Reviewer ’s report  
 
Title: Alcoholic vs Non -Alcoholic Fatty Liver in Rats: Distinct Differences in Endocytosis and 
Vesicle Trafficking despite Similar Pathology  
 
 
Version: 0 Date: 30 Nov 2015  
 
 
Reviewer: Pamela Tuma  
 
 
Reviewer's report:  
 
 
This manuscript examines whether distinct differences in altered hepatic function are present in 
two models of fatty liver that are histologically similar: non -alcoholic and alcoholic fatty liver 
(NALF and ALF).  They tested several parameters and determined  that the extent of liver injury 
as measured by ALT and ALP, lipid accumulation and oxidative stress was similar in the two 
disease models.  Interestingly, ALF was associated with additional impairments in asialo -
glycoprotein receptor (ASGPR) endocytosis, rab GTPase levels and possible changes in 
cholesterol synthesis.  Although the study is highly descriptive, it is important.  Despite their 
similarity in histology, the mechanisms by which NALF and ALF progress to end -point liver 
injury (fibrosis, hepatiti s, cirrhosis) are likely different and may point to differential therapeutic 
strategies for treating early stages of these diseases.   However, there are a few points that the 
authors might choose to address to enhance the overall impact of these studies.  
 
 
The selection of rab GTPases examined in the manuscript seems a bit arbitrary and incomplete.  
In their earlier studies, rabs 1, 2, 3d, 5, 7 and 18 were examined in post -nuclear supernatants and 
on purified lipid droplets.  In the PNS samples, changes wer e modest in total protein levels, but 
when lipid droplet associations were monitored, changes were much more apparent and 
numerous.  In an effort to be complete, it might be worthwhile to extend these studies to the 
same complement of rabs and their associ ations with lipid droplets from both ALF and NALF 
hepatocytes.   Also, because changes in levels don't necessarily correlate with altered function, it 
might be useful to determine by indirect immunofluorescence the subcellular distributions of the 
rabs (or  selected rabs) in the two disease models.  
 
 
 
Also, since rab7 has been identified as a key regulator of hepatic lipophagy, it would be 
informative to look at different lipophagic/autophagic markers (by immunoblotting and indirect 
immunofluorescence) in the two disease models to see if the decreased rab7 levels correlate with 
changes in lipophagy.  
Are the methods appropriate and well described?  
If not, please specify what is required in your comments to the authors.  
 
Yes 
 
Does the work include the necessary controls?  
If not, please specify which controls are required in your comments to the authors.  
 
Yes 
 
Are the conclusions drawn adequately supporte d by the data shown?  
If not, please explain in your comments to the authors.  
 
Yes 
 
Are you able to assess any statistics in the manuscript or would you recommend an  
additional statistical review?  
If an a dditional statistical review is recommended, please specify what aspects require further  
assessment in your comments to the editors.  
 
I am able to assess the statistics  
 
Quality of written English  
Please indicate the quality of language in the manuscript:  
 
Acceptable  
 
Declaration of competing interests  
Please complete a declaration of competing interests, considering the following questions:  
 
1. Have you in the past five years received reimbursements, fees, funding, or salary from 
an organisation that may in any way gai n or lose financially from the publication of this  
manuscript, either now or in the future?  
 
2. Do you hold any stocks or shares in an organisation that may in any way gain or lose  
financially from the publication of this manuscript, either now or in the fu ture?  
 
3. Do you hold or are you currently applying for any patents relating to the content of the  
manuscript?  
 
4. Have you received reimbursements, fees, funding, or salary from an organization that 
holds  or has applied for patents relating to the content of the manuscript?  
 
5. Do you have any other financial competing interests?  
 
6. Do you have any non -financial competing interests in relation to this paper?  
 
If you can answer no to all of the above, write 'I  declare that I have no competing interests' 
below.  If your reply is yes to any, please give details below.  
 
I declare that I have no competing interests.  
 
I agree to the open peer review policy of the journal. I understand that my name will be included  
on my  report to the authors and, if the manuscript is accepted for publication, my named report  
including any attachments I upload will be posted on the website along with the authors'  
responses. I agree for my report to be made available under an Open Access C reative Commons  
CC-BY licens e (http://creativecommons.org/licenses/by/4.0/ ). I understand that any comments  
which I do not wish to be included in my named report can be included as confidential comments  
to the editors, which will not be published.  
 
I agree to the open peer review policy of the journal  
Reviewer ’s report  
 
Title: Alcoholic vs Non -Alcoholic Fatty Liver in Rats: Distinct Differences in Endocytosis and 
Vesicle Trafficking despite Similar Pathology  
 
 
Version: 0 Date: 30 Nov 2015  
 
 
Reviewer: Pamela Tuma  
 
 
Reviewer's report:  
 
 
This manuscript examines whether distinct differences in altered hepatic function are present in 
two models of fatty liver that are histologically similar: non -alcoholic and alcoholic fatty liver 
(NALF and ALF).  They tested several parameters and determined  that the extent of liver injury 
as measured by ALT and ALP, lipid accumulation and oxidative stress was similar in the two 
disease models.  Interestingly, ALF was associated with additional impairments in asialo -
glycoprotein receptor (ASGPR) endocytosis, rab GTPase levels and possible changes in 
cholesterol synthesis.  Although the study is highly descriptive, it is important.  Despite their 
similarity in histology, the mechanisms by which NALF and ALF progress to end -point liver 
injury (fibrosis, hepatiti s, cirrhosis) are likely different and may point to differential therapeutic 
strategies for treating early stages of these diseases.   However, there are a few points that the 
authors might choose to address to enhance the overall impact of these studies.  
 
 
The selection of rab GTPases examined in the manuscript seems a bit arbitrary and incomplete.  
In their earlier studies, rabs 1, 2, 3d, 5, 7 and 18 were examined in post -nuclear supernatants and 
on purified lipid droplets.  In the PNS samples, changes wer e modest in total protein levels, but 
when lipid droplet associations were monitored, changes were much more apparent and 
numerous.  In an effort to be complete, it might be worthwhile to extend these studies to the 
same complement of rabs and their associ ations with lipid droplets from both ALF and NALF 
hepatocytes.   Also, because changes in levels don't necessarily correlate with altered function, it 
might be useful to determine by indirect immunofluorescence the subcellular distributions of the 
rabs (or  selected rabs) in the two disease models.  
 
 
 
Also, since rab7 has been identified as a key regulator of hepatic lipophagy, it would be 
informative to look at different lipophagic/autophagic markers (by immunoblotting and indirect 
immunofluorescence) in the two disease models to see if the decreased rab7 levels correlate with 
changes in lipophagy.  
Are the methods appropriate and well described?  
If not, please specify what is required in your comments to the authors.  
 
Yes 
 
Does the work include the necessary controls?  
If not, please specify which controls are required in your comments to the authors.  
 
Yes 
 
Are the conclusions drawn adequately supporte d by the data shown?  
If not, please explain in your comments to the authors.  
 
Yes 
 
Are you able to assess any statistics in the manuscript or would you recommend an  
additional statistical review?  
If an a dditional statistical review is recommended, please specify what aspects require further  
assessment in your comments to the editors.  
 
I am able to assess the statistics  
 
Quality of written English  
Please indicate the quality of language in the manuscript:  
 
Acceptable  
 
Declaration of competing interests  
Please complete a declaration of competing interests, considering the following questions:  
 
1. Have you in the past five years received reimbursements, fees, funding, or salary from 
an organisation that may in any way gai n or lose financially from the publication of this  
manuscript, either now or in the future?  
 
2. Do you hold any stocks or shares in an organisation that may in any way gain or lose  
financially from the publication of this manuscript, either now or in the fu ture?  
 
3. Do you hold or are you currently applying for any patents relating to the content of the  
manuscript?  
 
4. Have you received reimbursements, fees, funding, or salary from an organization that 
holds  or has applied for patents relating to the content of the manuscript?  
 
5. Do you have any other financial competing interests?  
 
6. Do you have any non -financial competing interests in relation to this paper?  
 
If you can answer no to all of the above, write 'I  declare that I have no competing interests' 
below.  If your reply is yes to any, please give details below.  
 
I declare that I have no competing interests.  
 
I agree to the open peer review policy of the journal. I understand that my name will be included  
on my  report to the authors and, if the manuscript is accepted for publication, my named report  
including any attachments I upload will be posted on the website along with the authors'  
responses. I agree for my report to be made available under an Open Access C reative Commons  
CC-BY licens e (http://creativecommons.org/licenses/by/4.0/ ). I understand that any comments  
which I do not wish to be included in my named report can be included as confidential comments  
to the editors, which will not be published.  
 
I agree to the open peer review policy of the journal  
Please review this paper.
user
Reviewer ’s report  
 
Title: Alcoholic vs Non -Alcoholic Fatty Liver in Rats: Distinct Differences in Endocytosis and 
Vesicle Trafficking despite Similar Pathology  
 
 
Version: 0 Date: 30 Nov 2015  
 
 
Reviewer: Pamela Tuma  
 
 
Reviewer's report:  
 
 
This manuscript examines whether distinct differences in altered hepatic function are present in 
two models of fatty liver that are histologically similar: non -alcoholic and alcoholic fatty liver 
(NALF and ALF).  They tested several parameters and determined  that the extent of liver injury 
as measured by ALT and ALP, lipid accumulation and oxidative stress was similar in the two 
disease models.  Interestingly, ALF was associated with additional impairments in asialo -
glycoprotein receptor (ASGPR) endocytosis, rab GTPase levels and possible changes in 
cholesterol synthesis.  Although the study is highly descriptive, it is important.  Despite their 
similarity in histology, the mechanisms by which NALF and ALF progress to end -point liver 
injury (fibrosis, hepatiti s, cirrhosis) are likely different and may point to differential therapeutic 
strategies for treating early stages of these diseases.   However, there are a few points that the 
authors might choose to address to enhance the overall impact of these studies.  
 
 
The selection of rab GTPases examined in the manuscript seems a bit arbitrary and incomplete.  
In their earlier studies, rabs 1, 2, 3d, 5, 7 and 18 were examined in post -nuclear supernatants and 
on purified lipid droplets.  In the PNS samples, changes wer e modest in total protein levels, but 
when lipid droplet associations were monitored, changes were much more apparent and 
numerous.  In an effort to be complete, it might be worthwhile to extend these studies to the 
same complement of rabs and their associ ations with lipid droplets from both ALF and NALF 
hepatocytes.   Also, because changes in levels don't necessarily correlate with altered function, it 
might be useful to determine by indirect immunofluorescence the subcellular distributions of the 
rabs (or  selected rabs) in the two disease models.  
 
 
 
Also, since rab7 has been identified as a key regulator of hepatic lipophagy, it would be 
informative to look at different lipophagic/autophagic markers (by immunoblotting and indirect 
immunofluorescence) in the two disease models to see if the decreased rab7 levels correlate with 
changes in lipophagy.  
Are the methods appropriate and well described?  
If not, please specify what is required in your comments to the authors.  
 
Yes 
 
Does the work include the necessary controls?  
If not, please specify which controls are required in your comments to the authors.  
 
Yes 
 
Are the conclusions drawn adequately supporte d by the data shown?  
If not, please explain in your comments to the authors.  
 
Yes 
 
Are you able to assess any statistics in the manuscript or would you recommend an  
additional statistical review?  
If an a dditional statistical review is recommended, please specify what aspects require further  
assessment in your comments to the editors.  
 
I am able to assess the statistics  
 
Quality of written English  
Please indicate the quality of language in the manuscript:  
 
Acceptable  
 
Declaration of competing interests  
Please complete a declaration of competing interests, considering the following questions:  
 
1. Have you in the past five years received reimbursements, fees, funding, or salary from 
an organisation that may in any way gai n or lose financially from the publication of this  
manuscript, either now or in the future?  
 
2. Do you hold any stocks or shares in an organisation that may in any way gain or lose  
financially from the publication of this manuscript, either now or in the fu ture?  
 
3. Do you hold or are you currently applying for any patents relating to the content of the  
manuscript?  
 
4. Have you received reimbursements, fees, funding, or salary from an organization that 
holds  or has applied for patents relating to the content of the manuscript?  
 
5. Do you have any other financial competing interests?  
 
6. Do you have any non -financial competing interests in relation to this paper?  
 
If you can answer no to all of the above, write 'I  declare that I have no competing interests' 
below.  If your reply is yes to any, please give details below.  
 
I declare that I have no competing interests.  
 
I agree to the open peer review policy of the journal. I understand that my name will be included  
on my  report to the authors and, if the manuscript is accepted for publication, my named report  
including any attachments I upload will be posted on the website along with the authors'  
responses. I agree for my report to be made available under an Open Access C reative Commons  
CC-BY licens e (http://creativecommons.org/licenses/by/4.0/ ). I understand that any comments  
which I do not wish to be included in my named report can be included as confidential comments  
to the editors, which will not be published.  
 
I agree to the open peer review policy of the journal  
assistant