Reviewer’s report
Title: 
Short chain fatty acids and colon motility in a mouse model of irritable
bowel syndrome
Version: 
0 
Date: 
20 Jul 2020
Reviewer's report:
The authors used a well established model (acetic acid exposure in neonatal
period) to create a mouse with visceral hypersensitivity. They show that this
model 
has higher amplitude contractions in an isolated colon preparation but less
response to carbacol 
Sodium acetate, sodium propionate and butyric acid induced a dose dependent
decrease of tonic tension, amplitude and frequency of spontaneous in 
the control
group
The effect was reduced in the IBS model particularly so for butyrate. 
Similarly
the effect was less on carbacol induced contraction
The results are clear cut and nicely illustrated and the paper well written though
the discussion seems a bit less well structured.
Specific Comments
1. Abstract: Should mention concentrations of SCFA used
2. Methods: The SCFAs were assessed in faeces but not clear why you did not
choose to measure caecal levels which might be more relevant to the proximal
colon motility which you measured
3. Although you demonstrate hypersensitivity, transit is more relevant to the
microbiota which produce the SCFAs so I think it would have been useful to
assess transit as well. Do we know if it is altered in this model? If so you should
reference this.
4. Results: In control mice the ratio of acetate (0.75±0.07, n=5) propionate
(0.15±0.03, n=5) and butyrate (0.11±0.02, n=5) was 6.5 : 1.3 : 1 (Fig. 5 a). Not
clear in text what units are 
but the figure gives ug/g which is less helpful than
mmol/l as is conventional. Please change in the text.
5. The decrease in 
anaerobic index from control values of -0.352±0.052 ru 
to -
0.437±0.052 ru in the model is fascinating but needs a bit more explanation in
methods 
or introduction to understand why the ratio of C3&C4/C2 is a valid
measure of anaerobicity. The decrease presumably reflects lower acetate values .
Is this related to faster transit?
6. Discussion: Given the contradictory results in the literature I would like to see
a discussion of why this is so. You already mentioned that species differences,
diet and different preparations may all play a role but here it would be useful to
be more specific particularly about your preparation which lacks the input from
enteroendocrine cells and mucosal nerves and is exposed normally to much
lower concentration of SCFAs which are orders of magnitude lower in blood
than the gut lumen. You should discuss how the concentrations you used match
blood levels in mice 
7. You mention the role of ICCs as potential mediators It would have been
helpful to assess these in your model Did you retain any sanples which could be
tested ? It would add a lot to the paper if you could
8. The sentence Incubation of the proximal colon with SCFAs (10
mM)decreased carbachol-induced contractions with a higher potency compared
to butyric acid 
does not make sense . What is being compared with butyrate? 
Minor points
9. Introduction Reference 24 seems wrong Perhaps an error as it refers to a
clinical trial
10. Page 9 line 12 Needs editing 
Mice do not have IBS Please use the term
"mouse 
IBS model" as more accurate.
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Yes
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 supported by the data shown?
If not, please explain in your comments to the authors.
No
Are you able to assess any statistics in the manuscript or would you
recommend an additional statistical review?
If an additional 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 gain 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 future?
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 I have no competing interests
Reviewer Publication Consent.
 I agree for my report to be made available
under an Open Access Creative Commons CC-BY License (
http://creativecommons.org/licenses/by/4.0
) if this manuscript is accepted for
publication. Any comments that I do not wish to be included in the published
report have been included as confidential comments to the editor, which will not
be published.
If you are not happy for us to publish this report, please contact the editorial
office before completing the review.
If you wish, you can include your name in your published report. Please note
you must decide whether to include your name at the start of the process and
confirm this decision whenever prompted. If you change your decision later, we
will not be able to publish your name.
I agree to the terms of the CC-BY 4.0 license; please 
do not
 publish my name
with my report. (default)
Reviewer’s report
Title: 
Short chain fatty acids and colon motility in a mouse model of irritable
bowel syndrome
Version: 
0 
Date: 
20 Jul 2020
Reviewer's report:
The authors used a well established model (acetic acid exposure in neonatal
period) to create a mouse with visceral hypersensitivity. They show that this
model 
has higher amplitude contractions in an isolated colon preparation but less
response to carbacol 
Sodium acetate, sodium propionate and butyric acid induced a dose dependent
decrease of tonic tension, amplitude and frequency of spontaneous in 
the control
group
The effect was reduced in the IBS model particularly so for butyrate. 
Similarly
the effect was less on carbacol induced contraction
The results are clear cut and nicely illustrated and the paper well written though
the discussion seems a bit less well structured.
Specific Comments
1. Abstract: Should mention concentrations of SCFA used
2. Methods: The SCFAs were assessed in faeces but not clear why you did not
choose to measure caecal levels which might be more relevant to the proximal
colon motility which you measured
3. Although you demonstrate hypersensitivity, transit is more relevant to the
microbiota which produce the SCFAs so I think it would have been useful to
assess transit as well. Do we know if it is altered in this model? If so you should
reference this.
4. Results: In control mice the ratio of acetate (0.75±0.07, n=5) propionate
(0.15±0.03, n=5) and butyrate (0.11±0.02, n=5) was 6.5 : 1.3 : 1 (Fig. 5 a). Not
clear in text what units are 
but the figure gives ug/g which is less helpful than
mmol/l as is conventional. Please change in the text.
5. The decrease in 
anaerobic index from control values of -0.352±0.052 ru 
to -
0.437±0.052 ru in the model is fascinating but needs a bit more explanation in
methods 
or introduction to understand why the ratio of C3&C4/C2 is a valid
measure of anaerobicity. The decrease presumably reflects lower acetate values .
Is this related to faster transit?
6. Discussion: Given the contradictory results in the literature I would like to see
a discussion of why this is so. You already mentioned that species differences,
diet and different preparations may all play a role but here it would be useful to
be more specific particularly about your preparation which lacks the input from
enteroendocrine cells and mucosal nerves and is exposed normally to much
lower concentration of SCFAs which are orders of magnitude lower in blood
than the gut lumen. You should discuss how the concentrations you used match
blood levels in mice 
7. You mention the role of ICCs as potential mediators It would have been
helpful to assess these in your model Did you retain any sanples which could be
tested ? It would add a lot to the paper if you could
8. The sentence Incubation of the proximal colon with SCFAs (10
mM)decreased carbachol-induced contractions with a higher potency compared
to butyric acid 
does not make sense . What is being compared with butyrate? 
Minor points
9. Introduction Reference 24 seems wrong Perhaps an error as it refers to a
clinical trial
10. Page 9 line 12 Needs editing 
Mice do not have IBS Please use the term
"mouse 
IBS model" as more accurate.
Publons Reviewer Recognition
. Springer Nature can send verification of this
review directly to Publons (a subsidiary of Clarivate Analytics). If you would
like to take advantage of this service, please click on the “Yes” option below.
Your name, email address, title of the reviewed manuscript, name of the journal,
and date of your review submission (the “Review Data”) will then be transmitted
to Publons upon publication of the manuscript. If you have already registered at
Publons, they will notify you of the receipt of this review and update your profile
as per your settings and their policy. If you are not registered with Publons, you
will receive an email from them asking you to register in order for them to be
able to recognize your review on your new profile page. Publons may use the
Review Data to generate derivative metadata for the benefit of Publons and you
as a reviewer, carefully considering the sensitivity of such information. For
example, Publons may verify your record as a reviewer by updating your profile
published on its webservice if you have registered for such service or help
editors to identify candidate reviewers. Please find the details of processing in
Publons’ privacy policy 
https://publons.com/about/terms
Yes
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 supported by the data shown?
If not, please explain in your comments to the authors.
No
Are you able to assess any statistics in the manuscript or would you
recommend an additional statistical review?
If an additional 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 gain 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 future?
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 I have no competing interests
Reviewer Publication Consent.
 I agree for my report to be made available
under an Open Access Creative Commons CC-BY License (
http://creativecommons.org/licenses/by/4.0
) if this manuscript is accepted for
publication. Any comments that I do not wish to be included in the published
report have been included as confidential comments to the editor, which will not
be published.
If you are not happy for us to publish this report, please contact the editorial
office before completing the review.
If you wish, you can include your name in your published report. Please note
you must decide whether to include your name at the start of the process and
confirm this decision whenever prompted. If you change your decision later, we
will not be able to publish your name.
I agree to the terms of the CC-BY 4.0 license; please 
do not
 publish my name
with my report. (default)
Please review this paper.
user
Reviewer’s report
Title: 
Short chain fatty acids and colon motility in a mouse model of irritable
bowel syndrome
Version: 
0 
Date: 
20 Jul 2020
Reviewer's report:
The authors used a well established model (acetic acid exposure in neonatal
period) to create a mouse with visceral hypersensitivity. They show that this
model 
has higher amplitude contractions in an isolated colon preparation but less
response to carbacol 
Sodium acetate, sodium propionate and butyric acid induced a dose dependent
decrease of tonic tension, amplitude and frequency of spontaneous in 
the control
group
The effect was reduced in the IBS model particularly so for butyrate. 
Similarly
the effect was less on carbacol induced contraction
The results are clear cut and nicely illustrated and the paper well written though
the discussion seems a bit less well structured.
Specific Comments
1. Abstract: Should mention concentrations of SCFA used
2. Methods: The SCFAs were assessed in faeces but not clear why you did not
choose to measure caecal levels which might be more relevant to the proximal
colon motility which you measured
3. Although you demonstrate hypersensitivity, transit is more relevant to the
microbiota which produce the SCFAs so I think it would have been useful to
assess transit as well. Do we know if it is altered in this model? If so you should
reference this.
4. Results: In control mice the ratio of acetate (0.75±0.07, n=5) propionate
(0.15±0.03, n=5) and butyrate (0.11±0.02, n=5) was 6.5 : 1.3 : 1 (Fig. 5 a). Not
clear in text what units are 
but the figure gives ug/g which is less helpful than
mmol/l as is conventional. Please change in the text.
5. The decrease in 
anaerobic index from control values of -0.352±0.052 ru 
to -
0.437±0.052 ru in the model is fascinating but needs a bit more explanation in
methods 
or introduction to understand why the ratio of C3&C4/C2 is a valid
measure of anaerobicity. The decrease presumably reflects lower acetate values .
Is this related to faster transit?
6. Discussion: Given the contradictory results in the literature I would like to see
a discussion of why this is so. You already mentioned that species differences,
diet and different preparations may all play a role but here it would be useful to
be more specific particularly about your preparation which lacks the input from
enteroendocrine cells and mucosal nerves and is exposed normally to much
lower concentration of SCFAs which are orders of magnitude lower in blood
than the gut lumen. You should discuss how the concentrations you used match
blood levels in mice 
7. You mention the role of ICCs as potential mediators It would have been
helpful to assess these in your model Did you retain any sanples which could be
tested ? It would add a lot to the paper if you could
8. The sentence Incubation of the proximal colon with SCFAs (10
mM)decreased carbachol-induced contractions with a higher potency compared
to butyric acid 
does not make sense . What is being compared with butyrate? 
Minor points
9. Introduction Reference 24 seems wrong Perhaps an error as it refers to a
clinical trial
10. Page 9 line 12 Needs editing 
Mice do not have IBS Please use the term
"mouse 
IBS model" as more accurate.
Publons Reviewer Recognition
. Springer Nature can send verification of this
review directly to Publons (a subsidiary of Clarivate Analytics). If you would
like to take advantage of this service, please click on the “Yes” option below.
Your name, email address, title of the reviewed manuscript, name of the journal,
and date of your review submission (the “Review Data”) will then be transmitted
to Publons upon publication of the manuscript. If you have already registered at
Publons, they will notify you of the receipt of this review and update your profile
as per your settings and their policy. If you are not registered with Publons, you
will receive an email from them asking you to register in order for them to be
able to recognize your review on your new profile page. Publons may use the
Review Data to generate derivative metadata for the benefit of Publons and you
as a reviewer, carefully considering the sensitivity of such information. For
example, Publons may verify your record as a reviewer by updating your profile
published on its webservice if you have registered for such service or help
editors to identify candidate reviewers. Please find the details of processing in
Publons’ privacy policy 
https://publons.com/about/terms
Yes
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 supported by the data shown?
If not, please explain in your comments to the authors.
No
Are you able to assess any statistics in the manuscript or would you
recommend an additional statistical review?
If an additional 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 gain 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 future?
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 I have no competing interests
Reviewer Publication Consent.
 I agree for my report to be made available
under an Open Access Creative Commons CC-BY License (
http://creativecommons.org/licenses/by/4.0
) if this manuscript is accepted for
publication. Any comments that I do not wish to be included in the published
report have been included as confidential comments to the editor, which will not
be published.
If you are not happy for us to publish this report, please contact the editorial
office before completing the review.
If you wish, you can include your name in your published report. Please note
you must decide whether to include your name at the start of the process and
confirm this decision whenever prompted. If you change your decision later, we
will not be able to publish your name.
I agree to the terms of the CC-BY 4.0 license; please 
do not
 publish my name
with my report. (default)
assistant