scholarly journals Evaluation of growth, stemness, and angiogenic properties of dental pulp stem cells cultured in cGMP xeno-/serum-free medium

2019 ◽  
Vol 380 (1) ◽  
pp. 93-105 ◽  
Author(s):  
Chengjuan Qu ◽  
Maria Brohlin ◽  
Paul J Kingham ◽  
Peyman Kelk
Keyword(s):  
Stem Cells ◽  
Human Serum ◽  
Dental Pulp ◽  
Dental Pulp Stem Cells ◽  
In Vitro Assays ◽  
Serum Free Medium ◽  
Free Medium ◽  
Serum Free ◽  
Cells Cultured

AbstractThis study was aimed to investigate the effects of cGMP xeno-/serum-free medium (XSF, Irvine Scientific) on the properties of human dental pulp stem cells (DPSCs). DPSCs, from passage 2, were cultured in XSF or fetal bovine serum (FBS)-supplemented medium, and sub-cultured up to passage 8. Cumulative population doublings (PDs) and the number of colony-forming-units (CFUs) were determined. qRT-PCR, ELISA, and in vitro assays were used to assess angiogenic capacity. Flow cytometry was used to measure CD73, CD90, and CD105 expression. Differentiation into osteo-, adipo-, and chondrogenic cell lineages was performed. DPSCs showed more elongated morphology, a reduced rate of proliferation at later passages, and lower CFU counts in XSF compared with FBS. Expression of angiogenic factors at the gene and protein levels varied in the two media and with passage number, but cells grown in XSF had more in vitro angiogenic activity. The majority of early and late passage DPSCs cultured in XSF expressed CD73 and CD90. In contrast, the percentage of CD105 positive DPSCs in XSF medium was significantly lower with increased passage whereas the majority of cells cultured in FBS were CD105 positive. Switching XSF-cultured DPSCs to medium supplemented with human serum restored the expression of CD105. The tri-lineage differentiation of DPSCs cultured under XSF and FBS conditions was similar. We showed that despite reduced CD105 expression levels, DPSCs expanded in XSF medium maintained a functional MSC phenotype. Furthermore, restoration of CD105 expression is likely to occur upon in vivo transplantation, when cells are exposed to human serum.

Dental pulp stem cells in serum-free medium for regenerative medicine

2019 ◽  
Vol 50 (1) ◽  
pp. 80-90 ◽  
Author(s):  
Dawn E. Coates ◽  
Mohammad Alansary ◽  
Lara Friedlander ◽  
Diogo G. Zanicotti ◽  
Warwick J. Duncan
Keyword(s):  
Stem Cells ◽  
Regenerative Medicine ◽  
Dental Pulp ◽  
Dental Pulp Stem Cells ◽  
Serum Free Medium ◽  
Free Medium ◽  
Serum Free

High-purity Hepatic Lineage Differentiated from Dental Pulp Stem Cells in Serum-free Medium

2012 ◽  
Vol 38 (4) ◽  
pp. 475-480 ◽  
Author(s):  
Nikolay Ishkitiev ◽  
Ken Yaegaki ◽  
Toshio Imai ◽  
Tomoko Tanaka ◽  
Taka Nakahara ◽  
...  
Keyword(s):  
Stem Cells ◽  
High Purity ◽  
Dental Pulp ◽  
Dental Pulp Stem Cells ◽  
Serum Free Medium ◽  
Free Medium ◽  
Serum Free

scholarly journals Cellular Properties and Expression of Pluripotent Markers in Human Dental Pulp Stem Cells Cultured in Serum-Free Medium

2021 ◽  
Author(s):  
Chethan Kumar ◽  
Basan Gowda Sharanappa Kurkalli ◽  
Shishir Shetty ◽  
Akshay Bairapura Manjappa ◽  
Veena Shetty ◽  
...  
Keyword(s):  
Stem Cells ◽  
Dental Pulp ◽  
Dental Pulp Stem Cells ◽  
Population Doubling ◽  
Animal Origin ◽  
Serum Free Medium ◽  
Free Medium ◽  
Serum Free ◽  
Alp Activity ◽  
Human Dental Pulp

Introduction: The standard isolation and expansion of human Dental Pulp Stem Cells (DPSCs) under invitro conditions normally involve the usage of Fetal Bovine Serum (FBS). However, its animal-origin poses possible concerns for clinically relevant procedures. This critical issue compels the use of Xenogeneic-Free (XF) or human-origin alternatives to FBS for culture expansion and differentiation of DPSCs to determine the usefulness for translating into therapeutic clinical applications. Aim: To evaluate the cellular characteristics and expression of pluripotent markers in DPSCs cultured using Serum-Containing Medium (SCM-DPSCs) and Serum-Free Medium (SFM-DPSCs). Materials and Methods: This in-vitro descriptive study was conducted at NITTE (Deemed to be University), Mangaluru, Karnataka, India, from June 2019 to August 2020. DPSCs were isolated from impacted third molars. The culture expanded DPSCs in serum-containing and serum-free media were analysed on their morphology, viability, proliferation rate, Population Doubling Time (PDT), Alkaline Phosphatase (ALP) activity, cell surface markers expression, osteogenic and adipogenic potential, and the relative expression of selected pluripotent genes. Results: The primary culture of DPSCs established in SCM and SFM showed spindle shaped fibroblastic morphology with >80% viability from passage 1 (P1) to P4. A significant (p-value<0.05) difference in the proliferation rates in terms of cell numbers between SCM-DPSCs and SFM-DPSCs was observed (day 6: 3×105 vs 0.8×105; day 9: 5.8×105 vs 1.27×105; day 12: 7.8×105vs 1.56×105, respectively). The average PDT values recorded in SCM- and SFM-DPSCs were 44.33 hours and 58.41 hours, respectively. A slightly higher expression of ALP activity was observed in SCM-DPSCs than in SFM-DPSCs. Flow cytometry analysis showed that both DPSCs were positive for CD29, CD73, CD90, and negative for CD34 and CD45. The expression of OCT4 and NANOG was relatively higher in SCM-DPSCs compared to SFM-DPSCs. Further, SCM-DPSCs showed the higher levels of SOX2 and SSEA4, but did not exhibit any significant differences in their expression levels. Conclusion: The results showed that DPSCs in FBS displayed better growth kinetics and stemness markers expression along with more propensities towards lineage differentiation. SFM can be used to establish and expand DPSCs with characteristics of multipotent stem cells, but needs further research for its optimisation.

Effect of noradrenaline and dopamine on progesterone and estradiol secretion of human granulosa cells

1993 ◽  
Vol 129 (2) ◽  
pp. 165-168 ◽  
Author(s):  
József Bódis ◽  
Hans R Tinneberg ◽  
Attila Török ◽  
Philippe Cledon ◽  
Volker Hanf ◽  
...  
Keyword(s):  
Granulosa Cells ◽  
Direct Action ◽  
Serum Free Medium ◽  
Free Medium ◽  
Vitro Fertilization ◽  
Serum Free ◽  
Human Granulosa Cells ◽  
Estradiol Secretion ◽  
Cells Cultured

The aim of this study was to explore the direct action of noradrenaline and dopamine on progesterone and estradiol secretion of human granulosa cells cultured in serum-free medium. Progesterone and estradiol production was measured in the presence and absence of noradrenaline, dopamine or propranolol using radioimmunoassays; statistical analysis was performed by analysis of variance and Newman-Keul's multiple range test. Twenty-six women aged 31±3 years undergoing in vitro fertilization and embryo transfer for infertility treatment at University Women's Hospital, University of Tübingen, Germany, took part in this study. Noradrenaline significantly inhibited progesterone production by human granulosa cells in a dose-related manner at a concentration of 10−4–10−6 mol/l. Dopamine significantly stimulated estradiol secretion by granulosa cells in an inverse dose-related manner. Both effects were blocked by propranolol. The results suggest that catecholaminergic actions switch over the steroid production of human granulosa cells cultured in serum-free medium from progesterone to estradiol.

Ex Vivo Expansion of Human Marrow Mesenchymal Stem Cells: A Serum-Free Medium Compared to Classical α-MEM Medium.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4257-4257 ◽  
Author(s):  
Nathalie Meuleman ◽  
Tatiana Tondreau ◽  
Alain Delforge ◽  
Marielle Dejeneffe ◽  
Martine Massy ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Culture Conditions ◽  
Adherent Cells ◽  
Serum Free Medium ◽  
Free Medium ◽  
Serum Free ◽  
Marrow Mesenchymal Stem Cells ◽  
Serum Substitute ◽  
Cells Cultured

Abstract BACKGROUND: Bone Marrow mesenchymal stem cells (MSC) are pluripotent cells that have the capacity to differentiate into several tissue lineages and also the ability to support hematopoiesis. Their use in gene and cell therapy requires their in vitro expansion. Maintenance and proliferation of MSC are strongly dependent on the culture conditions such as properties of the bovine serum added in the culture medium. However, duration and culture conditions are critical for the successful clinical use of MSC. OBJECTIVE: We have evaluated the efficiency of a commercial serum-free medium (UltraCulture, Bio-Whittaker, Walkersville, MD) supplemented with a serum substitute (Ultroser, BioSepra, Cergy-Saint-Christophe, France) in order to work without FBS and to have a more constant composition. This medium (UC) was compared to the classical medium a-MEM containing 10% FBS (Invitrogen, Merelbeke, Belgium). METHODS: BM-mononuclear cells collected from 11 healthy donors were plated in petri dishes at a concentration of 105/cm2. After 3 days, non adherent cells were removed and culture media were added to adherent cells which were maintained at 37°C until they reached confluence. MSC expansion was analysed after the primoculture (PM) and after the first passage (P1). CFU-F (colony forming units-fibroblastic) number, phenotypic analysis and differentiation potential were also evaluated. RESULTS::The mean culture duration was 13±2 and 8±2 days respectively for PM and P1 but the confluence was reached more rapidly for cells cultured in UC. After PM, 0.35x106 and 1.21x106 cells were obtained for MEM and UC respectively (p<0.005). Moreover, around 20% of cells cultured in MEM were CD45+ while the level of CD45+ cells was frequently < 5% in UC indicating that UC medium favored the rapid elimination of hematopoietic cells. After P1, the expansion rate was significantly higher in UC than MEM: respectively 5.13 ± 1.2 and 22.6 ± 3 (p<0.0005). The numbers of CFU-F were always higher in UC demonstrating the enhanced proliferation in serum-free medium. The phenotype of MSC was similar in the both media: SH2+, SH3+, CD44+, CD45−, CD34−, HLA-DR−. More importantly, these cells remained able to differentiate into osteocytes, chondrocytes, adipocytes and neuron-like cells in the both media confirming their multipotentiality. CONCLUSIONS: Our data strongly support UltraCulture medium supplemented with a serum substitute as an optimal medium for MSC culture. Indeed, it allows a better cell expansion, better proliferation and preserves multipotentiality. This medium, reducing culture period and containing low concentration of serum substitute, is of major interest for clinical production of MSC.

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