Journal: Scientific Reports
Article Title: Regulation of autophagy and its role in late preimplantation during mouse embryo development
doi: 10.1038/s41598-025-11359-2
Figure Lengend Snippet: Rescue of autophagy inhibition by AA supplementation. Developmental rates ( a ) and developmental results ( b ) following culture with 10 μM CQ treatment from the 4/8-cell stages to the blastocyst stage with/without AA supplementation. Scale bars represent 20 μm. ( c ) The embryos at the morula stage, cultured with CQ and AA as shown in panel ‘a,’ were immunostained for Cdx2 and Nanog, as well as with DAPI. Scale bars represent 50 μm. ( d ) The graph shows the numbers of DAPI-positive cells counted in the embryos, as shown in ( c ). Tukey–Kramer’s HSD test, CZB vs. CZB + AA + CQ p = 0.007, CZB + CQ vs. CZB + AA + CQ p = 0.0015. ( e ) The graph shows the proportion of Cdx2- and Nanog-positive cells per total cell number counted in the embryos, as shown in ( c ). Tukey–Kramer’s HSD test, Cdx2: CZB + CQ vs. CZB + AA + CQ p = 0.0012. Nanog: CZB vs. CZ + AA + CQ p = 0.0418, CZB + AA vs. CZB + CQ p = 0.0286, CZB + CQ vs. CZB + AA + CQ p = 0.0005. ( f ) The proportion of positive/negative cells in each embryo, as shown in ( c ). The cells are classified into four types: Nanog-positive (Nanog + , green), Cdx2-positive (Cdx2 + , red), double positive for Nanog and Cdx2 (Nanog + /Cdx2 + , yellow), and double negative (Nanog-/Cdx2-, gray). Tukey–Kramer’s HSD test, Cdx2-/Nanog-: CZB vs. CZB + CQ p = 0.0034, CZB + CQ vs. CZB + AA + CQ p = 0.0045. Cdx2 + /Nanog + : CZB vs. CZB + CQ p = 0.0035. ( g ) The embryos at the morula stage, cultured with CQ and AA as shown in ( a ), were immunostained for TFAP2C and Nanog, as well as with DAPI. Scale bars represent 50 μm. ( h ) The graph shows the proportion of TFAP2C- and Nanog-positive cells per total cell number counted in the embryos, as shown in ( g ). Tukey–Kramer’s HSD test, TFAP2C: no significant differences were observed . Nanog: CZB + CQ vs. CZB + AA + CQ p = 0.0322. ( i ) Relative fluorescence intensity of Nanog (green) and TFAP2C (red) in embryos shown in ( g ). Calibration was performed using DAPI fluorescence intensity.
Article Snippet: The anti-CDX2 monoclonal antibody( 1:500; BioGenex, San Ramon, CA, USA, MU392A-UC) to detect TE cells, anti-Nanog rabbit polyclonal antibody (1:500; Abcam, Cambridge, UK, ab80892) to detect the ICM cells and anti-TFAP2C(1:500; AP-2γ antibody; Santa Cruz Biotechnology, TX, USA, sc-12762) were the primary antibodies used.
Techniques: Inhibition, Cell Culture, Fluorescence
