C282Y hemochromatosis: a reality of potential pathophysiological and clinical importance

HEPATOLOGY, Vol. 38, No. 4, Suppl. 1, 2003

AASLD ABSTRACTS

1050 H E P C I D I N EXPRESSION IS DOWN-REGULATED IN

ALCOHOL-FED RATS: A POSSIBLE PATHOGENIC FACTOR IN ALCOHOL RELATED HEPATIC SIDEROSIS. Ting Kin

Cheung, Linda Fletcher, Princess Alexandra Hospital, Woolloongabba, Australia; Kim Bridle, University of Queensland, Woolloongabba, Australia; Therese Murphy, Darrell Crawford, Pm'ncessAlexandra Hospital, Woolloongabba, Australia Introduction: Both alcohol and excess iron in the liver can induce tissue damage and lead to fibrosis and ultimately cirrhosis. Hepatic siderosis is often seen in patients with alcoholic liver disease. The pathogenic mechanisms underlying this p h e n o m e n o n have not been well characterized. Hepcidin, a recently discovered liverderived circulating peptide, has been implicated in the homeostasis of iron metabolism and in the regulation of iron absorption. Hepcidin is believed to be a negative regulator of intestinal iron absorption. Divalent metal transporter I (DMT1) and iron-regulated protein (IREG1) are the major iron transporters in duodenal enterocytes, with DMT1 responsible for apical uptake and IREG1 for basolateral export of iron. The effect of alcohol on the expression of hepcidin, DMT1 and IREG1 is not known. Aim: To study the hepatic gene expression of hepcidin, DMT1 and IREG1 in a rat model of alcoholic liver disease. Methods: Male Sprague Dawley rats (n-10) were pair-fed an alcoholic (Lieber-deCarli) liquid diet or a control liquid diet for 12 weeks. Blood ethanol levels and serum AST were measured. Total liver RNA was extracted using Trizol(tm). cDNA was constructed using Superscript II(tm) following the manufacturer's instructions. Quantitative (real-time) PCR was performed using SYBR green and specific primers for the hepcidin gene, DMT1 and IREG1 with GAPDH as the control gene. Hepatic steatosis, inflammation and fibrosis were assessed histologically and hepatic iron concentration was measured colorimetrically. Results: Blood ethanol levels in alcohol-fed rats were significantly higher than controls (229.2_+32.1 vs. 4.5_+0.4 mg/dL; p<0.002). Serum AST concentration was mildly increased in the alcohol-fed group but did not achieve statistical significance (101_+10.1 vs. 87.8_+3.3 U/ml). Histological examination revealed extensive steatosis in the alcohol-fed rats with minimal inflammation and fibrosis. Serum ferritin (656.6_+41.2 vs. 973.2_+186.2/~g/L) and hepatic iron concentrations (6.2_+0.8 vs. 6.8_+0.7 ~mol/g dry weight) were not significantly different. Hepcidin mRNA expression was significantly decreased (7.4 fold reduction, p-0.009) in alcohol-fed rats compared to their pair-fed controls, however, hepatic DMT1 and IREG1 remained unchanged. Conclusion: Hepcidin expression is significantly decreased in the alcohol-fed rats. Altered hepcidin expression may be an important contributing factor to hepatic siderosis often seen in h u m a n alcoholic liver disease. Disclosures: Kim Bridle - No relationships to disclose Ting Kin Cheung - No relationships to disclose Darrell Crawford - No relationships to disclose Linda Fletcher - No relationships to disclose Therese Murphy - No relationships to disclose

661A

1051

LABILE PLASMA IRON IN C282Y/C282Y H E M O C H R O M A T O S I S : A REALITY OF POTENTIAL PATHOPHYSIOLOGICAL AND CLINICAL IMPORTANCE.

Caroline Le Lan, University Hospital Pontchaillou and INS~ RM U522, Rennes, France; Tally Cohen, Institute of Life Sciences, Jerusalem, Israel; Martine Ropert, University Hospital Pontchaillou, Rennes, France; Hava Glickstein, Institute of Life Sciences, Jerusalem, Israel; Michel Pouchard, Centre d~ xamens de Santd CPAM, Rennes, France; Andre Le Treut, Laboratoire de Biochime Gdn&ale et nzymologie, University Hospital Pontchaillou, Rennes, France; Yves Deugnier, Olivier Loreal, University Hospital Pontchaillou and INS~ RM U-522, Rennes, France; William Breuer, Ioav Cabantchik, Institute of Life Sciences, JerusMem, Israel; Pierre Brissot, University Hospital Pontchaillou and INS~ RM U-522, Rennes, France Labile Plasma Iron (LPI) is the redox active component of NonTransferrin-Bound Iron (NTBI) and therefore the potential circulating iron species responsible for tissue damage in iron overload conditions. Its presence in thalassemic patients has been recently reported (Esposito et al, Blood, 2003). The aim of the present study was to evaluate the presence and quantity of LPI in C282Y/C282Y Genetic Hemochromatosis (GH) as compared to Dysmetabolic Hepatosiderosis (DHS) (a condition with mild iron excess but high serum ferritin levels), alcoholic cirrhosis (in which low serum transferrinemia may generate by itself the appearance of NTBI), and normal controls. We studied 159 male subjects subdivided into: i) 23 iron overloaded Genetic Hemochromatosis (GH) ; ii) 14 iron depleted GH ; iii) 26 DHS ; iv) 33 Alcoholic Cirrhosis ; v) 63 strictly defined Normal Controls. NTBI was measured by a fluorescence-based one step assay (Breuer and Cabantchik, Anal Biochem 2001;299:194). LPI was determined with the fluorogenic dihydrorhodamine 123 by monitoring the generation of reactive radicals p r o m p t e d by ascorbate but blocked by iron chelators. The results indicated: i) in iron overloaded GH: a significant increase in both NTBI and LPI (0.761_+0.504 ~M and 0.250_+0.289 ~M, respectively) versus iron depleted GH (0.221_+0.559 and 0.019_+0.127) and normal controls (0.154_+0.328 and 0). Ii) in DHS: a significant increase in NTBI (0.381_+0.381) versus normal controls whereas no LPI elevation was observed, iii) in alcoholic cirrhosis : increased NTBI (0.385_+0.560) -which however did not reach significance- without LPI elevation (0.095_+0.401). These data demonstrate : i) the presence of LPI in iron overloaded hemochromatosis ; ii) the molecular heterogeneity of circulating NTBI since no LPI component could be found in the other tested conditions despite increased circulating NTBI. Taking into account the potential pathogenic importance of LPI, its determination may represent a new important diagnostic tool in the field of iron overload and liver diseases. (Supported by: European Grant QLRT-2001-00444 ; Israeli Ministry of Industry and Resources ; Aferrix Company). Disclosures: William Breuer - Aferrix Company: Consultant/Advisor Pierre Brissot - No relationships to disclose Ioav Cabantchik - Aferrix Company: Consultant/Advisor Tally Cohen - No relationships to disclose Yves Deugnier - No relationships to disclose Hava Glickstein - No relationships to disclose Caroline Le Lan - No relationships to disclose Andre Le Treut - No relationships to disclose Olivier Loreal - No relationships to disclose Michel Pouchard - No relationships to disclose Martine Ropert - No relationships to disclose