Fig. 1: Construction of iPSCs with a novel mutation in TIMM8A.
A Schematic diagram illustrating the generation of three induced pluripotent stem cell (iPSC) lines: MTS-iPSCs and control iPSCs (CTRL-iPSCs) were derived from an MTS patient carrying a novel mutation in TIMM8A (c.225-229del) and a healthy individual, respectively, while mutant iPSCs (MUT-iPSCs) were generated from CTRL-iPSCs using CRISPR/Cas9. All iPSCs were reprogrammed from peripheral blood mononuclear cells (PBMCs). B Partial Sanger sequencing chromatograms of TIMM8A in the CTRL-, MUT-, and MTS-iPSCs, showing the c.225-229del mutation. C mRNA levels of TIMM8A in CTRL- and MTS-iPSCs. n = 3 independent biological replicates. Data are represented as mean ± SD. Significance was determined by two-tailed unpaired Student’s t test: ns, not significant. D Western blot analysis showing the absence of TIMM8a protein with a frameshift mutation starting at the 75th amino acid residue (p.Q75fs95*), encoded by the mutant TIMM8A gene, in MUT- and MTS-iPSCs. E Hematoxylin and eosin staining of the three germ layers successfully differentiated from iPSCs in vivo. Scale bar: 50 μm. F Immunostaining confirming the generation of CTRL-, MUT-, and MTS-iPSCs using pluripotency markers, including NANOG (green), OCT4 (green), SOX2 (red), and SSEA4 (red). Nuclei were counterstained with DAPI (blue). Scale bar: 50 μm.
