Reviewer's report
Title:Activation of the Prostaglandin D2 metabolic pathway in Crohn's disease:
involvement of the enteric nervous system
Version:3Date:20 April 2015
Reviewer: Fernand-Pierre Gendron
Reviewer's report:
The work presented in this manuscript was to demonstrate that PGD2 metabolic
pathway in Crohn’s disease was affecting the enteric nervous system. The
presented results are supportive of the conclusion, but a number of minor
concerns must be address.
The abbreviation must be defined prior to first use.
Line 214. « (Fig 1A). Five patients with quiescent disease also presented with an
increase in TNF-# expression. This increase was not associated with any
relevant clinical characteristics or treatment. » How can this been explained?
There are clearly two groups for the quiescent CD group.
Line 224. “Cytoplasmic Phospholipase A2 (cPLA2) and COX1 were up-regulated
in the mucosa of patients with active CD and the mucosa of five patients with a
quiescent disease (Fig 2A-B). » Same comment has above and are these five
patients always the same 5 patients? This will need to be addressed in the
discussion. The mean values (bars) of Panels A and B appear a bit elevated
given that most of the measured values are in the bottom range.
Line 258 – Fig. 6A. The effect of AT56 strongly and significantly abolished the
LPS-induced PDG2 secretion but this compound did not abolish the PDG2
secretion as the concentration of released PGD2 is still significant as compared
to control (no LPS and no AT56). Appropriate correction/comment is needed.
Line 259. LPS is used to induce an inflammatory stress, but it is a not an
inflammatory conditions. This sentence and corresponding conclusion should
clearly state this difference. To mimic inflammatory conditions, cytokines such as
TNFalpha, IL1beta should be added as well. In fact, it would appropriate to show
that if any of these pro-inflammatory cytokines stimulate PGD2 release. Under in
vivo condition, ENS will be exposed to pro-inflammatory cytokines and only
weakly to bacterial products such as LPS.
Figure 6B. Is there a reason for not using AT56 at least in the co-culture model?
Such experiment should be provided to demonstrate that PGD2 secretion
involved LPGDS.
Line 271. « Our work showed for the first time that there is activation of the entire
PGD2 metabolic pathway involving cPLA2, COX1, COX2 and LPGDS in the
inflamed zone of the colonic mucosa in active CD. Activation leads to increased
PGD2 levels in inflamed colonic. » The authors did not show activation but rather
an increase in mRNA expression that is quite different than measuring protein
levels and enzyme activity.
Line 283. “Among quiescent patients, these five patients did not show any
obvious clinical differences from other quiescent patients. However, the different
profile could be predictive of recurrence of an acute disease. » Is this an
assumption by the authors or is it supported by the literature?
Line 300. “…we showed up-regulation of IL1-# and COX2 301 expression in the
inflamed zone of the colon in patients with active CD leading to 302 activation of
prostaglandin pathways. » Not supported by the presented results. The provided
data and the current literature are, however, supporting this conclusion.
Accordingly, the authors must re-write part of the discussion and conclusion.
Line 319. The discussion about the possible involvement of the different PGD2
receptor is very interesting and made me wonder as why the authors did not
quantify the expression levels of these receptors in their models and biopsies.
They have all the tools in place. By doing so, the manuscript would gain in
importance. Hence, these new results would support this paragraph (line
317-340) of the discussion.
Figure legends. Given that 3 different tests were used to determine the statistical
significance, the authors must write for each figure that one was use in particular.
Hence, all of the gene names need to be written according to the HUGO Gene
Nomenclature Committee. Corrections are required throughout the manuscript.
Figure 4. Typical micrograph of how many different specimens.
Figure 6 legend. AT56 was used at what concentration and how was determined
this effective concentration. Appropriate concentration-response must be
provided. Hence, the effect of this pharmacological compound could be better
supported by the use of interfering RNA to block LPGDS expression.
Level of interest: An article of importance in its field
Quality of written English: Acceptable
Statistical review: Yes, but I do not feel adequately qualified to assess the
statistics.
Declaration of competing interests:
I declare that I have no competing interests'
Reviewer's report
Title:Activation of the Prostaglandin D2 metabolic pathway in Crohn's disease:
involvement of the enteric nervous system
Version:3Date:20 April 2015
Reviewer: Fernand-Pierre Gendron
Reviewer's report:
The work presented in this manuscript was to demonstrate that PGD2 metabolic
pathway in Crohn’s disease was affecting the enteric nervous system. The
presented results are supportive of the conclusion, but a number of minor
concerns must be address.
The abbreviation must be defined prior to first use.
Line 214. « (Fig 1A). Five patients with quiescent disease also presented with an
increase in TNF-# expression. This increase was not associated with any
relevant clinical characteristics or treatment. » How can this been explained?
There are clearly two groups for the quiescent CD group.
Line 224. “Cytoplasmic Phospholipase A2 (cPLA2) and COX1 were up-regulated
in the mucosa of patients with active CD and the mucosa of five patients with a
quiescent disease (Fig 2A-B). » Same comment has above and are these five
patients always the same 5 patients? This will need to be addressed in the
discussion. The mean values (bars) of Panels A and B appear a bit elevated
given that most of the measured values are in the bottom range.
Line 258 – Fig. 6A. The effect of AT56 strongly and significantly abolished the
LPS-induced PDG2 secretion but this compound did not abolish the PDG2
secretion as the concentration of released PGD2 is still significant as compared
to control (no LPS and no AT56). Appropriate correction/comment is needed.
Line 259. LPS is used to induce an inflammatory stress, but it is a not an
inflammatory conditions. This sentence and corresponding conclusion should
clearly state this difference. To mimic inflammatory conditions, cytokines such as
TNFalpha, IL1beta should be added as well. In fact, it would appropriate to show
that if any of these pro-inflammatory cytokines stimulate PGD2 release. Under in
vivo condition, ENS will be exposed to pro-inflammatory cytokines and only
weakly to bacterial products such as LPS.
Figure 6B. Is there a reason for not using AT56 at least in the co-culture model?
Such experiment should be provided to demonstrate that PGD2 secretion
involved LPGDS.
Line 271. « Our work showed for the first time that there is activation of the entire
PGD2 metabolic pathway involving cPLA2, COX1, COX2 and LPGDS in the
inflamed zone of the colonic mucosa in active CD. Activation leads to increased
PGD2 levels in inflamed colonic. » The authors did not show activation but rather
an increase in mRNA expression that is quite different than measuring protein
levels and enzyme activity.
Line 283. “Among quiescent patients, these five patients did not show any
obvious clinical differences from other quiescent patients. However, the different
profile could be predictive of recurrence of an acute disease. » Is this an
assumption by the authors or is it supported by the literature?
Line 300. “…we showed up-regulation of IL1-# and COX2 301 expression in the
inflamed zone of the colon in patients with active CD leading to 302 activation of
prostaglandin pathways. » Not supported by the presented results. The provided
data and the current literature are, however, supporting this conclusion.
Accordingly, the authors must re-write part of the discussion and conclusion.
Line 319. The discussion about the possible involvement of the different PGD2
receptor is very interesting and made me wonder as why the authors did not
quantify the expression levels of these receptors in their models and biopsies.
They have all the tools in place. By doing so, the manuscript would gain in
importance. Hence, these new results would support this paragraph (line
317-340) of the discussion.
Figure legends. Given that 3 different tests were used to determine the statistical
significance, the authors must write for each figure that one was use in particular.
Hence, all of the gene names need to be written according to the HUGO Gene
Nomenclature Committee. Corrections are required throughout the manuscript.
Figure 4. Typical micrograph of how many different specimens.
Figure 6 legend. AT56 was used at what concentration and how was determined
this effective concentration. Appropriate concentration-response must be
provided. Hence, the effect of this pharmacological compound could be better
supported by the use of interfering RNA to block LPGDS expression.
Level of interest: An article of importance in its field
Quality of written English: Acceptable
Statistical review: Yes, but I do not feel adequately qualified to assess the
statistics.
Declaration of competing interests:
I declare that I have no competing interests'
Please review this paper.
user
Reviewer's report
Title:Activation of the Prostaglandin D2 metabolic pathway in Crohn's disease:
involvement of the enteric nervous system
Version:3Date:20 April 2015
Reviewer: Fernand-Pierre Gendron
Reviewer's report:
The work presented in this manuscript was to demonstrate that PGD2 metabolic
pathway in Crohn’s disease was affecting the enteric nervous system. The
presented results are supportive of the conclusion, but a number of minor
concerns must be address.
The abbreviation must be defined prior to first use.
Line 214. « (Fig 1A). Five patients with quiescent disease also presented with an
increase in TNF-# expression. This increase was not associated with any
relevant clinical characteristics or treatment. » How can this been explained?
There are clearly two groups for the quiescent CD group.
Line 224. “Cytoplasmic Phospholipase A2 (cPLA2) and COX1 were up-regulated
in the mucosa of patients with active CD and the mucosa of five patients with a
quiescent disease (Fig 2A-B). » Same comment has above and are these five
patients always the same 5 patients? This will need to be addressed in the
discussion. The mean values (bars) of Panels A and B appear a bit elevated
given that most of the measured values are in the bottom range.
Line 258 – Fig. 6A. The effect of AT56 strongly and significantly abolished the
LPS-induced PDG2 secretion but this compound did not abolish the PDG2
secretion as the concentration of released PGD2 is still significant as compared
to control (no LPS and no AT56). Appropriate correction/comment is needed.
Line 259. LPS is used to induce an inflammatory stress, but it is a not an
inflammatory conditions. This sentence and corresponding conclusion should
clearly state this difference. To mimic inflammatory conditions, cytokines such as
TNFalpha, IL1beta should be added as well. In fact, it would appropriate to show
that if any of these pro-inflammatory cytokines stimulate PGD2 release. Under in
vivo condition, ENS will be exposed to pro-inflammatory cytokines and only
weakly to bacterial products such as LPS.
Figure 6B. Is there a reason for not using AT56 at least in the co-culture model?
Such experiment should be provided to demonstrate that PGD2 secretion
involved LPGDS.
Line 271. « Our work showed for the first time that there is activation of the entire
PGD2 metabolic pathway involving cPLA2, COX1, COX2 and LPGDS in the
inflamed zone of the colonic mucosa in active CD. Activation leads to increased
PGD2 levels in inflamed colonic. » The authors did not show activation but rather
an increase in mRNA expression that is quite different than measuring protein
levels and enzyme activity.
Line 283. “Among quiescent patients, these five patients did not show any
obvious clinical differences from other quiescent patients. However, the different
profile could be predictive of recurrence of an acute disease. » Is this an
assumption by the authors or is it supported by the literature?
Line 300. “…we showed up-regulation of IL1-# and COX2 301 expression in the
inflamed zone of the colon in patients with active CD leading to 302 activation of
prostaglandin pathways. » Not supported by the presented results. The provided
data and the current literature are, however, supporting this conclusion.
Accordingly, the authors must re-write part of the discussion and conclusion.
Line 319. The discussion about the possible involvement of the different PGD2
receptor is very interesting and made me wonder as why the authors did not
quantify the expression levels of these receptors in their models and biopsies.
They have all the tools in place. By doing so, the manuscript would gain in
importance. Hence, these new results would support this paragraph (line
317-340) of the discussion.
Figure legends. Given that 3 different tests were used to determine the statistical
significance, the authors must write for each figure that one was use in particular.
Hence, all of the gene names need to be written according to the HUGO Gene
Nomenclature Committee. Corrections are required throughout the manuscript.
Figure 4. Typical micrograph of how many different specimens.
Figure 6 legend. AT56 was used at what concentration and how was determined
this effective concentration. Appropriate concentration-response must be
provided. Hence, the effect of this pharmacological compound could be better
supported by the use of interfering RNA to block LPGDS expression.
Level of interest: An article of importance in its field
Quality of written English: Acceptable
Statistical review: Yes, but I do not feel adequately qualified to assess the
statistics.
Declaration of competing interests:
I declare that I have no competing interests'
assistant