Title: Non-alcoholic fatty liver disease in hemochromatosis probands with iron overload and
HFE p.C282Y/p.C282Y
Version: 3 Date: 13 Feb 2023
Reviewer's report:
In this manuscript, Barton et al report their examination the effect on selected outcomes of
the diagnosis of non-alcoholic fatty liver disease (NAFLD) in a selected population of iron
overloaded (per serum ferritin) probands homozygous for the HFE C282Y mutation. They
excluded those with other known risk factors for NAFLD, including self-reported heavy alcohol
consumption. They observe an association between NAFLD and ferritin levels as well as type II
diabetes. They also noted higher prevalence of an elevated ALT/AST in the group with NAFLD.
These observations are of interest in further characterizing  liver disease in HFE-associated
hereditary hemochromatosis. The following suggestions and questions are raised:
1) For clarity, in the Abstract Results section, second sentence. Change the first word from
"These ..." to "The following ..."
2) Regarding alcohol consumption. The investigators define “heavy” alcohol consumption as
>14g per day (self reported). The CDC defines heavy alcohol consumption as 8 or more drinks
(14g/drink) per week for women and 15 or more drinks per week for men. Please either use these
definitions or provide basis for the cutoff used in this study.
3) The manuscript states “Thirty-five probands (53.0%) underwent liver biopsy. Of these, 11 had
NAFLD (68.6% (11/15)) and 24 did not have NAFLD (48.0% (24/50)) (p = 0.1653).” Should
this not be 24/35 that did not have NAFLD?
4) Please provide a breakdown of the tests used to diagnose NAFLD (biopsy, ultrasound, CT)
in the subjects.
5) Please add a footnote to the reported transferrin saturation levels in the table to flag that these
were obtained without regard to fasting state.
6) The manuscript states (p13) “Odds ratios were SF (1.0-1.0) and T2DM (1.1-22.3).” Please
clarify in the sentence the groups compared, and please correct the reported OR for SF.
7) The authors postulate that “combined effects of increased hepatic iron and hepatic steatosis
account in part for higher prevalences of elevated ALT and elevated AST in hemochromatosis
probands with p.C282Y/p.C282Y who also have NAFLD.” Do the authors mean elevations in
each? The ALT/AST?
8) What measurements, in addition to serum ferritin, were performed to assess hepatic iron
concentration in the subjects? Are these informative in assessing the combined effect of liver
iron and NAFLD on transaminase levels?
9) Have the authors considered utilizing the NAFLD index (ANI) score? It might be informative
to assess the ANI in this population, and to consider its potential as a continuous variable.
10) It would be informative in the discussion for the authors to compare, where possible, each of
the relevant outcomes (and correlations) across four groups: C282Y/C282Y subjects +/- NAFLD
in their study and the general (preferably comparable) population +/- NAFLD from the literature.
11) Please provide figures for the significant  logistic regression analyses.
Title: Non-alcoholic fatty liver disease in hemochromatosis probands with iron overload and
HFE p.C282Y/p.C282Y
Version: 3 Date: 13 Feb 2023
Reviewer's report:
In this manuscript, Barton et al report their examination the effect on selected outcomes of
the diagnosis of non-alcoholic fatty liver disease (NAFLD) in a selected population of iron
overloaded (per serum ferritin) probands homozygous for the HFE C282Y mutation. They
excluded those with other known risk factors for NAFLD, including self-reported heavy alcohol
consumption. They observe an association between NAFLD and ferritin levels as well as type II
diabetes. They also noted higher prevalence of an elevated ALT/AST in the group with NAFLD.
These observations are of interest in further characterizing  liver disease in HFE-associated
hereditary hemochromatosis. The following suggestions and questions are raised:
1) For clarity, in the Abstract Results section, second sentence. Change the first word from
"These ..." to "The following ..."
2) Regarding alcohol consumption. The investigators define “heavy” alcohol consumption as
>14g per day (self reported). The CDC defines heavy alcohol consumption as 8 or more drinks
(14g/drink) per week for women and 15 or more drinks per week for men. Please either use these
definitions or provide basis for the cutoff used in this study.
3) The manuscript states “Thirty-five probands (53.0%) underwent liver biopsy. Of these, 11 had
NAFLD (68.6% (11/15)) and 24 did not have NAFLD (48.0% (24/50)) (p = 0.1653).” Should
this not be 24/35 that did not have NAFLD?
4) Please provide a breakdown of the tests used to diagnose NAFLD (biopsy, ultrasound, CT)
in the subjects.
5) Please add a footnote to the reported transferrin saturation levels in the table to flag that these
were obtained without regard to fasting state.
6) The manuscript states (p13) “Odds ratios were SF (1.0-1.0) and T2DM (1.1-22.3).” Please
clarify in the sentence the groups compared, and please correct the reported OR for SF.
7) The authors postulate that “combined effects of increased hepatic iron and hepatic steatosis
account in part for higher prevalences of elevated ALT and elevated AST in hemochromatosis
probands with p.C282Y/p.C282Y who also have NAFLD.” Do the authors mean elevations in
each? The ALT/AST?
8) What measurements, in addition to serum ferritin, were performed to assess hepatic iron
concentration in the subjects? Are these informative in assessing the combined effect of liver
iron and NAFLD on transaminase levels?
9) Have the authors considered utilizing the NAFLD index (ANI) score? It might be informative
to assess the ANI in this population, and to consider its potential as a continuous variable.
10) It would be informative in the discussion for the authors to compare, where possible, each of
the relevant outcomes (and correlations) across four groups: C282Y/C282Y subjects +/- NAFLD
in their study and the general (preferably comparable) population +/- NAFLD from the literature.
11) Please provide figures for the significant  logistic regression analyses.
Please review this paper.
user
Title: Non-alcoholic fatty liver disease in hemochromatosis probands with iron overload and
HFE p.C282Y/p.C282Y
Version: 3 Date: 13 Feb 2023
Reviewer's report:
In this manuscript, Barton et al report their examination the effect on selected outcomes of
the diagnosis of non-alcoholic fatty liver disease (NAFLD) in a selected population of iron
overloaded (per serum ferritin) probands homozygous for the HFE C282Y mutation. They
excluded those with other known risk factors for NAFLD, including self-reported heavy alcohol
consumption. They observe an association between NAFLD and ferritin levels as well as type II
diabetes. They also noted higher prevalence of an elevated ALT/AST in the group with NAFLD.
These observations are of interest in further characterizing  liver disease in HFE-associated
hereditary hemochromatosis. The following suggestions and questions are raised:
1) For clarity, in the Abstract Results section, second sentence. Change the first word from
"These ..." to "The following ..."
2) Regarding alcohol consumption. The investigators define “heavy” alcohol consumption as
>14g per day (self reported). The CDC defines heavy alcohol consumption as 8 or more drinks
(14g/drink) per week for women and 15 or more drinks per week for men. Please either use these
definitions or provide basis for the cutoff used in this study.
3) The manuscript states “Thirty-five probands (53.0%) underwent liver biopsy. Of these, 11 had
NAFLD (68.6% (11/15)) and 24 did not have NAFLD (48.0% (24/50)) (p = 0.1653).” Should
this not be 24/35 that did not have NAFLD?
4) Please provide a breakdown of the tests used to diagnose NAFLD (biopsy, ultrasound, CT)
in the subjects.
5) Please add a footnote to the reported transferrin saturation levels in the table to flag that these
were obtained without regard to fasting state.
6) The manuscript states (p13) “Odds ratios were SF (1.0-1.0) and T2DM (1.1-22.3).” Please
clarify in the sentence the groups compared, and please correct the reported OR for SF.
7) The authors postulate that “combined effects of increased hepatic iron and hepatic steatosis
account in part for higher prevalences of elevated ALT and elevated AST in hemochromatosis
probands with p.C282Y/p.C282Y who also have NAFLD.” Do the authors mean elevations in
each? The ALT/AST?
8) What measurements, in addition to serum ferritin, were performed to assess hepatic iron
concentration in the subjects? Are these informative in assessing the combined effect of liver
iron and NAFLD on transaminase levels?
9) Have the authors considered utilizing the NAFLD index (ANI) score? It might be informative
to assess the ANI in this population, and to consider its potential as a continuous variable.
10) It would be informative in the discussion for the authors to compare, where possible, each of
the relevant outcomes (and correlations) across four groups: C282Y/C282Y subjects +/- NAFLD
in their study and the general (preferably comparable) population +/- NAFLD from the literature.
11) Please provide figures for the significant  logistic regression analyses.
assistant