Figure 6

Exogenous, extracellular TG2 inhibits adipogenesis and activates β-catenin signaling and recovers Pref-1 protein levels. (a, b) Tgm2+/+ and Tgm2−/− MEF cultures were treated with increasing concentrations (0.5–5 μg/ml) of exogenous TG2 (ExoTG2) from days 0 to 8. Graphs show quantification of Oil Red O staining on day 8. Exogenous TG2 was able to reduce lipid accumulation in a significant manner in both Tgm2+/+ and Tgm2−/− MEFs. Results are mean values±SEM (n=3). ***P<0.001. *P<0.05; **P<0.01. (c) mRNA expression of Pparγ and Cebpα in Tgm2+/+ and Tgm2−/− MEFs on days 0 and 1 with or without ExoTG2 (5 μg/ml); DM-differentiation medium. A reduced expression was observed with ExoTG2. (d) Western blot analysis of total β-catenin levels in total cell lysate of Tgm2+/+ and Tgm2−/− MEFs on day 1 with or without ExoTG2 (5 μg/ml) show no difference; actin used as a loading control. (e, f) Western blot analysis and quantification of β-catenin levels in cytosolic (c) and nuclear (N) fractions of Tgm2+/+ and Tgm2−/− MEFs on day 1 with or without ExoTG2 (5 μg/ml). Normalization was done to loading controls α-tubulin and histone H3. Tgm2−/− MEFs show significantly increased levels of β-catenin in the nucleus. Error bars±SEM (n=3), *P<0.05; **P<0.01. (g) mRNA expression of Pref-1 in Tgm2+/+ and Tgm2−/− MEFs on days 0 and 1 with or without ExoTG2 (5 μg/ml). (h) Western blot analysis of total cell lysate for Pref-1 in Tgm2+/+ and Tgm2−/− MEFs on day 1 with or without ExoTG2 (5 μg/ml). ExoTG2 treatment recovered Pref-1 protein levels in Tgm2−/− MEFs