- Email: [email protected]
A in North American Vietnamese with and without phenotypic evidence of iron overload
Hemochromatosis (HFE) gene splice site mutation IVS5⫹1 G/A in North American Vietnamese with and without phenotypic evidence of iron overload MICHAEL STEINER, CATHERINE LEIENDECKER-FOSTER, GORDON D. MCLAREN, BEVERLY M. SNIVELY, CHRISTINE E. MCLAREN, PAUL C. ADAMS, and JOHN H. ECKFELDT ROSTOCK, GERMANY; MINNEAPOLIS, MINN; LONG BEACH AND IRVINE, CALIF; WINSTON-SALEM, NC; LONDON, ONTARIO, CANADA; AND MINNEAPOLIS, MINN
Homozygosity for a novel hemochromatosis (HFE) gene splice site mutation (IVS5ⴙ1 G/A) was previously reported in a 48-year-old Vietnamese man residing in Germany who had an elevated serum ferritin (SF) and transferrin saturation (TS) and severe iron overload on liver biopsy. This mutation was not found in 222 controls of central European origin but has been found in Southeast Asians living in Vietnam without evidence of iron overload. Hemochromatosis and iron overload screening (HEIRS) Study is an ongoing, multiethnic, primary care-based study of 101,168 North American adults, including 12,772 Asians, a group that the HEIRS Study found has a significantly higher than expected prevalence of elevated serum TS and SF but very low prevalence of the common C282Y and H63D HFE alleles usually associated with hereditary hemochromatosis. It was hypothesized that the IVS5ⴙ1 G/A splice site mutation might explain some elevated biochemical iron measures in North American Asians. Overall, 200 Vietnamese subjects from the Los Angeles Field Center who had TS and SF values greater than the 75th percentile of all HEIRS Study participants after adjusting for covariates and 149 controls randomly selected to represent this Vietnamese population were genotyped. Among cases, 1 homozygous mutant and 7 heterozygotes were found; among controls, 1 homozygous mutant and 4 heterozygotes were found yielding an allele frequency of 2.32% for cases and 2.04% for controls (P > 0.5). This finding suggests that the HFE IVS5ⴙ1 G/A splice site mutation is not the major explanation for unexpectedly high prevalence of TS and SF in North American Asians. (Translational Research 2007;149:92–95) Abbreviations: HEIRS ⫽ hemochromatosis and iron overload screening; HFE ⫽ hemochromatosis; PCR ⫽ polymerase chain reaction; SD ⫽ standard deviation; SF ⫽ serum ferritin; TS ⫽ transferrin saturation
From the Institute of Clinical Chemistry and Laboratory Medicine, University of Rostock, Rostock, Germany; the Department of Laboratory Medicine and Pathology, University of Minnesota, Minneapolis, Minn USA; Veterans Affairs Long Beach Healthcare System, Long Beach, Calif and Division of Hematology/Oncology, Department of Medicine, University of California, Irvine, Calif USA; the Department of Public Health Sciences, Wake Forest University School of Medicine, Winston-Salem, NC USA; Epidemiology Division, Department of Medicine, University of California, Irvine, Calif USA; the Department of Medicine, London Health Sciences Center, London, Ontario, Canada; and the Department of Laboratory Medicine and Pathology, University of Minnesota, Minneapolis, Minn USA.
92
Submitted for publication July 21, 2006; revision submitted August 28, 2006; accepted for publication August 30, 2006. Reprint requests: Catherine Leiendecker-Foster, MS, CLSp(MB), University of Minnesota, Department of Laboratory Medicine and Pathology, 420 Delaware St. S.E., Minneapolis, Minn 55455. e-mail: [email protected]. 1931-5244/$ – see front matter © 2007 Mosby, Inc. All rights reserved. doi:10.1016/j.trsl.2006.08.007
Translational Research Volume 149, Number 2
Hereditary hemochromatosis is an autosomal recessive disorder that can lead to iron accumulation in the body. The hemochromatosis (HFE) gene, which is located on the short arm of chromosome 6, has 2 mutations, C282Y and H63D, that are found in homozygous or compound heterozygous states in almost all Caucasians of northern European ancestry with clinical iron overload.1– 4 The Hemochromatosis and Iron Overload Screening (HEIRS) Study5 is an ongoing, multiethnic, primary care-based study of 101,168 North American adults enrolled at 5 Field Centers in the United States and Canada, including 12,772 Asians. In the Asian subgroup, a significantly higher than expected prevalence of elevated serum transferrin saturation (TS) and serum ferritin (SF) was found, but a very low frequency of the C282Y and H63D HFE mutations usually associated with hereditary hemochromatosis was found.6 It was hypothesized that other HFE mutations might explain these elevated TS and SF findings in this ethnic subgroup. Homozygosity for a novel HFE splice site mutation (IVS5⫹1 G/A) had been reported in a 48-year-old Vietnamese patient residing in Germany who had a markedly elevated SF of 2478 g/L, TS of 96%, and severe iron overload documented by liver biopsy.7 This mutation was not found in 222 controls of central European origin7 but subsequently was identified by Pointon et al in Vietnamese patients,8 although they were heterozygous and did not have any evidence of iron overload. Pointon et al8 estimated an allele frequency of 0.003 for HFE IVS5⫹1 G/A in the Vietnamese population. As the IVS5⫹1 G/A splice site mutation predicts 2 potential messenger RNA products, both of which result in the deletion of the transmembrane domain of the resulting HFE protein potentially influencing the regulatory interaction of it with the transferrin receptor,7 it was hypothesized that this mutation might provide a partial explanation for the unexpectedly frequent elevations in biochemical iron measures that were found in North American Vietnamese enrolled in the HEIRS Study. METHODS Study population. Cases and controls were selected from participants at the Southern California Field Center who stated that their preferred language is Vietnamese. A simple random sample (N ⫽ 149) was selected as a control group, which allowed the straightforward estimation of prevalence of the IVS5⫹1 G/A splice site mutation in this population. Cases (N ⫽ 200) were defined as those in the population who ranked in the top 25% of all HEIRS participants for both TS and SF, after adjusting for covariates (HFE C282Y and H63D genotype, gender, age, Field Center, and race/ethnicity) in multiple linear regression models (1 each for TS and SF). Overlap was unlikely between cases and the control samples,
Steiner et al
93
Fig 1. Electropherogram of HFE gene splice site mutation IVS 5⫹1 PCR products. Lane 1: size ladder; Lane 2: sample 1 wild-type allele; Lane 3: sample 1 mutant allele; Lane 4: sample 2 wild-type allele; Lane 5: sample 2 mutant allele; Lane 6: Heterozygous control wildtype allele; Lane 7: Heterozygous control mutant allele; Lane 8: Negative control wild-type allele; Lane 9: Negative control mutant allele.
and none occurred. In compliance with the principles of the Declaration of Helsinki, this study was approved by the ethics committees of all participating institutions and all participants signed an informed consent. DNA isolation and genotyping. DNA was isolated from ethylene diamine tetraacetic acid blood using the Puregene kit (Gentra Systems, Minneapolis, Minn). Genotyping was performed using an allele-specific polymerase chain reaction (PCR) reaction in which the reaction contains a forward primer and either a wild-type-specific or a mutant-specific reverse primer as described previously.7 Figure 1 demonstrates an agarose gel electrophoresis of the PCR products obtained from allele-specific amplifications. RESULTS
Of the 200 cases, 194 were successfully genotyped for the IVS5⫹1 G/A splice site mutation. Among cases, 1 homozygote for this mutation and 7 heterozygotes were found (mean SF of 484 ⫾ 198 g/L [1 standard deviation (SD)]; mean TS of 60.6 ⫾ 11.2% [1 SD]). The homozygote, a man in his late 50s with hepatitis C, had markedly elevated serum iron test results (SF 1870 g/L, TS 99% at the initial screening visit). Seven months later, at the time of further clinical evaluation, he had an SF of 2777 g/L and TS of 99%. He subsequently underwent venesection therapy to remove excess iron and had a total of 4.95-L blood removed during the course of 11 phlebotomy sessions (450 mL each), corresponding to about 2.6-g storage iron, well below the amount deemed clinically significant by most experts. After phlebotomy, the SF fell to 20 g/L. It is certainly possible that hepatitis C is a contributing factor to the elevated iron tests in this participant. Of the 149 controls, 147 were successfully genotyped yielding 1 homozygote for the splice site mutation (a woman in her mid-50s with SF 59 g/L, TS 32% who was never referred for liver biopsy or phlebotomy ther-
94
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Table I. Gender, phenotype, and splice site genotype of cases and controls Splice site genotype
Case/control
Gender
TS, % ( ⴞSD)
SF, ug/L ( ⴞSD)
N
Homozygous Heterozygous Wild-type Heterozygous Wild-type Heterozygous Wild-type Homozygous Heterozygous Wild-type
Case Case Case Control Control Case Case Control Control Control
Male Male Male Male Male Female Female Female Female Female
99 65.8 (11.9) 65.7 (13.8) 28.7 (6.8) 35.0 (9.0) 53.7 (6.4) 53.7 (10.1) 32 31 28.6 (9.9)
1870 594 (172) 936 (971) 295 (40) 348 (195) 338 (131) 464 (415) 59 27 153 (112)
1 4 92 3 50 3 94 1 1 92
apy) and 4 heterozygotes (mean SF of 228 ⫾ 138 g/L [1 SD]; mean TS of 29.0 ⫾ 5.7% [1 SD]). The normal SF and TS found in the 1 homozygous female control participant could potentially be explained by multiple pregnancies; however, a medical history was not obtained from this participant. Details of the gender, TS, and SF for the cases and controls are shown in Table I. These data indicate comparable allele frequencies of 2.32% and 2.04% for the IVS5⫹1 G/A splice site mutation among cases and controls, respectively (chisquare test, P ⬎ 0.5). DISCUSSION
The HFE IVS5⫹1 G/A splice site mutation seems to be found in Vietnamese populations at a fairly low frequency. In a pilot study of HEIRS Asian participants having marked elevations of both SF and TS, which included 58 participants from the greater Los Angeles area who were mostly Vietnamese and 47 from the Toronto area who were mostly Chinese, this particular splice site mutation was found only in the Vietnamese participants (data not shown). In this report, data are presented for only the second and third homozygotes with this splice site mutation appearing in the scientific literature. The original HFE IVS5⫹1 G/A splice site mutation homozygote was a Vietnamese man living in Germany with marked iron overload. In this report, a second homozygote was described, a male in his late 50s who was also positive for hepatitis C virus with borderline iron overload, and a third homozygote, a woman in her mid-50s with no evidence of iron overload. Interestingly, the allele frequency in cases and controls is about 10-fold higher than that found by Pointon et al.8 However, in their study, 95% of the Vietnamese participants were of the Kinh ethnic subgroup. Although 5% of their participants were from the Tay, S’Tieng, Nung, and Muong ethnic subgroups combined, the number of participants in
each of these subgroups was much too small to make any reliable assessment of mutation allele frequency. Data regarding the geographic and ethnic origin of the HEIRS Study United States Vietnamese participants was not collected, but the 10-fold difference in allele frequencies found between this study and Pointon’s study strongly suggests that this mutation does vary substantially across ethnic subgroups of Vietnamese. As the allele frequencies of the cases and controls are similar in the subjects, the authors conclude that the unexpectedly high prevalence of elevated serum TS and SF in HEIRS Vietnamese participants cannot be explained on the basis of the HFE IVS5⫹1 G/A splice site mutation. The absence of marked iron overload in the 2 homozygous HEIRS participants identified in this study is in contrast to the earlier report of severe iron overload in a Vietnamese homozygote living in Germany, whom is estimated had roughly 6 g of storage iron (25 phlebotomies ⫻ 0.5-L blood/phlebotomy ⫻ ⬃0.5-g iron/L blood with a hemoglobin of approximately 150 g/L).7 This variable expression is analogous to the very variable expression that has been observed in homozygotes for the HFE C282Y mutation, which is most common in Caucasians. It has been proposed that environmental influences or interactions involving modifier genes may contribute to the phenotypic and clinical variability in C282Y homozygotes.1,3,8 –10 It is very likely that similar factors may play a role in determining the phenotype and clinical manifestation in those homozygotes for the IVS5⫹1 G/A splice site mutation as well. As a result of poor understanding of the long-term expression of any HFE mutations, the value of broad population screening for hemochromatosis in clinical practice remains controversial.11 The authors wish to thank their colleagues from the HEIRS Study.
Translational Research Volume 149, Number 2
Steiner et al
REFERENCES
1. Ajioka RS, Kushner JP. Hereditary hemochromatosis. Semin Hematol 2002;39:235– 41. 2. Andersen RV, Tybjaerg-Hansen A, Appleyard M, Birgens H, Nordestgaard BG. Hemochromatosis mutations in the general population: iron overload progression rate. Blood 2004;103: 2914 –9. 3. Beutler E, Felitti VJ, Koziol JA, Ho NJ, Gelbart T. Penetrance of 845G¡A (C282Y) HFE hereditary haemochromatosis mutation in the USA. Lancet 2002;359:211–18. 4. Le Gac G, Ferec C. The molecular genetics of haemochromatosis. Eur J Hum Genet 2005;13:1172– 85. 5. McLaren C, Barton J, Adams P, Harris E, Acton R, Press N, et al. Hemochromatosis and Iron Overload Screening (HEIRS) study design for an evaluation of 100,000 primary care-based adults. Am J Med Sci 2003;325:53– 62. 6. Adams PC, Reboussin DR, Barton JC, McLaren CE, Eckfeldt JH, McLaren GD, et al. Screening of a primary care population
7.
8.
9. 10.
11.
95
of 101,168 persons of diverse ethnic background. N Engl J Med 2005;352:1769 –78. Steiner M, Ochran K, Genschel J, Meier P, Gerl H, Ventz M, et al. A homozygous HFE gene splice site mutation IVS5⫹1 G/A in a hereditary hemochromatosis patient of Vietnamese origin. Gastroenterology 2002;122:789 –95. Pointon JJ, Viprakasit V, Miles KL, Livesey KJ, Steiner M, O’Riordan S, et al. Hemochromatosis gene (HFE) mutations in South East Asia: a potential for iron overload. Blood Cells, Molecules & Diseases 2003;30:302– 6. Ajioka RS, Kushner JP. Clinical consequences of iron overload in hemochromatosis homozygotes. Blood 2003;101:3351– 4. Beutler E. The HFE Cys282Tyr mutation as a necessary but not sufficient cause of clinical hereditary hemochromatosis. Blood 2003;101:3347–50. Whitlock EP, Garlitz BA, Harris EM, Beil TL, Smith PR. Screening for hereditary hemochromatosis: a systematic review for the U.S. Preventive Services Task Force. Ann Intern Med 2006;145:209 –23.
Homozygosity for a novel hemochromatosis (HFE) gene splice site mutation (IVS5ⴙ1 G/A) was previously reported in a 48-year-old Vietnamese man residing in Germany who had an elevated serum ferritin (SF) and transferrin saturation (TS) and severe iron overload on liver biopsy. This mutation was not found in 222 controls of central European origin but has been found in Southeast Asians living in Vietnam without evidence of iron overload. Hemochromatosis and iron overload screening (HEIRS) Study is an ongoing, multiethnic, primary care-based study of 101,168 North American adults, including 12,772 Asians, a group that the HEIRS Study found has a significantly higher than expected prevalence of elevated serum TS and SF but very low prevalence of the common C282Y and H63D HFE alleles usually associated with hereditary hemochromatosis. It was hypothesized that the IVS5ⴙ1 G/A splice site mutation might explain some elevated biochemical iron measures in North American Asians. Overall, 200 Vietnamese subjects from the Los Angeles Field Center who had TS and SF values greater than the 75th percentile of all HEIRS Study participants after adjusting for covariates and 149 controls randomly selected to represent this Vietnamese population were genotyped. Among cases, 1 homozygous mutant and 7 heterozygotes were found; among controls, 1 homozygous mutant and 4 heterozygotes were found yielding an allele frequency of 2.32% for cases and 2.04% for controls (P > 0.5). This finding suggests that the HFE IVS5ⴙ1 G/A splice site mutation is not the major explanation for unexpectedly high prevalence of TS and SF in North American Asians. (Translational Research 2007;149:92–95) Abbreviations: HEIRS ⫽ hemochromatosis and iron overload screening; HFE ⫽ hemochromatosis; PCR ⫽ polymerase chain reaction; SD ⫽ standard deviation; SF ⫽ serum ferritin; TS ⫽ transferrin saturation
From the Institute of Clinical Chemistry and Laboratory Medicine, University of Rostock, Rostock, Germany; the Department of Laboratory Medicine and Pathology, University of Minnesota, Minneapolis, Minn USA; Veterans Affairs Long Beach Healthcare System, Long Beach, Calif and Division of Hematology/Oncology, Department of Medicine, University of California, Irvine, Calif USA; the Department of Public Health Sciences, Wake Forest University School of Medicine, Winston-Salem, NC USA; Epidemiology Division, Department of Medicine, University of California, Irvine, Calif USA; the Department of Medicine, London Health Sciences Center, London, Ontario, Canada; and the Department of Laboratory Medicine and Pathology, University of Minnesota, Minneapolis, Minn USA.
92
Submitted for publication July 21, 2006; revision submitted August 28, 2006; accepted for publication August 30, 2006. Reprint requests: Catherine Leiendecker-Foster, MS, CLSp(MB), University of Minnesota, Department of Laboratory Medicine and Pathology, 420 Delaware St. S.E., Minneapolis, Minn 55455. e-mail: [email protected]. 1931-5244/$ – see front matter © 2007 Mosby, Inc. All rights reserved. doi:10.1016/j.trsl.2006.08.007
Translational Research Volume 149, Number 2
Hereditary hemochromatosis is an autosomal recessive disorder that can lead to iron accumulation in the body. The hemochromatosis (HFE) gene, which is located on the short arm of chromosome 6, has 2 mutations, C282Y and H63D, that are found in homozygous or compound heterozygous states in almost all Caucasians of northern European ancestry with clinical iron overload.1– 4 The Hemochromatosis and Iron Overload Screening (HEIRS) Study5 is an ongoing, multiethnic, primary care-based study of 101,168 North American adults enrolled at 5 Field Centers in the United States and Canada, including 12,772 Asians. In the Asian subgroup, a significantly higher than expected prevalence of elevated serum transferrin saturation (TS) and serum ferritin (SF) was found, but a very low frequency of the C282Y and H63D HFE mutations usually associated with hereditary hemochromatosis was found.6 It was hypothesized that other HFE mutations might explain these elevated TS and SF findings in this ethnic subgroup. Homozygosity for a novel HFE splice site mutation (IVS5⫹1 G/A) had been reported in a 48-year-old Vietnamese patient residing in Germany who had a markedly elevated SF of 2478 g/L, TS of 96%, and severe iron overload documented by liver biopsy.7 This mutation was not found in 222 controls of central European origin7 but subsequently was identified by Pointon et al in Vietnamese patients,8 although they were heterozygous and did not have any evidence of iron overload. Pointon et al8 estimated an allele frequency of 0.003 for HFE IVS5⫹1 G/A in the Vietnamese population. As the IVS5⫹1 G/A splice site mutation predicts 2 potential messenger RNA products, both of which result in the deletion of the transmembrane domain of the resulting HFE protein potentially influencing the regulatory interaction of it with the transferrin receptor,7 it was hypothesized that this mutation might provide a partial explanation for the unexpectedly frequent elevations in biochemical iron measures that were found in North American Vietnamese enrolled in the HEIRS Study. METHODS Study population. Cases and controls were selected from participants at the Southern California Field Center who stated that their preferred language is Vietnamese. A simple random sample (N ⫽ 149) was selected as a control group, which allowed the straightforward estimation of prevalence of the IVS5⫹1 G/A splice site mutation in this population. Cases (N ⫽ 200) were defined as those in the population who ranked in the top 25% of all HEIRS participants for both TS and SF, after adjusting for covariates (HFE C282Y and H63D genotype, gender, age, Field Center, and race/ethnicity) in multiple linear regression models (1 each for TS and SF). Overlap was unlikely between cases and the control samples,
Steiner et al
93
Fig 1. Electropherogram of HFE gene splice site mutation IVS 5⫹1 PCR products. Lane 1: size ladder; Lane 2: sample 1 wild-type allele; Lane 3: sample 1 mutant allele; Lane 4: sample 2 wild-type allele; Lane 5: sample 2 mutant allele; Lane 6: Heterozygous control wildtype allele; Lane 7: Heterozygous control mutant allele; Lane 8: Negative control wild-type allele; Lane 9: Negative control mutant allele.
and none occurred. In compliance with the principles of the Declaration of Helsinki, this study was approved by the ethics committees of all participating institutions and all participants signed an informed consent. DNA isolation and genotyping. DNA was isolated from ethylene diamine tetraacetic acid blood using the Puregene kit (Gentra Systems, Minneapolis, Minn). Genotyping was performed using an allele-specific polymerase chain reaction (PCR) reaction in which the reaction contains a forward primer and either a wild-type-specific or a mutant-specific reverse primer as described previously.7 Figure 1 demonstrates an agarose gel electrophoresis of the PCR products obtained from allele-specific amplifications. RESULTS
Of the 200 cases, 194 were successfully genotyped for the IVS5⫹1 G/A splice site mutation. Among cases, 1 homozygote for this mutation and 7 heterozygotes were found (mean SF of 484 ⫾ 198 g/L [1 standard deviation (SD)]; mean TS of 60.6 ⫾ 11.2% [1 SD]). The homozygote, a man in his late 50s with hepatitis C, had markedly elevated serum iron test results (SF 1870 g/L, TS 99% at the initial screening visit). Seven months later, at the time of further clinical evaluation, he had an SF of 2777 g/L and TS of 99%. He subsequently underwent venesection therapy to remove excess iron and had a total of 4.95-L blood removed during the course of 11 phlebotomy sessions (450 mL each), corresponding to about 2.6-g storage iron, well below the amount deemed clinically significant by most experts. After phlebotomy, the SF fell to 20 g/L. It is certainly possible that hepatitis C is a contributing factor to the elevated iron tests in this participant. Of the 149 controls, 147 were successfully genotyped yielding 1 homozygote for the splice site mutation (a woman in her mid-50s with SF 59 g/L, TS 32% who was never referred for liver biopsy or phlebotomy ther-
94
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Steiner et al
Table I. Gender, phenotype, and splice site genotype of cases and controls Splice site genotype
Case/control
Gender
TS, % ( ⴞSD)
SF, ug/L ( ⴞSD)
N
Homozygous Heterozygous Wild-type Heterozygous Wild-type Heterozygous Wild-type Homozygous Heterozygous Wild-type
Case Case Case Control Control Case Case Control Control Control
Male Male Male Male Male Female Female Female Female Female
99 65.8 (11.9) 65.7 (13.8) 28.7 (6.8) 35.0 (9.0) 53.7 (6.4) 53.7 (10.1) 32 31 28.6 (9.9)
1870 594 (172) 936 (971) 295 (40) 348 (195) 338 (131) 464 (415) 59 27 153 (112)
1 4 92 3 50 3 94 1 1 92
apy) and 4 heterozygotes (mean SF of 228 ⫾ 138 g/L [1 SD]; mean TS of 29.0 ⫾ 5.7% [1 SD]). The normal SF and TS found in the 1 homozygous female control participant could potentially be explained by multiple pregnancies; however, a medical history was not obtained from this participant. Details of the gender, TS, and SF for the cases and controls are shown in Table I. These data indicate comparable allele frequencies of 2.32% and 2.04% for the IVS5⫹1 G/A splice site mutation among cases and controls, respectively (chisquare test, P ⬎ 0.5). DISCUSSION
The HFE IVS5⫹1 G/A splice site mutation seems to be found in Vietnamese populations at a fairly low frequency. In a pilot study of HEIRS Asian participants having marked elevations of both SF and TS, which included 58 participants from the greater Los Angeles area who were mostly Vietnamese and 47 from the Toronto area who were mostly Chinese, this particular splice site mutation was found only in the Vietnamese participants (data not shown). In this report, data are presented for only the second and third homozygotes with this splice site mutation appearing in the scientific literature. The original HFE IVS5⫹1 G/A splice site mutation homozygote was a Vietnamese man living in Germany with marked iron overload. In this report, a second homozygote was described, a male in his late 50s who was also positive for hepatitis C virus with borderline iron overload, and a third homozygote, a woman in her mid-50s with no evidence of iron overload. Interestingly, the allele frequency in cases and controls is about 10-fold higher than that found by Pointon et al.8 However, in their study, 95% of the Vietnamese participants were of the Kinh ethnic subgroup. Although 5% of their participants were from the Tay, S’Tieng, Nung, and Muong ethnic subgroups combined, the number of participants in
each of these subgroups was much too small to make any reliable assessment of mutation allele frequency. Data regarding the geographic and ethnic origin of the HEIRS Study United States Vietnamese participants was not collected, but the 10-fold difference in allele frequencies found between this study and Pointon’s study strongly suggests that this mutation does vary substantially across ethnic subgroups of Vietnamese. As the allele frequencies of the cases and controls are similar in the subjects, the authors conclude that the unexpectedly high prevalence of elevated serum TS and SF in HEIRS Vietnamese participants cannot be explained on the basis of the HFE IVS5⫹1 G/A splice site mutation. The absence of marked iron overload in the 2 homozygous HEIRS participants identified in this study is in contrast to the earlier report of severe iron overload in a Vietnamese homozygote living in Germany, whom is estimated had roughly 6 g of storage iron (25 phlebotomies ⫻ 0.5-L blood/phlebotomy ⫻ ⬃0.5-g iron/L blood with a hemoglobin of approximately 150 g/L).7 This variable expression is analogous to the very variable expression that has been observed in homozygotes for the HFE C282Y mutation, which is most common in Caucasians. It has been proposed that environmental influences or interactions involving modifier genes may contribute to the phenotypic and clinical variability in C282Y homozygotes.1,3,8 –10 It is very likely that similar factors may play a role in determining the phenotype and clinical manifestation in those homozygotes for the IVS5⫹1 G/A splice site mutation as well. As a result of poor understanding of the long-term expression of any HFE mutations, the value of broad population screening for hemochromatosis in clinical practice remains controversial.11 The authors wish to thank their colleagues from the HEIRS Study.
Translational Research Volume 149, Number 2
Steiner et al
REFERENCES
1. Ajioka RS, Kushner JP. Hereditary hemochromatosis. Semin Hematol 2002;39:235– 41. 2. Andersen RV, Tybjaerg-Hansen A, Appleyard M, Birgens H, Nordestgaard BG. Hemochromatosis mutations in the general population: iron overload progression rate. Blood 2004;103: 2914 –9. 3. Beutler E, Felitti VJ, Koziol JA, Ho NJ, Gelbart T. Penetrance of 845G¡A (C282Y) HFE hereditary haemochromatosis mutation in the USA. Lancet 2002;359:211–18. 4. Le Gac G, Ferec C. The molecular genetics of haemochromatosis. Eur J Hum Genet 2005;13:1172– 85. 5. McLaren C, Barton J, Adams P, Harris E, Acton R, Press N, et al. Hemochromatosis and Iron Overload Screening (HEIRS) study design for an evaluation of 100,000 primary care-based adults. Am J Med Sci 2003;325:53– 62. 6. Adams PC, Reboussin DR, Barton JC, McLaren CE, Eckfeldt JH, McLaren GD, et al. Screening of a primary care population
7.
8.
9. 10.
11.
95
of 101,168 persons of diverse ethnic background. N Engl J Med 2005;352:1769 –78. Steiner M, Ochran K, Genschel J, Meier P, Gerl H, Ventz M, et al. A homozygous HFE gene splice site mutation IVS5⫹1 G/A in a hereditary hemochromatosis patient of Vietnamese origin. Gastroenterology 2002;122:789 –95. Pointon JJ, Viprakasit V, Miles KL, Livesey KJ, Steiner M, O’Riordan S, et al. Hemochromatosis gene (HFE) mutations in South East Asia: a potential for iron overload. Blood Cells, Molecules & Diseases 2003;30:302– 6. Ajioka RS, Kushner JP. Clinical consequences of iron overload in hemochromatosis homozygotes. Blood 2003;101:3351– 4. Beutler E. The HFE Cys282Tyr mutation as a necessary but not sufficient cause of clinical hereditary hemochromatosis. Blood 2003;101:3347–50. Whitlock EP, Garlitz BA, Harris EM, Beil TL, Smith PR. Screening for hereditary hemochromatosis: a systematic review for the U.S. Preventive Services Task Force. Ann Intern Med 2006;145:209 –23.