scholarly journals Differentiation Potential of Early- and Late-Passage Adipose-Derived Mesenchymal Stem Cells Cultured under Hypoxia and Normoxia

2020 ◽  
Vol 2020 ◽  
pp. 1-11
Author(s):  
Ashley G. Zhao ◽  
Kiran Shah ◽  
Julien Freitag ◽  
Brett Cromer ◽  
Huseyin Sumer
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Large Scale ◽  
Clinical Applications ◽  
Bone Diseases ◽  
Stem Cell Marker ◽  
Marker Genes ◽  
Cell Marker ◽  
Late Passage

With an increasing focus on the large-scale expansion of mesenchymal stem cells (MSCs) required for clinical applications for the treatment of joint and bone diseases such as osteoarthritis, the optimisation of conditions for in vitro MSC expansion requires careful consideration to maintain native MSC characteristics. Physiological parameters such as oxygen concentration, media constituents, and passage numbers influence the properties of MSCs and may have major impact on their therapeutic potential. Cells grown under hypoxic conditions have been widely documented in clinical use. Culturing MSCs on large scale requires bioreactor culture; however, it is challenging to maintain low oxygen and other physiological parameters over several passages in large bioreactor vessels. The necessity to scale up the production of cells in vitro under normoxia may affect important attributes of MSCs. For these reasons, our study investigated the effects of normoxic and hypoxic culture condition on early- and late-passage adipose-derived MSCs. We examined effect of each condition on the expression of key stem cell marker genes POU5F1, NANOG, and KLF4, as well as differentiation genes RUNX2, COL1A1, SOX9, COL2A1, and PPARG. We found that expression levels of stem cell marker genes and osteogenic and chondrogenic genes were higher in normoxia compared to hypoxia. Furthermore, expression of these genes reduced with passage number, with the exception of PPARG, an adipose differentiation marker, possibly due to the adipose origin of the MSCs. We confirmed by flow cytometry the presence of cell surface markers CD105, CD73, and CD90 and lack of expression of CD45, CD34, CD14, and CD19 across all conditions. Furthermore, in vitro differentiation confirmed that both early- and late-passage adipose-derived MSCs grown in hypoxia or normoxia could differentiate into chondrogenic and osteogenic cell types. Our results demonstrate that the minimal standard criteria to define MSCs as suitable for laboratory-based and preclinical studies can be maintained in early- or late-passage MSCs cultured in hypoxia or normoxia. Therefore, any of these culture conditions could be used when scaling up MSCs in bioreactors for allogeneic clinical applications or tissue engineering for the treatment of joint and bone diseases such as osteoarthritis.

Glial cell derived neurotrophic factor induces spermatogonial stem cell marker genes in chicken mesenchymal stem cells

2016 ◽  
Vol 48 (3) ◽  
pp. 235-241 ◽  
Author(s):  
Sohrab Boozarpour ◽  
Maryam M. Matin ◽  
Madjid Momeni-Moghaddam ◽  
Hesam Dehghani ◽  
Naser Mahdavi-Shahri ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Stem Cell ◽  
Glial Cell ◽  
Neurotrophic Factor ◽  
Spermatogonial Stem Cell ◽  
Stem Cell Marker ◽  
Marker Genes ◽  
Cell Marker

scholarly journals Environmental Benzopyrene Attenuates Stemness of Placenta-Derived Mesenchymal Stem Cells via Aryl Hydrocarbon Receptor

2019 ◽  
Vol 2019 ◽  
pp. 1-12 ◽  
Author(s):  
June Seok Heo ◽  
Ja-Yun Lim ◽  
Sangshin Pyo ◽  
Dae Wui Yoon ◽  
Dongsook Lee ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Aryl Hydrocarbon Receptor ◽  
Molecular Mechanisms ◽  
Cell Activity ◽  
Bone Diseases ◽  
Purine Derivative ◽  
Aryl Hydrocarbon ◽  
Polycyclic Aromatic

The toxic effects of particulate matter have been linked to polycyclic aromatic hydrocarbons (PAHs) such as benzopyrene. PAHs are potent inducers of the aryl hydrocarbon receptor (AhR), which is an expressed nuclear receptor that senses environmental stimuli and modulates gene expression. Even though several studies have shown that the benzopyrene (BP) of chemical pollutants significantly impaired stem cell activity, the exact molecular mechanisms were not clearly elucidated. In the present study, we aimed to investigate the effects of BP on placenta-derived mesenchymal stem cells (PD-MSCs) in vitro. We found that the AhR in PD-MSCs was expressed under the treatment of BP, and its activation markedly disrupted osteogenic differentiation through the alteration of stemness activity of PD-MSCs. Moreover, BP treatment significantly reduced the proliferation activity of PD-MSCs and expression of pluripotent markers through the induction of AhR. Treatment with StemRegenin 1 (SR1), a purine derivative that antagonizes the AhR, effectively prevented BP-induced reduction of the proliferation and differentiation activity of PD-MSCs. In this study, we found that BP treatment in PD-MSCs markedly obstructs PD-MSC stemness through AhR signaling. Noteworthy, SR1-mediated MSC application will contribute to new perspectives on MSC-based therapies for air pollution-related bone diseases.

191 Canine Mesenchymal Stem Cells as Therapeutic Agents in Spinal Cord Injured Dog

2018 ◽  
Vol 30 (1) ◽  
pp. 236 ◽  
Author(s):  
Y.-H. Choe ◽  
H.-J. Lee ◽  
S.-L. Lee ◽  
J.-H. Lee ◽  
B.-W. Park ◽  
...  
Keyword(s):  
Spinal Cord ◽  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Therapeutic Agents ◽  
Clinical Applications ◽  
Pain Sensation ◽  
Differentiation Potential ◽  
Na Currents ◽  
Spinal Cord Injured

In the recent era of veterinary research, stem cells have gained special attention due to their efficiency and use in clinical applications. Mesenchymal stem cells (MSC) have been extensively studied over decades, and their prospect for clinical application is recognised in human medicine. Despite numerous reports in veterinary clinical trials of stem cells, few studies have been presented regarding the in vitro characterisation of canine mesenchymal stem cells (cMSC). Therefore, their efficacy as therapeutic agents in vitro has not been much elucidated. Canine adipose-derived mesenchymal stem cells (cAMSC) were characterised as per International Society for Cellular Therapy guidelines. Culturing cells showed spindle-like morphology and high proliferation rate. They displayed positive expression of mesenchymal markers CD44, CD90, and CD105, and lacked expression of CD34 and CD45. They were also positive for expression of pluripotency-related transcription factors (Oct3/4, Nanog, and Sox2) and showed differentiation potential towards mesodermal lineages. The cAMSC were further analysed for the neuronal trans-differentiation potential. Under appropriate differentiation conditions, cAMSC displayed distinctive dendritic morphology along with axon projections. Neuronal specific genes including Nestin, β-tubulin, neurofilament protein (NF-M, NF-H), and nerve growth factor (NGF) were also positively expressed. Nevertheless, functional analysis of neuronal differentiated cAMSC displayed voltage dependence and kinetics for transient K+ and Na+ currents (Ito). Both K+ and Na+ currents were recorded in differentiated MSC by voltage steps (between −120 and +60 mV for K+ currents, −40 and +50 mV for Na+ currents), whereas control undifferentiated MSC lacked the currents. Taken together, we concluded that the cAMSC have potential to differentiate into neuron-like cells. Based on these findings, we transplanted cAMSC into the spinal cord injured dogs to evaluate their clinical efficiency under approved medical guidelines set by Gyeongsang National University Animal Medical Center (Korea). Neurological examination showed that the injured dog had undergone hind limb paralysis and lost deep pain sensation due to an L2 spinal cord lesion, as detected by CT and MRI. The dog was diagnosed with traumatic L2 intradural spinal cord contusion, and decompression surgery was performed, but deep pain sensation did not recover. Therefore, each cAMSC (diluted in 0.5 mL of saline) was transplanted into spinal cord segment (L2~L3) 5 times at 1-week intervals. The dog showed mild recovery of deep pain sensation by neurological examinations and exhibited gradual improvement in hind limb function. Finally, we concluded that transplantation of cAMSC has a beneficial therapeutic effect on spinal cord injury. This study also provides a significant advantage in understanding the potential of MSC-based products in veterinary clinical applications.

scholarly journals Platelet-Derived Growth Factor Receptor Alpha as a Marker of Mesenchymal Stem Cells in Development and Stem Cell Biology

2015 ◽  
Vol 2015 ◽  
pp. 1-8 ◽  
Author(s):  
Ramin M. Farahani ◽  
Munira Xaymardan
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Stem Cell ◽  
Cell Biology ◽  
Growth Factor Receptor ◽  
Stem Cell Marker ◽  
Factor Receptor Alpha ◽  
Lines Of Evidence

Three decades on, the mesenchymal stem cells (MSCs) have been intensively researched on the bench top and used clinically. However, ambiguity still exists in regard to their anatomical locations, identities, functions, and extent of their differentiative abilities. One of the major impediments in the quest of the MSC research has been lack of appropriatein vivomarkers. In recent years, this obstacle has been resolved to some degree as PDGFRαemerges as an important mesenchymal stem cell marker. Accumulating lines of evidence are showing that the PDGFRα+cells reside in the perivascular locations of many adult interstitium and fulfil the classic concepts of MSCsin vitroandin vivo. PDGFRαhas long been recognised for its roles in the mesoderm formation and connective tissue development during the embryogenesis. Current review describes the lines of evidence regarding the role of PDGFRαin morphogenesis and differentiation and its implications for MSC biology.

Large-scale production of human mesenchymal stem cells for clinical applications

2012 ◽  
Vol 59 (2) ◽  
pp. 106-120 ◽  
Author(s):  
Sunghoon Jung ◽  
Krishna M. Panchalingam ◽  
Reynold D. Wuerth ◽  
Lawrence Rosenberg ◽  
Leo A. Behie
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Large Scale ◽  
Human Mesenchymal Stem Cells ◽  
Clinical Applications ◽  
Scale Production ◽  
Large Scale Production

scholarly journals qPCR analysis of mesenchymal stem cell marker expression during the long-term culture of canine adipocyte derived stem cells

2020 ◽  
Vol 8 (4) ◽  
pp. 139-145
Author(s):  
Rut Bryl ◽  
Claudia Dompe ◽  
Maurycy Jankowski ◽  
Katarzyna Stefańska ◽  
Afsaneh Golkar Narenji ◽  
...  
Keyword(s):  
Stem Cells ◽  
Molecular Mechanisms ◽  
Cultured Cells ◽  
Stem Cell Marker ◽  
Pcr Analysis ◽  
Marker Genes ◽  
Long Term Culture ◽  
Marker Expression

AbstractDue to its availability and accessibility, adipose tissue has been the subject of various studies in many different medical fields and is believed to be a useful source of stem cells. The ability of ASCs to differentiate towards different cell lineages, with possibility of directing this differentiation, increases their possible clinical applications, and they have been widely employed in multiple therapies and treatment of different pathologies. However, a deeper understanding of the molecular mechanisms underlying the ASCs osteoblastic and chondrocyte differentiation may lead to novel applications treating a multitude of different bone-related diseases through techniques more likely meeting worldwide consensus. In this study, the RT-qPCR method was used to determine the changes in expression of ASC specific markers (CD105, CD73, CD14, CD34, CD90 and CD45) before and after long-term (14-day) in vitro cultures. To confirm the identity of the investigated cells, flow cytometry was used to evaluate the presence of positive (CD44, CD90) and negative (CD45, CD34) ASC markers. Overall, the results of the PCR analysis showed a significant change in expression of most of the marker genes, indicating significant changes in the cultured cells caused by their long-term culture, potentially altering their original stem-like characteristics.Running title: ASC marker expression during long-term in vitro culture

scholarly journals Manufacturing and Banking Canine Adipose-Derived Mesenchymal Stem Cells for Veterinary Clinical Application

2021 ◽  
Author(s):  
Huina Luo ◽  
Dongsheng Li ◽  
Zhisheng Chen ◽  
Bingyun Wang ◽  
Shengfeng Chen
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Normal Karyotype ◽  
Clinical Applications ◽  
Therapeutic Application ◽  
Differentiation Potential ◽  
Safety And Efficacy

Abstract BACKGROUND: Mesenchymal stem cells (MSCs) have generated a great amount of interest in recent years as a novel therapeutic application for improving the quality of pet life and helping them free from painful conditions and diseases. It has now become critical to address the challenges related to the safety and efficacy of MSCs expanded in vitro. In this study, we establish a standardized process for manufacture of canine adipose-derived MSCs (AD-MSCs), including tissue sourcing, cell isolation and culture, cryopreservation, thawing and expansion, quality control and testing, and evaluate the safety and efficacy of those cells for clinical applications. RESULTS: After expansion, the viability of AD-MSCs manufactured under our standardized process was above 90 %. Expression of surface markers and differentiation potential was consistent with ISCT standards. Sterility, mycoplasma, and endotoxin tests were consistently negative. AD-MSCs presented normal karyotype, and did not form in vivo tumors. No adverse events were noted in two cases treated with intravenously AD-MSCs. CONCLUSION: Herein we demonstrated the establishment of a feasible bioprocess for manufacturing and banking canine AD-MSCs for veterinary clinical use.

scholarly journals Genetically Modified Porcine Mesenchymal Stem Cells by Lentiviral Tbx18 Create a Biological Pacemaker

2019 ◽  
Vol 2019 ◽  
pp. 1-9
Author(s):  
Yannan Hu ◽  
Ning Li ◽  
Liang Liu ◽  
Hao Zhang ◽  
Xiang Xue ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Right Ventricle ◽  
Large Scale ◽  
Genetically Modified ◽  
Pacemaker Activity ◽  
Bone Mesenchymal Stem Cells ◽  
The Right ◽  
Biological Pacemaker

Background. Tbx18 is a vital transcription factor involved in embryonic sinoatrial node (SAN) formation process but is gradually vanished after birth. Myocardial injection of lentiviral Tbx18 converts cardiomyocytes into pacemaker-like cells morphologically and functionally. In this in vitro and in vivo study, genetical modification of porcine bone mesenchymal stem cells (BMSCs) by recapturing the Tbx18 expression creates a biological pacemaker which was examined. Methods. The isolated porcine BMSCs were transfected with lentiviral Tbx18, and the induced pacemaker-like cells were analyzed using real-time polymerase chain reaction and western blotting to investigate the efficiency of transformation. Then, the induced pacemaker-like cells were implanted into the right ventricle of the SAN dysfunction porcine model after the differentiation process. Biological pacemaker activity and ectopic pacing region were tested by an electrocardiograph (ECG) monitor. Results. The isolated porcine BMSCs expressed specific surface markers of stem cells; meanwhile, the expression of myocardial markers was upregulated significantly after lentiviral Tbx18 transfection. The porcine SAN dysfunction model was constructed by electrocoagulation using a surgical electrotome. The results showed that the mean heart beat (HR) of BMSCs-Tbx18 was significantly higher than that of BMSCs-GFP. An ectopic pacing region was affirmed into the right ventricle by ECG after implantation of BMSCs-Tbx18. Conclusion. It was verified that Lenti-Tbx18 is capable of transducing porcine BMSCs into pacemaker-like cells. Genetically modified porcine BMSCs by lentiviral Tbx18 could create a biological pacemaker. However, further researches in large-scale animals are required to rule out unexpected complications prior to application in clinical practice.

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