Reviewer's report
Title:Activation of Liver FGF21 in Hepatocarcinogenesis and During Hepatic
Stress
Version: 2Date:30 January 2013
Reviewer: Morichika Konishi
Reviewer's report:
This manuscript is interesting studies investigating the hepatic Fgf21 expression
in liver on perturbation and disease development. I found the research question
interesting and the experimental methods and most obtained data seem solid.
However I found some of the data are not clear (especially in IHC results) and
some additional data are required.
Major
1. The authors found that FGF21 expression is induced in in the livers under
many stressed conditions in mouse models and human patient samples and
found the elevation of serum Fgf21 using soluble KLB and Ni-chelating beads.
However, the method is not common to examine the serum Fgf21 levels. At least
in mouse PHx models and HCC models, the levels of Fgf21 must be determined
using commercial ELISA kit (for example Millipore Rat / Mouse FGF-21 ELISA
Kit).
2. The figures of IHC study are somewhat unclear. Using the asterisks, arrows,
arrowheads, and broken lines, the authors should indicate the representative
cells with and without Fgf21 IHC signal, focal steatosis and cirrhosis, HCC
tissues, and adjacent hepatic tissues clearly in the figures.
Especially, the authors explained the Fig. 4B as "FGF21 expression is lost in
cells with abnormal and irregular expansion of nucleus accompanied by focal
stetosis and cirrhosis in surrounding hepatocytes" (page 15, lines 9-12). In
addition, the authors explained Fig. 6F as " The cells within cirrhotic and fibrotic
foci do not express FGF21" (page 16, line 21-page 17, line 1). But I found hard to
understand them because the authors did not indicate the cell types in the
figures.
3. The FGFR4 and KLB KO mice should not be used as the model for
non-tumorigenic genetic deficiency of liver function (page 13, lines 17-22).
FGFR4 and KLB are potential component of FGF21 signaling in liver or white
adipose tissue. Therefore, the induction of liver Fgf21 in their KO mice may not
be caused by the change of bile acid metabolism but may compensate for the
decrease in Fgf21 signaling in white adipose tissue (or liver).
4. p53 is the negative regulator of Fgf21 expression. p53 is also known to inhibit
carcinogenesis. In a diseased liver, Fgf21 expression was lost at cells progress
to malignancy and normal hepatocytes adjacent to tumors expressed FGF21 at
highly elevated levels (page 15, lines 14-16). I think the function of p53 might be
inhibited rather in the former cells than the latter cells, so please discuss the role
of p53 in the Fgf21 induction during HCC.
Minor
1. I think that Fig, 6E and Figure 6F (page 16, lines 21-22) are typos.
2. FGFR4-/- and KLB-/- should be written as FGFR4<sup>-/-</sup> and
KLB<sup>-/-</sup>.
3. in Figure 1A legend, I found "mRNA levels are expressed as fold changes
relative to the internal b-actin levels." (page 34, lines 6-7). Is "mRNA levels are
expressed as fold changes relative to that in muscle? in the fed state" correct?
4. in Figure 1A legend, " *p ,(n=6) <0.05 (page 34, line 8)" is a typo.
Discretionary Revisions
I would recommend to examine the expression of Fgf21 in wild-type liver,
p53+/-LKB+/- liver and LKB+/- liver by IHC to make clear the role of p53 in Fgf21
induction on the cellular levels.
Level of interest: An article of importance in its field
Quality of written English: Acceptable
Statistical review: No, the manuscript does not need to be seen by a
statistician.
Declaration of competing interests:
I declare that I have no competing interests.
Reviewer's report
Title:Activation of Liver FGF21 in Hepatocarcinogenesis and During Hepatic
Stress
Version: 2Date:30 January 2013
Reviewer: Morichika Konishi
Reviewer's report:
This manuscript is interesting studies investigating the hepatic Fgf21 expression
in liver on perturbation and disease development. I found the research question
interesting and the experimental methods and most obtained data seem solid.
However I found some of the data are not clear (especially in IHC results) and
some additional data are required.
Major
1. The authors found that FGF21 expression is induced in in the livers under
many stressed conditions in mouse models and human patient samples and
found the elevation of serum Fgf21 using soluble KLB and Ni-chelating beads.
However, the method is not common to examine the serum Fgf21 levels. At least
in mouse PHx models and HCC models, the levels of Fgf21 must be determined
using commercial ELISA kit (for example Millipore Rat / Mouse FGF-21 ELISA
Kit).
2. The figures of IHC study are somewhat unclear. Using the asterisks, arrows,
arrowheads, and broken lines, the authors should indicate the representative
cells with and without Fgf21 IHC signal, focal steatosis and cirrhosis, HCC
tissues, and adjacent hepatic tissues clearly in the figures.
Especially, the authors explained the Fig. 4B as "FGF21 expression is lost in
cells with abnormal and irregular expansion of nucleus accompanied by focal
stetosis and cirrhosis in surrounding hepatocytes" (page 15, lines 9-12). In
addition, the authors explained Fig. 6F as " The cells within cirrhotic and fibrotic
foci do not express FGF21" (page 16, line 21-page 17, line 1). But I found hard to
understand them because the authors did not indicate the cell types in the
figures.
3. The FGFR4 and KLB KO mice should not be used as the model for
non-tumorigenic genetic deficiency of liver function (page 13, lines 17-22).
FGFR4 and KLB are potential component of FGF21 signaling in liver or white
adipose tissue. Therefore, the induction of liver Fgf21 in their KO mice may not
be caused by the change of bile acid metabolism but may compensate for the
decrease in Fgf21 signaling in white adipose tissue (or liver).
4. p53 is the negative regulator of Fgf21 expression. p53 is also known to inhibit
carcinogenesis. In a diseased liver, Fgf21 expression was lost at cells progress
to malignancy and normal hepatocytes adjacent to tumors expressed FGF21 at
highly elevated levels (page 15, lines 14-16). I think the function of p53 might be
inhibited rather in the former cells than the latter cells, so please discuss the role
of p53 in the Fgf21 induction during HCC.
Minor
1. I think that Fig, 6E and Figure 6F (page 16, lines 21-22) are typos.
2. FGFR4-/- and KLB-/- should be written as FGFR4<sup>-/-</sup> and
KLB<sup>-/-</sup>.
3. in Figure 1A legend, I found "mRNA levels are expressed as fold changes
relative to the internal b-actin levels." (page 34, lines 6-7). Is "mRNA levels are
expressed as fold changes relative to that in muscle? in the fed state" correct?
4. in Figure 1A legend, " *p ,(n=6) <0.05 (page 34, line 8)" is a typo.
Discretionary Revisions
I would recommend to examine the expression of Fgf21 in wild-type liver,
p53+/-LKB+/- liver and LKB+/- liver by IHC to make clear the role of p53 in Fgf21
induction on the cellular levels.
Level of interest: An article of importance in its field
Quality of written English: Acceptable
Statistical review: No, the manuscript does not need to be seen by a
statistician.
Declaration of competing interests:
I declare that I have no competing interests.
Please review this paper.
user
Reviewer's report
Title:Activation of Liver FGF21 in Hepatocarcinogenesis and During Hepatic
Stress
Version: 2Date:30 January 2013
Reviewer: Morichika Konishi
Reviewer's report:
This manuscript is interesting studies investigating the hepatic Fgf21 expression
in liver on perturbation and disease development. I found the research question
interesting and the experimental methods and most obtained data seem solid.
However I found some of the data are not clear (especially in IHC results) and
some additional data are required.
Major
1. The authors found that FGF21 expression is induced in in the livers under
many stressed conditions in mouse models and human patient samples and
found the elevation of serum Fgf21 using soluble KLB and Ni-chelating beads.
However, the method is not common to examine the serum Fgf21 levels. At least
in mouse PHx models and HCC models, the levels of Fgf21 must be determined
using commercial ELISA kit (for example Millipore Rat / Mouse FGF-21 ELISA
Kit).
2. The figures of IHC study are somewhat unclear. Using the asterisks, arrows,
arrowheads, and broken lines, the authors should indicate the representative
cells with and without Fgf21 IHC signal, focal steatosis and cirrhosis, HCC
tissues, and adjacent hepatic tissues clearly in the figures.
Especially, the authors explained the Fig. 4B as "FGF21 expression is lost in
cells with abnormal and irregular expansion of nucleus accompanied by focal
stetosis and cirrhosis in surrounding hepatocytes" (page 15, lines 9-12). In
addition, the authors explained Fig. 6F as " The cells within cirrhotic and fibrotic
foci do not express FGF21" (page 16, line 21-page 17, line 1). But I found hard to
understand them because the authors did not indicate the cell types in the
figures.
3. The FGFR4 and KLB KO mice should not be used as the model for
non-tumorigenic genetic deficiency of liver function (page 13, lines 17-22).
FGFR4 and KLB are potential component of FGF21 signaling in liver or white
adipose tissue. Therefore, the induction of liver Fgf21 in their KO mice may not
be caused by the change of bile acid metabolism but may compensate for the
decrease in Fgf21 signaling in white adipose tissue (or liver).
4. p53 is the negative regulator of Fgf21 expression. p53 is also known to inhibit
carcinogenesis. In a diseased liver, Fgf21 expression was lost at cells progress
to malignancy and normal hepatocytes adjacent to tumors expressed FGF21 at
highly elevated levels (page 15, lines 14-16). I think the function of p53 might be
inhibited rather in the former cells than the latter cells, so please discuss the role
of p53 in the Fgf21 induction during HCC.
Minor
1. I think that Fig, 6E and Figure 6F (page 16, lines 21-22) are typos.
2. FGFR4-/- and KLB-/- should be written as FGFR4<sup>-/-</sup> and
KLB<sup>-/-</sup>.
3. in Figure 1A legend, I found "mRNA levels are expressed as fold changes
relative to the internal b-actin levels." (page 34, lines 6-7). Is "mRNA levels are
expressed as fold changes relative to that in muscle? in the fed state" correct?
4. in Figure 1A legend, " *p ,(n=6) <0.05 (page 34, line 8)" is a typo.
Discretionary Revisions
I would recommend to examine the expression of Fgf21 in wild-type liver,
p53+/-LKB+/- liver and LKB+/- liver by IHC to make clear the role of p53 in Fgf21
induction on the cellular levels.
Level of interest: An article of importance in its field
Quality of written English: Acceptable
Statistical review: No, the manuscript does not need to be seen by a
statistician.
Declaration of competing interests:
I declare that I have no competing interests.
assistant