The formation of neurospheres was important in demonstrating that neurogenesis in the adult brain may be fuelled by a stem cell population. The olfactory mucosa is another site of neurogenesis which, in humans, has been observed to contain a stem cell population through the formation of neurospheres (Murrell et al., 2005). Stem cells can be defined as cells capable of self-renewal and multipotency. The aim of this study was to investigate the potential of rat olfactory stem cells growing as neurospheres. The hypothesis is that the rat olfactory mucosa contains a 'true' stem cell population that can be cultured as neurospheres and that will demonstrate multipotency by differentiating into 'non-olfactory' cell types and possess the capacity for self-renewal, if provided with the appropriate environmental niche. Here it was found that adult rat olfactory mucosa is capable of generating neurospheres when cultured in EGF and bFGF. Evidence of self-renewal was provided by the formation of six generations of neurospheres, the formation of neurospheres from single cells and the expression of markers associated with self-renewal by neurosphere cells. The multipotency of olfactory neurosphere cells was demonstrated through manipulation of the stem cell niche. In defined culture conditions, extracellular matrix molecules and growth factors were able to induce the differentiation of neurosphere cells down the dopaminergic lineage pathway. When co-cultured with differentiating cells, neonatal myoblasts and 3T3-L1 cells, olfactory neurosphere cells were able to differentiate and incorporate into a skeletal muscle myotube and differentiate into adipocytes, respectively. In conclusion it was found that the adult rat olfactory mucosa is capable of generating neurospheres. When presented with an appropriate niche neurosphere cells are able to self-renew and demonstrate multipotency.

No description available.

Treatments for skeletal tissue injuries include surgery and rehabilitation but in adult patients, the healing process is slow and incomplete, and the underlying biological mechanisms are largely unknown. Skeletal tissues contain stem cells responsible for their maintenance and repair, but the identity and location of these stem cells, and what molecular mechanisms regulate their fate decisions remain unclear. To design more effective regenerative therapies for skeletal conditions, understanding the fundamental biology of skeletal stem cells (SSC) in postnatal organisms is required. Our project aims at identifying and characterizing these SSC populations in postnatal murine and human tissues using lineage tracing techniques, combined with multicolor 3D confocal microscopy and computational image analysis, in vitro assays, and single cell transcriptomics. We hypothesized that the postnatal skeleton contains self-renewing and multipotent Sox9+ SSCs that persist in adulthood. We showed that the adult mouse skeleton contains Sox9+ cells self-renewing, multipotent skeletal stem cells (SSCs) with osteogenic and chondrogenic potential. They are located adjacent to the growth plates and in periosteum and persist in adulthood. Transcriptome analysis revealed that these cells express other putative SSCs markers, as well as genes involved in skeletal development, stem cell self-renewal, and fate decision. This data provides testable drug targets to pharmacologically manipulate SSCs fate decisions in situ. In addition, we showed that human tissues contain SSCs similar to murine tissues. This is the first experimental proof of self-renewal in postnatal Sox9+ SSCs in vivo. These findings provide actionable insights for the use of culture-expanded stem cell product for regenerative medicine product or pharmacological targeting of these stem cells in situ. We believe our data will help improve stem-cell based and tissue engineering therapies, increasing success rate of regenerative orthopaedic surgeries while reducing reoccurrence of injuries.

Origin of mammalian somatic cells in the developing testes remains unclear. This origin could be explained by established cell culture derived from testes of Xenopus tropicalis juvenile male. The expression profile of the cell culture showed transcription of some pluripotency genes, somatic Sertoli and peritubular myoid cell markers and last but not least, the mesenchymal stem cell markers. Conversely, germ cell genes were downregulated. Immunocytochemical analysis revealed expression of Vimentin, Sox9 and α-smooth muscle actin, indicating that the testicular cell culture is a common mesenchymal progenitor of the Sertoli and peritubular myoid cells and that the cell culture did not arise from spermatogonial stem cells undergoing incomplete reprogramming in vitro. Testing of X. tropicalis cell culture during induction of differentiation in vitro revealed that these cells are probably multipotent with the ability to differentiate into adipocytes, chondroblasts and osteoblasts. The ability to derive multipotent stem cells from the juvenile testes opens new possibilities of using these cells for biotechnology and medicine. Keywords: Testicular somatic cells, Xenopus tropicalis, progenitor, mesenchymal stem cells, induction of differentiation, multipotency

No description available.

Les cellules souches assurent le développement des tissus, leur renouvellement ainsique leur réparation suite à des blessures. L’une des questions clés du domaine de labiologie des cellules souches est l’identification des différents types cellulaires qu’unecellule souche peut donner. Les cellules souches peuvent être multipotentes, c’est-à-direcapables de donner naissance à plusieurs types cellulaires différents, ou unipotentes,c’est-à-dire qu’elles ne peuvent alors se différencier qu’en un seul type cellulaire. Lesexpériences de traçage cellulaire sont réalisées quotidiennement en biologie dudéveloppement et en biologie des cellules souches afin d’évaluer le devenir des cellulessouches in vivo. Cependant, il n’existe à ce jour aucune méthode rigoureuse permettantd’interpréter les résultats d’expériences de traçage cellulaire de manière non ambigüe etde déterminer la multipotence ou l’unipotence des cellules souches avec grande précisionet de manière statistiquement fiable. Nous avons développé de nouvelles méthodes afind’évaluer avec une très grande précision le caractère unipotent ou multipotent descellules souches du sein et de la prostate. Ces nouvelles découvertes démontrent demanière non ambigüe que la prostate provient de cellules souches multipotentes, alorsque seules des cellules souches unipotentes contribuent au développement et auremodelage de la glande mammaire au stade adulte. D’autre part, nous montrons que cesont des cellules souches multipotentes qui sont responsables des phases précoces dudéveloppement embryonnaire de la glande mammaire, et que ces cellules deviennentunipotentes peu avant la naissance. Nous avons étudié les mécanismes régulant le passagede l’état multipotent à l’état unipotent et démontrons que le facteur de transcription p63joue un rôle crucial dans la restriction du potentiel de différenciation des cellules souchesmammaires embryonnaires. Enfin, nous montrons que les cellules souches mammairesadultes, normalement unipotentes, peuvent redevenir multipotentes en conditionsphysiopathologiques telles que l’ablation spécifique d’une lignée cellulaire mammaire ouau cours de l’initiation tumorale. Nous essayons donc de comprendre de manière généraleles mécanismes impliqués dans le passage de l’état unipotent à l’état multipotent descellules souches mammaires adultes, et d’élucider les similarités existant entre lesdifférentes conditions dans lesquelles des cellules souches mammaires multipotentessont observées. / Doctorat en Sciences biomédicales et pharmaceutiques (Médecine) / info:eu-repo/semantics/nonPublished

Sertoli cells (SCs) are somatic cells of testicular tissue that are involved in spermatogenesis and maturation of germ cells. They are currently being extensively studied for their immunomodulatory abilities, and recent studies have shown that they share some properties with mesenchymal stromal cells (MSCs). Detailed characterization of SCs and clarification of their role in testicular tissue is crucial for potential use of SCs as a therapeutic tool in regenerative medicine. Cell culture of Xenopus tropicalis immature Sertoli cells (XtiSCs) and Mus musculus (mSCs) Sertoli cells were established in the Laboratories of Developmental Biology and Immunoregulations, Faculty of Science, Charles University. Previous research has characterized XtiSCs and demonstrated their multipotent potential by in vitro differentiation into a mesodermal line. Following this research, one of the goals of the diploma project was the induction of in vitro differentiation of XtiSC into other cell types, which would verify the differentiation potential of XtiSCs. The mSC expression profile confirmed the somatic origin of this culture as well as the transcription of Sertoli cell gene markers. Differentiation of mSCs along the mesodermal line into osteoblasts, chondrocytes and adipocytes has been successfully induced in vitro....

Hair follicle outer root sheath (ORS) is a putative source of stem cells with therapeutic capacity. ORS contains several multipotent stem cell populations, primarily in the distal compartment of the bulge region. However, the bulge is routinely obtained using invasive isolation methods, which require human scalp tissue ex vivo. Non-invasive sampling has been standardized by means of the plucking procedure, enabling to reproducibly obtain the mid-ORS part. The mid-ORS shows potential for giving rise to multiple stem cell populations in vitro. To demonstrate the phenotypic features of distal, middle, and proximal ORS parts, gene and protein expression profiles were studied in physically separated portions. The mid-part of the ORS showed a comparable or higher NGFR, nestin/NES, CD34, CD73, CD44, CD133, CK5, PAX3, MITF, and PMEL expression on both protein and gene levels, when compared to the distal ORS part. Distinct subpopulations of cells exhibiting small and round morphology were characterized with flow cytometry as simultaneously expressing CD73/CD271, CD49f/CD105, nestin, and not CK10. Potentially, these distinct subpopulations can give rise to cultured neuroectodermal and mesenchymal stem cell populations in vitro. In conclusion, the mid part of the ORS holds the potential for yielding multiple stem cells, in particular mesenchymal stem cells.