Abstract
BackgroundExpansion-mediated replicative senescence and age-related natural senescence have adverse effects on mesenchymal stem cell (MSC) regenerative capability and functionality, thus severely impairing the extensive applications of MSC-based therapies. Emerging evidences suggest that microRNA-34a (miR-34a) has been implicated in the process of MSC senescence; however, the molecular mechanisms with regard to how miR-34a influencing MSC senescence remain largely undetermined.MethodsMiR-34a expression in MSCs was evaluated utilizing RT-qPCR. The functional effects of miR-34a exerting on MSC senescence were investigated via gene manipulation. Relevant gene and protein expression levels were analyzed by RT-qPCR and western blot. Luciferase reporter assays were applied to confirm that Nampt is a direct target of miR-34a. The underlying regulatory mechanism of miR-34a targeting Nampt in MSC senescence was further explored by measuring intracellular NAD+ content, NAD+/NADH ratio and Sirt1 activity.ResultsIn contrast to Nampt expression, miR-34a expression incremented in senescent MSCs. MiR-34a overexpression in young MSCs resulted in senescence-associated characteristics as displayed by senescence-like morphology, prolonged cell proliferation, declined osteogenic differentiation potency, heightened senescence-associated-β-galactosidase activity, and upregulated expression levels of the senescence-associated factors. Conversely, miR-34a suppression in replicative senescent and natural senescent MSCs contributed to diminished senescence-related phenotypic features. We identified Nampt as a direct target gene of miR-34a. In addition, miR-34a repletion resulted in prominent reductions in Nampt expression levels, NAD+ content, NAD+/NADH ratio, and Sirt1 activity, whereas anti-miR-34a treatment exerted the opposite effects. Furthermore, miR-34a-mediated MSC senescence was evidently rescued following the co-treatment with Nampt overexpression.ConclusionThis study identifies a significant role of miR-34a playing in MSC replicative senescence and natural senescence via targeting Nampt and further mediating by NAD+-Sirt1 pathway, carrying great implications for optimal strategies for MSC therapeutic applications.
Highlights
Expansion-mediated replicative senescence and age-related natural senescence have adverse effects on mesenchymal stem cell (MSC) regenerative capability and functionality, severely impairing the extensive applications of mesenchymal SCs (MSCs)-based therapies
P3MSCs displayed fibroblasticlike morphology with elongated and spindle-shaped cell bodies; both P10MSCs and OMSCs exhibited senescence-like morphology with flattened, enlarged, and irregular-shaped cell bodies, less stereoscopic and visible granules, and particles in the cytoplasm
Quantitative analysis (Fig. 1e) showed that the percentage of SAβ-gal-positive cells in P10MSCs or OMSCs was significantly more abundant than that in P3MSCs. These observations displayed that P10MSCs and OMSCs showed the senescent alterations unlike young P3MSCs/YMSCs, suggesting that replicative senescence appeared with extensive passages and natural senescence occurred with advancing age
Summary
Expansion-mediated replicative senescence and age-related natural senescence have adverse effects on mesenchymal stem cell (MSC) regenerative capability and functionality, severely impairing the extensive applications of MSC-based therapies. Cultivated primary cells in vitro undergo replicative senescence, which is telomere-initiated senescence [8, 9] Another type of cellular senescence arising from in vivo chronological aging process of individuals, inescapably accompanied by distinctive senescence-related phenotypic characterization, is named as natural senescence [10]. These are considered as major impediments to applications in basic scientific research and clinical MSC-based therapeutic strategies. Promising strategies to rejuvenate replicative and natural senescent MSCs merit urgent exploration
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
