An Investigation on Mitochondrial DNA Deletions and Telomere Shortening during Multiple Passages of Adult Stem Cells

BACKGROUND Limited resources for adult stem cells necessitate their in vitro culture prior to clinical use. Investigating mitochondrial DNA (mtDNA) and telomere shortening has proved to be important indications of stem cell validity. This study was designed to investigate these indicators in multiple passages of three adult stem cell lines which were produced in our stem cell laboratory. METHODS In this study, Dental Pulp Stem Cells (DPSCs), Periapical Follicle Stem Cells (PAFSCs) and Human Foreskin Fibroblast (HFF) cell lines were expanded for 20 passages. After 1, 5, 10, 15 and 20 passages, expanded cells were harvested and DNA was extracted for further studies. Common mtDNA mutation was detected by multiplex PCR and telomere shortening was tested by Southern blot analysis. RESULTS The common deletion was not detected in any of the stem cells or cell lines after several passages. In addition, Southern blot analysis indicated that the mean difference of telomere length between first and last passage was 0.25 kb in DPSC, 0.1 kb in PAFSC and 0.32 kb in HFF which indicates that the mean telomere length in various passages of the samples showed insignificant changes. CONCLUSION Absence of mtDNA mutations in adult stem cell lines indicates good mitochondrial function even after 20 passages. In addition, absence of telomere shortening indicates stem cells validity after multiple passages. It is hoped this information could pave the way for using in vitro expansion of adult stem cells for future clinical applications.