Review for 10.1186/1471-230X-14-9 (2014)

Reviewer's report
Title:Nucleophosmin/B23 expression in gastric carcinogenesis
Version: 1Date:1 April 2013
Reviewer: Daniel P Barry
Reviewer's report:
The article is straightforward and presents findings relevant to the gastric
carcinogenesis field. However, to be acceptable for publication a number of
revisions, detailed below, are necessary.
--Major Compulsory Revisions--
1) In the abstract, the authors should touch on why they chose to focus on NPM1
as their target.
2) Protein levels were only measured in 17/22 patients. Why is this so if DNA,
RNA and protein were isolated “simultaneously”? Please explain.
3) In the first paragraph of Results, if samples are standardized to controls, why
is the control value not equal to 1.00? What was used to normalize?
4) In Results (paragraph 1), it is unclear what is meant by the following
statement: “The protein level of NPM1 was reduced 1.5-fold in 35% of GC
samples compared to their paired non-neoplastic gastric tissue.” Is a 1.5-fold
reduction used as a threshold of significance? This should be explained and
justified somewhere.
5) There is no reference to Figure 1B in the text of the manuscript. It must be
mentioned somewhere in the Results. In addition, it is unclear what is being
shown in panel B. Why are there only three pairs? How were they chosen?
Where is the calibrating control sample? These questions should be addressed.
Also, more blots showing more of the 17 pairs should be included.
6) In Figure 1D, what are the units of the two axes? Also, why do RNA values run
to ~2.0 in 1A but go to nearly 15 in 1D? Was some type of transformation
applied? In addition, the fit line from which the correlation values are determined
should be plotted on the graph.
7) Although analysis of NPM1 expression in different clinicopathological
categories revealed no differences (apart from those discussed in the
manuscript), if they appear in Table 1 they must be mentioned somewhere in the
Results.
8) The presentation of the data in Table 1 and the corresponding text in the
Results are confusing. It is unclear where the numbers “0.94 ± 0.58 vs 1.37 ±
0.39, p=0.036” come from. Are there four values: tumor/type 1, non-tumor/type 1,
tumor/type 2, non-tumor/type 2? And are these the groups compared by
ANOVA? Dividing 0.94 by 1.37 gives a result of 0.69, while the corresponding
value in the Table appears to be 0.66—why? Reformatting the table to include all
the relevant data could most likely clear up the confusion, or perhaps two tables
could be made, one containing clinical parameters and the other containing the
relevant values for those variables that showed differences.
9) The most significant finding in the article, a possible connection between
distant metastasis and lower expression of NPM1, is barely remarked on. More
discussion of this topic would increase the paper’s relevance.
--Minor Essential Revisions--
1) The title should be made more descriptive to interest potential readers. Some
hint of the findings should be included.
2) Spell out abbreviations on first use (e.g. NPM1, ACTB, ARF, MDM2), even in
the abstract.
3) In Methods/Tissue samples, the meaning of “distant location of primary tumor”
is unclear. The description of the tissue source should be reworded.
4) In Methods/Tissue samples, the authors equate the vacuolating cytotoxin with
CagA. These are not the same. VacA is the vacuolating cytotoxin and CagA is
cytotoxin-associated gene A. It is not clear which gene was analyzed. Please
rewrite this sentence.
5) In the Methods, a better description or reference of the techniques used to
confirm the presence of H. pylori should be given.
6) In Methods/NPM1 protein expression by Western blot, what was the “control
sample applied to all gels”? Was it one of the 22 patients or was it from another
source?
7) In Methods/ NPM1 mRNA expression by reverse transcription quantitative
polymerase chain, the authors cited for reference [27] are not that paper’s
authors. One or the other should be changed.
8) In the last sentence of Methods/ NPM1 mRNA expression by reverse
transcription quantitative polymerase chain reaction (qRT-PCR), the authors
mention clinicopathological analysis. This should be moved to the Tissue
samples subsection, above.
9) Throughout the text and Table 1 the use of “P” or “p” should be consistent.
10) Somewhere in the text, it should be explained what the ± represents. Is it
standard deviation?
11) In Methods/Statistical analysis, the authors describe using one-way ANOVA
for some analyses. Since all data seem to be paired (by tumor/non-tumor or
various clinical parameters) why aren’t t-tests sufficient? Please explain or
reanalyze.
12) In the Results (paragraph 1), the authors should refer to individual subpanels
of Figure 1 as the results are described to improve comprehension.
13) There are several minor English mistakes throughout the paper, mostly
involving the use of articles (a, an, the), that should be corrected.
--Discretionary Revisions--
None.
Level of interest: An article whose findings are important to those with closely
related research interests
Quality of written English: Needs some language corrections before being
published
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.