Impact of isolation method on doubling time and the quality of chondrocyte and osteoblast differentiated from murine dental pulp stem cells

Background Stem cells are normally isolated from dental pulps using the enzymatic digestion or the outgrowth method. However, the effects of the isolation method on the quality of the isolated stem cells are not studied in detail in murine models. The aim of this study was to compare the matrices secreted by osteoblast and chondrocytes differentiated from dental pulp stem cells isolated through different means. Method DPSC from murine incisors were isolated through either the outgrowth (DPSC-OG) or the enzymatic digestion (DPSC-ED) method. Cells at passage 4 were used in this study. The cells were characterized through morphology and expression of cell surface markers. The cells’ doubling time when cultured using different seeding densities was calculated and analyzed using one-way ANOVA and Tukey’s multiple comparison post-test. The ability of cells to differentiate to chondrocyte and osteoblast was evaluated through staining and analysis on the matrices secreted. Results Gene expression analysis showed that DPSC-OG and DPSC-ED expressed dental pulp mesenchymal stem cell markers, but not hematopoietic stem cell markers. The least number of cells that could have been used to culture DPSC-OG and DPSC-ED with the shortest doubling time was 5 × 102 cells/cm2 (11.49 ± 2.16 h) and 1 × 102 cells/cm2 (10.55 h ± 0.50), respectively. Chondrocytes differentiated from DPSC-ED produced 2 times more proteoglycan and at a faster rate than DPSC-OG. FTIR revealed that DPSC-ED differentiated into osteoblast also secreted matrix, which more resembled a calvaria. Discussion Isolation approaches might have influenced the cell populations obtained. This, in turn, resulted in cells with different proliferation and differentiation capability. While both DPSC-OG and DPSC-ED expressed mesenchymal stem cell markers, the percentage of cells carrying each marker might have differed between the two methods. Regardless, enzymatic digestion clearly yielded cells with better characteristics than outgrowth.

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Comparison of Mesenchymal Stem Cell Markers in Multiple Human Adult Stem Cells

International Journal of Stem Cells ◽

10.15283/ijsc.2014.7.2.118 ◽

2014 ◽

Vol 7 (2) ◽

pp. 118-126 ◽

Cited By ~ 104

Author(s):

Masoud Maleki ◽

Farideh Ghanbarvand ◽

Mohammad Reza Behvarz ◽

Mehri Ejtemaei ◽

Elham Ghadirkhomi

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Mesenchymal Stem Cell ◽

Adult Stem Cells ◽

Stem Cell Markers ◽

Cell Markers ◽

Human Adult

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Characterization of deciduous teeth stem cells isolated from crown dental pulp

Vojnosanitetski pregled ◽

10.2298/vsp1408735d ◽

2014 ◽

Vol 71 (8) ◽

pp. 735-741 ◽

Cited By ~ 3

Author(s):

Jasmina Debeljak-Martacic ◽

Jelena Francuski ◽

Tijana Luzajic ◽

Nemanja Vukovic ◽

Slavko Mojsilovic ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Stem Cell ◽

Mesenchymal Stem Cell ◽

Dental Pulp ◽

Doubling Time ◽

Deciduous Teeth ◽

Population Doubling ◽

Population Doubling Time ◽

Differentiation Potential

Background/Aim. The last decade has been profoundly marked by persistent attempts to use ex vivo expanded and manipulated mesenchymal stem cells (MSCs), as a tool in different types of regenerative therapy. In the present study we described immunophenotype and the proliferative and differentiation potential of cells isolated from pulp remnants of exfoliated deciduous teeth in the final phase of root resorption. Methods. The initial adherent cell population from five donors was obtained by the outgrowth method. Colony forming unit-fibroblast (CFU-F) assay was performed in passage one. Cell expansion was performed until passage three and all tests were done until passage eight. Cells were labeled for early mesenchymal stem cells markers and analysis have been done using flow cytometry. The proliferative potential was assessed by cell counting in defined time points and population doubling time was calculated. Commercial media were used to induce osteoblastic, chondrogenic and adipogenic differentiation. Cytology and histology methods were used for analysis of differentiated cell morphology and extracellular matrix characteristics. Results. According to immunophenotype analyses all undifferentiated cells were positive for the mesenchymal stem cell markers: CD29 and CD73. Some cells expressed CD146 and CD106. The hematopoietic cell marker, CD34, was not detected. In passage one, incidence of CFU-F was 4.7 ? 0.5/100. Population doubling time did not change significantly during cell subcultivation and was in average 25 h. After induction of differentiation, the multicolony derived cell population had a tri-lineage differentiation potential, since mineralized matrix, cartilage-like tissue and adipocytes were successfully formed after three weeks of incubation. Conclusion. Altogether, these data suggest that remnants of deciduous teeth dental pulp contained cell populations with mesenchymal stem cell-like features, with a high proliferation and trilineage differentiation potential and that these cultures are suitable for further in vitro evaluation of cell based therapies.

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Comparison of Immuno-Phenotypes of Stem Cells from Human Dental Pulp and Periodontal Ligament

International Journal of Immunopathology and Pharmacology ◽

10.1177/039463201202500115 ◽

2012 ◽

Vol 25 (1) ◽

pp. 127-134 ◽

Cited By ~ 23

Author(s):

D. Ponnaiyan ◽

K.M. Bhat ◽

G.S. Bhat

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Cell Population ◽

Dental Pulp ◽

Periodontal Ligament ◽

Embryonic Stem ◽

Stem Cell Population ◽

Stem Cell Markers ◽

Cell Markers ◽

Human Dental Pulp

It has been established that human dental pulp and periodontal ligament contain a population of mesenchymal stem cells (MSCs). However, the phenotypic analysis in terms of putative stem cell markers expressed by these stem cell populations is incomplete. It is relevant to understand whether stem cells derived from closely related tissues are programmed differently. The aim of the present study is to analyze whether these stem cells depict distinct characteristics by gaining insight into differences in their immunophenotype. Dental pulp and periodontal ligament tissue samples were obtained from extracted impacted wisdom teeth. Cell cultures were analyzed for surface and intracellular markers by indirect immunoflourescence. Detailed immunophenotype analysis was carried out by flow cytometry using relevant markers. The present study data shows dental pulp stem cells (DPSCs) and periodontal ligament stem cells (PDLSCs) expressed embryonic stem (ES) cell markers Oct-4, Nanog and mesodermal marker Vimentin by indirect immunoflourescence. PDLSCs, however, had a weak expression of Nanog. Immunophenotyping revealed strong expression of MSC markers (CD73, CD90) in DPSCs and PDLSCs. Differences were observed in expression of sternness-related markers. DPSCs displayed increased percentages of SSEA4, CD13 and CD166 and decreased CD9 expression compared to PDLSCs. Both stem cells express common MSC markers, different levels of expression suggests there might be more than one stem cell population existing within these tissues which differ in their embryonic status, and DPSCs are a more primitive stem cell population in comparison to PDLSCs.

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Isolation and Characterization of a Population of Immature Dental Pulp Stem Cells Expressing OCT-4 and Other Embryonic Stem Cell Markers

Cells Tissues Organs ◽

10.1159/000099617 ◽

2006 ◽

Vol 184 (3-4) ◽

pp. 105-116 ◽

Cited By ~ 264

Author(s):

Irina Kerkis ◽

Alexandre Kerkis ◽

Dmitri Dozortsev ◽

Gaëlle Chopin Stukart-Parsons ◽

Sílvia Maria Gomes Massironi ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Embryonic Stem Cell ◽

Dental Pulp ◽

Embryonic Stem ◽

Stem Cell Markers ◽

Cell Markers ◽

Isolation And Characterization ◽

Embryonic Stem Cell Markers

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Mesenchymal stem cell properties of dental pulp cells from deciduous teeth

Archives of Biological Sciences ◽

10.2298/abs1104933n ◽

2011 ◽

Vol 63 (4) ◽

pp. 933-942 ◽

Cited By ~ 9

Author(s):

N. Nikolic ◽

A. Krstic ◽

D. Trivanovic ◽

S. Mojsilovic ◽

J. Kocic ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Dental Pulp ◽

Adult Stem Cells ◽

Mesenchymal Cell ◽

Deciduous Teeth ◽

Proliferative Potential ◽

Stem Cell Markers ◽

Hematopoietic Stem ◽

Cell Markers

In the present study we have isolated and identified mesenchymal stem cells (MSCs) from the exfoliated deciduous teeth dental pulp (DP-MSCs), as plastic-adherent, spindle-shaped cells with a high proliferative potential. Immunophenotype analyses revealed that DP-MSCs were positive for mesenchymal cell markers (CD90, CD44, CD105, STRO-1, vimentin and ?-SMA), and negative for hematopoietic stem cell markers (CD11b, CD33, CD34, CD45, CD235a). DPMSCs were also capable of differentiating into adipogenic, chondrogenic, myogenic and osteogenic lineages, fulfilling the functional criterion for their characterization. These results demonstrate that DP-MSCs offer a valuable, readily accessible source to obtain and store adult stem cells for future use.

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Effect of laser phototherapy on gene expression of stem cell markers of human dental pulp stem cells

Medicina Oral Patología Oral y Cirugia Bucal ◽

10.4317/medoral.17643599 ◽

2012 ◽

pp. S100-. ◽

Cited By ~ 1

Author(s):

LS Ferreira ◽

MFSS Destro ◽

CMC Maranduba ◽

SPH Miyagi ◽

MM. Marques

Keyword(s):

Gene Expression ◽

Stem Cells ◽

Stem Cell ◽

Dental Pulp ◽

Dental Pulp Stem Cells ◽

Stem Cell Markers ◽

Cell Markers ◽

Laser Phototherapy ◽

Human Dental Pulp

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Mesenchymal Stem Cells in the Treatment of Cartilage Defects of the Knee: A Systematic Review of the Clinical Outcomes

The American Journal of Sports Medicine ◽

10.1177/0363546520986812 ◽

2021 ◽

pp. 036354652098681

Author(s):

Monketh Jaibaji ◽

Rawan Jaibaji ◽

Andrea Volpin

Keyword(s):

Systematic Review ◽

Stem Cells ◽

Mesenchymal Stem Cells ◽

Stem Cell ◽

Mesenchymal Stem Cell ◽

Osteochondral Lesions ◽

Common Source ◽

Significant Heterogeneity ◽

Osteochondral Defects

Background: Osteochondral lesions are a common clinical problem and their management has been historically challenging. Mesenchymal stem cells have the potential to differentiate into chondrocytes and thus restore hyaline cartilage to the defect, theoretically improving clincal outcomes in these patients. They can also be harvested with minimal donor site morbidity. Purpose: To assess the clinical and functional outcomes of mesenchymal stem cell implantation to treat isolated osteochondral defects of the knee. A secondary purpose is to assess the quality of the current available evidence as well as the radiological and histological outcomes. We also reviewed the cellular preparation and operative techniques for implantation. Study Design: Systematic review. Methods: A comprehensive literature search of 4 databases was carried out: CINAHL, Embase, MEDLINE, and PubMed. We searched for clinical studies reporting the outcomes on a minimum of 5 patients with at least 12 months of follow-up. Clinical, radiological, and histological outcomes were recorded. We also recorded demographics, stem cell source, culture technique, and operative technique. Methodological quality of each study was assessed using the modified Coleman methodology score, and risk of bias for the randomized controlled studies was assessed using the Cochrane Collaboration tool. Results: Seventeen studies were found, encompassing 367 patients. The mean patient age was 35.1 years. Bone marrow was the most common source of stem cells utilized. Mesenchymal stem cell therapy consistently demonstrated good short- to medium-term outcomes in the studies reviewed with no serious adverse events being recorded. There was significant heterogeneity in cell harvesting and preparation as well as in the reporting of outcomes. Conclusion: Mesenchymal stem cells demonstrated a clinically relevant improvement in outcomes in patients with osteochondral defects of the knee. More research is needed to establish an optimal treatment protocol, long-term outcomes, and superiority over other therapies. Registration: CRD42020179391 (PROSPERO).

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