Alberto Piperno

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.

Natural history of juvenile haemochromatosis

Summary. Juvenile haemochromatosis or haemochromatosis type 2 is a rare autosomal recessive disorder which causes iron overload at a young age, affects both sexes equally and is characterized by a prevalence of hypogonadism and cardiopathy. Patients with haemochromatosis type 2 have been reported in different ethnic groups. Linkage to chromosome 1q has been established recently, but the gene remains unknown. We report the analysis of the phenotype of 29 patients from 20 families of different ethnic origin with a juvenile 1q‐associated disease. We also compared the clinical expression of 26 juvenile haemochromatosis patients with that of 93 C282Y homozygous males and of 11 subjects with haemochromatosis type 3. Patients with haemochromatosis type 2 were statistically younger at presentation and had a more severe iron burden than C282Y homozygotes and haemochromatosis type 3 patients. They were more frequently affected by cardiopathy, hypogonadism and reduced glucose tolerance. In contrast cirrhosis was not statistically different among the three groups. These data suggest that the rapid iron accumulation in haemochromatosis type 2 causes preferential tissue damage. Our results clarify the natural history of the disease and are compatible with the hypothesis that the HFE2 gene has greater influence on iron absorption than other haemochromatosis‐associated genes.

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.

Early impairment of large artery structure and function in Type I diabetes mellitus

Aims/hypothesis. Diabetes mellitus is associated with an increased incidence of atherosclerosis. How early functional and structural alterations of large arteries that may preceed atherosclerosis occur in the course of this disease has, however, never been conclusively documented. Methods. We evaluated arterial wall distensibility in the radial artery, common carotid artery and abdominal aorta in 133 patients (aged 35.4 ± 0.9 years, means ± SEM) with Type I (insulin-dependent) diabetes mellitus and no macrovascular complications. Arterial distensibility was derived from continuous measurements of arterial diameter through echotracking techniques and use of either the Langewouters (radial artery) or the Reneman (carotid artery and aorta) formula. The same echotracking techniques enabled us to obtain radial artery and carotid artery wall thickness. Data were compared with those from 70 age-matched normotensive control subjects. Results. In diabetic patients arterial distensibility was consistently less (p < 0.01) than in control subjects, the reduction averaging 26 %, 14 % and 25 % for the radial artery, carotid artery and aorta, respectively. This was accompanied by an increase (p < 0.01) in both radial and carotid artery wall thickness. The changes were more pronounced in patients with microalbuminuria, retinopathy or neuropathy or both. They were evident also in those without microvascular complications. This was the case also when subjects in whom diabetes was associated with hypertension (n = 30) were excluded from data analysis. Carotid and aortic wall abnormalities showed a relation with the duration of disease and blood pressure whereas radial artery abnormalities showed a relation with glycated haemoglobin. Conclusion/interpretation. Type I diabetes is characterised by diffuse arterial wall stiffening and thickening which progress with the severity of the disease but can clearly be seen also in the absence of any diabetic-related complication. This suggests that in diabetes stiffening and thickening are an early marker of vascular damage. [Diabetologia (1999) 42: 987–994]

Time course of circulatory and humoral effects of rapid total paracentesis in cirrhotic patients with tense, refractory ascites

BACKGROUND/AIMS Tense ascites of cirrhosis can be treated with total paracentesis; however, the short-term effects of this procedure are poorly defined. METHODS The circulatory and humoral changes induced by total paracentesis (250 mL/min) were studied in 12 cirrhotics with tense, refractory ascites. Data were collected before, during, and after paracentesis and 24 hours later (after albumin infusion). Hormonal parameters were recorded again 48 hours and 6 days thereafter. RESULTS Paracentesis (10.7 +/- 4.4 L; 64 +/- 20 minutes) caused marked reduction of intra-abdominal, intrathoracic, right atrial, and pulmonary pressures. Heart rate did not change. Cardiac output and heart volumes increased. Systemic vascular resistances and mean arterial pressure slightly decreased. Baseline plasma renin and aldosterone levels were markedly increased; a reduction was already evident during paracentesis with the lowest values at the end of the procedure. All changes were maintained 24 hours later. Hormones regained baseline levels 6 days later. CONCLUSIONS Rapid total paracentesis is accompanied by marked cardiovascular and humoral changes. Some of these changes can be explained by mechanical factors that are directly or indirectly related to the relief of abdominal pressure. However, other changes (systemic vasodilatation, humoral deactivation) have a non-mechanical nature and may depend on reflexes originating from cardiac volume receptor stimulation. Most changes may beneficially (albeit transiently) influence the cardiovascular system of cirrhotic patients with tense ascites.

Modulation of hepcidin production during hypoxia-induced erythropoiesis in humans in vivo: data from the HIGHCARE project

Iron is tightly connected to oxygen homeostasis and erythropoiesis. Our aim was to better understand how hypoxia regulates iron acquisition for erythropoiesis in humans, a topic relevant to common hypoxia-related disorders. Forty-seven healthy volunteers participated in the HIGHCARE project. Blood samples were collected at sea level and after acute and chronic exposure to high altitude (3400-5400 m above sea level). We investigated the modifications in hematocrit, serum iron indices, erythropoietin, markers of erythropoietic activity, interleukin-6, and serum hepcidin. Hepcidin decreased within 40 hours after acute hypoxia exposure (P < .05) at 3400 m, reaching the lowest level at 5400 m (80% reduction). Erythropoietin significantly increased (P < .001) within 16 hours after hypoxia exposure followed by a marked erythropoietic response supported by the increased iron supply. Growth differentiation factor-15 progressively increased during the study period. Serum ferritin showed a very rapid decrease, suggesting the existence of hypoxia-dependent mechanism(s) regulating storage iron mobilization. The strong correlation between serum ferritin and hepcidin at each point during the study indicates that iron itself or the kinetics of iron use in response to hypoxia may signal hepcidin down-regulation. The combined and significant changes in other variables probably contribute to the suppression of hepcidin in this setting.

Breakthrough during recombinant interferon alfa therapy in patients with chronic hepatitis C virus infection : prevalence, etiology, and management

Recombinant interferon alfa (r-IFN alpha 2) has been shown to normalize the aminotransferase levels in approximately 50% of patients with chronic hepatitis C virus (HCV). Few patients experience a relapse during the treatment, in spite of a complete initial response (breakthrough). We studied 191 HCV Ab-positive patients with histologically proven chronic hepatitis. All of them were treated with r-IFN alpha 2 (3 MU three times a week). A complete response was seen in 54.4%. However, 12 of 104 responders experienced a breakthrough. At the time of breakthrough, neutralizing IFN antibodies were positive in 6 of 12 patients. Binding IFN antibodies were positive in all of these 12 patients. Continued treatment with r-IFN alpha 2, even at higher doses, did not restore the previous response in any patient. All of them were then switched to natural lymphoblastoid IFN, and this rapidly restored a complete response in all of the patients.

HCV genotypes in Northern Italy: a survey of 1368 histologically proven chronic hepatitis C patients

BACKGROUND/AIMS Hepatitis C virus (HCV) easily undergoes genomic changes, thus accounting for the presence of different genotypes, with different geographic distributions and different outcomes of chronic hepatitis. Type 1b is frequently found in advanced diseases; however, since this genotype is the most prevalent in older patients, the association with advanced age and severity of the disease is confounding. The aim of this study was to assess changes in the prevalence of HCV genotypes by surveying a large population of chronic hepatitis C patients in Northern Italy, and to assess if the high prevalence of genotype 1b in older patients with advanced diseases simply reflects the duration of HCV infection, rather than intrinsic biological properties of HCV. METHODS We studied 1368 HCV-RNA positive patients, with histologically proven chronic hepatitis. Drug addiction, blood transfusions and sporadically acquired infections represented the risk factors. RESULTS Genotype 1b, the most prevalent isolate, and genotype 2a were associated with older age, cirrhosis, sporadically-acquired infections and blood transfusion, while types 1a, 3a, and 4 were associated with younger age, chronic persistent hepatitis and drug addiction. Patients with a history of transfusions were divided into four groups depending on the period of transfusion. The prevalence of genotype 1b decreased with time. Type 3a appeared only after 1979. CONCLUSION The severity of chronic hepatitis C could be related more to the duration of the infection rather than to the intrinsic pathogenicity of HCV genotypes.

Hepcidin and iron-related gene expression in subjects with dysmetabolic hepatic iron overload

BACKGROUND/AIMS Many patients with hepatic iron overload do not have identifiable mutations and often present with metabolic disorders and hepatic steatosis. Since the pathophysiology of Dysmetabolic Hepatic Iron Overload (DHIO) is still obscure, the aim of this study was to evaluate, in these patients, possible alterations in iron-related molecule expression. METHODS Iron-related gene mRNA levels were determined by quantitative-PCR in liver biopsies of subjects with NAFLD without iron overload and patients with HFE-hemochromatosis, beta-thalassemia major and DHIO. Urinary hepcidin was measured by immunoblotting. RESULTS No alterations in mRNA expression of either iron transporters or exporters were found in DHIO. mRNA and urinary hepcidin levels normalized for the amount of iron overload showed a significantly lower ratio than in controls, although not as low as in hemochromatosis or beta-thalassemia. Differently from what observed in hemochromatosis, hepcidin mRNA did not correlate with urinary hepcidin. CONCLUSIONS Patients with DHIO show appropriate regulation of mRNAs encoding proteins involved in iron uptake and efflux but dysregulation of hepcidin production. The relatively elevated urinary hepcidin can explain the iron phenotype in DHIO (more macrophage iron retention and low/normal transferrin saturation).

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.