Gene expression profiles in murine hematopoietic stem cells revisited: Analysis of cDNA libraries reveals high levels of translational and metabolic activities

Gene expression studies from hematopoietic stem cell (HSC) populations purified to variable degrees have defined a set of sternness genes. Unexpectedly, results also hinted toward a HSC chromatin poised in a wide-open state. With the aim of providing a robust tool for further studies into the molecular biology of HSCs, the studies herein describe the construction and comparative molecular analysis of A-phage cDNA libraries from highly purified HSCs that retained their long-term repopulating activities (long-term HSCs [LT-HSCs]) and from short-term repopulating HSCs that were largely depleted of these activities. Microarray analysis of the libraries confirmed the previous results but also revealed an unforeseen preferential expression of translation- and metabolism-associated genes in the LT-HSCs. Therefore, these data indicate that HSCs are quiescent only in regard of proliferative activities but are in a state of readiness to provide the metabolic and translational activities required after induction of proliferation and exit from the HSC pool.

Stem cells in the periodontal ligament

The ability to identify and manipulate stem cells has been a significant advancement in regenerative medicine and has contributed to the development of tissue engineering-based clinical therapies. Difficulties associated with achieving predictable periodontal regeneration, means that novel techniques such as tissue engineering need to be developed in order to regenerate the extensive soft and hard tissue destruction that results from periodontitis. One of the critical requirements for a tissue engineering approach is the delivery of ex vivo expanded progenitor populations or the mobilization of endogenous progenitor cells capable of proliferating and differentiating into the required tissues. By definition, stem cells fulfill these requirements and the recent identification of stem cells within the periodontal ligament represents a significant development in the progress toward predictable periodontal regeneration. In order to explore the importance of stem cells in periodontal wound healing and regeneration, this review will examine contemporary concepts in stem cell biology, the role of periodontal ligament progenitor cells in the regenerative process, recent developments in identifying periodontal stem cells and the clinical implications of these findings.

A side order of stem cells: The SP phenotype

A defining property of murine hematopoietic stein cells (HSCs) is low fluorescence after staining with Hoechst 33342 and Rhodamine 123. These dyes have proven to be remarkably powerful tools in the purification and characterization of HSCs when used alone or in combination with antibodies directed against stem cell epitopes. Hoechst low cells are described as side population (SP) cells by virtue of their typical profiles in Hoechst red versus Hoechst blue bivariate fluorescent-activated cell sorting dot plots. Recently, excitement has been generated by the findings that putative stem cells from solid tissues may also possess this SP phenotype. SP cells have now been isolated from a wide variety of mammalian tissues based on this same dye efflux phenomenon, and in many cases this cell population has been shown to contain apparently multipotent stem cells. What is yet to be clearly addressed is whether cell fusion accounts for this perceived SP multipotency. Indeed, if low fluorescence after Hoechst staining is a phenotype shared by hematopoietic and organ-specific stem cells, do all resident tissue SP cells have bone marrow origins or might the SP phenotype be a property common to all stem cells? Subject to further analysis, the SP phenotype may prove invaluable for the initial isolation of resident tissue stem cells in the absence of definitive cell-surface markers and may have broad-ranging applications in stem cell biology, from the purification of novel stem cell populations to the development of autologous stem cell therapies.

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.

Transplantation of mesenchymal stem cells promotes an alternative pathway of macrophage activation and functional recovery after spinal cord injury

Abstract Mesenchymal stem cells (MSC) derived from bone marrow can potentially reduce the acute inflammatory response in spinal cord injury (SCI) and thus promote functional recovery. However, the precise mechanisms through which transplanted MSC attenuate inflammation after SCI are still unclear. The present study was designed to investigate the effects of MSC transplantation with a special focus on their effect on macrophage activation after SCI. Rats were subjected to T9-T10 SCI by contusion, then treated 3 days later with transplantation of 1.0×10(6) PKH26-labeled MSC into the contusion epicenter. The transplanted MSC migrated within the injured spinal cord without differentiating into glial or neuronal elements. MSC transplantation was associated with marked changes in the SCI environment, with significant increases in IL-4 and IL-13 levels, and reductions in TNF-a and IL-6 levels. This was associated simultaneously with increased numbers of alternatively activated macrophages (M2 phenotype: arginase-1- or CD206-positive), and decreased numbers of classically activated macrophages (M1 phenotype: iNOS- or CD16/32-positive). These changes were associated with functional locomotion recovery in the MSC-transplanted group, which correlated with preserved axons, less scar tissue formation, and increased myelin sparing. Our results suggested that acute transplantation of MSC after SCI modified the inflammatory environment by shifting the macrophage phenotype from M1 to M2, and that this may reduce the effects of the inhibitory scar tissue in the subacute/chronic phase after injury to provide a permissive environment for axonal extension and functional recovery.

Microspheres as a vehicle for biomolecule delivery to neural stem cells

Neural stem cells (NSC) are a valuable model system for understanding the intrinsic and extrinsic controls for self-renewal and differentiation choice. They also offer a platform for drug screening and neurotoxicity studies, and hold promise for cell replacement therapies for the treatment of neurodegenerative diseases. Fully exploiting the potential of this experimental tool often requires the manipulation of intrinsic cues of interest using transfection methods, to which NSC are relatively resistant. In this paper, we show that mouse and human NSC readily take up polystyrene-based microspheres which can be loaded with a range of chemical or biological cargoes. This uptake can take place in the undifferentiated stage without affecting NSC proliferation and their capacity to give rise to neurons and glia. We demonstrate that ß-galactosidase-loaded microspheres could be efficiently introduced into NSC with no apparent toxic effect, thus providing proof-of-concept for the use of microspheres as an alternative biomolecule delivery system.

Bone marrow-derived mesenchymal stem cells become anti-angiogenic when chondrogenically or osteogenically differentiated:implications for bone and cartilage tissue engineering

Osteochondral tissue repair requires formation of vascularized bone and avascular cartilage. Mesenchymal stem cells stimulate angiogenesis both in vitro and in vivo but it is not known if these proangiogenic properties change as a result of chondrogenic or osteogenic differentiation. We investigated the angiogenic/antiangiogenic properties of equine bone marrow-derived mesenchymal stem cells (eBMSCs) before and after differentiation in vitro. Conditioned media from chondrogenic and osteogenic cell pellets and undifferentiated cells was applied to endothelial tube formation assays using Matrigel™. Additionally, the cell secretome was analysed using LC-MS/MS mass spectrometry and screened for angiogenesis and neurogenesis-related factors using protein arrays. Endothelial tube-like formation was supported by conditioned media from undifferentiated eBMSCs. Conversely, chondrogenic and osteogenic conditioned media was antiangiogenic as shown by significantly decreased length of endothelial tube-like structures and degree of branching compared to controls. Undifferentiated cells produced higher levels of angiogenesis-related proteins compared to chondrogenic and osteogenic pellets. In summary, eBMSCs produce an array of angiogenesis-related proteins and support angiogenesis in vitro via a paracrine mechanism. However, when these cells are differentiated chondrogenically or osteogenically, they produce a soluble factor(s) that inhibits angiogenesis. With respect to osteochondral tissue engineering, this may be beneficial for avascular articular cartilage formation but unfavourable for bone formation where a vascularized tissue is desired. © Copyright 2014, Mary Ann Liebert, Inc.

Influence of Egr-1 in cardiac tissue-derived mesenchymal stem cells in response to glucose variations

Mesenchymal stem cells (MSCs) represent a promising cell population for cell therapy and regenerative medicine applications. However, how variations in glucose are perceived by MSC pool is still unclear. Since, glucose metabolism is cell type and tissue dependent, this must be considered when MSCs are derived from alternative sources such as the heart. The zinc finger transcription factor Egr-1 is an important early response gene, likely to play a key role in the glucose-induced response. Our aim was to investigate how short-term changes in in vitro glucose concentrations affect multipotent cardiac tissue-derived MSCs (cMSCs) in a mouse model of Egr-1 KO (Egr-1-/-). Results showed that loss of Egr-1 does not significantly influence cMSC proliferation. In contrast, responses to glucose variations were observed in wt but not in Egr-1 -/- cMSCs by clonogenic assay. Phenotype analysis by RT-PCR showed that cMSCs Egr-1-/- lost the ability to regulate the glucose transporters GLUT-1 and GLUT-4 and, as expected, the Egr-1 target genes VEGF, TGFβ-1, and p300. Acetylated protein levels of H3 histone were impaired in Egr-1-/- compared to wt cMSCs. We propose that Egr-1 acts as immediate glucose biological sensor in cMSCs after a short period of stimuli, likely inducing epigenetic modifications. © 2014 Daniela Bastianelli et al.