The Quintessence of Quiescence
Authors: Odessa Yabut, Samuel J. Pleasure
Journal: Neuron (2014)

Neurogenic niches in the adult mammalian brain are composed of heterogeneous populations of neural stem cells. In this issue of Neuron, Codega et al., 2014Codega P. Silva-Vargas V. Paul A. Maldonado-Soto A.R. DeLeo A.M. Pastrana E. Doetsch F. Neuron. 2014; 82 (this issue): 545-559Abstract Full Text Full Text PDF PubMed Scopus (389) Google Scholar isolate quiescent neural stem cells from the adult subventricular zone and demonstrate their stem cell characteristics. Neurogenic niches in the adult mammalian brain are composed of heterogeneous populations of neural stem cells. In this issue of Neuron, Codega et al., 2014Codega P. Silva-Vargas V. Paul A. Maldonado-Soto A.R. DeLeo A.M. Pastrana E. Doetsch F. Neuron. 2014; 82 (this issue): 545-559Abstract Full Text Full Text PDF PubMed Scopus (389) Google Scholar isolate quiescent neural stem cells from the adult subventricular zone and demonstrate their stem cell characteristics. Adult neurogenesis is responsible for the continuous generation and integration of new neurons in existing neural networks. These neurons are derived from multipotent neural stem cells (NSCs) residing in specialized neurogenic niches. The subventricular zone (SVZ) is the largest of these niches in the adult brain and is capable of generating at least 30,000 new neurons in rodents that then migrate through the rostral migratory stream (RMS) to integrate into networks in the olfactory bulb (OB) (Lois and Alvarez-Buylla, 1994Lois C. Alvarez-Buylla A. Science. 1994; 264: 1145-1148Crossref PubMed Scopus (1958) Google Scholar). One of the characteristics of tissue stem cell niches is the presence of quiescent stem cells, but unfortunately these quiescent cells are usually difficult to study since attempts to culture them or isolate them usually lead to activation. The study by Codega et al., 2014Codega P. Silva-Vargas V. Paul A. Maldonado-Soto A.R. DeLeo A.M. Pastrana E. Doetsch F. Neuron. 2014; 82 (this issue): 545-559Abstract Full Text Full Text PDF PubMed Scopus (389) Google Scholar in this issue of Neuron provides new insights into the nature of quiescent NSCs in the SVZ and also important tools that will facilitate future studies of these cells. In the SVZ, type B1 cells make up the primary NSC population; they divide to either self-renew or produce a secondary progenitor population, the intermediate progenitors (transit-amplifying or type C cells) that generate various neuronal subtypes destined for the OB, as well as progenitors that generate glial cell types (see Figure 1). Type B1 cells express glial fibrillary acidic protein (GFAP+) and maintain a bipolar morphology, with specialized apical processes that extend to receive signals from the CSF in the lateral ventricles and basal processes that maintain contact with blood vessels (Merkle et al., 2004Merkle F.T. Tramontin A.D. García-Verdugo J.M. Alvarez-Buylla A. Proc. Natl. Acad. Sci. USA. 2004; 101: 17528-17532Crossref PubMed Scopus (643) Google Scholar, Mirzadeh et al., 2008Mirzadeh Z. Merkle F.T. Soriano-Navarro M. Garcia-Verdugo J.M. Alvarez-Buylla A. Cell Stem Cell. 2008; 3: 265-278Abstract Full Text Full Text PDF PubMed Scopus (754) Google Scholar, Shen et al., 2008Shen Q. Wang Y. Kokovay E. Lin G. Chuang S.-M. Goderie S.K. Roysam B. Temple S. Cell Stem Cell. 2008; 3: 289-300Abstract Full Text Full Text PDF PubMed Scopus (823) Google Scholar, Tavazoie et al., 2008Tavazoie M. Van der Veken L. Silva-Vargas V. Louissaint M. Colonna L. Zaidi B. Garcia-Verdugo J.M. Doetsch F. Cell Stem Cell. 2008; 3: 279-288Abstract Full Text Full Text PDF PubMed Scopus (820) Google Scholar). Additionally, GFAP+ type B1 cells exist in both activated (as determined by the expression of epidermal growth factor receptor [EGFR+]) and quiescent states (Doetsch et al., 1999Doetsch F. Caillé I. Lim D.A. García-Verdugo J.M. Alvarez-Buylla A. Cell. 1999; 97: 703-716Abstract Full Text Full Text PDF PubMed Scopus (3226) Google Scholar, Morshead et al., 1994Morshead C.M. Reynolds B.A. Craig C.G. McBurney M.W. Staines W.A. Morassutti D. Weiss S. van der Kooy D. Neuron. 1994; 13: 1071-1082Abstract Full Text PDF PubMed Scopus (1216) Google Scholar, Pastrana et al., 2009Pastrana E. Cheng L.-C. Doetsch F. Proc. Natl. Acad. Sci. USA. 2009; 106: 6387-6392Crossref PubMed Scopus (288) Google Scholar). The characterization of the structure of GFAP+ type B1 cells, along with lineage-tracing studies, established that type B1 NSCs in the SVZ are the source of postnatally generated glial and neuronal cell subtypes. However, the relative contributions of active NSCs (aNSCs) versus quiescent NSCs (qNSCs) in the replenishment of type B1 cells and the production of specific neuronal and glial cell subtypes under normal and pathological states have not been elucidated. In addition, direct observations examining the potential precursor-product transitions between qNSCs and aNSCs have been lacking, with most understanding of their relationship being inferred. This is largely due to the lack of markers distinguishing qNSCs from aNSCs hampering examination of the lineage potentials and regenerative capacities of the heterogeneous population of type B1 cells in the SVZ. In this issue of Neuron, Codega et al., 2014Codega P. Silva-Vargas V. Paul A. Maldonado-Soto A.R. DeLeo A.M. Pastrana E. Doetsch F. Neuron. 2014; 82 (this issue): 545-559Abstract Full Text Full Text PDF PubMed Scopus (389) Google Scholar utilize a unique combination of markers to isolate and purify qNSCs and aNSCs from the mouse SVZ. Building on previous findings that a subset of purified CD133-positive (CD133+) cells from the SVZ is capable of generating neurons in vivo and neurospheres in vitro (Coskun et al., 2008Coskun V. Wu H. Blanchi B. Tsao S. Kim K. Zhao J. Biancotti J.C. Hutnick L. Krueger Jr., R.C. Fan G. et al.Proc. Natl. Acad. Sci. USA. 2008; 105: 1026-1031Crossref PubMed Scopus (266) Google Scholar), as well as coexpressing GFAP (Beckervordersandforth et al., 2010Beckervordersandforth R. Tripathi P. Ninkovic J. Bayam E. Lepier A. Stempfhuber B. Kirchhoff F. Hirrlinger J. Haslinger A. Lie D.C. et al.Cell Stem Cell. 2010; 7: 744-758Abstract Full Text Full Text PDF PubMed Scopus (253) Google Scholar, Mirzadeh et al., 2008Mirzadeh Z. Merkle F.T. Soriano-Navarro M. Garcia-Verdugo J.M. Alvarez-Buylla A. Cell Stem Cell. 2008; 3: 265-278Abstract Full Text Full Text PDF PubMed Scopus (754) Google Scholar), Codega et al., 2014Codega P. Silva-Vargas V. Paul A. Maldonado-Soto A.R. DeLeo A.M. Pastrana E. Doetsch F. Neuron. 2014; 82 (this issue): 545-559Abstract Full Text Full Text PDF PubMed Scopus (389) Google Scholar postulated and found that cells expressing GFAP (labeled using the transgenic mouse line GFAP::GFP in which GFP is under the control of the GFAP promoter) and CD133 identify both qNSCs and aNSCs among SVZ cells. A key finding is the observation that EGFR, a protein expressed in aNSCs (Pastrana et al., 2009Pastrana E. Cheng L.-C. Doetsch F. Proc. Natl. Acad. Sci. USA. 2009; 106: 6387-6392Crossref PubMed Scopus (288) Google Scholar), is expressed in a much lower percentage of GFAP+CD133+ cells (GFAP+CD133+EGFR+). This indicates that GFAP+ and CD133+ cells that do not express EGFR (GFAP+CD133+EGFR+) may belong to a distinct population of type B1 SVZ cells, the qNSC fraction. Indeed, unlike GFAP+CD133+EGFR+ cells, GFAP+CD133+EGFR− cells do not express proliferation markers, fail to incorporate bromodeoxyuridine (BrdU) after a pulse, and survive infusion of cytosine-β-D-arabinofuranoside (Ara-C), which efficiently eliminates dividing B1 cells. Moreover, BrdU incorporation is only observed in GFAP+CD133+EGFR− cells after prolonged administration, indicating that GFAP+CD133+EGFR− cells are slowly dividing. These findings cumulatively indicate that the cell cycle kinetics of GFAP+CD133+EGFR− cells differ significantly from GFAP+CD133+EGFR+ cells, consistent with quiescent stem cells found in other tissues. Therefore, the differential expression of GFAP, CD133, and EGFR enabled Codega et al., 2014Codega P. Silva-Vargas V. Paul A. Maldonado-Soto A.R. DeLeo A.M. Pastrana E. Doetsch F. Neuron. 2014; 82 (this issue): 545-559Abstract Full Text Full Text PDF PubMed Scopus (389) Google Scholar to reliably distinguish and isolate qNSCs (GFAP+CD133+EGFR− cells) and aNSCs (GFAP+CD133+EGFR+ cells). The ability to isolate and purify these two populations is a gateway to understanding the molecular and cellular characteristics of qNSCs for the first time. Stem cells are defined by their capacity for self-renewal and generate specialized cell types through differentiation. Through in vivo transplantation studies, Codega et al., 2014Codega P. Silva-Vargas V. Paul A. Maldonado-Soto A.R. DeLeo A.M. Pastrana E. Doetsch F. Neuron. 2014; 82 (this issue): 545-559Abstract Full Text Full Text PDF PubMed Scopus (389) Google Scholar show that GFAP+CD133+EGFR− qNSCs possess these defining characteristics of stem cells. One month after transplantation, progeny of transplanted qNSCs were found in the SVZ and within the OB neural circuitry, indicating that transplanted qNSCs are able to self-renew and differentiate into mature interneurons. Additionally, qNSCs are multipotent as they are capable of generating oligodendrocytes and mature astrocytes in addition to neuronal lineages. One surprising observation is the absence of Nestin expression in qNSCs. The expression of Nestin, an intermediate filament protein, has been a widely used marker of embryonic and adult neural stem cells. Codega and colleagues took advantage of this additional difference between qNSCs, aNSCs, and Nestin-expressing ependymal cells (which have been claimed to be able to differentiate into neuronal or glial cells [Carlén et al., 2009Carlén M. Meletis K. Göritz C. Darsalia V. Evergren E. Tanigaki K. Amendola M. Barnabé-Heider F. Yeung M.S.Y. Naldini L. et al.Nat. Neurosci. 2009; 12: 259-267Crossref PubMed Scopus (335) Google Scholar, Coskun et al., 2008Coskun V. Wu H. Blanchi B. Tsao S. Kim K. Zhao J. Biancotti J.C. Hutnick L. Krueger Jr., R.C. Fan G. et al.Proc. Natl. Acad. Sci. USA. 2008; 105: 1026-1031Crossref PubMed Scopus (266) Google Scholar]) to examine the regenerative properties of qNSCs in vivo. This was accomplished by utilizing an inducible Cre transgenic mouse line carrying a reporter transgene, tdTomato (GFAP::CreERT2;CAG::tdTomato). In this experiment, GFAP-derived SVZ cells were labeled by administration of tamoxifen for 10 days, followed by a 10-day infusion of Ara-C to eliminate all dividing cells. Codega et al., 2014Codega P. Silva-Vargas V. Paul A. Maldonado-Soto A.R. DeLeo A.M. Pastrana E. Doetsch F. Neuron. 2014; 82 (this issue): 545-559Abstract Full Text Full Text PDF PubMed Scopus (389) Google Scholar find that many of the tdTomato+ cells in the SVZ are eliminated after Ara-C infusion and that the remaining tdTomato+ cells do not express Nestin. However, 6 days after Ara-C removal, fractions of tdTomato+ cells began to express Nestin, and/or doublecortin, a marker for neuroblasts, implying that the remaining tdTomato+ cells are regenerative, as they are able to replenish the depleted aNSC population and contribute to the production of neuronal lineages. Thus, qNSCs retain long-term neurogenic potential and begs for further investigation of the intrinsic and extrinsic events that trigger qNSCs to enter the activated state and differentiate into neuronal or glial cell lineages. To start to address these questions, Codega et al., 2014Codega P. Silva-Vargas V. Paul A. Maldonado-Soto A.R. DeLeo A.M. Pastrana E. Doetsch F. Neuron. 2014; 82 (this issue): 545-559Abstract Full Text Full Text PDF PubMed Scopus (389) Google Scholar performed microarray analyses that will help characterize the molecular signature of qNSCs. These experiments show that factors commonly expressed by quiescent stem cell populations in tissues outside of the nervous system are also upregulated in qNSCs, including genes encoding cell surface proteins/receptors and extracellular matrix proteins. Codega et al., 2014Codega P. Silva-Vargas V. Paul A. Maldonado-Soto A.R. DeLeo A.M. Pastrana E. Doetsch F. Neuron. 2014; 82 (this issue): 545-559Abstract Full Text Full Text PDF PubMed Scopus (389) Google Scholar then went on to show that ligands for two of these differentially expressed receptors, the G protein-coupled receptors for Sphingosine-1-Phosphate (S1P) and Prostaglandin D2, effectively inhibit the activation of qNSCs. Notably, S1P selectively acts on qNSCs but not aNSCs, indicating that qNSCs can be specifically targeted by extrinsic factors. Since the ligands for these receptors are found in the CSF, they may be physiologic regulators of NSC quiescence. These findings, along with the observations that qNSCs are unable to form neurospheres in vitro using currently established protocols, underscore the need to further identify and characterize factors that regulate qNSCs, as well as other NSC populations, within the SVZ neurogenic niche at different phases of adult neurogenesis. It is also worth noting that an S1P agonist was recently approved as an oral treatment for multiple sclerosis (Bolli et al., 2011Bolli M.H. Lescop C. Nayler O. Curr. Top. Med. Chem. 2011; 11: 726-757Crossref PubMed Scopus (35) Google Scholar). The main mechanism of action for this agent (fingolimod—trade name Gilenya) is believed to be a modulation of lymphocyte trafficking; however, it is quite possible that this agent might affect qNSC dynamics. Since fingolimod is thought of as a first in family compound with other agents in various pipelines, it is quite possible that S1P agonists and antagonists could be studied for their effects on SVZ NSCs in future studies and in disease models. Codega et al., 2014Codega P. Silva-Vargas V. Paul A. Maldonado-Soto A.R. DeLeo A.M. Pastrana E. Doetsch F. Neuron. 2014; 82 (this issue): 545-559Abstract Full Text Full Text PDF PubMed Scopus (389) Google Scholar provide a powerful method that separates two major populations of type B1 NSCs in the SVZ. Given that the SVZ is composed of molecularly heterogeneous NSCs (Giachino et al., 2014Giachino C. Basak O. Lugert S. Knuckles P. Obernier K. Fiorelli R. Frank S. Raineteau O. Alvarez-Buylla A. Taylor V. Stem Cells. 2014; 32: 70-84Crossref PubMed Scopus (87) Google Scholar), future experiments must determine and characterize the existence of lineage-restricted qNSCs, and how their fate potential is affected over time under homeostatic and pathological conditions. In addition to intracellular and extracellular molecular factors, it would also be interesting to examine how other cell populations within the SVZ affect the behavior and maintenance of qNSCs. Advancing our knowledge on the biology and plasticity of qNSCs is especially valuable since the adult human brain harbors largely quiescent NSC populations (van den Berge et al., 2010van den Berge S.A. Middeldorp J. Zhang C.E. Curtis M.A. Leonard B.W. Mastroeni D. Voorn P. van de Berg W.D.J. Huitinga I. Hol E.M. Aging Cell. 2010; 9: 313-326Crossref PubMed Scopus (105) Google Scholar). Thus, by harnessing the regenerative potential of quiescent NSCs, new approaches may be developed to effectively treat neurological diseases, particularly those caused by cellular dysfunction or tissue injury. Prospective Identification and Purification of Quiescent Adult Neural Stem Cells from Their In Vivo NicheCodega et al.NeuronMay 07, 2014In BriefCodega et al. prospectively purify quiescent neural stem cells (qNSCs) from the adult ventricular-subventricular neurogenic niche. qNSCs exhibit strikingly different in vivo and in vitro functional properties and molecular signatures from actively dividing NSCs. Full-Text PDF Open Archive