A Decline in Wnt3a Signaling is Necessary for Mesenchymal Stem Cells to Proceed to Replicative Senescence

Abstract

Umbilical cord blood-derived mesenchymal stem cells are a promising source of cells for regeneration therapy due to their multipotency, high proliferative capacity, relatively noninvasive collection, and ready availability. However, extended cell culture inevitably triggers cellular senescence-the irreversible arrest of cell divisionthereby limiting the proliferative lifespan of adult stem cells. Wnt/b-catenin signaling plays a functional role as a key regulator of self-renewal and differentiation in mesenchymal stem cells (MSCs), and thus Wnt/b-catenin signaling and cellular senescence might be closely connected. Here, we show that the expression levels of canonical Wnt families decrease as MSCs age during subculture. Activation of the Wnt pathway by treatment with Wnt3a-conditioned medium or glycogen synthase kinase 3b inhibitors, such as SB-216763 and 6-bromoindirubin-3¢-oxime, delays the progression of cellular senescence as shown by the decrease in the senescence effectors p53 and pRb, lowered senescence-associated b-galactosidase activity, and increased telomerase activity. In contrast, suppression of the Wnt pathway by treatment with dickkopf-1 (an antagonist of the Wnt coreceptor) and bcatenin siRNA transfection promotes senescence in MSCs. Interestingly, the magnitude of the response to enhanced Wnt3a/b-catenin signaling appears to depend on the senescent state during extended culture, particularly after multiple passages. These results suggest that Wnt3a signaling might be a predominant factor that could be used to overcome senescence in long-term cultured MSCs by directly intervening in the proliferative capacity and MSC senescence. The functional role of Wnt3a/b-catenin signaling in hedging cellular senescence may allow the development of new approaches for stem cell-based therapies.

Wnt Signaling Inhibits High-Density Cell Sheet Culture Induced Mesenchymal Stromal Cell Aging by Targeting Cell Cycle Inhibitor p27

“…More importantly, Wnt signaling effector β-catenin showed a similar expression pattern to p27 suggesting a possible interaction between these two factors. A previous study also supports that Wnt signaling might be a predominant factor that could be used to overcome extensive passaging-induced cell aging in longterm cultured MSCs by directly intervening in the proliferative capacity and MSC senescence (Jeoung et al, 2015). In this study, we further discovered the underlying mechanism by using a unique cell sheet culture-induced cell aging system.…”

Adipose tissue-derived extracellular fraction characterization: biological and clinical considerations in regenerative medicine

“…Additionally, we evaluated the presence of α-MSH and SCF, two important factors in the regulation of melanocyte differentiation and survival [ 43 , 44 ]. We also demonstrated that two modulators of the Wnt/β-catenin pathway, Wnt3a and Wnt10b, implicated in replicative senescence regulation [ 45 ] and in wound healing processes [ 46 , 47 ], respectively, are abundantly present in lipoaspirates. Overall, our data demonstrated a specific signature of adipose tissue secretome that does not overlap with circulating factors.…”

Downregulation of Melanoma Cell Adhesion Molecule (MCAM/CD146) Accelerates Cellular Senescence in Human Umbilical Cord Blood-Derived Mesenchymal Stem Cells

“…A histochemical staining kit (Sigma-Aldrich) was used according to the manufacturer's instructions to qualitatively assess SA b-gal activity, and cells were examined on an inverted microscope. To assess senescent-cell formation, the overall percentage of stained cells in the cell populations was averaged from four fields [24].…”

“…Telomerase activity was analyzed by a telomeric repeat amplification protocol (TRAP) assay using a telomerase polymerase chain reaction (PCR) enzyme-linked immunosorbent assay kit (Roche, Mannheim, Germany, http://www.roche.com), according to manufacturer's protocol [24]. Briefly, cells (2 3 10 5 ) were harvested for each reaction and centrifuged at 3,000g for 10 minutes at 4°C, washed twice with PBS, incubated for 20 minutes at 4°C with 200 ml lysis buffer, and centrifuged at 16,000g for 20 minutes.…”