scholarly journals The Effect of Cultivation Passaging on the Relative Telomere Length and Proliferation Capacity of Dental Pulp Stem Cells

Biomolecules ◽  
2021 ◽  
Vol 11 (3) ◽  
pp. 464
Author(s):  
Nela Pilbauerova ◽  
Tomas Soukup ◽  
Tereza Suchankova Kleplova ◽  
Jan Schmidt ◽  
Jakub Suchanek
Keyword(s):  
Stem Cells ◽  
Telomere Length ◽  
Dna Sequences ◽  
Dental Pulp ◽  
Dental Pulp Stem Cells ◽  
Population Doubling ◽  
In Vitro Cultivation ◽  
Telomere Attrition ◽  
Proliferation Capacity ◽  
Relative Telomere Length

Telomeres are repetitive nucleoprotein DNA sequences that shorten with each cell division. The stem cells activate telomerase to compensate for the telomere loss. This study aimed to evaluate the effect of cultivation passaging on the relative telomere length and proliferation capacity of dental pulp stem cells. We used ten dental pulp stem cell (DPSC) lineages stored for 12 months using uncontrolled-rate freezing to reach the study’s goal. We analyzed their proliferation rate, phenotype using flow cytometry, multipotency, and relative telomere length using a qPCR analysis. We determined the relative telomere length in the added study by performing analysis after one, two, and three weeks of cultivation with no passaging. We documented the telomere attrition with increasing passaging. The shorter the relative telomere length, the lower reached population doublings, and longer population doubling time were observed at the end of the cultivation. We observed the telomere prolongation in DPSCs cultivated for two weeks with no passaging in the added subsequent study. We concluded that excessive proliferation demands on DPSCs during in vitro cultivation result in telomere attrition. We opened the theory that the telomerase might be more efficient during cell cultivation with no passaging. This observation could help in preserving the telomere length during ex vivo DPSC expansion.

scholarly journals Telomere Attrition Occurs during Ex Vivo Expansion of Human Dental Pulp Stem Cells

2010 ◽  
Vol 2010 ◽  
pp. 1-11 ◽  
Author(s):  
Jaroslav Mokry ◽  
Tomas Soukup ◽  
Stanislav Micuda ◽  
Jana Karbanova ◽  
Benjamin Visek ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Telomere Length ◽  
Dental Pulp ◽  
Ex Vivo ◽  
Ex Vivo Expansion ◽  
Dental Pulp Stem Cells ◽  
Cell Markers ◽  
Relative Telomere Length ◽  
Human Dental Pulp

We provide a detailed characteristic of stem cells isolated and expanded from the human dental pulp. Dental pulp stem cells express mesenchymal cell markers STRO-1, vimentin, CD29, CD44, CD73, CD90, CD166, and stem cell markers Sox2, nestin, and nucleostemin. They are multipotent as shown by their osteogenic and chondrogenic potential. We measured relative telomere length in 11 dental pulp stem cell lines at different passages by quantitative real-time PCR. Despite their large proliferative capacity, stable viability, phenotype, and genotype over prolonged cultivation, human dental pulp stem cells suffer from progressive telomere shortening over time they replicate in vitro. Relative telomere length (T/S) was inversely correlated with cumulative doubling time. Our findings indicate that excessive ex vivo expansion of adult stem cells should be reduced at minimum to avoid detrimental effects on telomere maintenance and measurement of telomere length should become a standard when certificating the status and replicative age of stem cells prior therapeutic applications.

Analysis of growth kinetics and colony-forming unit efficiency of human dental pulp stem cells cultured in different media and supplements

2021 ◽  
Vol 16 (7) ◽  
pp. 203-210
Author(s):  
Kumar Chethan ◽  
Shishir Shetty ◽  
Basan Gowda Kurkalli ◽  
Veena Shetty ◽  
Kumar Basavarajappa Mohana
Keyword(s):  
Stem Cells ◽  
Growth Kinetics ◽  
Dental Pulp ◽  
Culture Conditions ◽  
Dental Pulp Stem Cells ◽  
Population Doubling ◽  
Animal Origin ◽  
Dental Tissues ◽  
Colony Forming Unit ◽  
Human Dental Pulp

Dental tissues are considered as ideal autologous sources of multipotent stem cells. Presently, human dental pulp stem cells (DPSCs) are largely being isolated and expanded in media containing fetal bovine serum (FBS). However, the use of FBS has limitations due to its animal origin. Therefore, the present study evaluated the morphology, proliferation rate, population doubling time (PDT) and colony-forming unit fibroblast (CFU-F) efficiency of DPSCs cultured in animal serum-containing medium (SCM) and serumfree medium (SFM) in addition to serum-free culture conditions by supplementing human blood-derivatives such as platelet lysate (PL), fresh frozen plasma (FFP) and umbilical cord blood serum (UCS) at 2.5%, 5% and 7.5% concentrations. Established DPSCs had spindle-shape during primary culture but acquired characteristic fibroblast-like features when cultured in PL, FFP and UCS. DPSCs in SCM, SFM and PL had significantly (P<0.05) higher proliferative potential than those in UCS and FFP and these observations were supported by PDT values. The CFU efficiency of DPSCs was confirmed in all culture conditions with a slightly varied clonogenic potential in blood-derived components. Based on the growth kinetics and CFU ability, it is concluded that PL could be considered as a suitable alternative to FBS for the ex vivo expansion of DPSCs.

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.

scholarly journals The Effects of Cryogenic Storage on Human Dental Pulp Stem Cells

2021 ◽  
Vol 22 (9) ◽  
pp. 4432
Author(s):  
Nela Pilbauerova ◽  
Jan Schmidt ◽  
Tomas Soukup ◽  
Romana Koberova Koberova Ivancakova ◽  
Jakub Suchanek
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Dental Pulp ◽  
Dental Pulp Stem Cells ◽  
Great Promise ◽  
Cell Recovery ◽  
Proliferation Capacity ◽  
Human Dental Pulp ◽  
Ease Of Access ◽  
One Year

Dental pulp stem cells (DPSCs) are a type of easily accessible adult mesenchymal stem cell. Due to their ease of access, DPSCs show great promise in regenerative medicine. However, the tooth extractions from which DPSCs can be obtained are usually performed at a period of life when donors would have no therapeutic need of them. For this reason, it is imperative that successful stem cell storage techniques are employed so that these cells remain viable for future use. Any such techniques must result in high post-thaw stem cell recovery without compromising stemness, proliferation, or multipotency. Uncontrolled-rate freezing is not a technically or financially demanding technique compared to expensive and laborious controlled-rate freezing techniques. This study was aimed at observing the effect of uncontrolled-rate freezing on DPSCs stored for 6 and 12 months. Dimethyl sulfoxide at a concentration of 10% was used as a cryoprotective agent. Various features such as shape, proliferation capacity, phenotype, and multipotency were studied after DPSC thawing. The DPSCs did not compromise their stemness, viability, proliferation, or differentiating capabilities, even after one year of cryopreservation at −80 °C. After thawing, they retained their stemness markers and low-level expression of hematopoietic markers. We observed a size reduction in recovery DPSCs after one year of storage. This observation indicates that DPSCs can be successfully used in potential clinical applications, even after a year of uncontrolled cryopreservation.

Tranylcypromine Attenuates Senescence in Dental Pulp Stem Cells

2017 ◽  
Vol 14 (7) ◽  
Author(s):  
Junjun Liu ◽  
Zhi Liu ◽  
Chunyan Wang ◽  
Fang Yu ◽  
Wenping Cai ◽  
...  
Keyword(s):  
Stem Cells ◽  
Dental Pulp ◽  
Dental Pulp Stem Cells
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