Proliferative Capacities and Differentiation Potentials of Human Periodontal Ligament Stem Cells after Slow-freezing Cryopreservation

@#Introduction: The cryopreservation of periodontal ligament stem cells (PDLSCs) required a good combination of CPA composition as a step in the preparation of PDLSCs. This study aimed to analyze the proliferative capacities and differentiation potentials of PDLSCs after slow-freezing cryopreservation with CPA in different combinations. Methods: The fourth passage of the primary PDL cells were examined their fibroblast-like morphology and colony forming unit-fibroblast (CFU-F), and characterized by surface markers for mesenchymal stem cells using flow cytometry. PDLSCs were divided into two groups of freshly-PDLSCs (fPDLSCs) and cryopreserved-PDLSCs (cPDLSCs). The PDLSCs were cryopreserved using slow freezing method with CPA in different combinations: 1) 90%FBS+10%DMEM (FD-group), 2) 90%DMEM+10%DMSO (DDs-group), 3) 90%FBS+10%DMSO (FDs-group), and 4) 100% Cell Banker (CB-group) as positive control. The proliferation of fPDLSCs and cPDLSCs were evaluated by trypan blue dye exclusion method. The multipotency of cells was assessed by Oil Red O, Alizarin Red, and Alcian Blue staining. Results: The primary PDL cells had fibroblast-like morphology and CFU-F ability. They expressed more than 95% positive MSC surface markers of CD90, CD73, CD150, and CD44, but showed less than 2% hematopoietic cell markers of CD11b/CD19/CD34/CD45 and HLA-DR. The cPDLSCs viability of FDs-group was 81.5% and 80% in -80oC and LN2, respectively. The fPDLSCs and cPDLSCs proliferation and doubling time were no statistically significant difference (p>0.05). They could differentiate into adipogenic, osteogenic, and chondrogenic differentiation. Conclusion: The cPDLSCs could maintain their proliferative capacities and differentiation potentials after slow-freezing cryopreservation with 90%FBS+10%DMSO in -80oC.

Improvement of Cell Cycle Lifespan and Genetic Damage Susceptibility of Human Mesenchymal Stem Cells by Hypoxic Priming

Hypoxic culture is widely recognized as a method to efficiently expand human mesenchymal stem cells (MSCs) without loss of stem cell properties. However, the molecular basis of how hypoxia priming benefits MSC expansion remains unclear. We report that hypoxic priming markedly extends the cell cycle lifespan rather than augmenting the multipotency of MSC differentiation lineage. Hypoxic priming does not affect to chromosome damage but significantly attenuates the susceptibility of chromosome damage. Our results provide important evidence that multipotency of human MSCs by hypoxic priming is determined by cell cycle lifespan.

Effect of donor age on the proliferation and multipotency of canine adipose-derived mesenchymal stem cells

Research into adipose tissue-derived mesenchymal stem cells (AD-MSCs) has demonstrated the feasibility of their use in clinical applications due to their ease of isolation and abundance in adipose tissue. We isolated AD-MSCs from young and old dogs, and the cells were subjected to sequential sub-passaging from passage 1 (P1) to P7. Canine AD-MSCs (cAD-MSCs) were examined for proliferation kinetics, expression of molecules associated with self-renewal, expression of cell surface markers, and differentiation potentials at P3. Cumulative population doubling level was significantly higher in cAD-MSCs of young donors than in those of old donors. In addition, expressions of CD73, CD80, Oct3/4, Nanog, cell survival genes and differentiation potentials were significantly higher in young donors than in old donors. The present study suggests that donor age should be considered when developing cell-based therapies for clinical application of cAD-MSCs.

Elimination of the geomagnetic field stimulates the proliferation of mouse neural progenitor and stem cells

Living organisms are exposed to the geomagnetic field (GMF) throughout their lifespan. Elimination of the GMF, resulting in a hypogeomagnetic field (HMF), leads to central nervous system dysfunction and abnormal development in animals. However, the cellular mechanisms underlying these effects have not been identified so far. Here, we show that exposure to an HMF (<200 nT), produced by a magnetic field shielding chamber, promotes the proliferation of neural progenitor/stem cells (NPCs/NSCs) from C57BL/6 mice. Following seven-day HMF-exposure, the primary neurospheres (NSs) were significantly larger in size, and twice more NPCs/NSCs were harvested from neonatal NSs, when compared to the GMF controls. The self-renewal capacity and multipotency of the NSs were maintained, as HMF-exposed NSs were positive for NSC markers (Nestin and Sox2), and could differentiate into neurons and astrocyte/glial cells and be passaged continuously. In addition, adult mice exposed to the HMF for one month were observed to have a greater number of proliferative cells in the subventricular zone. These findings indicate that continuous HMF-exposure increases the proliferation of NPCs/NSCs, in vitro and in vivo. HMF-disturbed NPCs/NSCs production probably affects brain development and function, which provides a novel clue for elucidating the cellular mechanisms of the bio-HMF response.

Cancer stem cell: A rogue responsible for tumor development and metastasis.

Cancer stem cells are a small population of cells in a tumor. They have the ability to self-renew and maintain the tumor. The most apt and accepted hypothesis for tumor development is Cancer Stem Cells. This review focuses on this concept of cancer stem cells, serving their purpose and leading to the development of tumor. There are many cell biomarkers which have been described for the identification and characterization of cancer stem cells. The most prominent of the cellular markers for the detection of cancer stem cells; CD133, CD44, ALDH-1 along with some others have been discussed in detail in this review.

Multipotency of cultured olfactory epithelium neural stem cells / 第二军医大学学报

Objective: To investigate the culture method for adult olfactory epithelium neural stem cells and their multipotency during in vitro differentiation. Methods: Olfactory epithelium was sequentially digested with neutral protease and collagenase I A, then the olfactory epithelium neural stem cells were enriched and amplified. Neurospheres were formed by culturing the cells with serum free medium containing bFGF and EGF. Differentiation of neurospheres was induced with medium containing serum; the differentiated cells were identified by immunocytochemistry. Results: Primary cultured olfactory epithelium neural stem cells proliferated and formed colonies. Cells in the colonies expressed the in vivo markers GBC2 and cytokeratin 5. Passaged cells proliferated and formed neurospheres when cultured with serum free medium containing bFGF and EGF. Medium containing serum induced the differentiation of neurospheres. TUJ1 positive neurons, P75NGFR positive olfactory ensheathing cells, GFAP positive astrocytes and NG2 positive oligodendrocyte progenitors were detected in the highly heterogeneous differentiated cells; besides, the cells also contained nestin positive neural stem cells. Conclusion: Cultured adult olfactory epithelium neural stem cells can proliferate and form neurospheres. The neurosphere cells have the multipotency to differentiate into neurons, astrocytes and oligodendrocyte progenitors.

Stem cell properties of cells derived from canine periodontal ligament / 대한치주과학회지

In spite of the attention given to the study of mesenchymal stem cells derived periodontal ligament (PDL), there is a lack of information about canine PDL cells. In this study, we characterized canine PDL cells to clarify their stem cell properties, including self renewal, proliferate rate, stem cell markers and multipotency. PDL cells were obtained from extracted premolars of canines, following a colony forming assay and proliferation rate of sub-confluent cultures of cells for self-renewal, immunostaining for STRO-1 and CD146/MUC18 and a differentiation assay for multipotency. Canine PDL cells formed single-cells colonies and 25% of the PDL cells displayed positive staining for BrdU. The cells expressed the mesenchymal stem-cell markers, STRO-1 and CD146/MUC18. Under defined culture conditions, the cells differentiated into osteoblasts and adipocytes, but the cells didn't differentiated into chondrocytes. The findings of this study indicated that the canine PDL cells possess crucial stem cells properties, such as self-renewal and multipotency, and express the mesenchymal stem cell markers on their surface. The isolation and characterization of canine PDL cells makes it feasible to pursue preclinical models of periodontal regeneration in canine.

Neural Stem Cells

Multipotent neural stem cells (NSCs) are operationally defined by their ability to self-renew, to differentiate into cells of all glial and neuronal lineages throughout the neuraxis, and to populate developing or degenerating CNS regions. Thus their use as a graft material can be considered analogous to hematopoietic stem cell-mediated reconstitution and gene transfer. The recognition that NSCs propagated in culture could be reimplanted into mammalian brain, where they might integrate appropriately throughout the mammalian CNS and stably express foreign genes, has unveiled a new role for neural transplantation and gene therapy and a possible strategy for addressing the CNS manifestations of diseases that heretofore has been refractory to intervention. We have tracked the response of host and transplanted NSCs to brain or spinal cord injury and explored the therapeutic potential of NSCs injected into the animal CNS subjected to focal hypoxic-ische-mic (HI) brain or spinal cord injury. Such cells integrated appropriately into the degenerating CNS, showed robust engraftment and foreign gene expression within the region of CNS injury, and appeared to have migrated preferentially to the site of injury, experienced limited proliferation, and differentiated into neural cells lost to injury, trying to repopulate the damaged CNS area. The transplantation of exogenous NSCs may, in fact, augment a natural self-repair process in which the damaged CNS "attempts" to mobilize its own pool of stem cells. Providing additional NSCs and trophic factors may optimize this response. Therefore, NSCs may provide a novel approach to reconstituting CNS damaged by HI brain or spinal cord injury. Preliminary data in animal models of hypoxic-ischemic brain injury or contusive spinal cord injury lend support to these hypotheses.