Ryan J. Felling

Neural Stem/Progenitor Cells Participate in the Regenerative Response to Perinatal Hypoxia/Ischemia

Perinatal hypoxia/ischemia (H/I) is the leading cause of neurologic injury resulting from birth complications. Recent advances in critical care have dramatically improved the survival rate of infants suffering this insult, but ∼50% of survivors will develop neurologic sequelae such as cerebral palsy, epilepsy or cognitive deficits. Here we demonstrate that tripotential neural stem/progenitor cells (NSPs) participate in the regenerative response to perinatal H/I as their numbers increase 100% by 3 d and that they alter their intrinsic properties to divide using expansive symmetrical cell divisions. We further show that production of new striatal neurons follows the expansion of NSPs. Increased proliferation within the NSP niche occurs at 2 d after perinatal H/I, and the proliferating cells express nestin. Of those stem-cell related genes that change, the membrane receptors Notch1, gp-130, and the epidermal growth factor receptor, as well as the downstream transcription factor Hes5, which stimulate NSP proliferation and regulate stem cellness are induced before NSP expansion. The mechanisms for the reactive expansion of the NSPs reported here reveal potential therapeutic targets that could be exploited to amplify this response, thus enabling endogenous precursors to restore a normal pattern of brain development after perinatal H/I.

Neurobiology of tourette syndrome: current status and need for further investigation.

Tourette syndrome (TS) is a common, chronic neuropsychiatric disorder characterized by the presence of fluctuating motor and phonic tics. The typical age of onset is ∼5–7 years, and the majority of children improve by their late teens or early adulthood. Affected individuals are at increased risk for the development of various comorbid conditions, such as obsessive–compulsive disorder, attention deficit hyperactivity disorder, school problems, depression, and anxiety. There is no cure for tics, and symptomatic therapy includes behavioral and pharmacological approaches. Evidence supports TS being an inherited disorder; however, the precise genetic abnormality remains unknown. Pathologic involvement of cortico-striatal-thalamo-cortical (CSTC) pathways is supported by neurophysiological, brain imaging, and postmortem studies, but results are often confounded by small numbers, age differences, severity of symptoms, comorbidity, use of pharmacotherapy, and other factors. The primary site of abnormality remains controversial. Although numerous neurotransmitters participate in the transmission of messages through CSTC circuits, a dopaminergic dysfunction is considered a leading candidate. Several animal models have been used to study behaviors similar to tics as well as to pursue potential pathophysiological deficits. TS is a complex disorder with features overlapping a variety of scientific fields. Despite description of this syndrome in the late 19th century, there remain numerous unanswered neurobiological questions.

Enhanced neurogenesis following stroke.

Each year hundreds of thousands of people must cope with the severe neurological consequences of a stroke. Current therapeutic strategies for stroke focus on acute treatment and neuroprotection. Unfortunately, these practices do little to reduce the long‐term morbidity associated with the injury. To develop effective therapies that promote regeneration, we must have an understanding of the cellular and molecular events involved in the recovery from an insult. Neural stem and progenitor cells are likely to be affected during this period. Here we review how the proliferation, migration, and maturation of these precursors are affected by ischemia. Furthermore, we summarize data available on the underlying mechanisms and the therapeutic implications of these studies. The studies that we review provide compelling evidence that neural precursors resident in the brain initiate a compensatory response to stroke that results in the production of new neurons. Moreover, administration of growth factors can enhance this compensatory response. Based on these encouraging results, we may eventually be able to manipulate these precursors to improve recovery of function in individuals afflicted by this devastating injury.

Brain Injury Expands the Numbers of Neural Stem Cells and Progenitors in the SVZ by Enhancing Their Responsiveness to EGF

There is an increase in the numbers of neural precursors in the SVZ (subventricular zone) after moderate ischaemic injuries, but the extent of stem cell expansion and the resultant cell regeneration is modest. Therefore our studies have focused on understanding the signals that regulate these processes towards achieving a more robust amplification of the stem/progenitor cell pool. The goal of the present study was to evaluate the role of the EGFR [EGF (epidermal growth factor) receptor] in the regenerative response of the neonatal SVZ to hypoxic/ischaemic injury. We show that injury recruits quiescent cells in the SVZ to proliferate, that they divide more rapidly and that there is increased EGFR expression on both putative stem cells and progenitors. With the amplification of the precursors in the SVZ after injury there is enhanced sensitivity to EGF, but not to FGF (fibroblast growth factor)-2. EGF-dependent SVZ precursor expansion, as measured using the neurosphere assay, is lost when the EGFR is pharmacologically inhibited, and forced expression of a constitutively active EGFR is sufficient to recapitulate the exaggerated proliferation of the neural stem/progenitors that is induced by hypoxic/ischaemic brain injury. Cumulatively, our results reveal that increased EGFR signalling precedes that increase in the abundance of the putative neural stem cells and our studies implicate the EGFR as a key regulator of the expansion of SVZ precursors in response to brain injury. Thus modulating EGFR signalling represents a potential target for therapies to enhance brain repair from endogenous neural precursors following hypoxic/ischaemic and other brain injuries.

Molecular features of neural stem cells enable their enrichment using pharmacological inhibitors of survival-promoting kinases

Isolating a pure population of neural stem cells (NSCs) has been difficult since no exclusive surface markers have been identified for panning or FACS purification. Moreover, additional refinements for maintaining NSCs in culture are required, since NSCs generate a variety of neural precursors (NPs) as they proliferate. Here, we demonstrate that post‐natal rat NPs express low levels of pro‐apoptotic molecules and resist phosphatidylinositol 3′OH kinase and extracellular regulated kinase 1/2 inhibition as compared to late oligodendrocyte progenitors. Furthermore, maintaining subventricular zone precursors in LY294002 and PD98059, inhibitors of PI3K and ERK1/2 signaling, eliminated lineage‐restricted precursors as revealed by enrichment for Nestin+/SOX‐2+ cells. The cells that survived formed neurospheres and 89% of these neurospheres were tripotential, generating neurons, astrocytes, and oligodendrocytes. Without this enrichment step, less than 50% of the NPs were Nestin+/SOX‐2+ and 42% of the neurospheres were tripotential. In addition, neurospheres enriched using this procedure produced 3‐times more secondary neurospheres, supporting the conclusion that this procedure enriches for NSCs. A number of genes that enhance survival were more highly expressed in neurospheres compared to late oligodendrocyte progenitors. Altogether, these studies demonstrate that primitive neural precursors can be enriched using a relatively simple and inexpensive means that will facilitate cell replacement strategies using stem cells as well as other studies whose goal is to reveal the fundamental properties of primitive neural precursors.

Astrocyte-produced leukemia inhibitory factor expands the neural stem/progenitor pool following perinatal hypoxia–ischemia

Brain injuries, such as cerebral hypoxia–ischemia (H‐I), induce a regenerative response from the neural stem/progenitors (NSPs) of the subventricular zone (SVZ); however, the mechanisms that regulate this expansion have not yet been fully elucidated. The Notch‐ Delta‐Serrate‐Lag2 (DSL) signaling pathway is considered essential for the maintenance of neural stem cells, but it is not known if it is necessary for the expansion of the NSPs subsequent to perinatal H‐I injury. Therefore, the aim of this study was to investigate whether this pathway contributes to NSP expansion in the SVZ after H‐I and, if so, to establish whether this pathway is directly induced by H‐I or regulated by paracrine factors. Here we report that Notch1 receptor induction and one of its ligands, Delta‐like 1, precedes NSP expansion after perinatal H‐I in P6 rat pups and that this increase occurs specifically in the most medial cell layers of the SVZ where the stem cells reside. Pharmacologically inhibiting Notch signaling in vivo diminished NSP expansion. With an in vitro model of H‐I, Notch1 was not induced directly by hypoxia, but was stimulated by soluble factors, specifically leukemia inhibitory factor, produced by astrocytes within the SVZ. These data confirm the importance both of the Notch‐DSL signaling pathway in the expansion of NSPs after H‐I and in the role of the support cells in their niche. They further support the body of evidence that indicates that leukemia inhibitory factor is a key injury‐induced cytokine that is stimulating the regenerative response of the NSPs.

Responses of the SVZ to Hypoxia and Hypoxia/Ischemia

Stroke is a devastating injury caused by interruption of the blood supply to the brain. Each year, about 700,000 people in the United States suffer a stroke incurring healthcare costs of more than

Roles of the mammalian subventricular zone in cell replacement after brain injury

50 billion (Heart Disease and Stroke Statistics 2003 Update, 2002). While advances in the acute treatment of stroke have improved the survival rates, little success has been realized in decreasing the associated morbidity. Stroke survivors suffer a wide range of neurological deficits depending on the location in the brain where the stroke occurs. These effects include, but are not limited to, paralysis, sensory deficits, memory loss and personality changes. Additionally, hypoxic/ischemic (H/I) injury is the single most important cause of brain damage resulting from complications during birth, leading to permanent neurological deficits. Every year, perinatal H/I afflicts approximately 1-2 per 1000 term births and roughly half of surviving preterm infants. Many of these infants suffer long-term handicaps that include learning disabilities, mental retardation, epilepsy and cerebral palsy (Volpe, 2001). Formore than 20 years, experimental animal models have beenwidely used to study the effects of ischemic brain injury. The most common model is transient middle cerebral artery occlusion (MCAo) used primarily in adult animals to generate unilateral focal cerebral infarcts (Longa et al., 1989).