Fig. 2.

Biochemical assessment of OGT and O-GlcNAcylation in mouse and human placentas. (A) Representative Western blot images of OGT levels from male and female mouse placentas. Histogram is the maximum likelihood estimate for the normalized optical densities of each group (n = 7) ± the 95% confidence interval for that estimate. Asterisk indicates measurable difference between groups as determined by nonoverlapping confidence intervals for the estimates. (B) Representative Western blot images of OGT levels from control (n = 7) and EPS (n = 8) male mouse placentas. Histogram was derived and annotated as in A. (C) Representative Western blot images of total O-GlcNAcylated proteins from male and female mouse placentas. Histogram was derived and annotated as in A (n = 7). (D) Representative Western blot images of total O-GlcNAcylated proteins from male control (n = 7) and EPS (n = 8) placentas. The image is annotated to highlight the bands visibly identified with differential O-GlcNAcylation between treatment groups excised for proteomic analyses. (E) Amino acid sequence coverage of LC-MS/MS where the band at 28 kDA was identified as peroxiredoxin-1 (PRDX1) and the band at 37 kDA was identified as annexin A1 (ANXA1) via LC-MS/MS with greater than 99.9% certainty. (F) Representative spectrum image of peptide fragments used to identify both ANXA1 and PRDX1. (G) Expression of OGT by RT-PCR in human placental tissue associated with male births. Data were normalized to XY levels. Bars are the maximum likelihood estimate for each group (n = 4) ± the 95% confidence interval for that estimate. XX, maternal; XY, fetal. (H) Representative Western blot images of total O-GlcNAcylated proteins from both XX and XY placental contributions from human placentas. Histogram was derived as in A (n = 4). Asterisk indicates a measurable difference between groups with different X-chromosome complement as determined by nonoverlapping confidence intervals for each estimate.