scholarly journals Multipotent cells can be generated in vitro from several adult human organs (heart, liver, and bone marrow)

Blood ◽  
2007 ◽  
Vol 110 (9) ◽  
pp. 3438-3446 ◽  
Author(s):  
Antonio P. Beltrami ◽  
Daniela Cesselli ◽  
Natascha Bergamin ◽  
Patrizia Marcon ◽  
Silvia Rigo ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Cell Lines ◽  
Adult Stem Cells ◽  
Gene Expression Signature ◽  
Differentiation Potential ◽  
Adult Human ◽  
Wide Range ◽  
Functional Features

Abstract The aims of our study were to verify whether it was possible to generate in vitro, from different adult human tissues, a population of cells that behaved, in culture, as multipotent stem cells and if these latter shared common properties. To this purpose, we grew and cloned finite cell lines obtained from adult human liver, heart, and bone marrow and named them human multipotent adult stem cells (hMASCs). Cloned hMASCs, obtained from the 3 different tissues, expressed the pluripotent state–specific transcription factors Oct-4, NANOG, and REX1, displayed telomerase activity, and exhibited a wide range of differentiation potential, as shown both at a morphologic and functional level. hMASCs maintained a human diploid DNA content, and shared a common gene expression signature, compared with several somatic cell lines and irrespectively of the tissue of isolation. In particular, the pathways regulating stem cell self-renewal/maintenance, such as Wnt, Hedgehog, and Notch, were transcriptionally active. Our findings demonstrate that we have optimized an in vitro protocol to generate and expand cells from multiple organs that could be induced to acquire morphologic and functional features of mature cells even embryologically not related to the tissue of origin.

scholarly journals Are MSCs Stem Cells? Demonstration of in Vitro and in Vivo Stem Cell Properties of Highly Enriched linneg/CD45neg/CD271pos/CD140alow/Neg Mesenchymal Stromal Cells from Adult Human Bone Marrow

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 4374-4374
Author(s):  
Roshanak Ghazanfari ◽  
Hongzhe Li ◽  
Dimitra Zacharaki ◽  
Simón Méndez-Ferrer ◽  
Stefan Scheding
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Human Bone ◽  
Human Bone Marrow ◽  
Differentiation Potential ◽  
Self Renewal ◽  
Adult Human

Abstract Human bone marrow contains a rare population of non-hematopoietic mesenchymal stromal cells (BM-MSC) with multilineage differentiation capacity, which are essential constituents of the hematopoietic microenvironment. Self-renewal and differentiation are the two key properties of somatic stem cells, however, stem cell properties of human adult BM-MSC have not been demonstrated conclusively yet. We have previously shown that low/negative expression of PDGFRα on linneg/CD45neg/CD271pos cells identified a highly enriched population of primary BM-MSC in adult human bone marrow (Li et al. Blood, 2013, 122:3699). Based on this work, the current study aimed to investigate the in-vitro and in-vivo stem cell properties of this putative stromal stem cell population. The in-vitro clonogenic potential of freshly sorted human linneg/CD45neg/CD271pos/PDGFRlow/neg cells was evaluated by utilizing the CFU-F assay as well as the recently-developed mesensphere assay, which enables MSC amplification while preserving an immature phenotype (Isern et al, Cell Reports 2013, 30: 1714-24). Comparable colony frequencies were obtained with both assays (19.3 ± 2 and 17.5 ± 2.3 CFU-F and spheres per 100 plated cells, respectively, n=6, p=0.19). In order to test whether both assays identified the same population of clonogenic cells, colonies and spheres were replated under both conditions for up to three generations. The results showed comparable capacities of CFU-F and mesenspheres to form secondary and tertiary CFU-F and spheres. In-vitro self-renewal as indicated by increasing numbers of CFU-F and spheres (416.6 ± 431.7-fold and 49.5 ± 65.7-fold, respectively, n=3) was observed up to the third generation and decreased thereafter. The total number of generations was five (CFU-F) and six (spheres). In-vitro differentiation assays with both, CFU-F- and sphere-derived cells (tested until passage three) demonstrated tri-lineage differentiation potential (adipocytes, osteoblasts, chondrocytes). In addition, CFU-Fs and spheres had comparable surface marker profiles (CD73, CD90, CD105, and HLA-ABC positive; CD31, CD34 and HLA-DR negative), except for CD90, which was higher expressed on CFU-Fs. To investigate in-vivo self-renewal and differentiation potential of the putative stromal stem cells, linneg/CD45neg/CD271pos/PDGFRlow/neg -derived CFU-F and spheres were serially transplanted s.c into NSG mice. After 8 weeks, implants were harvested, human cells were FACS-isolated (CD90 and CD105 expression), and re-assayed under CFU-F and sphere conditions. Whereas in-vivo self-renewal of CFU-F could not be shown (111.5 ± 36 –fold decrease in total CFU-F numbers after primary transplantation, n=3), sphere self-renewal was clearly demonstrated by increased numbers of spheres after primary as well as secondary transplantation (1.13 ± 0.05 and 2.06 ± 0.26 –fold, respectively, n=3), which is remarkable given the fact that the number of recovered human cells is underestimated due to the isolation approach. Here, confirming GFP-marking experiments are ongoing. Finally, preliminary data indicate that linneg/CD45neg/CD271pos/PDGFRlow/neg –derived spheres display full in-vivo differentiation capacity in primary and secondary transplantations. Taken together, our data demonstrate - for the first time - that primary human linneg/CD45neg/CD271pos/PDGFRlow/neg cells meet stringent stem cell criteria, i.e. in-vitro and in-vivo self-renewal and differentiation. These findings answer the long-open question of the potential stem cell properties of adult human MSC and will enable to better understand the properties of native BM-MSC and their biological role in the bone marrow. Disclosures No relevant conflicts of interest to declare.

scholarly journals Cell Source-Dependent In Vitro Chondrogenic Differentiation Potential of Mesenchymal Stem Cell Established from Bone Marrow and Synovial Fluid of Camelus dromedarius

Animals ◽  
2021 ◽  
Vol 11 (7) ◽  
pp. 1918
Author(s):  
Young-Bum Son ◽  
Yeon Ik Jeong ◽  
Yeon Woo Jeong ◽  
Mohammad Shamim Hossein ◽  
Per Olof Olsson ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Synovial Fluid ◽  
Cartilage Regeneration ◽  
Cartilage Tissue ◽  
Chondrocyte Differentiation ◽  
Differentiation Potential ◽  
Proliferation Capacity

Mesenchymal stem cells (MSCs) are promising multipotent cells with applications for cartilage tissue regeneration in stem cell-based therapies. In cartilage regeneration, both bone marrow (BM-MSCs) and synovial fluid (SF-MSCs) are valuable sources. However, the cellular characteristics and chondrocyte differentiation potential were not reported in either of the camel stem cells. The in vitro chondrocyte differentiation competence of MSCs, from (BM and SF) sources of the same Camelus dromedaries (camel) donor, was determined. Both MSCs were evaluated on pluripotent markers and proliferation capacity. After passage three, both MSCs showed fibroblast-like morphology. The proliferation capacity was significantly increased in SF-MSCs compared to BM-MSCs. Furthermore, SF-MSCs showed an enhanced expression of transcription factors than BM-MSCs. SF-MSCs exhibited lower differentiation potential toward adipocytes than BM-MSCs. However, the osteoblast differentiation potential was similar in MSCs from both sources. Chondrogenic pellets obtained from SF-MSCs revealed higher levels of chondrocyte-specific markers than those from BM-MSCs. Additionally, glycosaminoglycan (GAG) content was elevated in SF-MSCs related to BM-MSCs. This is, to our knowledge, the first study to establish BM-MSCs and SF-MSCs from the same donor and to demonstrate in vitro differentiation potential into chondrocytes in camels.

scholarly journals Sustained, retransplantable, multilineage engraftment of highly purified adult human bone marrow stem cells in vivo

Blood ◽  
1996 ◽  
Vol 88 (11) ◽  
pp. 4102-4109 ◽  
Author(s):  
CI Civin ◽  
G Almeida-Porada ◽  
MJ Lee ◽  
J Olweus ◽  
LW Terstappen ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Cell Population ◽  
Mononuclear Cells ◽  
Cell Subset ◽  
Fetal Sheep ◽  
Adult Human

Abstract Data from many laboratory and clinical investigations indicate that CD34+ cells comprise approximately 1% of human bone marrow (BM) mononuclear cells, including the progenitor cells of all the lymphohematopoietic lineages and lymphohematopoietic stem cells (stem cells). Because stem cells are an important but rare cell type in the CD34+ cell population, investigators have subdivided the CD34+ cell population to further enrich stem cells. The CD34+/CD38-cell subset comprises less than 10% of human CD34+ adult BM cells (equivalent to < 0.1% of marrow mononuclear cells), lacks lineage (lin) antigens, contains cells with in vitro replating capacity, and is predicted to be highly enriched for stem cells. The present investigation tested whether the CD34+/CD38-subset of adult human marrow generates human hematopoiesis after transfer to preimmune fetal sheep. CD34+/ CD38- cells purified from marrow using immunomagnetic microspheres or fluorescence-activated cell sorting generated easily detectable, long- term, multilineage human hematopoiesis in the human-fetal sheep in vivo model. In contrast, transfer of CD34+/CD38+ cells to preimmune fetal sheep generated only short-term human hematopoiesis, possibly suggesting that the CD34+/CD38+ cell population contains relatively early multipotent hematopoletic progenitor cells, but not stem cells. This work extends the prior in vitro evidence that the earliest cells in fetal and adult human marrow lack CD38 expression. In summary, the CD34+/ CD38-cell population has a high capacity for long-term multilineage hematopoietic engraftment, suggesting the presence of stem cells in this minor adult human marrow cell subset.

scholarly journals AGE-ASSOCIATED INCREASE IN KYNURENINE INHIBITS AUTOPHAGY AND PROMOTES SENESCENCE IN BONE MARROW STEM CELLS

2019 ◽  
Vol 3 (Supplement_1) ◽  
pp. S956-S956
Author(s):  
Dmitry Kondrikov ◽  
Ahmed Elmansi ◽  
Xing-ming Shi ◽  
Sadanand Fulzele ◽  
Meghan mcGee-Lawrence ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Cell Lines ◽  
Free Radical Oxidation ◽  
Autophagic Flux ◽  
Galactosidase Activity ◽  
Activity Increase ◽  
Age Related ◽  
Old Mice

Abstract Aging is characterized by progressive decline of tissue functionality and age-related accumulation of cellular and molecular damage leading to multiple pathological conditions including osteoporosis and increased fracture rates. Bone marrow mesenchymal stem cells (BMSCs) play an essential role in bone development and regeneration with their ability to undergo differentiation into osteogenic, chondrogenic, myogenic, and adipogenic cell lines cell lines. Proliferation rate of MSC is declined with ages leading to misbalance between bone resorption and osteogenesis. A recently identified age-related change in bone and bone marrow is an accumulation of tryptophan metabolite, kynurenine (KYN), catalyzed by indoleamine-2,3-dioxygenase (IDO) or free-radical oxidation. We previously reported that KYN suppresses autophagy in BMSC. We now investigated the effect of KYN on BMSC cellular function. In vitro treatment of murine BMSC isolated from 18 month old mice with kynurenine disrupted autophagy suppressing autophagic flux. KYN treatment also induces senescence in BMSC marked by increase in SA-beta-galactosidase activity as well as, increased expression of senescence marker p21. Inhibition of Aryl Hydrocarbon Receptor (AhR) by AhR inhibitors significantly reduced β-galactosidase activity increase and blocked p21 expression elevation suggesting that KYN induces senescence in BMSC through the AhR pathway. Interestingly, KYN treatment failed to up-regulate beta-gal activity in BMSC isolated from 6 month-old mice suggesting that KYN induction of senescence maybe potentiated with aging. Together those data support the idea that KYN shifts the homeostatic balance of BMSC during prolonged stress or in aging through downregulating survival autophagic pathway in favor of driving BMSCs to senescence.

296 IN VITRO DIFFERENTIATION OF PORCINE BONE MARROW-DERIVED MESENCHYMAL STEM CELLS INTO HEPATOCYTE-LIKE CELLS

2013 ◽  
Vol 25 (1) ◽  
pp. 295
Author(s):  
B. Mohana Kumar ◽  
W. J. Lee ◽  
Y. M. Lee ◽  
R. Patil ◽  
S. L. Lee ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Class Ii ◽  
In Vitro Differentiation ◽  
Rt Pcr ◽  
Hepatic Differentiation ◽  
Alizarin Red ◽  
Differentiation Potential

Mesenchymal stem cells (MSC) are isolated from bone marrow or other tissues, and have properties of self renewal and multilineage differentiation ability. The current study investigated the in vitro differentiation potential of porcine bone marrow derived MSCs into hepatocyte-like cells. The MSC were isolated from the bone marrow of adult miniature pigs (7 months old, T-type, PWG Micro-pig®, PWG Genetics, Seoul, Korea) and adherent cells with fibroblast-like morphology were cultured on plastic. Isolated MSCs were positive for CD29, CD44, CD73, CD90, and vimentin, and negative for CD34, CD45, major histocompatibility complex-class II (MHC-class II), and swine leukocyte antigen-DR (SLA-DR) by flow cytometry analysis. Further, trilineage differentiation of MSC into osteocytes (alkaline phosphatase, von Kossa and Alizarin red), adipocytes (Oil Red O), and chondrocytes (Alcian blue) was confirmed. Differentiation of MSC into hepatocyte-like cells was induced with sequential supplementation of growth factors, cytokines, and hormones for 21 days as described previously (Taléns-Visconti et al. 2006 World J. Gastroenterol. 12, 5834–5845). Morphological analysis, expression of liver-specific markers, and functional assays were performed to evaluate the hepatic differentiation of MSC. Under hepatogenic conditions, MSC acquired cuboidal morphology with cytoplasmic granules. These hepatocyte-like cells expressed α-fetoprotein (AFP), albumin (ALB), cytokeratin 18 (CK18), cytochrome P450 7A1 (CYP7A1), and hepatocyte nuclear factor 1 (HNF-1) markers by immunofluorescence assay. In addition, the expression of selected markers was demonstrated by Western blotting analysis. In accordance with these features, RT-PCR revealed transcripts of AFP, ALB, CK18, CYP7A1, and HNF-1α. Further, the relative expression levels of these transcripts were analysed by quantitative RT-PCR after normalizing to the expression of the endogenous control, glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Data were analysed statistically by one-way ANOVA using PASW statistics 18 (SPSS Inc., Chicago, IL, USA), and significance was considered at P < 0.05. The results showed that the relative expressions of selected marker genes in hepatocyte-like cells were significantly increased compared with that in untreated MSC. The generated hepatocyte-like cells showed glycogen storage as analysed by periodic acid-Schiff (PAS) staining. Moreover, the induced cells produced urea at Day 21 of culture compared with control MSC. In conclusion, our results indicate the potential of porcine MSC to differentiate in vitro into hepatocyte-like cells. Further studies on the functional properties of hepatocyte-like cells are needed to use porcine MSC as an ideal source for liver cell therapy and preclinical drug evaluation. This work was supported by Basic Science Research Program through the National Research Foundation (NRF), funded by the Ministry of Education, Science and Technology (2010-0010528) and the Next-Generation BioGreen 21 Program (No. PJ009021), Rural Development Administration, Republic of Korea.

scholarly journals The impact of cryopreservation on bone marrow-derived mesenchymal stem cells: a systematic review

2019 ◽  
Vol 17 (1) ◽  
Author(s):  
Soukaina Bahsoun ◽  
Karen Coopman ◽  
Elizabeth C. Akam
Keyword(s):  
Systematic Review ◽  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Cell Types ◽  
Surface Marker Expression ◽  
Wide Range ◽  
And Migration ◽  
The Impact

AbstractMesenchymal stem cells (MSCs) represent an invaluable asset for the field of cell therapy. Human Bone marrow-derived MSCs (hBM-MSCs) are one of the most commonly used cell types in clinical trials. They are currently being studied and tested for the treatment of a wide range of diseases and conditions. The future availability of MSCs therapies to the public will require a robust and reliable delivery process. Cryopreservation represents the gold standard in cell storage and transportation, but its effect on BM-MSCs is still not well established. A systematic review was conducted to evaluate the impact of cryopreservation on BM-MSCs and to attempt to uncover the reasons behind some of the controversial results reported in the literature. Forty-one in vitro studies were analysed, and their results organised according to the cell attributes they assess. It was concluded that cryopreservation does not affect BM-MSCs morphology, surface marker expression, differentiation or proliferation potential. However, mixed results exist regarding the effect on colony forming ability and the effects on viability, attachment and migration, genomic stability and paracrine function are undefined mainly due to the huge variabilities governing the cryopreservation process as a whole and to the lack of standardised assays.

scholarly journals Generation of multipotent cell lines from a distinct population of male germ line stem cells

Reproduction ◽  
2008 ◽  
Vol 135 (6) ◽  
pp. 771-784 ◽  
Author(s):  
Fariborz Izadyar ◽  
Francis Pau ◽  
Joel Marh ◽  
Natalia Slepko ◽  
Tracy Wang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Germ Cell ◽  
Cell Lines ◽  
Germ Line ◽  
Es Cells ◽  
Embryonic Stem ◽  
Complex Mixture ◽  
Neural Cells ◽  
Differentiation Potential

Spermatogonial stem cells (SSCs) maintain spermatogenesis by self-renewal and generation of spermatogonia committed to differentiation. Under certain in vitro conditions, SSCs from both neonatal and adult mouse testis can reportedly generate multipotent germ cell (mGC) lines that have characteristics and differentiation potential similar to embryonic stem (ES) cells. However, mGCs generated in different laboratories showed different germ cell characteristics, i.e., some retain their SSC properties and some have lost them completely. This raises an important question: whether mGC lines have been generated from different subpopulations in the mouse testes. To unambiguously identify and track germ line stem cells, we utilized a transgenic mouse model expressing green fluorescence protein under the control of a germ cell-specific Pou5f1 (Oct4) promoter. We found two distinct populations among the germ line stem cells with regard to their expression of transcription factor Pou5f1 and c-Kit receptor. Only the POU5F1+/c-Kit+ subset of mouse germ line stem cells, when isolated from either neonatal or adult testes and cultured in a complex mixture of growth factors, generates cell lines that express pluripotent ES markers, i.e., Pou5f1, Nanog, Sox2, Rex1, Dppa5, SSEA-1, and alkaline phosphatase, exhibit high telomerase activity, and differentiate into multiple lineages, including beating cardiomyocytes, neural cells, and chondrocytes. These data clearly show the existence of two distinct populations within germ line stem cells: one destined to become SSC and the other with the ability to generate multipotent cell lines with some pluripotent characteristics. These findings raise interesting questions about the relativity of pluripotency and the plasticity of germ line stem cells.

Inhibition of PDGFRβ by Imatinib Mesylate Suppresses Proliferation and Alters Differentiation of Human Mesenchymal Stem Cells In Vitro.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 2563-2563
Author(s):  
Fernando Fierro ◽  
Thomas Illmer ◽  
Duhoui Jing ◽  
Philip Le Coutre ◽  
Gerhard Ehninger ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Tyrosine Kinase ◽  
Hematopoietic Stem Cells ◽  
Imatinib Mesylate ◽  
Dependent Manner ◽  
Differentiation Potential ◽  
Hematopoietic Stem

Abstract Recent data show that the tyrosine kinase inhibitor Imatinib mesylate (IM) also affects normal hematopoietic stem cells (HSC), T lymphocyte activation and dendritic cell function not relying on the specific inhibition of bcr-abl activity. Mesenchymal stem cells (MSC) have been identified in the bone marrow (BM) as multipotent non-hematopoietic progenitor cells that differentiate into osteoblasts, adipocytes, chondrocytes, tenocytes, skeletal myocytes, and cells of visceral mesoderm. MSC interact with HSC, influencing their homing and differentiation through cell-cell contact and the production of factors including chemokines We evaluated possible effects of IM in vitro on human bone marrow-derived MSC. Screening the activity of fourty-two receptor tyrosine kinases by a phospho-receptor tyrosine kinase (RTK)-array revealed an exclusive inhibition of platelet-derived growth factor receptor (PDGFRβ) by IM which consequently affects downstream targets of PDGFRβ as Akt and Erk1/2 signalling pathways in a concentration and time dependent manner. Furthermore, perinuclear multivesicular bodies harbouring PDGFRβ were found within 18–20 hours culture of MSC in the presence of 5 μM IM. Cell proliferation and clonogenicity (evaluated as the capability to form colony forming units - fibroblasts (CFU-F)) of MSC were significantly inhibited by IM in a concentration dependent fashion. IM inhibits significantly the differentiation process of MSC into osteoblasts as evaluated by decreased alkaline phosphatase activity and reduced calcium phosphate precipitates. In contrary, differentiation of MSC into adipocytes was strongly favoured in presence of IM. All these functional deficits described, probably contribute to an observed 50% reduction in the support of clonogenic hematopoietic stem cells, as evaluated by a long term culture-initiating cells (LTC-IC)-based assay. In summary our experiments show that IM inhibits the capacity of human MSC to proliferate and to differentiate into the osteogenic lineage, favouring adipogenesis. This effect is mainly mediated by an inhibition of PDGFRβ autophosphorylation leading to a more pronounced inhibition of PI3K/Akt compared to Erk1/2 signalling. This work confirms the role of PDGFRβ recently described for the proliferation and differentiation potential of MSC and provides a first possible explanation for the altered bone metabolism found in certain patients treated with IM.

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