We report here that cells co-purifying with mesenchymal stem cells--termed here multipotent adult progenitor cells or MAPCs--differentiate, at the single cell level, not only into mesenchymal cells, but also cells with visceral mesoderm, neuroectoderm and endoderm characteristics in vitro. When injected into an early blastocyst, single MAPCs contribute to most, if not all, somatic cell types. On transplantation into a non-irradiated host, MAPCs engraft and differentiate to the haematopoietic lineage, in addition to the epithelium of liver, lung and gut. Engraftment in the haematopoietic system as well as the gastrointestinal tract is increased when MAPCs are transplanted in a minimally irradiated host. As MAPCs proliferate extensively without obvious senescence or loss of differentiation potential, they may be an ideal cell source for therapy of inherited or degenerative diseases.

/pdf/pluripotency-of-mesenchymal-stem-cells-derived-from-adult-200s4qlizf.pdf

Pluripotency of mesenchymal stem cells derived from adult marrow

Adult Cardiac Stem Cells Are Multipotent and Support Myocardial Regeneration

Mesenchymal stem cells (MSCs) are a heterogeneous subset of stromal stem cells that can be isolated from many adult tissues. They can differentiate into cells of the mesodermal lineage, such as adipocytes, osteocytes and chondrocytes, as well as cells of other embryonic lineages. MSCs can interact with cells of both the innate and adaptive immune systems, leading to the modulation of several effector functions. After in vivo administration, MSCs induce peripheral tolerance and migrate to injured tissues, where they can inhibit the release of pro-inflammatory cytokines and promote the survival of damaged cells. This Review discusses the targets and mechanisms of MSC-mediated immunomodulation and the possible translation of MSCs to new therapeutic approaches.

Mesenchymal stem cells in health and disease

We have used the pH-induced self-assembly of a peptide-amphiphile to make a nanostructured fibrous scaffold reminiscent of extracellular matrix. The design of this peptide-amphiphile allows the nanofibers to be reversibly cross-linked to enhance or decrease their structural integrity. After cross-linking, the fibers are able to direct mineralization of hydroxyapatite to form a composite material in which the crystallographic c axes of hydroxyapatite are aligned with the long axes of the fibers. This alignment is the same as that observed between collagen fibrils and hydroxyapatite crystals in bone.

http://science.sciencemag.org/content/sci/294/5547/1684.full.pdf

Self-assembly and mineralization of peptide-amphiphile nanofibers

Adult marrow-derived Mesenchymal Stem Cells (MSCs) are capable of dividing and their progeny are further capable of differentiating into one of several mesenchymal phenotypes such as osteoblasts, chondrocytes, myocytes, marrow stromal cells, tendon-ligament fibroblasts, and adipocytes. In addition, these MSCs secrete a variety of cytokines and growth factors that have both paracrine and autocrine activities. These secreted bioactive factors suppress the local immune system, inhibit fibrosis (scar formation) and apoptosis, enhance angiogenesis, and stimulate mitosis and differentiation of tissue-intrinsic reparative or stem cells. These effects, which are referred to as trophic effects, are distinct from the direct differentiation of MSCs into repair tissue. Several studies which tested the use of MSCs in models of infarct (injured heart), stroke (brain), or meniscus regeneration models are reviewed within the context of MSC-mediated trophic effects in tissue repair.

/pdf/mesenchymal-stem-cells-as-trophic-mediators-1sjekxtzx0.pdf

Mesenchymal stem cells as trophic mediators.

Myocardial infarction leads to loss of tissue and impairment of cardiac performance The remaining myocytes are unable to reconstitute the necrotic tissue, and the post-infarcted heart deteriorates with time1 Injury to a target organ is sensed by distant stem cells, which migrate to the site of damage and undergo alternate stem cell differentiation2,3,4,5; these events promote structural and functional repair6,7,8 This high degree of stem cell plasticity prompted us to test whether dead myocardium could be restored by transplanting bone marrow cells in infarcted mice We sorted lineage-negative (Lin-) bone marrow cells from transgenic mice expressing enhanced green fluorescent protein9 by fluorescence-activated cell sorting on the basis of c-kit expression10 Shortly after coronary ligation, Lin- c-kitPOS cells were injected in the contracting wall bordering the infarct Here we report that newly formed myocardium occupied 68% of the infarcted portion of the ventricle 9 days after transplanting the bone marrow cells The developing tissue comprised proliferating myocytes and vascular structures Our studies indicate that locally delivered bone marrow cells can generate de novo myocardium, ameliorating the outcome of coronary artery disease

Bone marrow cells regenerate infarcted myocardium

Animal development involves a complex progression of tissue induction and morphogenesis, expansion of precursor cell populations, and the death or terminal differentiation of these cells into specific functional types. Bone morphogenetic protein (BMP) signaling is repeatedly used in this dynamic process for both vertebrates and invertebrates. In gastrulating Xenopus and Drosophila, BMPs and the BMP2/4 homolog Decapentaplegic (Dpp) contribute to specifying the embryonic dorsoventral axis. High activity of BMP/Dpp induces blood/amnioserosa, whereas maximal inhibition of BMP/Dpp leads to neural ectoderm induction (Dale 2000; Nakayama et al. 2000). Subsequently, BMP/Dpp signaling contributes to the specification and/or expansion of many tissues such as vertebrate limb and Drosophila wing (Vogt and Duboule 1999; Day and Lawrence 2000) and neural ectoderm (Cornell and Ohlen 2000).

Vertebrate neural ectoderm undergoes a period of rapid proliferation and morphogenetic movements to form a neural tube, as signals from adjacent tissues induce dorsal–ventral and anterior-posterior identities on precursors (Altmann and Brivanlou 2001). BMPs are expressed at high levels in nonneural ectoderm and then in the roof plate of the neural tube (Liem et al. 1995; Furuta et al. 1997). A BMP activity gradient induces dorsal identity markers indicative of neural crest or dorsal interneuron precursors (Liem et al. 1997; Nguyen et al. 1998, 2000; Barth et al. 1999). However, BMPs also cause apoptosis in early central nervous system (CNS) precursor cells (Graham et al. 1996; Furuta et al. 1997), neuronal differentiation in mid-gestation CNS precursors (Li et al. 1998; Mehler et al. 2000), and glial differentiation in late embryonic or adult CNS precursors (Gross et al. 1996).

It is unclear how BMPs mediate such a wide variety of responses, but it may be a result of diversity in the components of signal transduction. Some BMP-related ligands are known to be involved in the generation of specific cell types (Shah et al. 1996; Lee et al. 1998), but there is evidence that many act in a redundant manner (Dudley and Robertson 1997; Solloway et al. 1998; Zhao et al. 1999). BMPs exert their effects by activating a complex of type I and type II receptors. The type II receptor is the primary ligand-binding component; both BMPRII and ActRIIB are functional type II receptors for BMPs. Type I receptors also have BMP-binding properties but are primarily responsible for transducing the signal into the cell: BMPR-IA (Alk3), BMPR-IB (Alk6), and ActR-I (Alk2) are all known to transduce BMP signals (Kawabata et al. 1998). The activated type I receptors in turn phosphorylate the DNA-binding proteins Smad1, Smad5, and Smad8, each of which can then heteromerize with Smad4 and translocate to the nucleus to regulate transcription of downstream target genes (Itoh et al. 2000).

The expression patterns of Bmpr-1a and Bmpr-1b in the CNS (Dewulf et al. 1995; Zhang et al. 1998) suggest that they have distinct roles in the transduction of BMP signals. In an effort to understand their role in neural development, we used multipotent CNS stem cells as a defined system to understand signals that control proliferation and fate choice (Johe et al. 1996; Studer et al. 1998). We have also developed a vector using POU-regulated genomic elements that control expression of the intermediate filament gene nestin in CNS precursor cells (Josephson et al. 1998). This vector, called pNERV, expresses mutant BMP receptors selectively in CNS stem cells in vitro and in transgenic mice. Using these tools, we show that most of the known actions of BMPs during neural precursor development can be attributed to the distinct actions of BMPR-IA and BMPR-IB. Furthermore, the activities of these receptors are linked sequentially, with BMPR-IA acting first and inducing the expression of Bmpr-1b. These results identify a feed-forward mechanism by which BMPs can control both the production and fate of precursor cells.

/pdf/sequential-actions-of-bmp-receptors-control-neural-precursor-28y1shsmz2.pdf

Sequential actions of BMP receptors control neural precursor cell production and fate

Multipotential stem cells throughout the developing central nervous system have common properties. Among these is expression of the intermediate filament protein nestin and the brain fatty acid binding protein (B-FABP). To determine if common mechanisms control transcription in CNS stem cells, the regulatory elements of these two genes were mapped in transgenic mice. A 257 basepair enhancer of the rat nestin gene is sufficient for expression throughout the embryonic neuroepithelium. This enhancer contains two sites bound by the class III POU proteins Brn-1, Brn-2, Brn-4, and Tst-1. Only one of the two POU sites is required for CNS expression. An adjacent hormone response element is necessary for expression in the dorsal midbrain and forebrain. The regulatory sites of the B-FABP gene are strikingly similar to those of the nestin gene. A hybrid POU/Pbx binding site is recognized in vitro by Pbx-1, Brn-1 and Brn-2. This site is essential for expression in most of the CNS. In addition, a hormone response element is necessary for forebrain expression. Both the nestin and B-FABP genes therefore depend on POU binding sites for general CNS expression, with hormone response elements additionally required for activity in the anterior CNS. These data indicate that regulation by POU proteins and hormone receptors is a general mechanism for CNS stem cell-specific transcription.

POU transcription factors control expression of CNS stem cell-specific genes

Adeno-associated viruses (AAVs) can enable robust and safe gene delivery to the mammalian central nervous system (CNS). While the scientific community has developed numerous neurotropic AAV variants for systemic gene-transfer to the rodent brain, there are few AAVs that efficiently access the CNS of higher order primates. We describe here AAV.CAP-Mac, an engineered AAV variant that enables systemic, brain-wide gene delivery in infants of two Old World primate species—the rhesus macaque (Macaca mulatta) and the green monkey (Chlorocebus sabaeus). We identified CAP-Mac using a multi-species selection strategy, initially screening our library in the adult common marmoset (Callithrix jacchus) and narrowing our pool of test-variants for another round of selection in infant macaques. In individual characterization, CAP-Mac robustly transduces human neurons in vitro and Old World primate neurons in vivo, where it targets all lobes of cortex, the cerebellum, and multiple subcortical regions of disease relevance. We use CAP-Mac for Brainbow-like multicolor labeling of macaque neurons throughout the brain, enabling morphological reconstruction of both medium spiny neurons and cortical pyramidal cells. Because of its broad distribution throughout the brain and high neuronal efficiency in infant Old World primates compared to AAV9, CAP-Mac shows promise for researchers and clinicians alike to unlock novel, noninvasive access to the brain for efficient gene transfer.

/pdf/intravenous-gene-transfer-throughout-the-brain-of-infant-old-2jk9r73h.pdf

Intravenous gene transfer throughout the brain of infant Old World primates using AAV

To determine whether hematopoietic stem cells (HSC) can transform into cardiomyocytes with the potential to repair dead myocardium after infarction, Lin/c-kitI-II HSC were harvested from transgenic mice expressing green fluorescent protein (GFP) and injected in the region bordering the infarct, 3–5 hours after coronary artery occlusion in mice. A band of closely packed cells was identified 7 to 17 days …

James Pickel

Papers

Bone marrow cells regenerate infarcted myocardium

Sequential actions of BMP receptors control neural precursor cell production and fate

POU transcription factors control expression of CNS stem cell-specific genes

Intravenous gene transfer throughout the brain of infant Old World primates using AAV

Exogenous Hematopoietic Stem Cells Can Regenerate Infarcted Myocardium