessive IV iron administra‐ tion can adversely affect the prognosis of hemodialysis patients by increasing mortality and cardiovascular events [52-54]. In a prospective cohort study conducted in Taiwan, 1239 hemodialysis patients were followed for one year: 583 patients not receiving iron therapy were compared to 656 patients treated with IV ferric chloride hexahydrate, the latter patients being divided into 3 subgroups according the cumulative dose of IV iron: 40-800 mg/6 months, 840-1600 mg/6 months and 1640-2400 mg/6 month [52]. Patients in the 2 subgroups with the largest cumulative iron dose had higher adjusted mortality (Hazard ratio (HR) 3.1 and 3.7) and more cardiovascular events (HR 3.5 and 5.1) than those not receiving IV iron and those having received less than 820 mg/6 months (136 mg/month) [52]. Similarly, Kuragano and coworkers prospectively followed 1086 Japanese hemodialysis patients during 2 years and compared 4 subgroups of patients: an oral iron group, an oral iron+very low IV iron group, a low IV iron group (< 200 mg/month), and a high IV iron group (> 200 mg/month) [53]. They observed more acute cardiocerebralvasculardisease (hazardratio 6.02) andhospitalizations (hazardratio 2.77) in the high IV iron group, whereas both low (hazard ratio 1.78) and high (hazard ratio 5.22) IV ironregimens increasedthe frequencyofinfectionsbut atdifferentrates [53].Highferritinlevels (consistently above 100 µg/L) were associated with an increase risk of acute cardiocerebral vascular disease (hazard ratio 2.22), infections (hazard ratio 1.76) and death (hazard ratio 2.28) [53]. Similarly, a jump in the ferritin level from low to high (from less to more than 100 µg/L) was associated with an increased risk of acute cardiocerebral vascular disease (hazard ratio 1.59) and death (hazard ratio 6.18) [53]. More recently, the DOPPS study, using Cox regres‐ sion models with multiple adjustments, analyzed associations between IV iron and outcomes in32435hemodialysispatientsfollowedin12countriesfrom2002to2011andfoundanincreased adjusted mortality rate among patients receiving 300-399 mg/month (HR: 1.13) and 400 mg/ month or more (HR: 1.18) as compared with those receiving no iron and those receiving 1-99, 100-199 and 200-299 mg of IV iron per month [54]. Similarly, the risk of hospitalization was elevated (HR: 1.12) in patients receiving 300 mg/month or more of IV iron as compared to those receiving 100-199 mg/month [54]. The results of the Japanese study on the risk of infection are convergent with recent results fom an American study showing that iron maintenance therapy at 200 mg/month is not associated with an increased short-term risk of infections, as encoun‐ tered with bolus characterized by monthly iron exposure of 700 mg [55].

Three mechanisms may act synergystically to increase mortality and cardiovascular events in iron-overloaded dialysis patients, namely increased levels of hepcidin and oxidative stress, and arterial structural changes.

Some authors recently advocated critical re-evaluation of hepcidin levels in renal failure patients, postulating that hepcidin is not intrinsically elevated in hemodialysis patients but rather reflects poor matching with healthy subjects and frequently excessive iron stores in these patients [56]. It thus seems that hepcidin elevation in fact represents a physiologic defense mechanism against iron overload that is preserved in CKD, even during dialysis [56]. More‐ over, increased levels of hepcidin-25 in patients with severe iron overload on MRI have been shown to normalize in parallel with LIC normalization [6]. As high levels of hepcidin-25 in dialysis patients have recently been linked to fatal and nonfatal cardiovascular events, it is tempting to postulate that the main pathophysiological pathway between iron overload and these events involves pleiotropic effects of hepcidin-25 [57]. The worsening of oxidative stress usually encountered in end-stage renal disease by IV iron infusions and iron overload (mediated by the release of labile, non transferrin-bound iron) may also adversely affect the vascular bed and act as a "second hit" [58, 59]. Finally, in the hemodialysis population, excess iron may also play a direct role in the high burden of cardiovascular complications by impairing endothelial function, as shown in patients with hereditary hemochromatosis [60], and also by favoring atherosclerosis [61, 62].

Given data on heterozygotous genetic hemochromatosis and secondary hemosideroses, the risk/benefit ratio of iron therapy may remain favorable in hemodialysis patients with mild iron overload (LIC < 100 µmol/g), whereas the risk in patients with moderate iron overload (LIC > 100 µmol/g and < LIC < 200 µmol/g) needs to be ascertained [6,7]. It is also tempting to postulate that hemodialysis patients with severe hepatic iron overload (e.g. > 200 µmol/g) are at risk of silent and gradual multiple organ dysfunctions due to hemosiderosis, together with a higher burden of cardiovascular complications [6,7].
