TY - JOUR
T1 - Characterization of a stem cell of apical papilla cell line
T2 - Effect of passage on cellular phenotype
AU - Ruparel, Nikita B.
AU - De Almeida, José Flávio Affonso
AU - Henry, Michael A
AU - Diogenes, Anibal R
N1 - Funding Information:
This study was supported by the University of Texas Health Science Center at San Antonio, Department of Endodontics and CAPES, BrazilCAPES , Brazil ( BEX3563/103 ).
Copyright:
Copyright 2014 Elsevier B.V., All rights reserved.
PY - 2013/3
Y1 - 2013/3
N2 - Introduction: Stem cells of the apical papilla (SCAP) have been identified as an important population of mesenchymal stem cells (MSCs) in regenerative endodontics. Preclinical studies that evaluate the various aspects of the regenerative process must use fully characterized MSCs. The phenotype of these cells when maintained in culture is crucial for the translational applicability of these studies. Thus, in this study, we aimed to characterize a SCAP cell line that preferentially expressed and maintained the required MSCs markers in culture, namely the RP-89 cell line. Methods: Apical papillae from extracted mandibular third molars from a single donor were processed for immunohistochemistry, cell culture, and RT-PCR. SCAP were successfully cultured and maintained in culture for up to 20 passages. The expression of MSC-related molecular markers was analyzed by laser confocal microscopy, flow cytometry, and real-time RT-PCR arrays. Results: Cells within the apical papillae tissue had a widespread expression of CD90, whereas the expression of CD105 and CD73 was compartmentalized to the blood vessels and periphery, respectively. The RP-89 cell line population coexpressed CD73, CD90, and CD105 in all passages evaluated. There was a dramatic change in gene expression when cells were cultured and maintained in culture with the up-regulation of MSCs markers, inhibitors of differentiation, and stemness markers. Conversely, genes involved in the differentiation of MSCs were suppressed. Conclusions: Collectively, these results highlight the need to use fully characterized cell lines in regenerative studies and provide the foundational knowledge of gene expression modulation that occurs in cultured SCAP.
AB - Introduction: Stem cells of the apical papilla (SCAP) have been identified as an important population of mesenchymal stem cells (MSCs) in regenerative endodontics. Preclinical studies that evaluate the various aspects of the regenerative process must use fully characterized MSCs. The phenotype of these cells when maintained in culture is crucial for the translational applicability of these studies. Thus, in this study, we aimed to characterize a SCAP cell line that preferentially expressed and maintained the required MSCs markers in culture, namely the RP-89 cell line. Methods: Apical papillae from extracted mandibular third molars from a single donor were processed for immunohistochemistry, cell culture, and RT-PCR. SCAP were successfully cultured and maintained in culture for up to 20 passages. The expression of MSC-related molecular markers was analyzed by laser confocal microscopy, flow cytometry, and real-time RT-PCR arrays. Results: Cells within the apical papillae tissue had a widespread expression of CD90, whereas the expression of CD105 and CD73 was compartmentalized to the blood vessels and periphery, respectively. The RP-89 cell line population coexpressed CD73, CD90, and CD105 in all passages evaluated. There was a dramatic change in gene expression when cells were cultured and maintained in culture with the up-regulation of MSCs markers, inhibitors of differentiation, and stemness markers. Conversely, genes involved in the differentiation of MSCs were suppressed. Conclusions: Collectively, these results highlight the need to use fully characterized cell lines in regenerative studies and provide the foundational knowledge of gene expression modulation that occurs in cultured SCAP.
KW - Apical papilla
KW - endodontics
KW - flow cytometry
KW - passage
KW - polymerase chain reaction array
KW - regenerative
KW - stem cell
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U2 - 10.1016/j.joen.2012.10.027
DO - 10.1016/j.joen.2012.10.027
M3 - Article
C2 - 23402507
AN - SCOPUS:84873701850
SN - 0099-2399
VL - 39
SP - 357
EP - 363
JO - Journal of Endodontics
JF - Journal of Endodontics
IS - 3
ER -