Transglutaminase 2 Prevents Premature Senescence and Promotes Osteoblastic Differentiation of Mesenchymal Stem Cells through NRF2 Activation

Figure 1

TG2 is required for cell proliferation but has no effect on surface marker expression in human umbilical cord-derived MSCs. (a) Human umbilical cord-derived MSCs were induced into replicative senescence after subculture for a long-term expansion (passage number 3, 7, and 15). The expression levels of TG2, NRF2, NRF2-responsive genes, and senescence marker (p16) were assessed by western blot analysis. β-Actin was used as the loading control, and the relative intensities were normalized using the loading control. (b) MSCs were transfected with lentivirus expressing shRNA for TG2. The level of TG2 protein was assessed by western blot analysis. β-Actin was used as the loading control in the experiment. (c, d) Cell proliferation rate of wild-type and TG2-knockdowned MSCs was estimated by measuring cumulative population doubling level (CPDL) through passage 3–20 (c) and the percentage of BrdU incorporated cells with flow cytometry (d). (e, f) Representative examples of flow cytometric profiles from wild-type and TG2-knockdowned MSCs. The percentage of cells expressing positive surface markers ((e), CD73, CD90, CD105 and CD44) and negative markers ((f), CD45, CD31, and CD11b) for MSCs was shown. The results for western blots are represented from three independent experiments. The measurement of growth rate in MSCs and flow cytometry images are representative results from three independent experiments performed. Statistical significance was tested by Student’s t-test (n = 3; ).

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