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WILT


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#1

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Posted 18 August 2005 - 06:05 AM


Aside from telomere maintenance telomerase reverse transcriptase appears to be also involved in stem cell mobilization suggesting that pertubations of its expression in normal tissues could have unforeseen consequences.

Effects of Telomerase and Telomere Length on Epidermal Stem Cell Behavior.
Flores I, Cayuela ML, Blasco MA.
Science. 2005 Jul 21

A key process in organ homeostasis is the mobilization of stem cells out of their niches. We show through analysis of mouse models that telomere length, as well as the catalytic component of telomerase, Tert, are critical determinants in the mobilization of epidermal stem cells. Telomere shortening inhibited mobilization of stem cells out of their niche, impaired hair growth, and resulted in suppression of stem cell proliferative capacity in vitro. In contrast, Tert overexpression in the absence of changes in telomere length promoted stem cell mobilization, hair growth, and stem cell proliferation in vitro. The effects of telomeres and telomerase on stem cell biology anticipate their role in cancer and aging.


Nature Reviews Molecular Cell Biology, published online 15 August 2005

More than just the 'maintenance man'

Shannon Amoils

In higher organisms, the proliferative and multipotent properties of adult stem cells enable the maintenance and regeneration of different tissues. These stem cells express high levels of telomerase, which is the ribonucleoprotein enzyme that extends telomeres during DNA duplication. As telomeres must be sufficiently long for cell division to take place, the increased activity of telomerase is probably one of the factors that endow these cells with their large proliferative capacity.

But does telomerase have other roles in the stem-cell compartment apart from the maintenance of chromosome ends? A group of scientists led by Maria Blasco at the Spanish National Cancer Center tackled this question by analysing the behaviour of epidermal stem cells in mouse models of defective telomerase expression.

The authors showed that, in G1 telomerase deficient (Terc-/-) mice, in which telomeres are only slightly reduced in length, and in G3 Terc-/- mice, which have critically short telomeres, epidermal stem cells accumulated in their niche in the bulge of the hair follicle. Cells from both G1 and G3 mice were unable to mobilize efficiently from the bulge and did not initiate appropriate hair growth in response to a proliferative stimulus. Also, the in vitro proliferative capacity of epidermal stem cells derived from both G1 and G3 Terc-/- mice was impaired.

By contrast, in K5-mTert mice, in which epidermal stem cells overexpress the protein component of telomerase, the hair follicle niche was depleted of stem cells and, compared to wild-type mice, an increased proportion of these cells was mobilized by a proliferative stimulus. These cells also showed a markedly increased capacity to proliferate in vitro.

Taken together, these results indicate that the role of telomerase is not merely to maintain chromosome ends — telomerase activity per se has a crucial role in regulating stem-cell turnover and mobilization. As Terc-/- mice have an ageing-resistant phenotype, and K5-mTert mice have a propensity to develop skin tumours, these effects of telomerase on stem-cell behaviour are crucial in the aetiology of both cancer and ageing.



#2 ag24

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Posted 18 August 2005 - 08:14 AM

Right - and this work is actually the main reason why Blasco is on the program for SENS2. Note also that the mice with deficient mobilisation are deficient for the RNA subunit (Terc), not the catalytic one, while the transgenic ones are overexpressing the catalytic one; this is strong evidence that the effect on mobilisation is indeed due to telomerase activity, rather than a telomere-elongation-independent function for one or other of the subunits (which has been proposed in other studies). However, the point to note is that mobilisation is something that should not happen too much or too little. Luckily, human stem cells already express only trace amounts of telomerase - it's the transit amplifying cells that express rather more - so deleting telomerase may make little difference. The most that can be said here is that this is another reason why WILT will be tricky to model in mice.

#3

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Posted 21 August 2005 - 06:43 AM

And here is another study reported in Nature on the intimacy between stem cell mobilization and telomerase.

Nature 436, 1048-1052 (18 August 2005)

Conditional telomerase induction causes proliferation of hair follicle stem cells

Kavita Y. Sarin, Peggie Cheung, Daniel Gilison, Eunice Lee, Ruth I. Tennen, Estee Wang, Maja K. Artandi, Anthony E. Oro and Steven E. Artandi

TERT, the protein component of telomerase1, 2, serves to maintain telomere function through the de novo addition of telomere repeats to chromosome ends, and is reactivated in 90% of human cancers. In normal tissues, TERT is expressed in stem cells and in progenitor cells3, but its role in these compartments is not fully understood. Here we show that conditional transgenic induction of TERT in mouse skin epithelium causes a rapid transition from telogen (the resting phase of the hair follicle cycle) to anagen (the active phase), thereby facilitating robust hair growth. TERT overexpression promotes this developmental transition by causing proliferation of quiescent, multipotent stem cells in the hair follicle bulge region. This new function for TERT does not require the telomerase RNA component, which encodes the template for telomere addition, and therefore operates through a mechanism independent of its activity in synthesizing telomere repeats. These data indicate that, in addition to its established role in extending telomeres, TERT can promote proliferation of resting stem cells through a non-canonical pathway


Aside from the obvious suggestion to also be associated with a possible cure for balding we see that telomerase may harbor the until now unknown function of increasing regeneration via promotion of stem cell proliferation.

#4 ag24

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Posted 21 August 2005 - 11:34 AM

> increasing regeneration via promotion of stem cell proliferation

No no, this is a common misunderstanding. Stem cells are not supposed to divide too often -- they divide much more slowly than the progenitor (i.e. partially differentiated) cells that they produce. They also, surprise surprise, express less telomerase than progenitor cells. The reasons why stem cells divide very slowly are (as is the way with metabolism in general) probably much more numerous than we know, but even what we know is suggestive that this is important: DNA replication is mutagenic so should be done as little as possible; DNA repair takes time so must be allowed time; cell division takes energy so rapid division may ramp up OXPHOS and thus ROS production. Thus the strong prediction is that overexpression of telomerase in stem cells will not increase regeneration at all in the long run, but rather cause depletion of stem cell pools in the long run by making stem cells die or differentiate in response to self-inflicted and unrepaired damage. Again, this is good news for WILT, because human stem cells express so little telomerase already that expressing none at all may make little difference to them - they have evidently already got ways (which we have not yet discovered) to keep their stemness with less telomerase than is present in mouse cells.

#5

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Posted 21 August 2005 - 03:01 PM

> increasing regeneration via promotion of stem cell proliferation

No no, this is a common misunderstanding. Stem cells are not supposed to divide too often -- they divide much more slowly than the progenitor (i.e. partially differentiated) cells that they produce.

There are numerous tissues that rely on a certain rate of cell replenishment, which if not met results in impairment of their function. In such cases the rate of stem cell (and/or progenitor as you mentioned) proliferation is a limiting factor. In aging tissues a depletion in the rate of stem cell replicative potential is observed. Is it not likely that a means of upregulating the proliferative potential in stem cell niches whose reserves are nearing age-related exhaustion would help in extending functional lifespan in those tissues?

Thus the strong prediction is that overexpression of telomerase in stem cells will not increase regeneration at all in the long run, but rather cause depletion of stem cell pools in the long run by making stem cells die or differentiate in response to self-inflicted and unrepaired damage.

This implies that the most rapidly dividing cells should be the most vulnerable to genomic damage, but this is not observed.


human stem cells express so little telomerase already that expressing none at all may make little difference to them

Is this speculation or are you aware of a way where stem cells can prevent telomeric degradation sans telomerase?

#6 ag24

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Posted 21 August 2005 - 06:10 PM

> Is it not likely that a means of upregulating the proliferative potential in stem cell niches
> whose reserves are nearing age-related exhaustion would help in extending functional
> lifespan in those tissues?

Proliferative potential, yes; proliferative rate, probably not, for the reasons I gave.

> This implies that the most rapidly dividing cells should be the most vulnerable to genomic
> damage, but this is not observed.

In what way is it not observed? Clearly one must take into account not only the rate at which damage is inflicted but the rate at which it is eliminated, whether by death of differentiated cells (e.g. skin cells) or by selection among stem cells.

> > human stem cells express so little telomerase already that expressing none at all may
> > make little difference to them
>
> are you aware of a way where stem cells can prevent telomeric degradation sans telomerase?

Of course I'm not aware of such a way. I'm saying that human stem cells express very little telomerase, only just enough to compensate for rather rare stem cell division. Obviously by "little difference" I was talking about regenerative capacity, not about indefinite survival, since WILT proposes periodic replenishment.

#7

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Posted 22 August 2005 - 09:37 AM

In what way is it not observed? Clearly one must take into account not only the rate at which damage is inflicted but the rate at which it is eliminated, whether by death of differentiated cells (e.g. skin cells) or by selection among stem cells.


Jay made an interesting post on the advantages of higher stem cell division rates here.

As you know, the advantages that a more rapidly dividing cell has in terms of genomic stability over slower counterparts include decreased concentration of ROS and other harmful aggregates due to dilution, increased turnover of critical organelles such mitochondria and access to DNA repair pathways that are only available during mitosis. The small intestine rarely develops cancer despite the substantial mount of stem cells produced to maintain the mucosal lining (1).


(1) Journal of Cell Science 115, 2381-2388 (2002)
Intestinal stem cells protect their genome by selective segregation of template DNA strands
Christopher S. Potten, Gary Owen and Dawn Booth

#8 ag24

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Posted 22 August 2005 - 09:41 PM

> Jay made an interesting post on the advantages of higher stem cell division rates here.

Heh - I must have read some of that thread at the time and not had time to reply - I was travelling a lot at that time. The logic is nice, but there are a couple of problems with it. First, the later parts of the differentiation pathway are neither stem cells nor totally differentiated and non-dividing cells but intermediate cells called "transit amplifying" cells, whose daughter cells on division are both more differentiated than the pre-division cell was. Second, stem cell divisions don't necessarily give rise to one daughter that is at the same state as the pre-division cell was and one that is more differentiated: symmetrical ("self-renewal") divisions can take place in which both daughters are just as stemmish as the pre-division cell was. Indeed, most people's models of the cell dynamics of the gut (including Potten's, though not mine) are that all stem cells are of equal stemmishness. This all means that DNA repair can be really quite sloppy once the transit amplifying cell stage is reached, because all progeny of the cell in question will be history quite soon. The original six-cell model of Potten (which was actually 4-8 cells - he never said exactly 6) needed to be revised, because evidence arose (and is now decisive) that most crypts are monoclonal, i.e. all cells in the crypt derive from a relatively recent single common ancestor cell; this is no problem when stem cells are replaced by self-renewal divisions, of course. Self-renewal divisions also make a bit of a mess of the "immortal strand" hypothesis, which is the core of the JCS paper you mention -- it only works if essentially all divisions are asymmetrical. Nonetheless, the disparity of cancer incidence in different parts of the intestine is certainly of interest. But the idea that cells can have high turnover but be kept cancer-free by death and replacement is not correct, because it only takes a mutation that cheats the system by turning off the apoptosis-sensitivity and then subsequent mutations will be able to create a cancer.

#9 John Schloendorn

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Posted 03 September 2005 - 08:16 AM

Has anyone actually made a direct comparison between late generation Tert-/- and Terc-/- phenotypes? Prometheus' view might predict that Terc-/- is less severe, while Aubrey's view might predict that Terc-/- might initially seem less severe but would become more severe in late life, right?

#10 jaydfox

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Posted 06 September 2005 - 02:40 PM

While we're on the subject of TERT and TERC:
http://www.futurepun...967.html#002967

The ability to turn on adult stem cell division by knocking out one component of telomerase while increasing the availability of another teolomerase component simultaneously increases the supply of adult stem cells available for stem cell therapy and decreases the odds of getting cancer from stem cells.

To be certain it was TERT alone that was triggering the stem cells, Artandi's team crossbred mice to eliminate the presence of the RNA component, TERC. Since TERC is crucial to rebuilding telomeres, when the mice grew shaggy even with no TERC around, it was clear the follicle stem cell stimulation was due solely to TERT and that the telomere repair function of telomerase played no role.

"This is really an unanticipated effect for TERT, one that's independent of the conventional telomerase complex," Artandi said.

I see this having some exciting implications for WILT, at least for the WILT scaffolding, if not the whole package.

#11 henri

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Posted 14 September 2005 - 12:59 PM

As an aside, I am somewhat unclear on the effects of WILT on the immune system. As the telomeres are depleted, memory cells can't expand. In his discussions on this, Aubrey de Grey has stated that it wouldn't matter, as the body and the immune system would work better on the whole, and young people are better at fending off disease even if not ecquipped with memory cells yet. This baffles me, as I was under the impression that vaccinations and immunization as a whole are based on memory cells.

#12 jaydfox

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Posted 14 September 2005 - 01:24 PM

This baffles me, as I was under the impression that vaccinations and immunization as a whole are based on memory cells.

Well, we can immunize the stem cell regular replacements against the standard battery of immunizations, achieving the same effect...

#13 Michael

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Posted 15 September 2005 - 02:01 PM

All:

henri: As an aside, I am somewhat unclear on the effects of WILT on the immune system. As the telomeres are depleted, memory cells can't expand. In his discussions on this, Aubrey de Grey has stated that it wouldn't matter, as the body and the immune system would work better on the whole, and young people are better at fending off disease even if not ecquipped with memory cells yet. This baffles me, as I was under the impression that vaccinations and immunization as a whole are based on memory cells.

jaydfox: Well, we can immunize the stem cell regular replacements against the standard battery of immunizations, achieving the same effect...

Right: remember that Aubrey's reading of the evidence (1,2) suggests that we will get a pretty solid decade out of each seeding, so that we will have plenty of time to administer vaccines per a standard schedule (or perhaps do something with the cell clones ex vivo).

In addition, the removal of memory cells will itself have rejuvenative effects on the immune system.

Dysregulated T cell-mediated immunity contributes materially to the increased susceptibility to infectious disease ... in the elderly. One hallmark of this state of "immunosenescence" is the predominance of large clones of peripheral T cells with limited antigen receptor heterogeneity and a corresponding contraction of diversity in the T cell antigen recognition repertoire.  ...  The human immune system is thus subjected to chronic antigenic stress caused by persistent activating virus, which can never be fully eliminated. This constant stimulation results in clonal exhaustion of the CMV-specific cells, a state of anergy which is similar in many respects, and, we believe for similar reasons, to that commonly seen in the tumour-specific T cells of cancer patients. The problem which we have identified in a subset of the elderly ... is not a lack of functional ... cells, but an enormous excess of persisting dysfunctional ... cells.

That is, these cells are apoptosis-resistant, anergic, and are passively suppressive by filling the "immunological space", as well as potentially actively suppressive via cytokine secretion. The accumulation of these dysfunctional cells is, we believe, one of the major problems in the elderly immune system. Moreover, because these cells by persisting maintain overall T cell numbers in the periphery, T cell homeostasis mechanisms feed back to the thymus to prevent the production of naive functional T cells specific for other antigens. This, coupled with the age-associated process of thymic involution, results in shrinkage of the T cell antigen repertoire in the elderly, and contributes to increased susceptibility to infections, especially to newly-arising infectious agents not experienced by that individuals "young" immune system in the past...

For those already at a more advanced age ... treatment targeting the dysregulated apoptosis-resistant CMV-specific cells would be an option. This would require distinguishing the functional CMV-specific cells (which are essential to maintain immunosurveillance) from the dysfunctional ones. We have discovered certain surface molecules on dysfunctional cells which may enable us to achieve this aim. Capitalising on technology now essentially routinely applied in hematopietic stem cell transplantation, functional anti-CMV effector cells could be first isolated and expanded in vitro, prior to re-infusion into the depleted individual, as a safety measure to ensure and enhance CMV immunosurveillance during and after specific depletion [emphasis mine]. (3)


(And see (4)). Pawalec et al are advocating a targeted deletion of the anergic CMV clones because f the evidence that dysfunctional CMV-targeted clones are a relatively widespread, specific problem in the elderly, but (a) there are doubtless less common and/or less severe variations on this theme (elsewhere (4), they list Epstein–Barr virus (EBV) and Varicella (the herpesvirus that cuases shingles) as other potential cases) and (b) this specificity is only necessary because we don't want to wipe out the rest of a person's immunological complement under normal circumstances -- but if (as under WILT) we will allow all T-cell clones to die, sequentially, and replace them with new cells, every decade or so, then the problem will never arise in the first place for any pathogen.

-Michael

1. de Grey AD, Campbell FC, Dokal I, Fairbairn LJ, Graham GJ, Jahoda CA, Porterg AC.
Total deletion of in vivo telomere elongation capacity: an ambitious but
possibly ultimate cure for all age-related human cancers.
Ann N Y Acad Sci. 2004 Jun;1019:147-70. Review.
PMID: 15247008 [PubMed - indexed for MEDLINE]
http://www.gen.cam.ac.uk/sens/WILT.pdf

2. de Grey AD.
Whole-body interdiction of lengthening of telomeres: a proposal for cancer prevention.
Front Biosci. 2005 Sep 1;10:2420-9.
PMID: 15970505 [PubMed - in process]
http://www.gen.cam.a...ns/WILT-FBS.pdf

3. Graham Pawelec [1], Sven Koch [1], Cécile Gouttefangeas [2] and Anders Wikby [3] .
Immunorejuvenation in the elderly.
Rejuv Res. 2005 Sep;8(Suppl 1):S44-5(Abs 88).
http://www.gen.cam.a...abs/Pawelec.htm

4. Pawelec G, Akbar A, Caruso C, Effros R, Grubeck-Loebenstein B, Wikby A.
Is immunosenescence infectious?
Trends Immunol. 2004 Aug;25(8):406-10. Review. No abstract available.
PMID: 15275638 [PubMed - indexed for MEDLINE]
http://dx.doi.org/10....it.2004.05.006

#14 John Schloendorn

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Posted 15 September 2005 - 05:42 PM

Hmm, as far as I understand you Henri, your concern is not merely the destruction of memory cells, but also the inability of lymphocytes in general to expand greatly without telomerase? Concerns about the ability of rapidly renewing tissues (blood, gut, lung ect.) to continue functioning in the absence of telomerase are not new. Were you suggesting that frequently challenged T-cell clones might be the first such tissue to be affected? If so, then vaccines might not do as much good as they do in the presence of telomerase, because they rely on the repeated expansion of the same clones.
Phenotypical studies in the telomerase-KO mouse would have missed this problem, because they did not perform adequate immunological tests (vaccination, repeated infectious challenge). I tend to think that youthful levels of naive thymopoiesis might compensate for such a problem to a degree. I am not sure how to make a quantitative prediction out of this idea, but I think it would deserve a second look.

#15 ag24

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Posted 15 September 2005 - 06:03 PM

I addressed this topic in the more recent WILT paper (Frontiers in Bioscience) - sections 6.3 and 9. Note the bit about telomere-elongating memory cells themselves.

#16 henri

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Posted 18 September 2005 - 03:26 PM

remember that Aubrey's reading of the evidence (1,2) suggests that we will get a pretty solid decade out of each seeding, so that we will have plenty of time to administer vaccines per a standard schedule (or perhaps do something with the cell clones ex vivo).


Yes, but as ag24 has noted, memory cells (formed in response to vaccination) might not have enough telomerase left to expand in response to the germ when it appears. "What would be prevented, ... is re-expansion of memory cells on re-exposure to the same antigen" (Frontiers in Bioscience article).

Hmm, as far as I understand you Henri, your concern is not merely the destruction of memory cells, but also the inability of lymphocytes in general to expand greatly without telomerase?


Yes.

Were you suggesting that frequently challenged T-cell clones might be the first such tissue to be affected?


According to what the Frontiers in Bioscience article says, memory cells could not do any good (unless one equips them with extra long telomeres, as suggested by ag24).

I addressed this topic in the more recent WILT paper (Frontiers in Bioscience) - sections 6.3 and 9. Note the bit about telomere-elongating memory cells themselves.


Okay, elongating telomeres would solve the problem.

The article says:

"Memory cells are essential for middle-aged
and elderly adults only because their advancing age renders
them more likely to succumb more quickly to a given
infection than a young adult would, and thus reliant on
memory cell expansion to eliminate the infection quickly.
Young people do not have memory cells for many antigens,
because they have not been exposed to many, so they rely
on their overall youthfulness to maintain function while the
slower naïve cell response builds up. Yet, young adults are
more resistant to death from infectious diseases than older
people"

But can even young people do without memory cells if we don't want them to get the disease in the first place, as is the case when vaccinations are given? No one can fend off something like polio without memory cells, right? But sure enough, if we could extend telomeres in vitro, the problem would be solved.

edit: quote tags

Edited by prometheus, 19 September 2005 - 02:03 AM.


#17 Mind

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Posted 23 October 2008 - 05:17 PM

Researchers uncover new links between stem cells, aging and cancer.

The three-year study of mouse brain cells also helped explain why human adult stem cells can't match the embryonic stem cell's potential to regenerate damaged tissues in patients, Morrison said.

"The genes identified in this study work together to reduce the function of adult stem cells as they age," he said. "Embryonic stem cells offer the advantage of not aging, not turning on this pathway. If you need to generate large numbers of cells to treat a major public health problem—such as juvenile diabetes—this is a big advantage."

The four genes examined in the study were Ink4a, Arf, Hmga2 and let-7b. The work involved breeding mice that lacked combinations of these genes, then measuring the effects on stem-cell function and brain-cell formation at different life stages.


I am not an expert on the WILT strategy but I understand enough to know this is an important finding. WILT is to address cancer and thus a deeper understanding of the genetic regulatory pathways is crucial. For those who haven't read about aging, cancer, and WILT, this would solidify the thesis that the process body shutting down stem cell action in old age is a defense mechanism against cancer. The author of this article makes this point:

The genes switch on and off in a coordinated fashion as cells age to reduce the risk of cancer. In the process, they also shut down stem-cell function in aging tissues, reducing their capacity to regenerate.


So if we had a 100% sure-fire cure for cancer (outside of WILT), then one simple strategy to reverse some of the effects of aging would be administer growth factors and other stem cell signaling chemicals. If you would happen to get cancer as your tissues regenerate - so what - there would be the sure-fire cancer treatment to help you out. Pardon me if I am making some grave error by being so simplistic. Nishino probably says it better:

Two years ago, Morrison's team demonstrated that Ink4a, well known for its role as a tumor suppressor, becomes increasingly active with age and shuts down stem-cell replication in older mice. Flicking that genetic switch likely serves as a defense against cancer-causing genetic mutations, which accumulate as cells repeatedly divide.

The main question remaining after the 2006 Nature paper was: What causes Ink4a to turn on with age?

In the new Cell paper, the U-M researchers show that Ink4a's activity in mouse neural stem cells is regulated by Hmga2, which in turn is controlled by let-7b. The same relationship is likely at work in humans, who possess the same four genes.

"The tumor-suppressor mechanisms ramp up with age," Morrison said. "And the good news is that it allows us to get older before getting cancer. The bad news is that your tissues lose their regenerative capacity, making you older



#18 Mind

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Posted 23 October 2008 - 05:31 PM

Other WILT related discussion at Imminst:

WILT light - using WILT only after the onset of cancer.

Should WILT be given a higher priority

Introduction to WILT

Gut stem cells suggest we can do better than WILT




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