Prelamin a

SCB-MSCs at passage 3 were seeded on glass coverslips placed in 24-well culture plates and fixed in 4% paraformaldehyde (PFA). Cells were examined by confocal fluorescence microscopy (Leica SP8, Confocal microscope). Immunofluorescence was performed using the following antibodies: prelamin A (sc-518013 C-3, Santa Cruz), phospho-histone H2AX (#2577, Cell Signaling Technology), 53BP1 (#4937, Cell Signaling Technology), Ki67 (ab15580, Abcam), calponin 1 (ab46794, Abcam), alpha smooth muscle actin (ab124964, Abcam), mCD31(#14-0311-82, eBioscence), and β-actin (sc-47778,Santa Cruz). 4′,6-diamidino-2-phenylindole (DAPI) (zli-9557, Zhongshan) was used to counterstain cell nuclei. Around 100 randomly selected cells were analyzed.

For immunostaining, cells were fixed in 4% paraformaldehyde, permeabilised in 0.5% Nonidet P-40 alternative in phosphate buffered saline (PBS) and blocked in 1% bovine serum albumin (BSA) in PBS for 1 h. Cells were incubated with prelamin A (Santa Cruz, Dallas, TX, USA) or vinculin (Sigma-Aldrich, St Louis, MO, USA) primary antibody in blocking solution overnight at 4 °C. After washing with PBS, cells were incubated in fluorescence-conjugated secondary antibody solution or Rhodamine phalloidin (Invitrogen) for 1 h before staining cell nuclei with 4′,6′-Diamidino-2-Pheylindoledihydrochloride (DAPI). Cells were washed and mounted onto slides using Mowiol/Dabco media. Images were captured using a Zeiss Axioplan 2 light microscope and Volocity software (Perkin Elmer, Waltham, MA, USA) was used to control image acquisition and measure morphological parameters (area and circularity) by fluorescence detection.