Rossella Galli

Isolation and characterization of tumorigenic, stem-like neural precursors from human glioblastoma.

Transformed stem cells have been isolated from some human cancers. We report that, unlike other brain cancers, the lethal glioblastoma multiforme contains neural precursors endowed with all of the critical features expected from neural stem cells. Similar, yet not identical, to their normal neural stem cell counterpart, these precursors emerge as unipotent (astroglial) in vivo and multipotent (neuronal-astroglial-oligodendroglial) in culture. More importantly, these cells can act as tumor-founding cells down to the clonal level and can establish tumors that closely resemble the main histologic, cytologic, and architectural features of the human disease, even when challenged through serial transplantation. Thus, cells possessing all of the characteristics expected from tumor neural stem cells seem to be involved in the growth and recurrence of adult human glioblastomas multiforme.

Injection of adult neurospheres induces recovery in a chronic model of multiple sclerosis

Widespread demyelination and axonal loss are the pathological hallmarks of multiple sclerosis. The multifocal nature of this chronic inflammatory disease of the central nervous system complicates cellular therapy and puts emphasis on both the donor cell origin and the route of cell transplantation. We established syngenic adult neural stem cell cultures and injected them into an animal model of multiple sclerosis—experimental autoimmune encephalomyelitis (EAE) in the mouse—either intravenously or intracerebroventricularly. In both cases, significant numbers of donor cells entered into demyelinating areas of the central nervous system and differentiated into mature brain cells. Within these areas, oligodendrocyte progenitors markedly increased, with many of them being of donor origin and actively remyelinating axons. Furthermore, a significant reduction of astrogliosis and a marked decrease in the extent of demyelination and axonal loss were observed in transplanted animals. The functional impairment caused by EAE was almost abolished in transplanted mice, both clinically and neurophysiologically. Thus, adult neural precursor cells promote multifocal remyelination and functional recovery after intravenous or intrathecal injection in a chronic model of multiple sclerosis.

CXCR4-activated astrocyte glutamate release via TNFalpha: amplification by microglia triggers neurotoxicity.

Astrocytes actively participate in synaptic integration by releasing transmitter (glutamate) via a calcium-regulated, exocytosis-like process. Here we show that this process follows activation of the receptor CXCR4 by the chemokine stromal cell-derived factor 1 (SDF-1). An extraordinary feature of the ensuing signaling cascade is the rapid extracellular release of tumor necrosis factor-α (TNFα). Autocrine/paracrine TNFα-dependent signaling leading to prostaglandin (PG) formation not only controls glutamate release and astrocyte communication, but also causes their derangement when activated microglia cooperate to dramatically enhance release of the cytokine in response to CXCR4 stimulation. We demonstrate that altered glial communication has direct neuropathological consequences and that agents interfering with CXCR4-dependent astrocyte–microglia signaling prevent neuronal apoptosis induced by the HIV-1 coat glycoprotein, gp120IIIB. Our results identify a new pathway for glia–glia and glia–neuron communication that is relevant to both normal brain function and neurodegenerative diseases.

Brain tumour stem cells

The dogma that the genesis of new cells is a negligible event in the adult mammalian brain has long influenced our perception and understanding of the origin and development of CNS tumours. The discovery that new neurons and glia are produced throughout life from neural stem cells provides new possibilities for the candidate cells of origin of CNS neoplasias. The emerging hypothesis is that alterations in the cellular and genetic mechanisms that control adult neurogenesis might contribute to brain tumorigenesis, thereby allowing the identification of new therapeutic strategies.

Isolation and cloning of multipotential stem cells from the embryonic human CNS and establishment of transplantable human neural stem cell lines by epigenetic stimulation

Stem cells that can give rise to neurons, astroglia, and oligodendroglia have been found in the developing and adult central nervous system (CNS) of rodents. Yet, their existence within the human brain has not been documented, and the isolation and characterization of multipotent embryonic human neural stem cells have proven difficult to accomplish. We show that the developing human CNS embodies multipotent precursors that differ from their murine counterpart in that they require simultaneous, synergistic stimulation by both epidermal and fibroblast growth factor-2 to exhibit critical stem cell characteristics. Clonal analysis demonstrates that human C NS stem cells are multipotent and differentiate spontaneously into neurons, astrocytes, and oligodendrocytes when growth factors are removed. Subcloning and population analysis show their extensive self-renewal capacity and functional stability, their ability to maintain a steady growth profile, their multipotency, and a constant potential for neuronal differentiation for more than 2 years. The neurons generated by human stem cells over this period of time are electrophysiologically active. These cells are also cryopreservable. Finally, we demonstrate that the neuronal and glial progeny of long-term cultured human CNS stem cells can effectively survive transplantation into the lesioned striatum of adult rats. Tumor formation is not observed, even in immunodeficient hosts. Hence, as a consequence of their inherent biology, human CNS stem cells can establish stable, transplantable cell lines by epigenetic stimulation. These lines represent a renewable source of neurons and glia and may significantly facilitate research on human neurogenesis and the development of clinical neural transplantation.

Skeletal myogenic potential of human and mouse neural stem cells

Distinct cell lineages established early in development are usually maintained throughout adulthood. Thus, adult stem cells have been thought to generate differentiated cells specific to the tissue in which they reside. This view has been challenged; for example, neural stem cells can generate cells that normally originate from a different germ layer. Here we show that acutely isolated and clonally derived neural stem cells from mice and humans could produce skeletal myotubes in vitro and in vivo, the latter following transplantation into adult animals. Myogenic conversion in vitro required direct exposure to myoblasts, and was blocked if neural cells were clustered. Thus, a community effect between neural cells may override such myogenic induction. We conclude that neural stem cells, which generate neurons, glia and blood cells, can also produce skeletal muscle cells, and can undergo various patterns of differentiation depending on exposure to appropriate epigenetic signals in mature tissues.

Gene therapy of experimental brain tumors using neural progenitor cells

Glioblastomas, the most frequent and malignant of primary brain tumors, have a very poor prognosis. Gene therapy of glioblastomas is limited by the short survival of viral vectors and by their difficulty in reaching glioblastoma cells infiltrating the brain parenchyma. Neural stem/progenitor cells can be engineered to produce therapeutic molecules and have the potential to overcome these limitations because they may travel along the white matter, like neoplastic cells, and engraft stably into the brain. Retrovirus-mediated transfer of the gene for interleukin-4 is an effective treatment for rat brain glioblastomas. Here, we transferred the gene for interleukin-4 into C57BL6J mouse primary neural progenitor cells and injected those cells into established syngeneic brain glioblastomas. This led to the survival of most tumor-bearing mice. We obtained similar results by implanting immortalized neural progenitor cells derived from Sprague-Dawley rats into C6 glioblastomas. We also documented by magnetic resonance imaging the progressive disappearance of large tumors, and detected 5-bromodeoxyuridine-labeled progenitor cells several weeks after the injection. These findings support a new approach for gene therapy of brain tumors, based on the grafting of neural stem cells producing therapeutic molecules.

Neural Stem Cells An Overview

Abstract— This review focuses on the nature and functional properties of stem cells of the adult mammalian central nervous system (CNS). It has recently been shown that cell turnover, including neurons, does occur in the mature CNS, thanks to the persistence of precursor cells that possess the functional characteristics of bona-fide neural stem cells (NSCs) within restricted brain areas. We discuss how the subventricular zone of the forebrain (SVZ) is the most active neurogenetic area and the richest source of NSCs. These NSCs ensure a life-long contribution of new neurons to the olfactory bulb and, when placed in culture, can be grown and extensively expanded for months, allowing the generation of stem cell lines, which maintain stable and constant functional properties. A survey of the differentiation potential of these NSCs, both in vitro and in vivo, outlines their extreme plasticity that seems to outstretch the brain boundaries, so that these neuroectodermal stem cells may give rise to cells that derive from developmentally distinct tissues. A critical discussion of the latest, controversial findings regarding this surprising phenomenon is provided.

Cardiomyocytes induce endothelial cells to trans-differentiate into cardiac muscle: Implications for myocardium regeneration

The concept of tissue-restricted differentiation of postnatal stem cells has been challenged by recent evidence showing pluripotency for hematopoietic, mesenchymal, and neural stem cells. Furthermore, rare but well documented examples exist of already differentiated cells in developing mammals that change fate and trans-differentiate into another cell type. Here, we report that endothelial cells, either freshly isolated from embryonic vessels or established as homogenous cells in culture, differentiate into beating cardiomyocytes and express cardiac markers when cocultured with neonatal rat cardiomyocytes or when injected into postischemic adult mouse heart. Human umbilical vein endothelial cells also differentiate into cardiomyocytes under similar experimental conditions and transiently coexpress von Willebrand factor and sarcomeric myosin. In contrast, neural stem cells, which efficiently differentiate into skeletal muscle, differentiate into cardiomyocytes at a low rate. Fibroblast growth factor 2 and bone morphogenetic protein 4, which activate cardiac differentiation in embryonic cells, do not activate cardiogenesis in endothelial cells or stimulate trans-differentiation in coculture, suggesting that different signaling molecules are responsible for cardiac induction during embryogenesis and in successive periods of development. The fact that endothelial cells can generate cardiomyocytes sheds additional light on the plasticity of endothelial cells during development and opens perspectives for cell autologous replacement therapies.

Mash1 specifies neurons and oligodendrocytes in the postnatal brain

Progenitors in the telencephalic subventricular zone (SVZ) remain mitotically active throughout life, and produce different cell types at embryonic, postnatal and adult stages. Here we show that Mash1, an important proneural gene in the embryonic telencephalon, is broadly expressed in the postnatal SVZ, in progenitors for both neuronal and oligodendrocyte lineages. Moreover, Mash1 is required at birth for the generation of a large fraction of neuronal and oligodendrocyte precursors from the olfactory bulb. Clonal analysis in culture and transplantation experiments in postnatal brain demonstrate that this phenotype reflects a cell‐autonomous function of Mash1 in specification of these two lineages. The conservation of Mash1 function in the postnatal SVZ suggests that the same transcription mechanisms operate throughout life to specify cell fates in this structure, and that the profound changes in the cell types produced reflect changes in the signalling environment of the SVZ.