I am looking for a specific marker that can distinguish between quiescent and activated stem cells in Subventricular zone in the adult brain. Preferably, a marker that is not expressed in transit amplifying progenitor cells. I looked into TLX and Sox2, but they are expressed in both progenitors and stem cells. Long term BrdU labelling is another option, but it is time consuming. Is there a specific marker, or a group of markers, that can be used for this purpose?
Dear Osama,
I advice you to read these two papers, published by Barbara Demeneix:
1) Inhibition of Sox2 Expression in the Adult Neural Stem Cell Niche In Vivo by Monocationic-based siRNA Delivery.
2)Thyroid hormone signaling acts as a neurogenic switch by repressing Sox2 in the adult neural stem cell niche.
You will find the answer :)
Good luck
Pierluigi
Thank you Peirluigi, I will get on it. But based on what I know, I think SOX2 is expressed in both stem and progenitors. But I will take a look.
Thanks
Hi Osama, you could also look at our recent paper looking at transcriptional regulation of neural stem cell quiescence. http://www.ncbi.nlm.nih.gov/pubmed/23964093 Mainly carried out in cell culture, but we suspect the key mechanisms and marker genes will also apply in vivo. Good luck!
Hi Osama,
Unfortunately, no there is no one marker that will identify quiescent neural stem cells. You would have better luck trying 2 or more markers.
I had the same question once and it led me to write this:
http://www.sciencedirect.com/science/article/pii/S0301008213000658
Hope that helps!
- Carlyn
I suggest using a combination of Ki67 (proliferation marker), BrdU, and a marker specific to progenitor population. I recommend replacing traditional BrdU labeling with Click-iT EdU by Invitrogen. It is MUCH less of a pain to work with. Using this combination of markers will allow you to determine not only if your progenitors are quiescent, but also when they became quiescent. For example, lets say you treated your cells with EdU a week prior to immunocytochemical staining of Ki67 in addition to a marker that allows you to identify your cells of intrest out of the heterogeneous population. Cells that are EdU+/Ki67- would have been proliferating at the time of EdU treatment but became quiescent at some point during the subsequent 7 days prior to analysis. Cells that are EdU-/Ki68- were quiescent at the time of EdU treatment and remained that way. Likewise, cells that are EdU+/Ki67+ have been proliferative the whole time. Cells that are EdU-/Ki67+ were quiescent at the time of EdU treatment but became proliferative at some point during the subsequent 7 days prior to analysis.
I hope this helps. Good luck!
Hello Osama,
I did a quick Google search for "quiescent neural stem cell marker" and found a very good paper that was published in Nature (volume 437:894-897, 2005). These cells respond to Shh signaling. I will try to attach the article. I think you can find a genetic marker to accurately identify these cells in vivo.
Hi!
I don't have experience with this but recently I read this paper and according to the authors GFAP+ and S100B- are quiescent stem cells. Look here: (Kim et al., 2009, Adult Mouse Subventricular Zone Stem and Progenitor Cells Are Sessile and Epidermal Growth Factor Receptor Negatively Regulates Neuroblast Migration).
Ah! And look this too: Cell cycle and lineage progression of neural progenitors in the ventricular-subventricular zones of adult mice, Giovanna Ponti, Kirsten Obernier, Cristina Guinto, Lingu Jose, Luca Bonfanti, and Arturo Alvarez-Buylla, 2013.
Have a good luck!
Thank you all for your responses!! I guess my reading list has just doubled! I will get back to you all with more specific responses. I really appreciate all the help
There is not a definitive marker for quiescent neural stem cell population. One way to make sure you have a relatively purified quiescent NSCs is to use CFSE. Cells that are actively proliferating will dilute out the dye but the quiescent NSC will retain the dye and can be sorted out using FACS. For details of the protocol I refer you to the following publication by our group published by JOVE:
http://www.ncbi.nlm.nih.gov/pubmed/22565048
Good luck
Dear Osama,
You have a good luck! I found this paper, published in Genes and Dev (Ben Martynoga, as first author):
Epigenomic enhancer annotation reveals a key role for NFIX in neural stem cell quiescence
Have fun
Dear Pierluigi,
Thank you for your help on this. Ben Martynoga pointed that paper out. I will get round to reading it at some point. I am sure it will be fun
Thank you Ben Martynoga, Your paper gave me great insight. Adhesion molecules and ECM componets are most important for quiescence, but the in vivo data from the hippocampus show wide-spread expression for NFIX, contrary to in vitro data, have you tried sorting quiescent cells from the SVZ or the hippocampus to do the ChIP-Seq and RNA-Seq? The Kokovay 2012 paper was very insightful in the discussion.
Carlyn Mamber, I will recommend your review to the members of my lab, it is very important and well-written! I think EphA4 and VCAM-1 are the most promising.
Brooke DeRosa, This double labelling technique is very useful and we use it in the lab, however, it takes sometime to get the quiescent cells, but I guess I will need to use a multitude of markers and methods to answer my questions.
Thank you all for the useful articles, it was an interesting couple of week!
Hi Osama, I wanted to ask you if you found the right combination of markers to distinguish quiescent from active neural stem cells in the SVZ.
Hi Lara, I think people use EGFR to label activated NSC and TAPs, but VCAM-1 was used to label Quiescent cells both In Kokovay 2015 and more recently in Arturo's last paper on the origin of postnatal NSCs (Fuentealba et al., 2015, Cell 161, 1644–1655)
Possibly a combination of CD133, EGFR and VCAM-1 can distinguish activated from quiescent NSCs, I on the other hand, have not tried that combination.
I hope that helps
Osama
Hi Osama, Thank you for your advice. I will try GFAP+EGFR- and GFAP+VCAM-1+ on tissues. Hopefully one of the two combinations will work. Thank you again! Best wishes, Lara
Dear All, does anyone know of a good antibody for IF on mouse tissues that label the SVZ progenitors (e.g. EGFR, Mash1, etc...)? I have tried some antibodies but I was not very lucky. Thank you for your help, Lara
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