Fanie Barnabé-Heider

Evidence for Cardiomyocyte Renewal in Humans

It has been difficult to establish whether we are limited to the heart muscle cells we are born with or if cardiomyocytes are generated also later in life. We have taken advantage of the integration of carbon-14, generated by nuclear bomb tests during the Cold War, into DNA to establish the age of cardiomyocytes in humans. We report that cardiomyocytes renew, with a gradual decrease from 1% turning over annually at the age of 25 to 0.45% at the age of 75. Fewer than 50% of cardiomyocytes are exchanged during a normal life span. The capacity to generate cardiomyocytes in the adult human heart suggests that it may be rational to work toward the development of therapeutic strategies aimed at stimulating this process in cardiac pathologies.

A dermal niche for multipotent adult skin-derived precursor cells

A fundamental question in stem cell research is whether cultured multipotent adult stem cells represent endogenous multipotent precursor cells. Here we address this question, focusing on SKPs, a cultured adult stem cell from the dermis that generates both neural and mesodermal progeny. We show that SKPs derive from endogenous adult dermal precursors that exhibit properties similar to embryonic neural-crest stem cells. We demonstrate that these endogenous SKPs can first be isolated from skin during embryogenesis and that they persist into adulthood, with a niche in the papillae of hair and whisker follicles. Furthermore, lineage analysis indicates that both hair and whisker follicle dermal papillae contain neural-crest-derived cells, and that SKPs from the whisker pad are of neural-crest origin. We propose that SKPs represent an endogenous embryonic precursor cell that arises in peripheral tissues such as skin during development and maintains multipotency into adulthood.

Spinal Cord Injury Reveals Multilineage Differentiation of Ependymal Cells

Spinal cord injury often results in permanent functional impairment. Neural stem cells present in the adult spinal cord can be expanded in vitro and improve recovery when transplanted to the injured spinal cord, demonstrating the presence of cells that can promote regeneration but that normally fail to do so efficiently. Using genetic fate mapping, we show that close to all in vitro neural stem cell potential in the adult spinal cord resides within the population of ependymal cells lining the central canal. These cells are recruited by spinal cord injury and produce not only scar-forming glial cells, but also, to a lesser degree, oligodendrocytes. Modulating the fate of ependymal progeny after spinal cord injury may offer an alternative to cell transplantation for cell replacement therapies in spinal cord injury.

Forebrain ependymal cells are Notch-dependent and generate neuroblasts and astrocytes after stroke

Neurons are continuously generated from stem cells in discrete regions in the adult mammalian brain. We found that ependymal cells lining the lateral ventricles were quiescent and did not contribute to adult neurogenesis under normal conditions in mice but instead gave rise to neuroblasts and astrocytes in response to stroke. Ependymal cell quiescence was actively maintained by canonical Notch signaling. Inhibition of this pathway in uninjured animals allowed ependymal cells to enter the cell cycle and produce olfactory bulb neurons, whereas forced Notch signaling was sufficient to block the ependymal cell response to stroke. Ependymal cells were depleted by stroke and failed to self-renew sufficiently to maintain their own population. Thus, although ependymal cells act as primary cells in the neural lineage to produce neurons and glial cells after stroke, they do not fulfill defining criteria for stem cells under these conditions and instead serve as a reservoir that is recruited by injury.

Origin of New Glial Cells in Intact and Injured Adult Spinal Cord

Several distinct cell types in the adult central nervous system have been suggested to act as stem or progenitor cells generating new cells under physiological or pathological conditions. We have assessed the origin of new cells in the adult mouse spinal cord by genetic fate mapping. Oligodendrocyte progenitors self-renew, give rise to new mature oligodendrocytes, and constitute the dominating proliferating cell population in the intact adult spinal cord. In contrast, astrocytes and ependymal cells, which are restricted to limited self-duplication in the intact spinal cord, generate the largest number of cells after spinal cord injury. Only ependymal cells generate progeny of multiple fates, and neural stem cell activity in the intact and injured adult spinal cord is confined to this cell population. We provide an integrated view of how several distinct cell types contribute in complementary ways to cell maintenance and the reaction to injury.

Evidence that Embryonic Neurons Regulate the Onset of Cortical Gliogenesis via Cardiotrophin-1

Precursor cells of the embryonic cortex sequentially generate neurons and then glial cells, but the mechanisms regulating this neurogenic-to-gliogenic transition are unclear. Using cortical precursor cultures, which temporally mimic this in vivo differentiation pattern, we demonstrate that cortical neurons synthesize and secrete the neurotrophic cytokine cardiotrophin-1, which activates the gp130-JAK-STAT pathway and is essential for the timed genesis of astrocytes in vitro. Our data indicate that a similar phenomenon also occurs in vivo. In utero electroporation of neurotrophic cytokines in the environment of embryonic cortical precursors causes premature gliogenesis, while acute perturbation of gp130 in cortical precursors delays the normal timed appearance of astrocytes. Moreover, the neonatal cardiotrophin-1-/- cortex contains fewer astrocytes. Together, these results describe a neural feedback mechanism; newly born neurons produce cardiotrophin-1, which instructs multipotent cortical precursors to generate astrocytes, thereby ensuring that gliogenesis does not occur until neurogenesis is largely complete.

An Essential Role for a MEK-C/EBP Pathway during Growth Factor-Regulated Cortical Neurogenesis

Mammalian neurogenesis is determined by an interplay between intrinsic genetic mechanisms and extrinsic cues such as growth factors. Here we have defined a signaling cascade, a MEK-C/EBP pathway, that is essential for cortical progenitor cells to become postmitotic neurons. Inhibition of MEK or of the C/EBP family of transcription factors inhibits neurogenesis while expression of a C/EBPbeta mutant that is a phosphorylation-mimic at a MEK-Rsk site enhances neurogenesis. C/EBP mediates this positive effect by direct transcriptional activation of neuron-specific genes such as Talpha1 alpha-tubulin. Conversely, inhibition of C/EBP-dependent transcription enhances CNTF-mediated generation of astrocytes from the same progenitor cells. Thus, activation of a MEK-C/EBP pathway enhances neurogenesis and inhibits gliogenesis, thereby providing a mechanism whereby growth factors can selectively bias progenitors to become neurons during development.

Stem Cells for Spinal Cord Repair

Spinal cord injury typically results in permanent disability. Many studies have indicated that transplantation of several different types of stem cells promotes functional recovery in animal models of spinal cord injury. A conceptually different approach to utilize stem cells for regenerative therapies may be recruitment of endogenous neural stem cells resident in the adult spinal cord. We discuss the possibilities, risks, and mechanisms for stem cells in spinal cord repair.

Analysis of the neurogenic potential of multipotent skin-derived precursors

Multipotent precursors similar to stem cells of the embryonic neural crest (NC) have been identified in several postnatal tissues, and are potentially useful for research and therapeutic purposes. However, their neurogenic potential, including their ability to produce electrophysiologically active neurons, is largely unexplored. We investigated this issue with regard to skin-derived precursors (SKPs), multipotent NC-related precursors isolated from the dermis of skin. SKP cultures follow an appropriate pattern and time-course of neuronal differentiation, with proliferating nestin-expressing SKPs generating post-mitotic neuronal cells that co-express pan-neuronal and peripheral autonomic lineage markers. These SKP-derived neuron-like cells survive and maintain their peripheral phenotype for at least 5 weeks when transplanted into the CNS environment of normal or kainate-injured hippocampal slices. Undifferentiated SKPs retain key neural precursor properties after multi-passage expansion, including growth factor dependence, nestin expression, neurogenic potential, and responsiveness to embryonic neural crest fate determinants. Despite undergoing an apparently appropriate neurogenic process, however, SKP-derived neuron-like cells possess an immature electrophysiological profile. These findings indicate that SKPs retain latent neurogenic properties after residing in a non-neural tissue, but that additional measures will be necessary to promote their differentiation into electrophysiologically active neurons.

Role for Runx1 in the Proliferation and Neuronal Differentiation of Selected Progenitor Cells in the Mammalian Nervous System

Neurogenesis requires factors that regulate the decision of dividing progenitors to leave the cell cycle and activate the neuronal differentiation program. It is shown here that the murine runt-related gene Runx1 is expressed in proliferating cells on the basal side of the olfactory epithelium. These include both Mash1+ olfactory receptor neuron (ORN) progenitors and NeuroD+ ORN precursors. Disruption of Runx1 function in vivo does not cause a change in Mash1 expression but leads to a decrease in the number of NeuroD+ neuronal precursors and an increase in differentiated ORNs. These effects result in premature and ectopic ORN differentiation. It is shown further that exogenous Runx1 expression in cultured olfactory neural progenitors causes an expansion of the mitotic cell population. In agreement with these findings, exogenous Runx1 expression also promotes cortical neural progenitor cell proliferation without inhibiting neuronal differentiation. These effects are phenocopied by a chimeric protein containing ETO, the eight twenty one transcriptional repressor, fused to the Runx1 DNA-binding domain, which suggests the involvement of transcription repression mechanisms. Consistent with this possibility, Runx1 represses transcription driven by the promoter of the cell cycle inhibitor p21Cip 1 in cortical progenitors. Together, these findings suggest a previously unrecognized role for Runx1 in coordinating the proliferation and neuronal differentiation of selected populations of neural progenitors.