Review for 10.1186/1471-230X-14-12 (2014)

Reviewer's report
Title:Protective effects of hydrogen enriched saline on liver ischemia reperfusion
injury by reducing oxidative stress and HMGB1 release
Version: 1Date:11 June 2013
Reviewer: Uta Dahmen
Reviewer's report:
This associative descriptive study explores the effect of the preoperative
treatment with hydrogen enriched saline on hepatic IRI. The authors report that
parameters of oxidative stress as well as HMGB1 release are lower in the
treatment groups compared to the control group.
Major comments:
1. Clinical relevance: The potential clinical relevance of the approach to reduce
IRI using hydrogen enriched saline is -besides the efficiency- also related to
profile of side effects and the effort needed to produce/store the reagent. This
topic is briefly mentioned in the discussion, but not really covered.
2. Experimental design: Since no specific blocking intervention was performed,
the mechanistic relationship is not proven, but at the most suggested.
3. Experimental set-up (Figure 1) is unclear: It remains elusive how many
observation time points were performed in the untreated IRI group: either animals
were sacrificed after 2h as stated in the experimental design or the whole kinetic
experiment was performed as stated in the figure legends. It remains also elusive
why the authors sacrificed the sham control and the IRI no treatment after 2h and
not after 6h, where the observed changes were most prominent.
4. Quantitation of results: Although quantitative results are presented, the
methods to reach the quantitative results are not or ill described in the method
section. This holds especially for the quantitation of immunohistochemical
evaluation.
5. Histological Images: Selection of the depicted regions of interest should
contain comparable histological areas, either portal field or central vein regions,
and should not be selected randomly.
6. Figure 3B/3G: Quantification based on counting cells with brown precipitation
at cellular membrane seems questionable since extremely hard to perform in a
reliable way. Therefore, the quantification method to detemine the relative
number of positive cells must be reconsidered. Sample size with only 100 cells
evaluated for the quantitative assessment as stated in the legends seems rather
small and not sufficient to reach a reliable and representative result. Either the
sample size must be enlarged or the authors should explain how they ensured
that the sample size is sufficient.
7. Figure 4: Distribution of HMGB1-staining signals seems questionable. In the
sham-operated group, one would not expect a blank slide like a negative control,
but would expect to find signals for HMGB1 in the hepatocyte nuclei, but not in
the cytoplasma. In contrast, in the saline group, where damage should be most
pronounced, one would expect translocation of HMGB1 from the nucleus to the
cytoplasma leading to nuclei without staining and strongly staining cytoplasma.
8. Number of samples employed in the different assays apparently varies as
stated in the legends. The reason for excluding individual samples in selected
assays should be indicated.
Requested answers to Questions of Editorial Board
1. Is the question posed by the authors well defined? yes
2. Are the methods appropriate and well described? No
3. Are the data sound? questionable
4. Does the manuscript adhere to the relevant standards for reporting and data
deposition? questionable
5. Are the discussion and conclusions well balanced and adequately supported
by the data? questionable
6. Are limitations of the work clearly stated? some
7. Do the authors clearly acknowledge any work upon which they are building,
both published and unpublished? yes
8. Do the title and abstract accurately convey what has been found? Somehow
yes
9. Is the writing acceptable? can be improved in terms of spelling errors and
basic grammatical issues. Figure legend should be improved and state the key
message which the authors want to visualize with a given figure. It is
unnecessary and annoying to repeat the same sentence describing the
experimental d
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: No, the manuscript does not need to be seen by a
statistician.
Declaration of competing interests:
'I declare that I have no competing interests'