Int J Med Sci. 2014 Sep 13;11(11):1201-7.

Oxidative stress-induced premature senescence in Wharton’s jelly-derived mesenchymal stem cells.

Choo KB1, Tai L2, Hymavathee KS2, Wong CY3, Nguyen PN2, Huang CJ4, Cheong SK5, Kamarul T6.
  • 1Centre for Stem Cell Research, Universiti Tunku Abdul Rahman, Selangor, Malaysia; ; 2. Department of Preclinical Sciences, Faculty of Medicine and Health Sciences, Universiti Tunku Abdul Rahman, Selangor, Malaysia;
  • 2Centre for Stem Cell Research, Universiti Tunku Abdul Rahman, Selangor, Malaysia;
  • 3Cytopeutics Sdn Bhd, Selangor, Malaysia;
  • 4Department of Animal Science & Graduate Institute of Biotechnology, Chinese Culture University, Taipei, Taiwan;
  • 5Centre for Stem Cell Research, Universiti Tunku Abdul Rahman, Selangor, Malaysia; ; 5. Dean’s Office, Faculty of Medicine and Health Sciences, Universiti Tunku Abdul Rahman, Selangor, Malaysia;
  • 6Tissue Engineering Group, National Orthopaedic Centre of Excellence for Research and Learning, Department of Orthopaedic Surgery, Faculty of Medicine, University of Malaya, 50603 Kuala Lumpur, Malaysia.

 

Abstract

BACKGROUND: On in vitro expansion for therapeutic purposes, the regenerative potentials of mesenchymal stem cells (MSCs) decline and rapidly enter pre-mature senescence probably involving oxidative stress. To develop strategies to prevent or slow down the decline of regenerative potentials in MSC culture, it is important to first address damages caused by oxidative stress-induced premature senescence (OSIPS). However, most existing OSIPS study models involve either long-term culture to achieve growth arrest or immediate growth arrest post oxidative agent treatment and are unsuitable for post-induction studies.

METHODS: In this work, we aimed to establish an OSIPS model of MSCs derived from Wharton’s Jelly by hydrogen peroxide (H2O2) treatment.

RESULTS: The optimal H2O2 concentration was determined to be 200 µM to achieve OSIPS when MSC reached growth arrest in 3 to 4 passages post-H2O2 treatment. H2O2-treated cells became heterogeneous in morphology, and were irregularly enlarged and flattened with granular cytoplasm. The cells were stained positive for SA-β-galactosidase, a senescence marker, and were shown to express elevated levels of other well-characterized senescence molecular markers, including p53, p21, p16 and lysosomal β-galactosidase (GLB1) in real-time RT-PCR analysis. The OSIPS-like features were confirmed with three independent WJ-MSC lines.

CONCLUSION: The establishment of an OSIPS model of WJ-MSC is a first step for subsequent investigation on molecular mechanisms of senescence and for screening potential anti-oxidative agents to delay or revert stressed-induced senescence.

KEYWORDS: hydrogen peroxide.; mesenchymal stem cell; oxidative stress; senescence

PMID: 25249788

 

Supplement

Mesenchymal stem cell (MSC) displays self-renewal ability and multilineage differentiation potential, and can be engineered to provide an important cell source in therapeutic applications and regenerative medicine, including cancer therapy, cartilage regeneration, peripheral nerve repair and cardiac differentiation. MSC exists in multiple tissues, in particular the bone marrow, adipose tissue, umbilical cord and peripheral blood. However, MSC isolated from the Wharton’s jelly (WJ) of the umbilical cord possesses greater proliferation and differentiation abilities, and contains more therapeutically active stem cells compared to MSC derived from adult tissues. Hence, WJ-MSC is thought to be a more promising cell type for transplantation treatments. Organismal aging due to long-term exposure to harmful environmental factors or endogenous metabolic stresses results in senescence, even in stem cells. Similarly, on in vitro expansion, MSC enters senescence gradually and, thus, impeding the stemness properties of the cell.

In our work, we demonstrated that oxidative stress by hydrogen peroxide treatment resulted in stress induced-premature senescence (OSIPS) in WJ-MSC, forcing the cells to enter senescence in a shorter duration. OSIPS WJ-MSC shares similar features with cells in replicative senescence, which is a state of permanent growth arrest after a finite cell division. OSIPS is, therefore, useful as a model to further study the mechanism(s) underlying senescence.

In our lab, we are currently using such an OSIPS senescence MSC model to investigate the role of microRNA (miRNA) and targets in regulating senescence. miRNAs are small endogenous non-coding RNAs that regulate expression of target genes at the post-transcriptional level through degradation or translational repression.

Through microarray analysis, we have now identified members of the miR-146 and miR-17/106b families that re dysregulated in H2O2-induced senescent WJ-MSC; the putative negatively-regulated G1/S-phase factors have also been identified (manuscript in preparation). Such modulatory miRNAs in senescence may subsequently be used to delay or reverse aging.

 

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