Review for 10.1186/1471-230X-13-90 (2013)

Reviewer's report
Title:TNF-alpha and LPA promote synergistic expression of COX-2 in human
colonic myofibroblasts: role of LPA-mediated transactivation of upregulated
EGFR
Version: 1Date:26 October 2012
Reviewer: Bernd Fiebich
Reviewer's report:
In the manuscript entitled “TNF-alpa and LPA promote synergistic expression of
COX-2 in human colonic myofibroblasts: role of LPA-mediated transactivation of
upregulated EGFR” Yoo et al. studied whether EGFR signalling plays a role in
the synergistic COX-2 expression induced by LPA and TNF-alpha. They found
that chronic exposure to TNF-alpha led to upregulation of EGFR in association
with sustained LPA-mediated EGFR phosphorylation at Y1068. TNF-alpha #and
LPA also led to sustained p42/44 MAPK phosphorylation and synergistic COX-2
expression, effects that were partially inhibited by the EGFR tyrosine kinase
inhibitor AG1478. The authors furthermore demonstrate that p42/44 MAPK
phosphorylation and COX-2 expression were inhibited to the same degree by the
MMP inhibitors GM6001 and BB-94, suggesting that LPA-mediated EGFR
transactivation involved MMP-mediated release of EGFR ligands from the cell
surface. Using the Src3 inhibitor SU6556, they found inhibition of
TNF-alpha/LPA-mediated EGFR phosphorylation at Y1068, p42/44 MAPK
phosphorylation, and COX-2 expression suggesting an upstream role of Src in
the transactivation of EGFR.
Although, the authors show very interesting data concerning the TNF/LPA/EGF
signalling in colonic fibroblasts, the following points need to be addressed:
Major points:
1) It would be very helpful for the reader of this manuscript if the data are
summarized in a signalling pathway diagram showing the crosstalk between the
receptors and signalling pathways.
2) COX-2 is one of the key enzymes of the prostanoid synthesis. However, there
are also other enzymes involved such as COX-1 and for the synthesis of PGE2,
the prostaglandin E synthases such as mPGES-1. It would be interesting to see,
whether the expression of those enzyme are regulated by TNF/EGF/LPA.
3) The most interesting prostanoid in respect to gut inflammation is PGE2. The
expression and regulation of COX-2, most likely but not necessarily, results in the
induction PGE2. Especially if the expression, like shown via TNF, is quite low,
PGe2 levels might not be affected. Data showing the release and synthesis of
PGE2 after stimulation with TNF/LTA/EGF would strengthen the conclusion in
respect to gut inflammation.
4) What was the statistics used in the manuscript to calculate the significance of
the densitographs?
5) The statement, that SU6656 inhibited p42/44MAPK and COX-2 in the same
degree then AG, GM and BB is not true by looking on the Western blots. More
representive Western blots should be shown, which fit to the densitographs.
6) Alpha-SMA is not the appropriate control for phosphorylated p42/44 MAPK. An
antibody recognizing total p42/44 MAPK should be used to avoid false positive
effects due to increased p42/44 MAPK protein levels.
Minor points
1) Please provide the specific information about the antibodies and the dilution
used for the Western blots. There are more antibodies for each target in each
company and the reader needs to know, which one was used.
2) In the densitograph in Fig. 1A, the time line should be hours instead of
minutes.
Level of interest: An article of importance in its field
Quality of written English: Acceptable
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