Eleanor Ryan

Defective bone morphogenic protein signaling underlies hepcidin deficiency in HFE hereditary hemochromatosis

Hereditary hemochromatosis (HH) is a common inherited iron overload disorder. The vast majority of patients carry the missense Cys282Tyr mutation of the HFE gene. Hepcidin, the central regulator of iron homeostasis, is deficient in HH, leading to unchecked iron absorption and subsequent iron overload. The bone morphogenic protein (BMP)/small mothers against decapentaplegic (Smad) signaling cascade is central to the regulation of hepcidin. Recent data from HH mice models indicate that this pathway may be defective in the absence of the HFE protein. Hepatic BMP/Smad signaling has not been characterized in a human HFE‐HH cohort to date. Hepatic expression of BMP/Smad‐related genes was examined in 20 HFE‐HH males with significant iron overload, and compared to seven male HFE wild‐type controls using quantitative real‐time reverse transcription polymerase chain reaction. Hepatic expression of BMP6 was appropriately elevated in HFE‐HH compared to controls (P = 0.02), likely related to iron overload. Despite this, no increased expression of the BMP target genes hepcidin and Id1 was observed, and diminished phosphorylation of Smad1/Smad5/Smad8 protein relative to iron burden was found upon immunohistochemical analysis, suggesting that impaired BMP signaling occurs in HFE‐HH. Furthermore, Smad6 and Smad7, inhibitors of BMP signaling, were up‐regulated in HFE‐HH compared to controls (P = 0.001 and P = 0.018, respectively). Conclusion: New data arising from this study suggest that impaired BMP signaling underlies the hepcidin deficiency of HFE‐HH. Moreover, the inhibitory Smads, Smad6, and Smad7 are identified as potential disruptors of this signal and, hence, contributors to the pathogenesis of this disease. (HEPATOLOGY 2010;)

Underdiagnosis of hereditary haemochromatosis: lack of presentation or penetration?

Background: The majority of hereditary haemochromatosis (HH) patients are homozygous for the C282Y mutation in the HFE gene. We have demonstrated a homozygote frequency of 1 in 83 for the C282Y mutation in a retrospective analysis of Irish neonates. However, a fully developed phenotype is not observed at the same frequency clinically, suggesting that a large proportion of Irish HH patients may remain undiagnosed. Aims: To determine whether underdiagnosis of HH results from the non-specific nature of early symptoms or incomplete penetrance of the C282Y mutation. Methods: Seventy nine C282Y homozygous individuals identified from family screening for HH and 30 HH probands were investigated. Non-specific symptoms (fatigue, arthropathy, and impotence) and their association with iron indices (transferrin saturation and serum ferritin) and hepatic iron deposition were analysed. Results: We found that 78% of men (mean age 42 years) and 36% of women (mean age 39 years) who were identified as C282Y homozygotes following family screening had iron overload, as defined by a transferrin saturation ≥52% combined with a serum ferritin ≥300 μg/l for men and ≥200 μg/l for women. The frequency of reports of non-specific symptoms in those individuals with iron overload was not significantly different from those who did not have iron overload. Conclusions: Our findings indicate that underdiagnosis of HH may be due to the non-specific nature of early symptoms and less frequently to the incomplete penetrance of the C282Y mutation.

Increased DMT1 but not IREG1 or HFE mRNA following iron depletion therapy in hereditary haemochromatosis

Background and aims: While upregulation of divalent metal transporter 1 (DMT1) and iron regulated gene 1 (IREG1) within duodenal enterocytes is reported in patients with hereditary haemochromatosis (HH), these findings are controversial. Furthermore, the effect of HFE, the gene mutated in HH, on expression of these molecules is unclear. This study examines duodenal expression of these three molecules in HH patients (prior to and following phlebotomy), in patients with iron deficiency (ID), and in controls. Methods: DMT1, IREG1, and HFE mRNA were measured in duodenal tissue of C282Y homozygous HH patients, in ID patients negative for the C282Y mutation with a serum ferritin concentration less than 20 μg/l, and in controls negative for C282Y and H63D mutations with normal iron indices, using real time polymerase chain reaction. Results: DMT1 and IREG1 mRNA levels were not significantly different in non-phlebotomised (untreated) HH patients compared with controls. DMT1 expression was significantly increased in HH patients who had undergone phlebotomy therapy (treated) and in patients with ID compared with controls. IREG1 was significantly increased in ID patients relative to controls, and while IREG1 expression was 1.8-fold greater in treated HH patients, this was not statistically significant. HFE mRNA expression was not significantly different in any of the groups investigated relative to controls. Conclusions: These findings demonstrate that untreated HH patients do not have increased duodenal DMT1 and IREG mRNA, but rather phlebotomy increases expression of these molecules, reflecting the effect of phlebotomy induced erythropoiesis. Finally, HFE appears to play a minor role in the regulation of iron absorption by the duodenal enterocyte.

GNPAT variant is not associated with severe iron overload in Irish C282Y homozygotes

Following the recent publication by McLaren et al. whereby the rs11558492 polymorphism in the GNPAT gene (p.D519G) was demonstrated to be a modifier of the hereditary hemochromatosis (HH) phenotype in a cohort of male C282Y homozygotes, we sought to examine this polymorphism in the Irish HH population. We looked at the polymorphism in our C282Y homozygous HH population as a whole (n 5 263), in males only (n 5 177), and in males categorized according to a serum ferritin value at diagnosis of (1) 1000 lg/L (n 5 57, median serum ferritin 1650 lg/L, interquartile range 1196-1947 lg/L) and (2) <1000 lg/L (n 5 100, median serum ferritin 558 lg/L, interquartile range 420-810 lg/L). Allele frequencies of GNPAT p.D519G were similar across all groups (chi-squared test, P 5 0.632). The allele frequency of GNPAT p.519G was 24.5% for the whole population and 23.7% for males only, in agreement with frequencies reported in 35 male C282Y homozygotes resident in the United States, Canada, and Australia (24.3%) and in 274 French male C282Y homozygotes (26.3%). Independent t tests did not demonstrate a significant difference in the allele frequency of p.D519G or mean age in males with a diagnostic serum ferritin of 1000 lg compared to those with a diagnostic serum ferritin of <1000 lg/L, 19.3% versus 25.5% (P 5 0.593), and mean age 50 6 10 years versus 47 6 13 years (P 5 0.094), respectively. The allele frequency of 19.3% in those with a diagnostic serum ferritin of 1000 lg/L contrasts with that reported by McLaren et al. of 38.6% in a cohort comprising 22 males with a diagnostic serum ferritin of >1000 lg/L. Excess alcohol intake was noted in nine individuals (two of whom had serum ferritin 1000 lg/L), and when these were removed from the analysis on the whole population, the p.D519G allele frequency was 24.2% The relationship between diagnostic serum ferritin and the GNPAT p.D519G variant was examined using Pearson product-moment correlation coefficient analysis. Correlation analysis failed to demonstrate any association between the presence of GNPAT p.D519G and diagnostic serum ferritin across all cohorts. In summary, our data show that GNPAT p.D519G is not associated with a high iron phenotype (diagnostic serum ferritin 1000 lg/L) in Irish C282Y homozygotes. Thus, our data disagree with the findings of McLaren et al. but agree with the findings of BardouJacquet et al. and indeed De Tayrac et al., who did not find the rs11558492 polymorphism to be a significant genetic modifier of the C282Y HH phenotype in a genome-wide association study. Further studies are warranted in order to clarify the role of the GNPAT p.D519G variant as a modifier of the HH phenotype.

Letter by Ryan et al Regarding Article, “Do Hemochromatosis Mutations Protect Against Iron-Mediated Atherogenesis?”

To the Editor:nnThe presence of iron within atheromatous plaques is thought to contribute to the pathogenesis of atherosclerosis. The “hemochromatosis paradox,” as referred to in the recent article by Sullivan,1 originates from the observation that despite significant iron overload, patients with hereditary hemochromatosis (HH) are not at increased risk for atherosclerosis. A number of reasons for this have been proposed, not least is the finding of favorable lipid profiles in C282Y homozygotes compared with wild-type controls.2nnThe …