Review for 10.1186/s12876-015-0248-8 (2015)

Reviewer's report
Title:Fluvastatin attenuates hepatic steatosis-induced fibrogenesis in rats
through inhibiting paracrine effect of hepatocyte on hepatic stellate cells
Version:2Date:11 September 2014
Reviewer: Stefania Grimaudo
Reviewer's report:
The manuscript is interesting because it is aimed at clarifying the effects of
statins on NASH mediated fibrogenesis. Even if the current standard of care for
the treatment of patients with NAFLD/NASH is focuses on lifestyle interventions,
particularly weight loss and exercise, pharmacological therapies will still be
required for the majority of patients with NASH. However there is a lack of
consensus regarding the most effective and appropriate treatment for
NAFLD/NASH.
In this work the paracrine effect of hepatocyte on HSCs activation is investigated
and an appreciable in vitro and in vivo integrate approach is used. However the
animal model for NAFLD/NASH chosen (Choline-deficient L-amino acid defined
diet) is able to cause rapid hepatic accumulation of lipids and development of
fibrosis due to an acute damage. This condition is deeply different from the
human’s progression from NAFLD to NASH, so be careful in extending the
results to human NASH.
Moreover the methods are generally appropriate and well described and the
discussion and the conclusions are adequately supported by the data. For these
reasons the writing is acceptable.
However the following Minor Essential Revisions are required:
• ABSTRACT - Results paragraph: “Flu-pretreated cells” instead of “Flu-treatment
group”.
• INTRODUCTION: “nuclear factor kappa (NF#B) comprises a family of inducible
transcription factors consisting among others of p65 and p50 subunit of Rel
protein family”. Remember that NF#B is a dimeric transcription factor comprised
of five family members RelA (p65), RelB, c-Rel, p50 and p52.
• MATERIALS AND METHODS – The title of Third paragraph must be
“Measurement of reactive oxygen species (ROS) production”.
• MATERIALS AND METHODS – Third paragraph- “Performed at indicated time
points by the excitation” instead of “Performed with”.
• MATERIALS AND METHODS – The title of Fourth paragraph must be
“Cytotoxicity assay”.
• MATERIALS AND METHODS – Fourth paragraph- Cellular plating
concentration must be indicate.
• MATERIALS AND METHODS – Fourth paragraph- “The absorption intensity at
540 nm” instead of “The A540 absorption intensity”.
• MATERIALS AND METHODS –Seventh paragraph- “levels were measured
following standard procedures using a colorimetric analyzer (Dri#Chem 3000).
• RESULTS –First paragraph, first sub- paragraph: “after 24 hours of exposure”
instead of culturing.
• RESULTS –First paragraph, second sub- paragraph: I disagree with the
assertion “Flu (1-20 µM) concentration-dependently attenuated NF#B p65
nuclear translocation in both cell types (Figure 2A)”. In fact, even though it is
visible that Pre-treatment with Flu for 2 hr reduced the NF#B p65 nuclear
translocation in PA-treated HepG2 cells and PRHs at 6 hr after treatment, it is not
evident a dose-dependent effect. The same assertion is found in the Discussion
section.
• RESULTS –First paragraph, second sub- paragraph: “Flu treatment inhibited
the mRNA expression levels of pro-inflammatory gene transcripts (ICAM-1, IL-8,
TNF-#) in both PA-treated HepG2 cells and PRHs”. It is not clearly visible in the
electrophoresis the difference between the line 2 and the others especially for
TNF-# (Figures 2B and 2C).
• FIGURE 1 (A): On the ordinate axis of the HepG2 diagram “% of control group”
must be use instead of “Fold of control group”.
• FIGURE 1(B),1(C): the figure must be more similar for HepG2 and PRH cells:
the istograms –PA, -Flu and 200 uM PA, -Flu must be repeat twice in HepG2 -6hr
and -12hr.
• FIGURE (2A): On the ordinate axis of the HepG2 diagram “P65/PCNA” must be
used instead of “P65/#-actin”.
• NOX could be added to abbreviations.
• Finally, I suggest the following Discretionary Revision:
The method used to evaluate mRNA expression is the Reverse Transcription and
semi-quantitative PCR. I think that an approach of relative quantification using
Reverse Transcription Real Time PCR could be more appropriate. It can provide
better sensitivity and precision respect to the results shown in FIGURE 2B and
2C.
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' below