trilineage differentiation
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Cartilage ◽  
2021 ◽  
pp. 194760352110424
Author(s):  
Elizabeth Vinod ◽  
Kawin Padmaja ◽  
Abel Livingston ◽  
Jithu Varghese James ◽  
Soosai Manickam Amirtham ◽  
...  

Purpose Chondrocytes, isolated from articular cartilage, are routinely utilized in cell-based therapeutics for the treatment of cartilage pathologies. However, restoration of the biological tissue faces hindrance due to the formation of primarily fibrocartilaginous repair tissue. Chondroprogenitors have been reported to display superiority in terms of their chondrogenic potential and lesser proclivity for hypertrophy. In line with our recent results, comparing chondroprogenitors and chondrocytes, we undertook isolation of progenitors from the general pool of chondrocytes, based on surface marker expression, namely, CD166, CD34, and CD146, to eliminate off-target differentiation and generate cells of stronger chondrogenic potential. This study aimed to compare chondrocytes, chondroprogenitors, CD34−CD166+CD146+ sorted chondrocytes, and CD34−CD166+CD146− sorted chondrocytes. Methods Chondrocytes obtained from 3 human osteoarthritic knee joints were subjected to sorting, to isolate CD166+ and CD34− subsets, and then were further sorted to obtain CD146+ and CD146− cells. Chondrocytes and fibronectin adhesion-derived chondroprogenitors served as controls. Assessment parameters included reverse transcriptase polymerase chain reaction for markers of chondrogenesis and hypertrophy, trilineage differentiation, and total GAG/DNA content. Results Based on gene expression analysis, CD34−CD166+CD146+ sorted chondrocytes and chondroprogenitors displayed comparability and significantly higher chondrogenesis with a lower tendency for hypertrophy when compared to chondrocytes and CD34−CD166+CD146− sorted chondrocytes. The findings were also reiterated in multilineage potential differentiation with the 146+ subset and chondroprogenitors displaying lower calcification and chondroprogenitors displaying higher total GAG/DNA content compared to chondrocytes and 146− cells. Conclusion This unique progenitor-like population based on CD34−CD166+CD146+ sorting from chondrocytes exhibits efficient potential for cartilage repair and merits further evaluation for its therapeutic application.


Membranes ◽  
2021 ◽  
Vol 11 (9) ◽  
pp. 687
Author(s):  
Nathalia Barth de Oliveira ◽  
Ana Carolina Irioda ◽  
Priscila Elias Ferreira Stricker ◽  
Bassam Felipe Mogharbel ◽  
Nádia Nascimento da Rosa ◽  
...  

Adipose tissue-derived mesenchymal stem cells (ADMSCs) are promising candidates for regenerative medicine, as they have good cell yield and can differentiate into several cell lines. When induced to the neuronal differentiation, they form neurospheres composed of neural precursors (NPs) that can be an alternative in treating neurodegenerative diseases. This study aimed to characterize NPs from neurospheres obtained after seeding ADMSCs on a natural polyisoprene-based membrane. The ADMSCs were isolated from adipose tissue by enzymatic dissociation, were subjected to trilineage differentiation, and were characterized by flow cytometry for specific ADMSC surface markers. For neuronal differentiation, the cells were seeded on polystyrene flasks coated with the membrane and were characterized by immunocytochemistry and RT-PCR. The results demonstrated that the isolated cells showed characteristics of ADMSCs. At 15 to 25 days, ADMSCs seeded on the natural membrane developed neurospheres. Then, after dissociation, the cells demonstrated characteristic neuronal markers expressed on NPs: nestin, ß-III tubulin, GFAP, NeuN, and the YAP1/AMOT in the cytoplasm. In conclusion, it was demonstrated that this membrane differentiates the ADMSCs to NPs without any induction factors, and suggests that their differentiation mechanisms are related to mechanotransduction regulated by the YAP and AMOT proteins.


Anemia ◽  
2021 ◽  
Vol 2021 ◽  
pp. 1-9
Author(s):  
Vandana Sharma ◽  
Sonali Rawat ◽  
Suchi Gupta ◽  
Sweta Tamta ◽  
Rinkey Sharma ◽  
...  

Background and Objective. Acquired aplastic anemia (aAA) is a bone marrow failure disorder characterized by pancytopenia and bone marrow aplasia. Bone marrow Mesenchymal Stem Cells (BM-MSCs) are an important component of BM microenvironment, associated with hematopoietic and immune homeostasis. Any alterations in BM microenvironment can disrupt the normal functioning and it needs to be assessed. Methods. In the current study, we investigated the morphological differences, proliferation capacity, population doubling time (PDT), surface marker profiling, trilineage differentiation potential, and immunosuppressive ability of BM Mesenchymal Stem Cells (BM-MSCs) from untreated aAA patients and in the same number of age- and gender-matched controls. Results. We observed similar morphology, proliferation capacity, phenotype, trilineage differentiation potential, and immunomodulatory properties of BM-MSCs in aAA patients and control subjects. Conclusion. Our results confirm that the basic and immunosuppressive properties of BM-MSCs from aAA patients do not differ from normal BM-MSCs. Our data suggest that BM-MSCs from aAA patients might not be involved in disease pathogenesis. However, owing to a smaller number of samples, it is not conclusive, and future studies with more exhaustive investigation at transcriptome level are warranted.


2021 ◽  
Vol 2021 ◽  
pp. 1-12
Author(s):  
Guanlin Qu ◽  
Yan Li ◽  
Lu Chen ◽  
Qin Chen ◽  
Duohong Zou ◽  
...  

Background. Mesenchymal stem cells (MSCs) have become promising candidates for regeneration medicine due to their multidifferentiation potential and immunomodulatory ability. Compared with classic MSCs derived from the bone marrow and fat, dental-derived MSCs show high plasticity, accessibility, and applicability. Therefore, they are considered alternative sources for regeneration medicine. Methods. Four types of MSCs were isolated from the dental pulp, periodontal ligament, dental follicle, and alveolar bone of the same donor, and there were five different individuals. We analyzed their morphology, immunophenotype, proliferation rate, apoptosis, trilineage differentiation potential, and the gene expression during osteogenic differentiation. Results. Our research demonstrated that DPSCs, PDLSCs, DFPCs and ABMMSCs exhibited similar morphology and immunophenotype. DFPCs showed a higher rate of proliferation and apoptosis. When cultured in the trilineage differentiation medium, all types of MSCs presented the differentiation potential of osteogenesis, adipogenesis, and chondrogenesis. Through staining and genetic analysis during osteogenic induction, ABMMSCs and PDLSCs showed the highest osteogenic ability, followed by DPSCs, and DFPCs were the lowest. Conclusions. Overall, our results indicated that different dental-derived stem cells possessed different biological characteristics. For bone tissue engineering, ABMMSCs and PDLSCs can be used as optimal candidates of seed cells.


Author(s):  
Gabriele Spohn ◽  
Anne-Sophie Witte ◽  
Anja Kretschmer ◽  
Erhard Seifried ◽  
Richard Schäfer

BackgroundMesenchymal stromal cells (MSCs), multipotent progenitors that can be isolated from a variety of different tissues, are becoming increasingly important as cell therapeutics targeting immunopathologies and tissue regeneration. Current protocols for MSC isolation from bone marrow (BM) rely on density gradient centrifugation (DGC), and the production of sufficient MSC doses is a critical factor for conducting clinical MSC trials. Previously, a Good Manufacturing Practice (GMP)–compatible non-woven fabric filter device system to isolate MSCs was developed to increase the MSC yield from the BM. The aim of our study was to compare high-resolution phenotypic and functional characteristics of BM-MSCs isolated with this device and with standard DGC technology.MethodsHuman BM samples from 5 donors were analyzed. Each sample was divided equally, processing by DGC, and with the filter device. Stem cell content was assessed by quantification of colony-forming units fibroblasts (CFU-F). Immunophenotype was analyzed by multicolor flow cytometry. In vitro trilineage differentiation potential, trophic factors, and IDO-1 production were assessed. Functionally, immunomodulatory potential, wound healing, and angiogenesis were assayed in vitro.ResultsThe CFU-F yield was 15-fold higher in the MSC preparations isolated with the device compared to those isolated by DGC. Consequently, the MSC yield that could be manufactured at passage 3 per mL collected BM was more than 10 times higher in the device group compared to DGC (1.65 × 109 vs. 1.45 × 108). The immunomodulatory potential and IDO-1 production showed donor-to-donor variabilities without differences between fabric filter-isolated and DGC-isolated MSCs. The results from the wound closure assays, the tube formation assays, and the trilineage differentiation assays were similar between the groups with respect to the isolation method. Sixty-four MSC subpopulations could be quantified with CD140a+CD119+CD146+ as most common phenotype group, and CD140a+CD119+CD146+MSCA-1–CD106–CD271– and CD140a+CD119+CD146–MSCA-1–CD106–CD271– as most frequent MSC subpopulations. As trophic factors hepatocyte growth factor, epidermal growth factor, brain-derived neurotrophic factor, angiopoietin-1, and vascular endothelial growth factor A could be detected in both groups with considerable variability between donors, but independent of the respective MSC isolation technique.ConclusionThe isolation of MSCs using a GMP-compatible fabric filter system device resulted in higher yield of CFU-F, producing substantially more MSCs with similar subpopulation composition and functional characteristics as MSCs isolated by DGC.


2021 ◽  
Vol 162 (6) ◽  
pp. 227-232
Author(s):  
Attila Zalatnai ◽  
Judit Tőke ◽  
Gergely Huszty ◽  
Katalin Müllner ◽  
Miklós Tóth

Összefoglaló. A szerzők egy különleges pancreaselváltozás esetét ismertetik, melyben az acinusok neuroendokrin jellegű transzformációja diffúz, atípusos megjelenésű szigetsejtes hyperplasiával társult, valamint a pancreas mindhárom sejtvonalát (acinaris, ductalis, insularis) tartalmazó nodulusok képződtek. A komplex megjelenés ellenére a kórfolyamat nem járt endokrin tünetekkel. Esetünkben a kiváltó ok hátterében a struktúrák kóros progenitorsejt-differenciációja állhatott. Az irodalomban ilyen közlés eddig nem ismert. Orv Hetil. 2021; 162(6): 227–232. Summary. The authors present a case of a peculiar pancreatic lesion, in which the neuroendocrine transformation of the acini was associated with a diffuse, atypical insular hyperplasia, and micronodules exhibiting trilineage differentiation. Despite the complex alteration, no endocrine symptoms were noted. The case may represent the result of an abnormal pancreatic differentiation raising the possibility of reprogramming of the progenitor cells. To the best of our knowledge, this is the first report of such a lesion in the literature. Orv Hetil. 2021; 162(6): 227–232.


2020 ◽  
Author(s):  
Kejia Li ◽  
Tong Ning ◽  
Hao Wang ◽  
Yangzi Jiang ◽  
Jue Zhang ◽  
...  

Abstract Background: Multiple strategies have been proposed to promote the differentiation potential of mesenchymal stem cells (MSCs), which is the fundamental property in tissue formation and regeneration. However, these strategies are relatively inefficient that limit the application. In this study, we reported a novel and efficient strategy, nanosecond pulsed electric fields (nsPEFs) stimulation, which can enhance the trilineage differentiation potential of MSCs; and further explained the mechanism behind.Methods: We used histological staining to screen out the nsPEFs parameters that promoted the trilineage differentiation potential of MSCs, and further proved the effect of nsPEFs by detecting the functional genes. In order to explore the corresponding mechanism, we examined the expression of pluripotency genes and the methylation status of their promoters. Finally, we targeted the DNA methyltransferase which was affected by nsPEFs. Results: The trilineage differentiation of bone marrow derived MSCs was significantly enhanced in vitro by simply pre-treated with 5 pulses of nsPEFs stimulation (energy levels as 10 ns, 20 kV/cm; 100 ns, 10 kV/cm), due to that the nsPEFs demethylated the promoters of stem cell pluripotency genes OCT4 and NANOG through instantaneous downregulation of DNA methylation transferase 1 (DNMT1), thereby increased the expression of OCT4 and NANOG for up to 3 days, and created a treatment window period of stem cells.Conclusions: In summary, nsPEFs can enhance MSCs differentiation via the epigenetic regulation, and could be a safe and effective strategy for future stem cell application


Author(s):  
Naomi Moris ◽  
Kerim Anlas ◽  
Julia Schroeder ◽  
Sabitri Ghimire ◽  
Tina Balayo ◽  
...  

Abstract Gastruloids are aggregates of Pluripotent Stem Cells (PSCs) which, when exposed to differentiation medium and plated within defined conditions, undergo trilineage differentiation to all three germ layers (mesoderm, ectoderm and endoderm) with constitutive cell types organised spatiotemporally along 3 axes (Becarri et. al. 2018). They also undergo morphological shape changes including axial elongation through convergent extension cell movements. The gastruloid method has been well-established using mouse Embryonic Stem Cells (mESCs) and mouse induced PSCs (iPSCs) (van den Brink et. al., 2014; Turner et. al. 2016a; Turner et. al. 2016b; Bailie-Johnson et. al. 2015; Turner et. al. 2017a; Turner et. al. 2017b, Beccari et. al., 2018, See method in Girgin et. al., 2018). Here, we describe a new method to generate equivalent gastruloids from human pluripotent stem cells (hPSCs). hPSCs are able to generate axially elongated human gastruloids with evidence of spatially organised germ layers and comparable features to their mouse counterparts (Moris et. al., 2020).


Cartilage ◽  
2020 ◽  
pp. 194760352091863 ◽  
Author(s):  
Upasana Kachroo ◽  
Shikha Mary Zachariah ◽  
Augustine Thambaiah ◽  
Aleya Tabasum ◽  
Abel Livingston ◽  
...  

Purpose Articular chondroprogenitors, a suitable contender for cell-based therapy in cartilage repair, routinely employ fetal bovine serum (FBS) for expansion and differentiation. The possibility of transplant rejections or zoonoses transmissions raise a need for xeno-free alternatives. Use of human platelet lysate (hPL), a nutrient supplement abundant in growth factors, has not been reported for human chondroprogenitor expansion thus far. Our aim was to compare the biological profile of chondroprogenitors grown in hPL versus FBS. Methods Chondroprogenitors were isolated from 3 osteoarthritic knee joints. Following differential fibronectin adhesion assay, passage 0 cells grown in (a) 10% FBS and (b) 10% hPL were considered for assessment of growth kinetics, surface marker expression, gene expression, and trilineage differentiation. Latent transforming growth factor–β1 (TGFβ1) levels were also measured for each culture medium used. Results Cellular proliferation was significantly higher in cells grown with hPL ( P < 0.01). Surface marker expression was comparable except in CD-146 where hPL group had significantly higher values ( P = 0.03). Comparison of mRNA expression revealed notably low values of collagen I, collagen X, aggrecan, and collagen II ( P < 0.05). Trilineage differentiation was seen in both groups with higher alizarin red uptake noted in hPL. There were also significantly higher levels of latent TGFβ1 in the medium containing hPL as compared to FBS. Conclusions This is the first in vitro xeno-free study to affirm that hPL can serve as an optimal growth supplement for expansion of articular chondroprogenitors, although an in-depth assessment of resident growth factors and evaluation of different dilutions of hPL is required to assess suitability for use in translational research.


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