**3. TGF-***β* **superfamily expression and function in normal adult brain: Role in neurogenesis**

Adult neurogenesis involves proliferation of neural stem cells (NSCs), cell cycle exit, differ‐ entiation, maturation, and integration into the neural circuits, in a process that is involved in learning and memory in the normal adult brain [68]. The neurogenic niche of the adult forebrain subventricular zone (SVZ) is comprised of three major proliferative cell types; A, B and C. Multipotent, self-renewing type B cells occur earliest in the neurogenic lineage of the SVZ and give rise to the rapidly dividing type C cells, or transit amplifying progenitors. Type A cells or neuroblasts differentiate from Type C cells and are migratory neuronal progenitors with proliferative capacity, which migrate to the olfactory bulb where they differentiate into interneurons (reviewed in [69-71]. In the subgranular zone (SGZ) of the hippocampal dentate gyrus (DG), type 1 and type 2 slowly-dividing progenitors give rise to more rapidly dividing intermediate progenitor cells, and these in turn differentiate into immature neuroblasts, which migrate into the granule cell layer, then differentiate into mature neurons and integrate with the existing hippocampal circuitry [71].

Within the CNS, all three isoforms of TGF-β are produced by both glial and neuronal cells [72]. Immunohistochemical studies show widespread expression of TGF-β2 and -β3 in the devel‐ oping CNS, and these proteins play a role in regulation of neuronal migration, glial prolifer‐ ation and differentiation [73-76]. In adult brain, TGF-β receptors are found in all areas of the CNS including the cortex, hippocampus, striatum, brainstem and cerebellum [77, 78]. Immu‐ noreactivity for TβRI and TβRII is detected on neurons, astrocytes and microglia and endo‐ thelial cells located in the cortical gray matter, suggesting that almost every cell type in the CNS is a potential target for TGF-β signaling [79].

The TGF-β superfamily and its downstream targets are capable of controlling proliferation, differentiation, maturation and survival of stem cells and precursors in the neurogenic niches of adult brain [18]. TβRI and TβRII are expressed by Nestin-positive type B and C cells in the SVZ [80, 81]. Our data show mRNA expression of TGF-β1, -β2, and -β3 in both the adult SVZ and DG [82]. In the adult human brain, TGF-β1 protein expression has been reported in the hippocampus, and the protein levels significantly increased with the age of the individual [83]. As neurogenesis declines with age [84], it has been suggested that TGF-β is a possible regulator of this age-related decline [83]. Signaling by the Smad2/3 pathway is high in the hippocampus and specifically the dentate gyrus, indicating a role for TGF-β and/or activin in regulation of neurogenesis [85, 86]. When TGF-β protein is overexpressed or infused directly into the lateral ventricles of uninjured animals, hippocampal neurogenesis is dramatically inhibited [81, 87]. This may be due to a direct anti-proliferative effect of TGF-β on type 1 and 2 primary NSCs [17]. A direct effect of TGF-β on NSCs is supported by *in vitro* studies showing that TGF-β1 treatment of cultured adult NSCs induces the cyclin-dependent kinase inhibitor (p21) and leads to cell cycle termination, without altering the differentiation choices of the NSCs [81]. Additionally, overexpression studies lead to increased TGF-β signaling in many different cell types within the neurogenic niche, making the exact contribution of more restricted, endoge‐ nous TGF-β difficult to determine. Recent data have suggested that TGF-β signaling at later stages of neurogenesis is critical for newborn neuron survival and maturation in the DG. Conditional deletion of the TβRI (ALK5) gene specifically in immature and mature neurons, leads to decreased neurogenesis and reduced survival of newborn neurons [85]. Thus, TGFβ potentially has opposing roles at different stages of neurogenesis, providing an additional example of the contextual nature of TGF-β action.

**Conventional knockout mouse model of TGF-β proteins**

**in neurogenesis**

the existing hippocampal circuitry [71].

CNS is a potential target for TGF-β signaling [79].

**Phenotype References**

[65]

[67]

Activin-βB Large litters but delayed parturition; nursing defects; Eye lid closure defects at birth

8 Trends in Cell Signaling Pathways in Neuronal Fate Decision

Noggin Perinatal lethal, cartilage hyperplasia [66]

**Table 1.** Phenotype of mice that do not express specific TGF-β ligands, receptors or signaling molecules.

**3. TGF-***β* **superfamily expression and function in normal adult brain: Role**

Adult neurogenesis involves proliferation of neural stem cells (NSCs), cell cycle exit, differ‐ entiation, maturation, and integration into the neural circuits, in a process that is involved in learning and memory in the normal adult brain [68]. The neurogenic niche of the adult forebrain subventricular zone (SVZ) is comprised of three major proliferative cell types; A, B and C. Multipotent, self-renewing type B cells occur earliest in the neurogenic lineage of the SVZ and give rise to the rapidly dividing type C cells, or transit amplifying progenitors. Type A cells or neuroblasts differentiate from Type C cells and are migratory neuronal progenitors with proliferative capacity, which migrate to the olfactory bulb where they differentiate into interneurons (reviewed in [69-71]. In the subgranular zone (SGZ) of the hippocampal dentate gyrus (DG), type 1 and type 2 slowly-dividing progenitors give rise to more rapidly dividing intermediate progenitor cells, and these in turn differentiate into immature neuroblasts, which migrate into the granule cell layer, then differentiate into mature neurons and integrate with

Within the CNS, all three isoforms of TGF-β are produced by both glial and neuronal cells [72]. Immunohistochemical studies show widespread expression of TGF-β2 and -β3 in the devel‐ oping CNS, and these proteins play a role in regulation of neuronal migration, glial prolifer‐ ation and differentiation [73-76]. In adult brain, TGF-β receptors are found in all areas of the CNS including the cortex, hippocampus, striatum, brainstem and cerebellum [77, 78]. Immu‐ noreactivity for TβRI and TβRII is detected on neurons, astrocytes and microglia and endo‐ thelial cells located in the cortical gray matter, suggesting that almost every cell type in the

The TGF-β superfamily and its downstream targets are capable of controlling proliferation, differentiation, maturation and survival of stem cells and precursors in the neurogenic niches of adult brain [18]. TβRI and TβRII are expressed by Nestin-positive type B and C cells in the SVZ [80, 81]. Our data show mRNA expression of TGF-β1, -β2, and -β3 in both the adult SVZ

Follistatin Neonatal lethal, craniofacial defects, growth retardation and skin defects retardation and skin defects

> Activin receptors are expressed throughout the brain, with strong expression in the neuronal layers of the hippocampus [88-90]. We have found that mRNA for activin-A and for activin's endogenous high affinity inhibitor, follistatin, are expressed in both the SVZ and DG of the adult mouse [82] and several recent reports have demonstrated that activin-A modulates adult neurogenesis [88, 91, 92]. Chronic overexpression of follistatin by neurons of the hippocampus almost entirely ablates adult DG neurogenesis, due to drastically lowered survival of adult-generated neurons [91], although short-term infusion of follistatin does not affect neurogenesis in uninjured animals [88]. Infusion of activin to the lateral ventri‐ cle of uninjured mice mildly increases the rate of NSC proliferation and neuron genera‐ tion in the DG, indicating that activin might stimulate division of NSCs. This effect may be indirect as activin has a potent anti-inflammatory effect in the CNS, and may modulate local microglia to stimulate neurogenesis [88]. Smad3 knockout mice have decreased levels of cell proliferation in the SVZ and along the rostral migratory stream, and decreased levels of olfactory bulb neurogenesis [93]. As these mice have defective signaling by both TGF-β and activin, these data suggest that activin signaling in the SVZ may be the predominant Smad3-utilizing cytokine in defining basal levels of neurogenesis. In the DG pSmad2 is normally absent from Sox2-positive type 1 and 2 primary NSCs in the DG of adult mice [17]. However, Smad3 knockout mice also have reduced proliferation in the DG potential‐ ly pointing to a different role for Smad2 and Smad3 in the DG [93].

> The BMP family of proteins regulates cell proliferation and fate commitment throughout development and within the adult neurogenic niches [19]. Expression of BMP2, -4 and -7 mRNAs have been reported in neurogenic regions of adult rodent brain [94], and the BMP receptors BMPRIA, -IB and -II are expressed abundantly in neurons, as well as in astrocytes

and ependymal cells [95]. All three of these receptors are expressed in type A cells of the SVZ, while type B and C cells express BMPRIA and BMPRII [96]. In the DG, radial stem cells of the SGZ marked with glial fibrillary acidic protein (GFAP) and Nestin or Sox2 primarily express BMPRIA but not BMPRIB, while mature neurons express only BMPRIB [97]. BMP ligands are also expressed in the adult rat brain [98, 99]. BMP2, -4, -6, and -7 are expressed by cells of the SVZ and DG [96, 97]. In the DG, the BMP signal transducer pSmad1 is strongly expressed in non-dividing primary NSCs and neuroblasts, but is absent in dividing primary NSCs [97], while in the SVZ, pSmad1/5/8 has been reported in primary NSCs and transit amplifying progenitors, but not in DCX-positive neuroblasts [40]. The soluble BMP inhibitor noggin is also expressed by ependymal cells of the SVZ [96] and by cells of the DG [100].

114]. Smad proteins are also upregulated after injury and were mainly located in the cerebral cortex,typicallyinthenucleusand/orinthecytoplasmofastrocytes,oligodendrocytesorneurons [86, 108, 115, 116]. We have summarized many studies that have examined changes in the TGF-

> **Expression in neurogenic niche**

Hippocampus, Subventricular

zone

**Cell types in which protein is expressed**

oligodendrocytes, endothelial cells, astrocytes, macrophages, and ependymal cells

Microglia, T cells, neuroblasts and neurons

\_ \_ \_ \_ \_ Neurons, neuroblasts mRNA,

and blood vessels

Microglia, astrocytes and neurons

\_ \_ \_ \_ \_ Astrocytes, neurons Protein [129]

\_ \_ \_ \_ \_ Astrocytes, Microglia

Stab wound Cerebral cortex \_ \_ \_ \_ \_ Neurons Protein [116]

Cerebral cortex Hippocampus Neurons mRNA,

meningeal

astrocytes

cells, choroid plexus

Subventricular

Cerebral cortex \_ \_ \_ \_ \_ Activated glia,

Irradiation Cerebral cortex \_ \_ \_ \_ \_ Macrophages and

Excitotoxic Injury \_ \_ \_ \_ \_ Hippocampus Neurons Protein [133]

zone

Dentate gyrus Neurons, vessels Protein [124, 125]

**mRNA and/or protein**

http://dx.doi.org/10.5772/53941

Role of TGF-β Signaling in Neurogenic Regions After Brain Injury

mRNA, protein

mRNA, protein

protein

mRNA, protein

protein

mRNA, protein

Protein [134]

Protein [126]

**References**

11

[107-110]

[117-120]

[121-123]

[82, 112, 127, 128]

[130, 131]

[132]

β superfamily of cytokines after central nervous system injury in Table 2.

TGF-β1 Ischemia Cerebral cortex \_ \_ \_ \_ \_ Microglia, neurons,

**Expression in Brain**

Cerebellum, Cerebral cortex

Cerebral cortex, Striatum

Corpus callosum

Gray matter surrounding the

lesion

Cerebral cortex Hippocampus,

Cerebellum, Cerebral cortex

Hypoxic-ischemic Cerebral cortex,

**TGF-β protein** **Acute brain Insult**

Transient ischemia

Permanent ischemia

Bilateral cerebral ischemia

Traumatic brain

Excitotoxic lesion (NMDA)

Triethyltin exposure

Penetrating brain Injury

injury

**(Animal model)**

Changing the ratio of BMP to noggin alters the rates of NSC proliferation and neurogenesis in adult animals, indicating that these proteins are primary regulators of basal adult neurogene‐ sis [96, 97, 100]. Administration of exogenous BMP4 or BMP7 potently inhibits the division of NSCs and generation of new neurons in vivo and in vitro [96, 97], as does inhibition of noggin expression [101]. Conversely, infusion of noggin or genetic deletion of the BMPRIA receptor causes an increase in NSC proliferation and generation of NeuN-expressing neurons in the DG [96, 97]. However this increase is transient, there is an eventual depletion of the primary NSC pool and a drastically reduced level of neurogenesis [97]. Decreased BMP signaling in the DG is thought to be responsible for increased neurogenesis driven by exercise [102]. It has been proposed that secretion of noggin from ependymal cells inhibits BMP signaling allowing a low level of basal neurogenesis to occur, while BMP signaling maintains the overall quiescence of the primary NSC pool [96, 97, 100]. Exogenous noggin infusion potentially has a different effect onSVZNSCs,leavingtheirproliferationrateunaffected,butcausinganincreaseinthegeneration of oligodendrocyte precursor cells from primary NSCs at the expense of immature neuro‐ blasts [40].Thisnoggininfusionphenocopies the effectof conditionallydeletingSmad4 inNSCs usingGLAST-cre [40] andis incontrasttothepro-neurogenic effectsofnoggindescribedbyLim et al [96]. Thus, although there is still some controversy in the field it its clear that the balance between BMP and noggin is critical to proper maintenance of the adult NSC population.
