Proliferation and Osteogenic Differentiation of Mesenchymal Stem Cells on Biodegradable Calcium-deficient Hydroxyapatite Tubular Bacterial Cellulose Composites

2014 ◽  
Vol 1621 ◽  
pp. 71-79 ◽  
Author(s):  
Pelagie Favi ◽  
Madhu Dhar ◽  
Nancy Neilsen ◽  
Roberto Benson
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Bacterial Cellulose ◽  
Periodate Oxidation ◽  
Osteogenic Potential ◽  
Alizarin Red ◽  
Native Bone ◽  
Cellulose Composites

ABSTRACTAdvanced biomaterials that mimic the structure and function of native tissues and permit stem cells to adhere and differentiate is of paramount importance in the development of stem cell therapies for bone defects. Successful bone repair approaches may include an osteoconductive scaffold that permits excellent cell adhesion and proliferation, and cells with an osteogenic potential. The objective of this study was to evaluate the cell proliferation, viability and osteocyte differentiation of equine-derived bone marrow mesenchymal stem cells (EqMSCs) when seeded onto biocompatible and biodegradable calcium-deficient hydroxyapatite (CdHA) tubular-shaped bacterial cellulose scaffolds (BC-TS) of various sizes. The biocompatible gel-like BC-TS was synthesized using the bacterium Gluconacetobacter sucrofermentans under static culture in oxygen-permeable silicone tubes. The BC-TS scaffolds were modified using a periodate oxidation to yield biodegradable scaffolds. Additionally, CdHA was deposited in the scaffolds to mimic native bone tissues. The morphological properties of the resulting BC-TS and its composites were characterized using scanning electron microscopy. The ability of the BC-TS and its composites to support and maintain EqMSCs growth, proliferation and osteogenic differentiation in vitro was also assessed. BC-TS and its composites exhibited aligned nanofibril structures. MTS assay demonstrated increasing proliferation and viability with time (days 1, 2 and 3). Cell-scaffold constructs were cultured for 8 days under osteogenic conditions and the resulting osteocytes were positive for alizarin red. In summary, biocompatible and biodegradable CdHA BC-TS composites support the proliferation, viability and osteogenic differentiation of EqMSCs cultured onto its surface in vitro, allowing for future potential use for tissue engineering therapies.

scholarly journals The Effects of Vancomycin on the Viability and Osteogenic Potential of Bone-derived Mesenchymal Stem Cells

2018 ◽  
Author(s):  
Elzaan Booysen ◽  
Hanél Sadie-Van Gijsen ◽  
Shelly M. Deane ◽  
William Ferris ◽  
Leon M.T. Dicks
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Osteogenic Potential ◽  
Short Term ◽  
Alizarin Red ◽  
Methicillin Resistant ◽  
Joint Infections ◽  
Diphenyltetrazolium Bromide ◽  
Vancomycin Treatment

ABSTRACTPeriprosthetic joint infections (PJI), caused by methicillin-resistant Staphylococcus aureus (MRSA), is the major cause of total hip arthroplasty (THA) failures. Traditionally, MRSA is treated with vancomycin, administrated intravenously or applied directly onto infected tissue. The effect of direct (as opposed to systemic) vancomycin treatment on bone formation and remodelling is largely unknown. The minimal inhibitory concentration (MIC) of vancomycin was determined by adding 200 μl of different concentrations (1 – 20 μg/ml) to actively growing cultures of S. aureus Xen 31 (methicillin-resistant) and S. aureus Xen 36 (methicillin-sensitive), respectively, and recording changes in optical density over 24 h. Bone marrow-derived and proximal femur-derived mesenchymal stem cells (bmMSCs and pfMSCs) from rat femora were exposed to 1 x MIC (5 μg/ml) and 4 x MIC (20 μg/ml) of vancomycin for 7 days. Cell viability was determined by staining with crystal violet and MTT (3-(4,5- dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide), respectively, and osteogenic differentiation by staining with Alizarin Red S. Vancomycin had no effect on the viability of bmMSCs and pfMSCs, even at high levels (20 μg/ml). The osteogenic differentiation of pfMSCs was partially inhibited, while osteogenesis in bmMSCs was not severely affected. The direct application of vancomycin to infected bone tissue, even at excessive levels, may preserve the viability of resident MSC populations. Short-term demineralization may thus be reversed after cessation of vancomycin treatment, improving the outcome of THA surgery.

scholarly journals Identification of WNT16 as a Predictable Biomarker for Accelerated Osteogenic Differentiation of Tonsil-Derived Mesenchymal Stem Cells In Vitro

2019 ◽  
Vol 2019 ◽  
pp. 1-10 ◽  
Author(s):  
Yu-Hee Kim ◽  
Kyung-Ah Cho ◽  
Hyun-Ji Lee ◽  
Minhwa Park ◽  
Han Su Kim ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Selection Criterion ◽  
Osteogenic Potential ◽  
Differentiation Potential ◽  
Real Time Quantitative Pcr ◽  
Single Cell Cloning

The application of mesenchymal stem cells (MSCs) for treating bone-related diseases shows promising outcomes in preclinical studies. However, cells that are isolated and defined as MSCs comprise a heterogeneous population of progenitors. This heterogeneity can produce variations in the performance of MSCs, especially in applications that require differentiation potential in vivo, such as the treatment of osteoporosis. Here, we aimed to identify genetic markers in tonsil-derived MSCs (T-MSCs) that can predict osteogenic potential. Using a single-cell cloning method, we isolated and established several lines of nondifferentiating (ND) or osteoblast-prone (OP) clones. Next, we performed transcriptome sequencing of three ND and three OP clones that maintained the characteristics of MSCs and determined the top six genes that were upregulated in OP clones. Upregulation of WNT16 and DCLK1 expression was confirmed by real-time quantitative PCR, but only WNT16 expression was correlated with the osteogenic differentiation of T-MSCs from 10 different donors. Collectively, our findings suggest that WNT16 is a putative genetic marker that predicts the osteogenic potential of T-MSCs. Thus, examination of WNT16 expression as a selection criterion prior to the clinical application of MSCs may enhance the therapeutic efficacy of stem cell therapy for bone-related complications, including osteoporosis.

scholarly journals Role of Fzd6 in Regulating the Osteogenic Differentiation of Adipose-derived Stem Cells in Osteoporosis

2021 ◽  
Author(s):  
Tianli Wu ◽  
Zhihao Yao ◽  
Gang Tao ◽  
Fangzhi Lou ◽  
Hui Tang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Osteogenic Differentiation ◽  
Wnt Signaling Pathway ◽  
Osteogenic Potential ◽  
Adipose Derived Stem Cells ◽  
Alizarin Red ◽  
Differentiation Potential

Abstract Objective: Although it has been demonstrated that adipose-derived stem cells (ASCs) from osteoporosis mice (OP-ASCs) exhibit impaired osteogenic differentiation potential, the molecular mechanism has not yet been elucidated. We found that Fzd6 was decreased in OP-ASCs compared with ASCs. This study investigates the effects and underlying mechanisms of Fzd6 in the osteogenic potential of OP-ASCs. Methods: Fzd6 expression in ASCs and OP-ASCs was measured by PCR gene chip. Fzd6 overexpression and silencing lentiviruses were used to evaluate the role of Fzd6 in the osteogenic differentiation of OP-ASCs. Real-time PCR (qPCR) and western blotting (WB) was performed to detect the expression of Fzd6 and bone-related molecules, including runt-related transcription factor 2 (Runx2) and osteopontin (Opn). Alizarin red staining and Alkaline phosphatase (ALP) staining was performed following osteogenic induction. Microscopic CT (Micro-CT), hematoxylin and eosin staining (H&E) staining, and Masson staining were used to assess the role of Fzd6 in osteogenic differentiation of osteoporosis (OP) mice in vivo.Results: Expression of Fzd6 was decreased significantly in OP-ASCs. Fzd6 silencing down-regulated the osteogenic ability of OP-ASCs in vitro. Overexpression of Fzd6 rescued the impaired osteogenic capacity in OP-ASCs in vitro. We obtained similar results in vivo.Conclusions: Fzd6 plays an important role in regulating the osteogenic ability of OP-ASCs both in vivo and in vitro. Overexpression of Fzd6 associated with the Wnt signaling pathway promotes the osteogenic ability of OP-ASCs, which provides new insights for the prevention and treatment of OP.

scholarly journals Effect of GARP on osteogenic differentiation of bone marrow mesenchymal stem cells via the regulation of TGFβ1 in vitro

PeerJ ◽  
2019 ◽  
Vol 7 ◽  
pp. e6993 ◽  
Author(s):  
Ruixue Li ◽  
Jian Sun ◽  
Fei Yang ◽  
Yang Sun ◽  
Xingwen Wu ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Transforming Growth Factor ◽  
Alizarin Red S ◽  
Bone Mesenchymal Stem Cells ◽  
Alizarin Red ◽  
Alp Activity

Mesenchymal stem cells (MSCs), which have multipotential differentiation and self-renewal potential, are possible cells for tissue engineering. Transforming growth factor β1 (TGFβ1) can be produced by MSCs in an inactive form, and the activation of TGFβ1 functions as an important regulator of osteogenic differentiation in MSCs. Recently, studies showed that Glycoprotein A repetitions predominant (GARP) participated in the activation of latent TGFβ1, but the interaction between GARP and TGFβ1 is still undefined. In our study, we successfully isolated the MSCs from bone marrow of rats, and showed that GARP was detected in bone mesenchymal stem cells (BMSCs). During the osteogenic differentiation of BMSCs, GARP expression was increased over time. To elucidate the interaction between GARP and TGFβ1, we downregulated GARP expression in BMSCs to examine the level of active TGFβ1. We then verified that the downregulation of GARP decreased the secretion of active TGFβ1. Furthermore, osteogenic differentiation experiments, alkaline phosphatase (ALP) activity analyses and Alizarin Red S staining experiments were performed to evaluate the osteogenic capacity. After the downregulation of GARP, ALP activity and Alizarin Red S staining significantly declined and the osteogenic indicators, ALP, Runx2, and OPN, also decreased, both at the mRNA and protein levels. These results demonstrated that downregulated GARP expression resulted in the reduction of TGFβ1 and the attenuation of osteoblast differentiation of BMSCs in vitro.

scholarly journals Cannabidiol and Vitamin D3 Impact on Osteogenic Differentiation of Human Dental Mesenchymal Stem Cells

Medicina ◽  
2020 ◽  
Vol 56 (11) ◽  
pp. 607
Author(s):  
Nausica B. Petrescu ◽  
Ancuta Jurj ◽  
Olga Sorițău ◽  
Ondine P. Lucaciu ◽  
Noemi Dirzu ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Stem Cell ◽  
Osteogenic Differentiation ◽  
Vitamin D3 ◽  
Bone Matrix ◽  
Third Molar ◽  
Osteogenic Potential ◽  
Rt Pcr ◽  
Alizarin Red

Background and objective: The aim of the present study was to establish a new differentiation protocol using cannabidiol (CBD) and vitamin D3 (Vit. D3) for a better and faster osteogenic differentiation of dental tissue derived mesenchymal stem cells (MSCs). Materials and methods: MSCs were harvested from dental follicle (DFSCs), dental pulp (DPSCs), and apical papilla (APSCs) of an impacted third molar of a 17-year old patient. The stem cells were isolated and characterized using flow cytometry; reverse transcription polymerase chain reaction (RT-PCR); and osteogenic, chondrogenic, and adipogenic differentiation. The effects of CBD and Vit. D3 on osteogenic differentiation of dental-derived stem cell were evaluated in terms of viability/metabolic activity by alamar test, expression of collagen1A, osteopontin (OP), osteocalcin (OC), and osteonectin genes and by quantification of calcium deposits by alizarin red assay. Results: Stem cell characterization revealed more typical stemness characteristics for DFSCs and DPSCs and atypical morphology and markers expression for APSCs, a phenotype that was confirmed by differences in multipotential ability. The RT-PCR quantification of bone matrix proteins expression revealed a different behavior for each cell type, APSCs having the best response for CBD. DPSCs showed the best osteogenic potential when treated with Vit. D3. Cultivation of DFSC in standard stem cell conditions induced the highest expression of osteogenic genes, suggesting the spontaneous differentiation capacity of these cells. Regarding mineralization, alizarin red assay indicated that DFSCs and APSCs were the most responsive to low doses of CBD and Vit. D3. DPSCs had the lowest mineralization levels, with a slightly better response to Vit. D3. Conclusions: This study provides evidence that DFSCs, DPSCs, and APSCs respond differently to osteoinduction stimuli and that CBD and Vit. D3 can enhance osteogenic differentiation of these types of cells under certain conditions and doses.

scholarly journals Osteogenic Differentiation of Miniature Pig Mesenchymal Stem Cells in 2D and 3D Environment

2011 ◽  
pp. 559-571 ◽  
Author(s):  
J. JUHÁSOVÁ ◽  
Š. JUHÁS ◽  
J. KLÍMA ◽  
J. STRNÁDEL ◽  
M. HOLUBOVÁ ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Collagen I ◽  
Miniature Pig ◽  
Plasma Clot ◽  
Alizarin Red ◽  
3D Environment ◽  
2D And 3D

Mesenchymal stem cells (MSCs) have been repeatedly shown to be able to repair bone defects. The aim of this study was to characterize the osteogenic differentiation of miniature pig MSCs and markers of this differentiation in vitro. Flow-cytometrically characterized MSCs were seeded on cultivation plastic (collagen I and vitronectin coated/uncoated) or plasma clot (PC)/plasma-alginate clot (PAC) scaffolds and differentiated in osteogenic medium. During three weeks of differentiation, the formation of nodules and deposition of calcium were visualized by Alizarin Red Staining. In addition, the production of alkaline phosphatase (ALP) activity was quantitatively detected by fluorescence. The expression of osteopontin, osteonectin and osteocalcin were assayed by immunohistochemistry and Western Blot analysis. We revealed a decrease of osteopontin expression in 2D and 3D environment during differentiation. The weak initial osteonectin signal, culminating on 7th or 14th day of differentiation, depends on collagen I and vitronectin coating in 2D system. The highest activity of ALP was detected on 21th day of osteogenic differentiation. The PC scaffolds provided better conditions for osteogenic differentiation of MSCs than PAC scaffolds in vitro. We also observed expected effects of collagen I and vitronectin on the acceleration of osteogenic differentiation of miniature pig MSC. Our results indicate similar ability of miniature pig MSCs osteogenic differentiation in 2D and 3D environment, but the expression of osteogenic markers in scaffolds and ECM coated monolayers started earlier than in the monolayers without ECM.

In vitro effect of prolactin on the osteogenic potential of bone marrow mesenchymal stem cells of rats

2013 ◽  
Author(s):  
Melo Ocarino Natalia de ◽  
Silvia Silva Santos ◽  
Lorena Rocha ◽  
Juneo Freitas ◽  
Reis Amanda Maria Sena ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Potential ◽  
Marrow Mesenchymal Stem Cells ◽  
Vitro Effect

scholarly journals Evaluation of the Impact of Different Pain Medication and Proton Pump Inhibitors on the Osteogenic Differentiation Potential of hMSCs Using 99mTc-HDP Labelling

Life ◽  
2021 ◽  
Vol 11 (4) ◽  
pp. 339
Author(s):  
Tobias Grossner ◽  
Uwe Haberkorn ◽  
Tobias Gotterbarm
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Bone Metabolism ◽  
Negative Impact ◽  
Pain Medication ◽  
Group 3 ◽  
Group 5 ◽  
The Impact

First-line analgetic medication used in the field of musculoskeletal degenerative diseases, like Nonsteroidal anti-inflammatory drugs (NSAIDs), reduces pain and prostaglandin synthesis, whereby peptic ulcers are a severe adverse effect. Therefore, proton pump inhibitors (PPI) are frequently used as a concomitant medication to reduce this risk. However, the impact of NSAIDs or metamizole, in combination with PPIs, on bone metabolism is still unclear. Therefore, human mesenchymal stem cells (hMSCs) were cultured in monolayer cultures in 10 different groups for 21 days. New bone formation was induced as follows: Group 1 negative control group, group 2 osteogenic differentiation media (OSM), group 3 OSM with pantoprazole (PAN), group 4 OSM with ibuprofen (IBU), group 5 OSM with diclofenac (DIC), group 6 OSM with metamizole (MET), group 7 OSM with ibuprofen and pantoprazole (IBU + PAN), group 8 OSM with diclofenac and pantoprazole (DIC + PAN), group 9 OSM with metamizole and pantoprazole (MET + PAN) and group 10 OSM with diclofenac, metamizole and pantoprazole (DIC + MET + PAN). Hydroxyapatite content was evaluated using high-sensitive radioactive 99mTc-HDP labeling. Within this study, no evidence was found that the common analgetic medication, using NSAIDs alone or in combination with pantoprazole and/or metamizole, has any negative impact on the osteogenic differentiation of mesenchymal stem cells in vitro. To the contrary, the statistical results indicate that pantoprazole alone (group 3 (PAN) (p = 0.016)) or diclofenac alone (group 5 (DIC) (p = 0.008)) enhances the deposition of minerals by hMSCS in vitro. There is an ongoing discussion between clinicians in the field of orthopaedics and traumatology as to whether post-surgical (pain) medication has a negative impact on bone healing. This is the first hMSC in vitro study that investigates the effects of pain medication in combination with PPIs on bone metabolism. Our in vitro data indicates that the assumed negative impact on bone metabolism is subsidiary. These findings substantiate the thesis that, in clinical medicine, the patient can receive every pain medication needed, whether or not in combination with PPIs, without any negative effects for the osteo-regenerative potential.

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