Anna Vergani

Heterogeneity of Hemochromatosis in Italy

BACKGROUND & AIMS Patients with hemochromatosis show variable phenotype expression. We evaluated the frequency of hemochromatosis gene (HFE) mutations and the contribution of HFE genotype, ancestral haplotype, ethnic background, and additional factors (alcohol intake, hepatitis viruses, and beta-thalassemia trait) to the severity of iron overload in a large series of Italian patients with a hemochromatosis phenotype. METHODS HFE genotype was studied in 188 patients. Phenotype evaluation was available in 153 men and 20 women and was based mainly on iron removed. HFE genotype was determined by a polymerase chain reaction restriction assay and ancestral haplotype through D6S265 and D6S105 microsatellite analysis. RESULTS The frequency of C282Y homozygotes was 64%, with a decreasing gradient from north to south. C282Y homozygotes showed more severe iron overload than the other HFE genotypes. In the same group, ancestral haplotype was associated with a more severe phenotype. Additional factors may favor the development of a relatively mild hemochromatosis phenotype in patients nonhomozygous for the C282Y mutation. CONCLUSIONS Hemochromatosis in Italy is a nonhomogenous disorder in which genetic and acquired factors are involved. In patients with a single or no HFE mutation, further studies will enable a differentiation between true genetic disorders and interactions between genetic and acquired factors.

Increased serum ferritin is common in men with essential hypertension

Objectives Insulin-resistance-associated hepatic iron overload syndrome (IRHIO) is characterized by high serum ferritin and presence of metabolic alterations that are part of insulin-resistance syndrome (IRS). Thus, clinical conditions characterized by a high prevalence of IRS may also be characterized by a high prevalence of IRHIO. Design and methods We studied 88 consecutive patients with essential hypertension, 62 patients with IRHIO and 102 healthy normotensive controls. Hemochromatosis, other conditions able to induce secondary iron overload or serum ferritin increase unrelated to body iron stores were excluded. Iron indices, metabolic profiles and hepatic tests in hypertensive with or without increased serum ferritin and in IRHIO with and without hypertension were studied. Metabolic variables, serum iron indices, liver function tests and hepatic ultrasound data were analysed. Data were compared by non-parametric tests. Results In men with hypertension, increased serum ferritin was more frequent than in controls (21 versus 0%, P = 0.001). Hypertensive men with increased serum ferritin had more frequent and pronounced metabolic alterations than those with normal serum ferritin, the metabolic abnormalities and serum ferritin being frequently positively correlated. In hypertensive men with increased serum ferritin, metabolic and iron data were similar to those of IRHIO patients with hypertension. Conclusions In males, hypertension is characterized by a higher prevalence of increased iron stores and metabolic abnormalities that are part of the IRHIO syndrome. This finding may have clinical implications due to the increased risk of IRHIO patients to develop hepatic cirrhosis and also for the role of iron in early atherogenesis.

Haemochromatosis in patients with beta-thalassaemia trait.

Severe iron overload has been reported in patients with the β‐thalassaemia trait. Studies performed before the discovery of the haemochromatosis gene (HFE) have yielded conflicting results: some suggest that iron overload might arise from the interaction of the β‐thalassaemia trait with heterozygosity for haemochromatosis, some with homozygosity for haemochromatosis and others that it was unrelated to haemochromatosis. We have studied the clinical phenotype, iron indices and HFE genotypes of 22 unrelated patients with the β‐thalassaemia trait and haemochromatosis, the inheritance of chromosome 6p and 1q haplotypes in families of non‐homozygous C282Y probands and serum measures of iron status in relatives heterozygous for C282Y with or without the β‐thalassaemia trait. We demonstrate that the β‐thalassaemia trait aggravates the clinical picture of C282Y homozygotes, favouring higher rates of iron accumulation and the development of severe iron‐related complications. We suggest that the coexistence of the β‐thalassaemia trait might also increase the risk of iron overload in patients with HFE genotypes at a mild risk of haemochromatosis. Our findings do not support the hypothesis that the association of the β‐thalassaemia trait with a single C282Y or H63D allele might lead to iron overload and suggest that other non‐HFE‐related inherited factors are present in haemochromatosis patients with incomplete HFE genotypes.

The mitochondrial superoxide dismutase A16V polymorphism in the cardiomyopathy associated with hereditary haemochromatosis

The A16V mitochondrial targeting sequence polymorphism influences the antioxidant activity of MnSOD, an enzyme involved in neutralising iron induced oxidative stress. Patients with hereditary haemochromatosis develop parenchymal iron overload, which may lead to cirrhosis, diabetes, hypogonadism, and heart disease. The objective of this study was to determine in patients with haemochromatosis whether the presence of the Val MnSOD allele, associated with reduced enzymatic activity, affects tissue damage, and in particular heart disease, as MnSOD knockout mice develop lethal cardiomyopathy. We studied 217 consecutive unrelated probands with haemochromatosis, and 212 healthy controls. MnSOD polymorphism was evaluated by restriction analysis. The frequency distribution of the polymorphism did not differ between patients and controls. Patients carrying the Val allele had higher prevalence of cardiomyopathy (A/A 4%, A/V 11%, V/V 30%, p = 0.0006) but not of cirrhosis, diabetes, or hypogonadism, independently of age, sex, alcohol misuse, diabetes, and iron overload (odds ratio 10.1 for V/V, p = 0.006). The frequency of the Val allele was higher in patients with cardiomyopathy (0.67 v 0.45, p = 0.003). The association was significant in both C282Y+/+ (p = 0.02), and in non-C282Y+/+ patients (p = 0.003), and for both dilated (p = 0.01) and non-dilated stage (p = 0.04) cardiomyopathy, but not for ischaemic heart disease. In patients with hereditary haemochromatosis, the MnSOD genotype affects the risk of cardiomyopathy related to iron overload and possibly to other known and unknown risk factors and could represent an iron toxicity modifier gene.

Effects of venesections and restricted diet in patients with the insulin-resistance hepatic iron overload syndrome.

Abstract: Goal: We evaluated the effect of venesections and restricted diet on iron and metabolic indices and liver function tests in patients with insulin‐resistance hepatic iron overload (IR‐HIO).

Iron accumulation in chronic hepatitis C: relation of hepatic iron distribution, HFE genotype, and disease course.

The aim of the present study was to describe the histopathologic features of hepatic iron accumulation in patients with chronic hepatitis C (CH-C) infection, the relation between HFE mutations and hepatic iron location and among iron distribution, HFE, and hepatic damage. We studied 206 patients with CH-C infection. Of 101 patients with hemosiderin deposits, 90.1% had iron deposits in hepatocytes (alone or with sinusoidal and/or portal involvement). The hepatic iron score increased significantly as iron accumulation involved sinusoidal and portal tract compartments and according to HFE genotypes. Severe fibrosis and cirrhosis were associated more markedly with the presence of hemosiderin iron in the 3 hepatic compartments, HFE mutations, and high alcohol intake. We suggest that part of the iron accumulation in CH-C infection derives from increased iron absorption and release from storage cells and that the amount and distribution of hepatic iron deposits is related to hepatic damage. HFE mutations favor both processes, but other factors, genetic or acquired, are involved.

Effects of Hematocrit Changes on Flow-Mediated and Metabolic Vasodilation in Humans

Endothelial function is noninvasively assessed by measuring nitric oxide–dependent increase in radial artery diameter accompanying the elevation in shear stress induced by increasing blood flow through a short-lasting ischemia of the hand. However, shear stress also depends on blood viscosity, whose changes might thus affect nitric oxide increase in a manner that is not properly reflected by blood flow changes. In 12 subjects with hemochromatosis, we measured ultrasonographically radial artery diameter and blood flow responses to a 4-minute ischemia of the hand. This was done also after removing 500 mL of blood (and concomitantly infusing 500 mL of saline), which significantly (P <0.01) reduced hemoglobin concentration and hematocrit. The increase in blood flow induced by the 4-minute ischemia was similar before and after blood removal (+76% and +80%), which, in contrast, markedly attenuated the accompanying increase in radial artery diameter (+25% versus +13%, P <0.01). Thus, in humans, blood viscosity is involved in the endothelial response to an increase in shear stress. This implies that this response may not be accurately assessed and compared by quantifying the stimulus only through an increase in blood flow.

Insulin resistance influences iron metabolism and hepatic steatosis in type II diabetes

Dear Sir: We read with interest the article by Mendler et al.1 who described the association of hepatic iron overload, steatosis, and presence of one or more components of the insulin resistance syndrome. They suggested a role for insulin resistance in the development of hepatic iron overload, steatosis, and increased serum ferritin levels, although the mechanisms whereby insulin resistance would induce alterations in iron metabolism remain to be elucidated. Insulin resistance is very common among subjects with metabolic disorders, but its prevalence varies substantially among clinical conditions.2 Type II diabetes is one of the metabolic conditions with the higher rate of insulin resistance and is frequently associated with increased serum ferritin levels.2,3 We have evaluated iron indices in patients with type II diabetes before and after improvement of metabolic control to provide some insights into the relationship between iron and metabolic disorders. From 1989 to 1990, in our Department of Medicine, 10 outpatients with poorly controlled diabetes were enrolled to evaluate the incidence of hepatic steatosis and its reversibility after correction of the glycemic control. Results of this study have not been published. Chronic alcoholism, acute and chronic liver diseases, malignancies, and inflammatory and iron overload disorders were excluded. After baseline evaluation, patients received a strict diet regimen for 12 months in addition to antidiabetic therapy. The study schedule included monthly measurements of mean blood glucose, fructosamine, glycosylated hemoglobin, cholesterol, and triglyceride levels; assessment of uricemia and liver function tests for 12 months; and hepatic ultrasound examination and liver biopsy at baseline and after 12 months. Small aliquots of the serum samples were collected and stored at 280°. The study was approved by the Hospital’s ethical committee. All patients gave their informed consent to the study. Four denied liver biopsy. We then retrospectively evaluated serum iron, transferrin, and ferritin levels of the patients on each sample and hepatic iron concentration (HIC) at baseline and at the end of study. HIC was determined by atomic absorption spectrophotometry (Perkin-Elmer S2380; Norwalk, CT) in deparaffinized specimens. Table 1 shows the main data of the patients at baseline and during the study. Significant improvement of glycemic control was obtained in each patient. At baseline, serum ferritin levels were significantly higher than those in 20 normal age-matched controls (223 6 139 vs. 122 6 66 μg/L; P , 0.01) and significantly decreased at the end of the study. Serum ferritin correlated with triglyceride at baseline (r 5 0.69; P 5 0.026) and with HIC only at the end of the study (r 5 0.87; P 5 0.024). Liver function test results were normal at baseline and did not change during the study. Hepatic steatosis, as defined by ultrasound examination and histology, was present in 80% of the patients at baseline and in 25% after 12 months; it was mild and often associated with very mild necroinflammatory activity. In 5 patients, HIC decreased after 12 months, but did not change in 1 patient. The results suggest the existence of a relationship between glucose metabolism, fatty liver, serum ferritin, and hepatic iron. Hyperinsulinemia, the main manifestation of insulin resistance, favors the accumulation of free fatty acid in the liver and increases the risk of steatosis.4 High serum ferritin levels are common in patients with metabolic disorders and nonalcoholic steatosis or steatohepatitis, and some of them also have increased HIC.4,5 In our patients, serum ferritin levels at baseline were increased to a degree disproportionate to liver iron stores because they significantly decreased after metabolic improvement when the expected correlation with HIC finally appeared. Also, the mild but significant decrease of HIC at the end of the study suggests a possible influence of diabetic-associated metabolic alterations on hepatocellular iron metabolism. The improvement of the glycemic control observed in the patients can be ascribed to increased insulin action in the liver and in the peripheral tissues. The concomitant improvement of steatosis and the decrease of serum ferritin and hepatic iron levels suggest that these alterations are distinct consequences of a common factor (probably insulin resistance in this case) and that they are at least partially reversible by the improvement of metabolic control. Hypertriglyceridemia is another condition typically associated with the insulin resistance syndrome,2 and the correlation observed in our patients at baseline between serum ferritin and triglyceride further supports the hypothesis of a relationship between insulin resistance and alterations in iron metabolism. Table 1. Iron and Metabolic Indices in 10 Patients With Poorly Controlled Type II Diabetes at Baseline and During Treatment

Iron Accumulation in Chronic Hepatitis C

The aim of the present study was to describe the histopathologic features of hepatic iron accumulation in patients with chronic hepatitis C (CH-C) infection, the relation between HFE mutations and hepatic iron location and among iron distribution, HFE, and hepatic damage. We studied 206 patients with CH-C infection. Of 101 patients with hemosiderin deposits, 90.1% had iron deposits in hepatocytes (alone or with sinusoidal and/or portal involvement). The hepatic iron score increased significantly as iron accumulation involved sinusoidal and portal tract compartments and according to HFE genotypes. Severe fibrosis and cirrhosis were associated more markedly with the presence of hemosiderin iron in the 3 hepatic compartments, HFE mutations, and high alcohol intake. We suggest that part of the iron accumulation in CH-C infection derives from increased iron absorption and release from storage cells and that the amount and distribution of hepatic iron deposits is related to hepatic damage. HFE mutations favor both processes, but other factors, genetic or acquired, are involved.

Type 3 hemochromatosis and β‐thalassemia trait

Type 3 hemochromatosis is a rare autosomal recessive disorder due to mutations of the TFR2 gene. We describe clinical, biochemical and histopathologic findings of a patient with type 3 hemochromatosis at presentation and during a follow‐up of more than 20 yr and we evaluate the effect of an associated β‐thalassemia trait on phenotypic expression. At the age of 33 yr the patient showed a marked iron overload and severe iron‐related complications. After removal of 26 g of iron by subcutaneous deferoxamine infusion a marked clinical improvement was observed. Liver biopsies, performed at the age of 34 and 49 yr, indicate that in type 3 hemochromatosis there is a progressive hepatocellular iron accumulation from Rappaports zone 1–3 and that iron loading in sinusoidal and portal macrophages occurs only in the more advanced stage. As observed in HFE hemochromatosis, the β‐thalassemia trait seems to aggravate the clinical picture of patients lacking TFR2, favoring higher rates of iron accumulation probably by activation of the erythroid iron regulator.