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Human stromal (mesenchymal) stem cells from bone marrow, adipose tissue and skin exhibit differences in molecular phenotype and differentiation potential.

ABSTRACT: Human stromal (mesenchymal) stem cells (hMSCs) are multipotent stem cells with ability to differentiate into mesoderm-type cells e.g. osteoblasts and adipocytes and thus they are being introduced into clinical trials for tissue regeneration. Traditionally, hMSCs have been isolated from bone marrow, but the number of cells obtained is limited. Here, we compared the MSC-like cell populations, obtained from alternative sources for MSC: adipose tissue and skin, with the standard phenotype of human bone marrow MSC (BM-MSCs). MSC from human adipose tissue (human adipose stromal cells (hATSCs)) and human skin (human adult skin stromal cells, (hASSCs) and human new-born skin stromal cells (hNSSCs)) grew readily in culture and the growth rate was highest in hNSSCs and lowest in hATSCs. Compared with phenotype of hBM-MSC, all cell populations were CD34(-), CD45(-), CD14(-), CD31(-), HLA-DR(-), CD13(+), CD29(+), CD44(+), CD73(+), CD90(+),and CD105(+). When exposed to in vitro differentiation, hATSCs, hASSCs and hNSSCs exhibited quantitative differences in their ability to differentiate into adipocytes and to osteoblastic cells. Using a microarray-based approach we have unveiled a common MSC molecular signature composed of 33 CD markers including known MSC markers and several novel markers e.g. CD165, CD276, and CD82. However, significant differences in the molecular phenotype between these different stromal cell populations were observed suggesting ontological and functional differences. In conclusion, MSC populations obtained from different tissues exhibit significant differences in their proliferation, differentiation and molecular phenotype, which should be taken into consideration when planning their use in clinical protocols.

SUBMITTER: Al-Nbaheen M

PROVIDER: S-EPMC3563956 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundMesenchymal stem cells (MSCs) are known to have different differentiation potential depending on the tissue of origin. Dedifferentiated fat cells (DFATs) are MSC-like multipotent cells that can be prepared from mature adipocytes by ceiling culture method. It is still unknown whether DFATs derived from adipocytes in different tissue showed different phenotype and functional properties. In the present study, we prepared bone marrow (BM)-derived DFATs (BM-DFATs), BM-MSCs, subcutaneous (SC) adipose tissue-derived DFATs (SC-DFATs), and adipose tissue-derived stem cells (ASCs) from donor-matched tissue samples. Then, we compared their phenotypes and multilineage differentiation potential in vitro. We also evaluated in vivo bone regeneration ability of these cells using a mouse femoral fracture model.MethodsBM-DFATs, SC-DFATs, BM-MSCs, and ASCs were prepared from tissue samples of knee osteoarthritis patients who received total knee arthroplasty. Cell surface antigens, gene expression profile, and in vitro differentiation capacity of these cells were determined. In vivo bone regenerative ability of these cells was evaluated by micro-computed tomography imaging at 28 days after local injection of the cells with peptide hydrogel (PHG) in the femoral fracture model in severe combined immunodeficiency mice.ResultsBM-DFATs were successfully generated at similar efficiency as SC-DFATs. Cell surface antigen and gene expression profiles of BM-DFATs were similar to those of BM-MSCs, whereas these profiles of SC-DFATs were similar to those of ASCs. In vitro differentiation analysis revealed that BM-DFATs and BM-MSCs had higher differentiation tendency toward osteoblasts and lower differentiation tendency toward adipocytes compared to SC-DFATs and ASCs. Transplantation of BM-DFATs and BM-MSCs with PHG enhanced bone mineral density at the injection sites compared to PHG alone in the mouse femoral fracture model.ConclusionsWe showed that phenotypic characteristics of BM-DFATs were similar to those of BM-MSCs. BM-DFATs exhibited higher osteogenic differentiation potential and bone regenerative ability compared to SC-DFATs and ASCs. These results suggest that BM-DFATs may be suitable sources of cell-based therapies for patients with nonunion bone fracture.

Project description:BackgroundOsteogenesis Imperfecta (OI) (OMIM %259450) is a heterogeneous group of inherited disorders characterized by increased bone fragility, with clinical severity ranging from mild to lethal. The majority of OI cases are caused by mutations in COL1A1 or COL1A2. Bruck Syndrome (BS) is a further recessively-inherited OI-like phenotype in which bone fragility is associated with the unusual finding of pterygia and contractures of the large joints. Notably, several studies have failed to show any abnormalities in the biosynthesis of collagen 1 in BS patientes. Evidence was obtained for a specific defect of the procollagen telopeptide lysine hydroxylation in BS, whereas mutations in the gene PLOD2 have been identified. Recently, several studies described FKBP10 mutations in OI-like and BS patients, suggesting that FKBP10 is a bonafide BS locus.MethodsWe analyzed the coding region and intron/exon boundaries of COL1A1, COL1A2, PLOD2 and FKBP10 genes by sequence analysis using an ABI PRISM 3130 automated sequencer and Big Dye Terminator Sequencing protocol. Mononuclear cells obtained from the bone marrow of BS, OI patients and healthy donors were cultured and osteogenic differentiation was induced. The gene expression of osteoblast specific markers were also evaluated during the osteoblastic differentiation of mesenchymal stem cell (MSC) by qRT-PCR using an ABI7500 Sequence Detection System.ResultsNo mutations in COL1A1, COL1A2 or PLOD2 were found in BS patient. We found a homozygous 1-base-pair duplication (c.831dupC) that is predicted to produce a translational frameshift mutation and a premature protein truncation 17 aminoacids downstream (p.Gly278ArgfsX95). The gene expression of osteoblast specific markers BGLAP, COL1A1, MSX2, SPARC and VDR was evaluated by Real Time RT-PCR during differentiation into osteoblasts and results showed similar patterns of osteoblast markers expression in BS and healthy controls. On the other hand, when compared with OI patients, the expression pattern of these genes was found to be different.ConclusionsOur work suggests that the gene expression profiles observed during mesenchymal stromal cell differentiation into osteoblast are distinct in BS patients as compared to OI patients. The present study shows for the first time that genes involved in osteogenesis are differentially expressed in BS and OI patients.

Project description:Mesenchymal stem cells (MSCs) are highly responsive to cues in the microenvironment (niche) that must be recapitulated ex vivo to study their authentic behavior. In this study, we hypothesized that native bone marrow (BM)- and adipose (AD)-derived extracellular matrices (ECM) were unique in their ability to control MSC behavior. To test this, we compared proliferation and differentiation of bone marrow (BM)-derived MSCs when maintained on native decellularized ECM produced by BM versus AD stromal cells (i.e. BM- versus AD-ECM). We found that both ECMs contained similar types of collagens but differed in the relative abundance of each. Type VI collagen was the most abundant (≈60% of the total collagen present), while type I was the next most abundant at ≈30%. These two types of collagen were found in nearly equal proportions in both ECMs. In contrast, type XII collagen was almost exclusively found in AD-ECM, while types IV and V were only found in BM-ECM. Physically and mechanically, BM-ECM was rougher and stiffer, but less adhesive, than AD-ECM. During 14 days in culture, both ECMs supported BM-MSC proliferation better than tissue culture plastic (TCP), although MSC-related surface marker expression remained relatively high on all three culture surfaces. BM-MSCs cultured in osteogenic (OS) differentiation media on BM-ECM displayed a significant increase in calcium deposition in the matrix, indicative of osteogenesis, while BM-MSCs cultured on AD-ECM in the presence of adipogenic (AP) differentiation media showed a significant increase in Oil Red O staining, indicative of adipogenesis. Further, culture on BM-ECM significantly increased BM-MSC-responsiveness to rhBMP-2 (an osteogenic inducer), while culture on AD-ECM enhanced responsiveness to rosiglitazone (an adipogenic inducer). These findings support our hypothesis and indicate that BM- and AD-ECMs retain unique elements, characteristic of their tissue-specific microenvironment (niche), which promote retention of MSC differentiation state (i.e. "stemness") during expansion and direct cell response to lineage-specific inducers. This study provides a new paradigm for precisely controlling MSC fate to a desired cell lineage for tissue-specific cell-based therapies.

Project description:BackgroundTherapeutic efficacy of various mesenchymal stromal cell (MSC) types for orthopaedic applications is currently being investigated. While the concept of MSC therapy is well grounded in the basic science of healing and regeneration, little is known about individual MSC populations in terms of their propensity to promote the repair and/or regeneration of specific musculoskeletal tissues. Two promising MSC sources, adipose and amnion, have each demonstrated differentiation and extracellular matrix (ECM) production in the setting of musculoskeletal tissue regeneration. However, no study to date has directly compared the differentiation potential of these 2 MSC populations.PurposeTo compare the ability of human adipose- and amnion-derived MSCs to undergo osteogenic and chondrogenic differentiation.Study designControlled laboratory study.MethodsMSC populations from the human term amnion were quantified and characterized via cell counting, histologic assessment, and flow cytometry. Differentiation of these cells in comparison to commercially purchased human adipose-derived mesenchymal stromal cells (hADSCs) in the presence and absence of differentiation media was evaluated via reverse transcription polymerase chain reaction (PCR) for bone and cartilage gene transcript markers and histology/immunohistochemistry to examine ECM production. Analysis of variance and paired t tests were performed to compare results across all cell groups investigated.ResultsThe authors confirmed that the human term amnion contains 2 primary cell types demonstrating MSC characteristics-(1) human amniotic epithelial cells (hAECs) and (2) human amniotic mesenchymal stromal cells (hAMSCs)-and each exhibited more than 90% staining for MSC surface markers (CD90, CD105, CD73). Average viable hAEC and hAMSC yields at harvest were 2.3 × 106 ± 3.7 × 105 and 1.6 × 106 ± 4.7 × 105 per milliliter of amnion, respectively. As well, hAECs and hAMSCs demonstrated significantly greater osteocalcin ( P = .025), aggrecan ( P < .0001), and collagen type 2 ( P = .044) gene expression compared with hADSCs, respectively, after culture in differentiation medium. Moreover, both hAECs and hAMSCs produced significantly greater quantities of mineralized ( P < .0001) and cartilaginous ( P = .0004) matrix at earlier time points compared with hADSCs when cultured under identical osteogenic and chondrogenic differentiation conditions, respectively.ConclusionAmnion-derived MSCs demonstrate a greater differentiation potential toward bone and cartilage compared with hADSCs.Clinical relevanceAmniotic MSCs may be the source of choice in the regenerative treatment of bone or osteochondral musculoskeletal disease. They show significantly higher yields and better differentiation toward these tissues than MSCs derived from adipose.

Project description:BackgroundMesenchymal stromal cells (MSC) hold promise for both cell replacement and immune modulation strategies owing to their progenitor and non-progenitor functions, respectively. Characterization of MSC from different sources is an important and necessary step before clinical use of these cells is widely adopted. Little is known about the biology and function of canine MSC compared to their mouse or human counterparts. This knowledge-gap impedes development of canine evidence-based MSC technologies.Hypothesis and objectivesWe hypothesized that canine adipose tissue (AT) and bone marrow (BM) MSC (derived from the same dogs) will have similar differentiation and immune modulatory profiles. Our objectives were to evaluate progenitor and non-progenitor functions as well as other characteristics of AT- and BM-MSC including 1) proliferation rate, 2) cell surface marker expression, 3) DNA methylation levels, 4) potential for trilineage differentiation towards osteogenic, adipogenic, and chondrogenic cell fates, and 5) immunomodulatory potency in vitro.Results1) AT-MSC proliferated at more than double the rate of BM-MSC (population doubling times in days) for passage (P) 2, AT: 1.69, BM: 3.81; P3, AT: 1.80, BM: 4.06; P4, AT: 2.37, BM: 5.34; P5, AT: 3.20, BM: 7.21). 2) Canine MSC, regardless of source, strongly expressed cell surface markers MHC I, CD29, CD44, and CD90, and were negative for MHC II and CD45. They also showed moderate expression of CD8 and CD73 and mild expression of CD14. Minor differences were found in expression of CD4 and CD34. 3) Global DNA methylation levels were significantly lower in BM-MSC compared to AT-MSC. 4) Little difference was found between AT- and BM-MSC in their potential for adipogenesis and osteogenesis. Chondrogenesis was poor to absent for both sources in spite of adding varying levels of bone-morphogenic protein to our standard transforming growth factor (TGF-β3)-based induction medium. 5) Immunomodulatory capacity was equal regardless of cell source when tested in mitogen-stimulated lymphocyte reactions. Priming of MSC with pro-inflammatory factors interferon-gamma and/or tumour necrosis factor did not increase the lymphocyte suppressive properties of the MSC compared to untreated MSC.Conclusions/significanceNo significant differences were found between AT- and BM-MSC with regard to their immunophenotype, progenitor, and non-progenitor functions. Both MSC populations showed strong adipogenic and osteogenic potential and poor chondrogenic potential. Both significantly suppressed stimulated peripheral blood mononuclear cells. The most significant differences found were the higher isolation success and proliferation rate of AT-MSC, which could be realized as notable benefits of their use over BM-MSC.

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Human stromal (mesenchymal) stem cells from bone marrow, adipose tissue and skin exhibit differences in molecular phenotype and differentiation potential.

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