Reviewer's report
Title:Deficiency in four and one half LIM domain protein 2 (FHL2) aggravates
liver fibrosis in mice
Version: 1Date:29 August 2012
Reviewer: Leo A van Grunsven
Reviewer's report:
Authors show a protective role for FHL2 during hepatic injury, showing an
aggravated fibrosis in the FHL2 KO mice when treated with CCl4. They then
focus on the Hepatic stellate cells since they are key in the fibrotic process.
However, in vitro, they do not document a difference between wt and FHL2-/-
HSCs.
Experiments are thoroughly documented.
However, to me, the link between the HSCs and the documented aggravated
hepatic fibrosis is not entirely clear and should be further described or discussed
Major compulsory revisions
Figure 4.
Isolated HSCs from wt and FLH2-/- mice are stained for actin filaments. It would
strengthen the paper if the genes that have been tested in vivo, with an emphasis
on ColIII gene expression, would be tested in a time course of wt and mutant
HSCs (2 and 7 days for example). While maybe phaloidin staining is identical,
FLH2 could still specifically regulate collagens and TGFb. This would corroborate
their in vivo data.
In addition to the staining shown in this figure, one could do stainings for FLH2
during in vitro HSC. Is the pattern of expression the same as in the human
samples, i.e. upregulation and localisation of the protein during activation?
Minor essential revisions.
Figure 2.
The ALT in the FHL2-/- mice treated with CCL4 seems to be higher than in the wt
mice. Although this doesn’t seem to be significant, the trend is clear. Authors
should address this. Since they don’t see an effect on intrinsic HSC
transdifferentiation capacity when comparing wt with FHL2-/- HSCs the
differences they see could be not HSC related.
Figure 3
Authors show that in total liver samples there is only a significant difference in
Col III and TGFb mRNA levels. It is strange that the difference in Laminin
between CCl4 wt and CCl4 FHL2-/- is not considered to be significant. To me,
the difference between sham and CCl4 FHL2-/- is as big as between wt and
FHL2-/- (both CCl4). Can the authors explain this discrepancy?
Who is responsible for this collagen transcription if there is no difference in HSC
activation (according to their in vitro HSC differentiation as well as SMA mRNA
levels in total liver). What about portal fibroblast?
Figure 5.
This figure indeed shows a clear localisation of FHL2 to fibrotic septa. Authors
state that there is a co-expression of FHL2 with SMA while not showing
costainings. One can replace the sentence “strong nuclear and cytoplasmatic
expression of FHL2 and strongly coexpressed #-SMA” maybe with “strong
nuclear and cytoplasmatic expression of FHL2 in fibrotic septa”
Discussion: The authors statement “our findings demonstrating that FHL2 is an
essential factor in controlling hepatic fibrogenesis via regulating HSC activation
and transdifferentiation” is not justified with the data they show. The authors do
not see a difference in HSC transdifferentiation in vitro, so one cannot ommit the
fact that expression of FHL2 in activated HSCs doesnot influence HSC activation
per se but rather has an influence on surrounding cells. This should be discussed
and the statement should be weakened.
Discretionary Revisions
P7: should probably be “FHL2 deficient mice show a different” instead of “FHL2
deficient show an different”
P8 One can ommit one “per se” in this sentence
“types were noted suggesting that the gene deficiency does not per se interfere
with
cellular activation or transdifferentiation per se”
Level of interest: An article whose findings are important to those with closely
related research interests
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:Deficiency in four and one half LIM domain protein 2 (FHL2) aggravates
liver fibrosis in mice
Version: 1Date:29 August 2012
Reviewer: Leo A van Grunsven
Reviewer's report:
Authors show a protective role for FHL2 during hepatic injury, showing an
aggravated fibrosis in the FHL2 KO mice when treated with CCl4. They then
focus on the Hepatic stellate cells since they are key in the fibrotic process.
However, in vitro, they do not document a difference between wt and FHL2-/-
HSCs.
Experiments are thoroughly documented.
However, to me, the link between the HSCs and the documented aggravated
hepatic fibrosis is not entirely clear and should be further described or discussed
Major compulsory revisions
Figure 4.
Isolated HSCs from wt and FLH2-/- mice are stained for actin filaments. It would
strengthen the paper if the genes that have been tested in vivo, with an emphasis
on ColIII gene expression, would be tested in a time course of wt and mutant
HSCs (2 and 7 days for example). While maybe phaloidin staining is identical,
FLH2 could still specifically regulate collagens and TGFb. This would corroborate
their in vivo data.
In addition to the staining shown in this figure, one could do stainings for FLH2
during in vitro HSC. Is the pattern of expression the same as in the human
samples, i.e. upregulation and localisation of the protein during activation?
Minor essential revisions.
Figure 2.
The ALT in the FHL2-/- mice treated with CCL4 seems to be higher than in the wt
mice. Although this doesn’t seem to be significant, the trend is clear. Authors
should address this. Since they don’t see an effect on intrinsic HSC
transdifferentiation capacity when comparing wt with FHL2-/- HSCs the
differences they see could be not HSC related.
Figure 3
Authors show that in total liver samples there is only a significant difference in
Col III and TGFb mRNA levels. It is strange that the difference in Laminin
between CCl4 wt and CCl4 FHL2-/- is not considered to be significant. To me,
the difference between sham and CCl4 FHL2-/- is as big as between wt and
FHL2-/- (both CCl4). Can the authors explain this discrepancy?
Who is responsible for this collagen transcription if there is no difference in HSC
activation (according to their in vitro HSC differentiation as well as SMA mRNA
levels in total liver). What about portal fibroblast?
Figure 5.
This figure indeed shows a clear localisation of FHL2 to fibrotic septa. Authors
state that there is a co-expression of FHL2 with SMA while not showing
costainings. One can replace the sentence “strong nuclear and cytoplasmatic
expression of FHL2 and strongly coexpressed #-SMA” maybe with “strong
nuclear and cytoplasmatic expression of FHL2 in fibrotic septa”
Discussion: The authors statement “our findings demonstrating that FHL2 is an
essential factor in controlling hepatic fibrogenesis via regulating HSC activation
and transdifferentiation” is not justified with the data they show. The authors do
not see a difference in HSC transdifferentiation in vitro, so one cannot ommit the
fact that expression of FHL2 in activated HSCs doesnot influence HSC activation
per se but rather has an influence on surrounding cells. This should be discussed
and the statement should be weakened.
Discretionary Revisions
P7: should probably be “FHL2 deficient mice show a different” instead of “FHL2
deficient show an different”
P8 One can ommit one “per se” in this sentence
“types were noted suggesting that the gene deficiency does not per se interfere
with
cellular activation or transdifferentiation per se”
Level of interest: An article whose findings are important to those with closely
related research interests
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:Deficiency in four and one half LIM domain protein 2 (FHL2) aggravates
liver fibrosis in mice
Version: 1Date:29 August 2012
Reviewer: Leo A van Grunsven
Reviewer's report:
Authors show a protective role for FHL2 during hepatic injury, showing an
aggravated fibrosis in the FHL2 KO mice when treated with CCl4. They then
focus on the Hepatic stellate cells since they are key in the fibrotic process.
However, in vitro, they do not document a difference between wt and FHL2-/-
HSCs.
Experiments are thoroughly documented.
However, to me, the link between the HSCs and the documented aggravated
hepatic fibrosis is not entirely clear and should be further described or discussed
Major compulsory revisions
Figure 4.
Isolated HSCs from wt and FLH2-/- mice are stained for actin filaments. It would
strengthen the paper if the genes that have been tested in vivo, with an emphasis
on ColIII gene expression, would be tested in a time course of wt and mutant
HSCs (2 and 7 days for example). While maybe phaloidin staining is identical,
FLH2 could still specifically regulate collagens and TGFb. This would corroborate
their in vivo data.
In addition to the staining shown in this figure, one could do stainings for FLH2
during in vitro HSC. Is the pattern of expression the same as in the human
samples, i.e. upregulation and localisation of the protein during activation?
Minor essential revisions.
Figure 2.
The ALT in the FHL2-/- mice treated with CCL4 seems to be higher than in the wt
mice. Although this doesn’t seem to be significant, the trend is clear. Authors
should address this. Since they don’t see an effect on intrinsic HSC
transdifferentiation capacity when comparing wt with FHL2-/- HSCs the
differences they see could be not HSC related.
Figure 3
Authors show that in total liver samples there is only a significant difference in
Col III and TGFb mRNA levels. It is strange that the difference in Laminin
between CCl4 wt and CCl4 FHL2-/- is not considered to be significant. To me,
the difference between sham and CCl4 FHL2-/- is as big as between wt and
FHL2-/- (both CCl4). Can the authors explain this discrepancy?
Who is responsible for this collagen transcription if there is no difference in HSC
activation (according to their in vitro HSC differentiation as well as SMA mRNA
levels in total liver). What about portal fibroblast?
Figure 5.
This figure indeed shows a clear localisation of FHL2 to fibrotic septa. Authors
state that there is a co-expression of FHL2 with SMA while not showing
costainings. One can replace the sentence “strong nuclear and cytoplasmatic
expression of FHL2 and strongly coexpressed #-SMA” maybe with “strong
nuclear and cytoplasmatic expression of FHL2 in fibrotic septa”
Discussion: The authors statement “our findings demonstrating that FHL2 is an
essential factor in controlling hepatic fibrogenesis via regulating HSC activation
and transdifferentiation” is not justified with the data they show. The authors do
not see a difference in HSC transdifferentiation in vitro, so one cannot ommit the
fact that expression of FHL2 in activated HSCs doesnot influence HSC activation
per se but rather has an influence on surrounding cells. This should be discussed
and the statement should be weakened.
Discretionary Revisions
P7: should probably be “FHL2 deficient mice show a different” instead of “FHL2
deficient show an different”
P8 One can ommit one “per se” in this sentence
“types were noted suggesting that the gene deficiency does not per se interfere
with
cellular activation or transdifferentiation per se”
Level of interest: An article whose findings are important to those with closely
related research interests
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