Review for 10.1186/1471-230X-8-57 (2008)

Reviewer's report
Title:Wnt-reporter expression pattern in the mouse intestine during homeostasis
Version: 1Date:13 August 2008
Reviewer: Stefano Piccolo
Reviewer's report:
This is an interesting report on beta-catenin activity in intestinal crypts that should
attract the readers of this journal.
The main novelty is the demonstration of beta-catenin-dependent transcription in
the colon/SI epithelium. This is unprecedented but also bears some surprises
that are not sufficiently highlighted by the discussion/presentation.
They show that a remarkably limited number of crypts is positive (with the criteria
to be counted as positive being the presence of one single positive cell).
The authors should discuss more critically why 85% of the small intestine and
99.2 (!) % of the colon crypts remain completely unstained. This is inconsistent
with the broad distribution of beta-catenin protein staining in the nucleus of cells
at the bottom of the crypts.
These are some ideas:
a) the biology of the small intestine and colon is different in term of Wnt
responsiveness and requirement for stem cells/homeostasis, at least in mice.
b) Beta-catenin may well enter in the nucleus after Wnt stabilization but this is
insufficient to trigger gene transcription. Provide as examples the known nuclear
restraining factors for the beta-catenin/TCF complex.
c) There may be another pathway, still unknown, that is permissive for Wnt
mediated transcription (through the canonical pathway)
The fact that the beta-gal staining of TOP-gal is localized in differentiated Paneth
cells is intriguing and puzzling at the same time. The authors should at least
discuss the possibility that Wnt may be involved in normal colon homeostasis but
only indirectly, namely by inducing an unknown factor (nurturing, as an example,
the interspersed LRG5 columnar stem cells).
Page 11. there is a mistake in the interpretation of the data. Finding cells positive
for lacZ and BrdU can be coincidental. By no mean it shows a common lineage.
This is also a point raised also later in the MS (discussion), but it is a conceptual
mistake.
Page 12. from a naive point of view: if the stem cells are retaining the label, how
can the Wnt responsive cells be “differentiated” Paneth?. Please discuss this for
the general audience.
Page 13: after tissue damage there is remarkable raise in lacZ activity. How does
this translate in numbers of positive crypts and average number of cells positive
for each crypt? The authors give total lacZ quantification whereas in the previous
Figures they offer histological stainings. It looks like that they are hiding
something, but there is no need of this. Readers really want to know if is the
massive 140 fold (!) increase corresponds to a massive transcription in just few
cells etc... Is there an increase in the stromal cells as well? In maretto et al, we
saw positive cells in the mesenchyme. Quantifications are essential for this point
Other important points.
the authors should show few pictures of BAT-gal as well.
It is important to know what is the ratio of positive crypts displaying cells at the
bottom vs those with cells in the TA or differentiating compartment. This is an
important point that is unclear from the present MS.
the conclusion is trivial and out of focus.
Level of interest: An article of importance in its field
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'