D*******8
发帖数: 4 | 1 Please don’t dig me out. I am writing this out of my conscience as a
scientist. We are using, largely, the tax payers’ money to explore
mechanisms underlying diseases that we don’t have a cure, yet. We are doing
this in the hope that little bit knowledge can accumulate till one day we
can find that magic bullet. However, there are people out there, their
purpose is to use these money to achieve their personal fames, sometimes
international ones, totally disregarding our original noble cause. I am
doing this because I believe a wrong message has to be corrected if it is
wrong, but I found I am now in no place to do that, I am expose this here in
the hope that somebody with power, can do this for us.
I am working in the inflammasomes field, where adaptor molecule ASC plays a
pivotal role by bridging interactions between NLR family members and the
proinflammatory Caspase-1. However, that paradigm was changed recently. In a
paper published in the 10th issue of volume 12 of Nature Immunology,
Ippagunta et al. claimed that they have identified an additional and
unexpected mechanism by which ASC, a core component of inflammasomes, can
control adaptive immune responses via regulation of the expression of the
gene encoding Dock2. The authors first demonstrated that primed dendritic
cells from Asc deficient mice are defective for uptaking of particulate
antigens. In addition, Asc deficient mice showed lower number of
lymphocytes as well as myeloid cells in peripheral lymphoid organs. More or
less, the phenotype presented in Asc deficient mice recapitulated the ones
discovered in Dock2 deficient mice. Gene expression array analysis followed
by Western blotting did indicate that Dock2 protein expression was abrogated
in Asc deficient lymphoid and myeloid cells. The authors further provided
evidence that messenger RNA for Dock2 has a much-shortened life span than
its wild type counterpart. Finally complementation of Asc deficient cells
with Dock2 protein did have rescued immune cell functions. The authors
concluded that they have identified a critical and unexpected role of Asc in
regulating the motility of lymphocytes and professional antigen presenting
cells independently of inflammasomes.
While the data presented in this paper are largely not flawed, the
interpretation of these data are quite questionable since the authors did
not provide evidence regarding complementation of Asc deficiency by forced
expression of ectopic Asc protein within these cells to determine whether
ASC regulates Dock2 expression.
We have the same Asc deficient mouse strain as the one the authors used in
their publication, which was originally generated by Millennium
Biopharmaceuticals. Inc. We have established stable bone marrow derived
macrophage cell lines (BMDM) from the original mouse line when we just
imported the mouse line. After a SNP analysis, we were not satisfied with
the amount of C57BL/6 genetic background in the mouse line and decided to
further back-cross them to B6 mice for two more generations.
We set out to explore the mechanism how Asc regulate Dock2 expression. When
we look at the stable BMDM Asc deficient cell line, we find that Dock2
protein expression is indeed absent. We took these cell lines and transfect
them with Asc expressing retroviral vectors. Ectopic Asc expression was
confirmed by Western blot, and pro-IL-1 processing, a hallmark of
inflammasomes activity, was restored by re-expression of Asc in these cells.
However, Asc protein re-expression was not able to rescue Dock2 expression
in these cells. We reasoned that the immortalization process by which we
have generated the cell line might have caused some changes that prevent
Dock2 re-expression after Asc complementation. We established primary BMDM
cultures with bone marrow cells from mice in our current Asc -/- mouse
colony. We were planning to work on these primary cells with the Asc
complementation experiment when, to our surprise, we found that these
primary cells are sufficient for Dock2 expression in the absence of Asc. We
took additional mice from our Asc-/- colony and found every one of them are
Asc deficient but Dock2 sufficient.
While this finding is surprising, we believe what really happened is that
the Asc-/- mouse strain originally from Millennium was a compound mutant
strain with mutations in both Asc and Dock2 locus. The nature of the Dock2
mutation in these mice remains mysterious to us. Our BMDM stable cell line
was established from the original Millennium mouse line, as a result these
cells are also deficient for Dock2 expression. However, after backcrossing
to B6 mice for two generations, that mutant Dock2 allele was segregated from
Asc deficient allele in our current Asc-/- mouse strain since Dock2 and Asc
encoding genes are located on different chromosomes. This explains why
reconstitution of Asc-/- cell lines with Asc protein expression vector
cannot rescue Dock2 expression.
It was a genuine mistake on the authors’ side, since they did not provide
complementation experimental data. It is also a mistake on the reviewers’
since they probably did not ask for such data. However, I believe we have
solid scientific evidence that the phenomenon observed by Ippagunta, et al
was no more than an artificial effect in an accidentally generated compound
mutant mouse line, thus the whole message in that paper does not stand.
It is truly an unfortunate situation for the authors, however, it is more
important to inform the community that they have made a mistake instead of
covering it up and let the wrong message be there to misled other scientists
or even finding its way into textbooks for the next generation. | d***y
发帖数: 8107 | | s*********t
发帖数: 600 | 3 。。。。。lz精神状况还好吗？
为何贴在这里，不跟你的supervisor讨论 | D*******8
发帖数: 4 | 4 I am fine. The result has been extensive discussed and a decision has been
made that we are not going to take any action since the senior author has
just reviewed one of the PI's grant in the department, and she is likely to
review our papers and grants in the future. Anyway, there are already a lot
of trash there in the literature, why bother with one more. | b*******n
发帖数: 8420 | 5 没办法，人家也要讨生活啊
Anyway, there are already a lot
【在 D*******8 的大作中提到】
: I am fine. The result has been extensive discussed and a decision has been
: made that we are not going to take any action since the senior author has
: just reviewed one of the PI's grant in the department, and she is likely to
: review our papers and grants in the future. Anyway, there are already a lot
: of trash there in the literature, why bother with one more.
| g********0
发帖数: 6201 | | S**********e
发帖数: 1789 | 7 赞“上访”。
【在 g********0 的大作中提到】
: LZ这是到生物版上访来啦？
| c********g
发帖数: 1106 | 8 不能在杂志上论仗么？来回几次，大家都看得过瘾。
生物研究，不就是盲人摸象么？谁嗓门大，谁就控制象的样子。很多时候，很难肯
定谁对谁错，是不是？
doing
in
【在 D*******8 的大作中提到】
: Please don’t dig me out. I am writing this out of my conscience as a
: scientist. We are using, largely, the tax payers’ money to explore
: mechanisms underlying diseases that we don’t have a cure, yet. We are doing
: this in the hope that little bit knowledge can accumulate till one day we
: can find that magic bullet. However, there are people out there, their
: purpose is to use these money to achieve their personal fames, sometimes
: international ones, totally disregarding our original noble cause. I am
: doing this because I believe a wrong message has to be corrected if it is
: wrong, but I found I am now in no place to do that, I am expose this here in
: the hope that somebody with power, can do this for us.
| F******p
发帖数: 2099 | 9 如果你们的新asc-/-BMDM能重复已发表的phenotype,那人家的文章一半数据是正确的。
另外如果dock2是假象的话，为什么人家data显示dock2能rescue asc-/- phenotype? | b******n
发帖数: 4225 | 10 呵呵，生物研究确实像盲人摸象
现在还在observation->explanation的阶段
至于这个explanation是不是可靠，参照中医对疾病的理论:)
【在 c********g 的大作中提到】
: 不能在杂志上论仗么？来回几次，大家都看得过瘾。
: 生物研究，不就是盲人摸象么？谁嗓门大，谁就控制象的样子。很多时候，很难肯
: 定谁对谁错，是不是？
:
: doing
: in
| | | b*******n
发帖数: 8420 | 11 上访这个比喻很贴切啊。。大多数上访的都是走正常程序走不通，被逼得上访的。
可惜上访的地方不对，应该去CNS的编辑部或者NIH门口上访。
【在 g********0 的大作中提到】
: LZ这是到生物版上访来啦？
| D*a
发帖数: 6830 | | d********m
发帖数: 3662 | 13 猛啊，能看完并且抓住中心思想
【在 F******p 的大作中提到】
: 如果你们的新asc-/-BMDM能重复已发表的phenotype,那人家的文章一半数据是正确的。
: 另外如果dock2是假象的话，为什么人家data显示dock2能rescue asc-/- phenotype?
| S***a
发帖数: 257 | 14 To fatsheep:
You gave a nice comment. I have the same question here.
If LZ suspect the phenotype is from Asc-/-Dock2-/- compound mutant mice,
then it explained why DOCK2 re-induction can rescue the ASC-/- (Asc-/-Dock2-
/- actually) phenotype.
If LZ have already get pure B6 background Asc-/- mice, then did you check
the phenotype of antigen presentation and DC migration? Does LZ's current
Asc-/- BMDCs have the same phenotype as NI paper reported? If LZ also see
the phenotype consistent with the NI paper, then the main point of the NI
paper is well supported. | D*******8
发帖数: 4 | 15 The NI paper reported that Asc-/- mice phenocopies Dock2-/- mice.
They concluded that Asc regulated Dock2 expression through controlling Dock2
mRNA stability.
They failed to provide Asc complementation experimental data which is
considered critical in a genetic approach.
We have the same mouse strain, but back crossed to B6 for couple more times.
We found that Asc-/- mice are no longer deficient for Dock2 and the defect
in lymphocyte migration and antigen presentation is gone in our mouse strain.
Our conclusion: the NI paper was reporting phenotypes observed in a Asc/
Dock2 double mutant, and the conclusion that Asc regulates Dock2 expression
is not true.
It is complicated and it took hours to read the original publication, which
most of the readers wouldn't like to spend to get to the bottom of this
issue. However, jumping to the wrong conclusion and misleading the others is
terrible. This really tells me why 95% of post-docs didn't make it to
independence eventually. | D*******8
发帖数: 4 | 16 To Sonma,
To answer your question:
Yes we did the same experiment, since the Asc-/- mice we have are sufficient
for Dock2, so there is no defect in lymphocyte migration or antigen
presentation in these mice. We have communicated with the authors already,
they have all the data we have and I would hope that the senior author will
do the right thing out of conscience. |
|
