Reviewer's report
Title:Neoplastic transformation of rat liver epithelial cells is enhanced by
non-transferrin-bound iron.
Version: 1Date:3 September 2007
Reviewer: Debbie Trinder
Reviewer's report:
General
Developing experimental models of liver cell transformation has been proved
difficult and previous studies have had variable success using non-biological
forms of iron to promote liver cell tumors. The authors of this study have
undertaken a very scientifically sound series of experiments that clearly
illustrated for the first time in the presence of the tumor initiator
N-methyl-N’-nitro-N-nitrosoguanidine (MNNG) that NTBI in a biological form
found in serum from humans with iron overload was able to promote
transformation of liver epithelial cells in culture. They treated a liver epithelial cell
line T51B with NTBI in the form of ferric ammonium citrate (FAC) for up to 20
weeks. Low FAC concentrations alone had little effect on proliferation or cell
transformation. However, when the cells were incubated with both MNNG and
FAC there was an increase in cell colony formation. This did not occur if FAC
was replaced with ammonium citrate or desferoxamine was added indicating that
iron was required for promotion of cell transformation. Changes in cyclin
expression indicate that disregulation of the cell cycle may be mechanism
responsible for the promotion of tumors.
-------------------------------------------------------------------------------
Major Compulsory Revisions (that the author must respond to before a decision
on publication can be reached)
Introduction p5 last sentence. “However, even in iron-overloaded livers, tumors
do not contain high levels of iron, implicating non-transferrin mechanism in
overload” Clarify this statement. It is not clear to me what this means.
Introduction p6 2nd paragraph, p7 1st paragraph and discussion page 14
paragraph 1 refers to FAC as a physiological form of NTBI which is misleading.
Although ferric citrate is the form of NTBI found in human serum it is usually
associated with iron overload indicating that it is a pathological rather than a
physiological form of iron.
Discussion p14, paragraph 1, 2nd sentence
“We also found tumor-promoting concentrations of FAC had anti-proliferative
effect on normal T51B cells.” This is not correct because 50µM and 200µM FAC
concentration promoted tumor growth but it did not have significant
anti-proliferative effects. While 500µM FAC had a slight effect on tumor
promotion which may not be significant but reduced proliferation. (Table 1 and
Fig 2)
In Table 1 and 2 the results are the means from 4 values. Results should be
expressed as mean ± SEM and p values stated.
Figure 2 and 3: p values need to be given.
-------------------------------------------------------------------------------
Minor Essential Revisions (such as missing labels on figures, or the wrong use of
a term, which the author can be trusted to correct)
Figure 1: Add units to Y-axis
-------------------------------------------------------------------------------
Discretionary Revisions (which the author can choose to ignore)
What next?: Accept after discretionary revisions
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:Neoplastic transformation of rat liver epithelial cells is enhanced by
non-transferrin-bound iron.
Version: 1Date:3 September 2007
Reviewer: Debbie Trinder
Reviewer's report:
General
Developing experimental models of liver cell transformation has been proved
difficult and previous studies have had variable success using non-biological
forms of iron to promote liver cell tumors. The authors of this study have
undertaken a very scientifically sound series of experiments that clearly
illustrated for the first time in the presence of the tumor initiator
N-methyl-N’-nitro-N-nitrosoguanidine (MNNG) that NTBI in a biological form
found in serum from humans with iron overload was able to promote
transformation of liver epithelial cells in culture. They treated a liver epithelial cell
line T51B with NTBI in the form of ferric ammonium citrate (FAC) for up to 20
weeks. Low FAC concentrations alone had little effect on proliferation or cell
transformation. However, when the cells were incubated with both MNNG and
FAC there was an increase in cell colony formation. This did not occur if FAC
was replaced with ammonium citrate or desferoxamine was added indicating that
iron was required for promotion of cell transformation. Changes in cyclin
expression indicate that disregulation of the cell cycle may be mechanism
responsible for the promotion of tumors.
-------------------------------------------------------------------------------
Major Compulsory Revisions (that the author must respond to before a decision
on publication can be reached)
Introduction p5 last sentence. “However, even in iron-overloaded livers, tumors
do not contain high levels of iron, implicating non-transferrin mechanism in
overload” Clarify this statement. It is not clear to me what this means.
Introduction p6 2nd paragraph, p7 1st paragraph and discussion page 14
paragraph 1 refers to FAC as a physiological form of NTBI which is misleading.
Although ferric citrate is the form of NTBI found in human serum it is usually
associated with iron overload indicating that it is a pathological rather than a
physiological form of iron.
Discussion p14, paragraph 1, 2nd sentence
“We also found tumor-promoting concentrations of FAC had anti-proliferative
effect on normal T51B cells.” This is not correct because 50µM and 200µM FAC
concentration promoted tumor growth but it did not have significant
anti-proliferative effects. While 500µM FAC had a slight effect on tumor
promotion which may not be significant but reduced proliferation. (Table 1 and
Fig 2)
In Table 1 and 2 the results are the means from 4 values. Results should be
expressed as mean ± SEM and p values stated.
Figure 2 and 3: p values need to be given.
-------------------------------------------------------------------------------
Minor Essential Revisions (such as missing labels on figures, or the wrong use of
a term, which the author can be trusted to correct)
Figure 1: Add units to Y-axis
-------------------------------------------------------------------------------
Discretionary Revisions (which the author can choose to ignore)
What next?: Accept after discretionary revisions
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:Neoplastic transformation of rat liver epithelial cells is enhanced by
non-transferrin-bound iron.
Version: 1Date:3 September 2007
Reviewer: Debbie Trinder
Reviewer's report:
General
Developing experimental models of liver cell transformation has been proved
difficult and previous studies have had variable success using non-biological
forms of iron to promote liver cell tumors. The authors of this study have
undertaken a very scientifically sound series of experiments that clearly
illustrated for the first time in the presence of the tumor initiator
N-methyl-N’-nitro-N-nitrosoguanidine (MNNG) that NTBI in a biological form
found in serum from humans with iron overload was able to promote
transformation of liver epithelial cells in culture. They treated a liver epithelial cell
line T51B with NTBI in the form of ferric ammonium citrate (FAC) for up to 20
weeks. Low FAC concentrations alone had little effect on proliferation or cell
transformation. However, when the cells were incubated with both MNNG and
FAC there was an increase in cell colony formation. This did not occur if FAC
was replaced with ammonium citrate or desferoxamine was added indicating that
iron was required for promotion of cell transformation. Changes in cyclin
expression indicate that disregulation of the cell cycle may be mechanism
responsible for the promotion of tumors.
-------------------------------------------------------------------------------
Major Compulsory Revisions (that the author must respond to before a decision
on publication can be reached)
Introduction p5 last sentence. “However, even in iron-overloaded livers, tumors
do not contain high levels of iron, implicating non-transferrin mechanism in
overload” Clarify this statement. It is not clear to me what this means.
Introduction p6 2nd paragraph, p7 1st paragraph and discussion page 14
paragraph 1 refers to FAC as a physiological form of NTBI which is misleading.
Although ferric citrate is the form of NTBI found in human serum it is usually
associated with iron overload indicating that it is a pathological rather than a
physiological form of iron.
Discussion p14, paragraph 1, 2nd sentence
“We also found tumor-promoting concentrations of FAC had anti-proliferative
effect on normal T51B cells.” This is not correct because 50µM and 200µM FAC
concentration promoted tumor growth but it did not have significant
anti-proliferative effects. While 500µM FAC had a slight effect on tumor
promotion which may not be significant but reduced proliferation. (Table 1 and
Fig 2)
In Table 1 and 2 the results are the means from 4 values. Results should be
expressed as mean ± SEM and p values stated.
Figure 2 and 3: p values need to be given.
-------------------------------------------------------------------------------
Minor Essential Revisions (such as missing labels on figures, or the wrong use of
a term, which the author can be trusted to correct)
Figure 1: Add units to Y-axis
-------------------------------------------------------------------------------
Discretionary Revisions (which the author can choose to ignore)
What next?: Accept after discretionary revisions
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