Fig. 4
LMW-DFR-GM and - DM prevent proliferation and aging of fibroblasts. (A) Confocal live images of live and dead assays show the CCD-986sk cells cultivated in con-DFR-GM and LMW-F-DFR-GM, with live cells indicated in green. Scale bars represent 200 μm. (B) CCD-986sk cells were incubated with LMW-F-DFR-GM for 7 days. Representative images show the harvested cell pellets of each group after 7 days. (C) The bar graph represents cell viability of CCD-986sk cells cultivated in con-DFR-GM and LMW-F-DFR-GM using a hemocytometer. (D) The histogram data of each group was determined by live PI staining and CytoFLEX FACS analysis. The cell population (purple area, blue line) indicates the positive PI staining to assess cell death. The overlaying histogram displays the cell viability of CCD-986sk cells cultivated in con-DFR-GM (green) and LMW-F-DFR-GM (red) at 7 days. (E) CCD-986sk cells were incubated with LMW-F-DFR-DM for 14 days. Brightfield images of cellular senescence-associated β-galactosidase staining of each group. The red arrows indicate the senescent cells positive for SA-β-gal staining. Scale bars, 200 μm. (F) The bar graph shows the number of SA-β-gal positive cells in CCD-986sk cells cultivated in con-DFR-GM (green) and LMW-F-DFR-GM (red) at 14 days. The data are represented as the mean ± SEM of triplicate assays expressed as a ratio of the con-DFR-DM. Statistical analysis was performed using Student’s t-test. Data was analyzed using Student’s t-test. **P < 0.01 or *P < 0.05 versus controls.
