Figure 7

Spatial and temporal profile of Umodl1 expression in ovarian follicles of aging Tg mice and disturbed ovary-specific gene regulatory pathways in response to Umodl1 overexpression in the 4-month-old Tg mice. (a) SYBR green-based quantitative PCR assay was employed to determine the relative levels of Umodl1 in mGCs, cGCs, oocytes and whole ovaries of WT and Tg female mice (n⩾3 per genotype) at various time points. Quantity of each sample was normalized to expression of GAPDH (Ct=14.8–15.6). No significant levels of Umodl1 were detected in the WT mGCs and cGCs, as well as the Tg mGCs (Ct>25). Umodl1 levels in Tg cGCs of different ages were compared with that of 2-month-old Tg cGCs; while levels of Umodl1 in oocytes and ovaries were compared with those from the 2-month-old WT samples, respectively. Data points represent mean±S.E.M. of three repeats. * and ** represent significant difference P<0.01 and P<0.05, respectively, as compared with the age-matched WT controls. (b) Representative western blots showing expression of Umodl1 in different ovarian cells at various ages before a complete degeneration of the Tg ovaries. Samples from age-matched WT cells per tissues were included. β-tubulin was used as the internal control. Tissue lysate from a 2-month-old WT spleen was used as the positive control for Umodl1(+Ctrl). The blot loaded with lysate from whole ovaries was stripped and re-probed with a Bax antibody to demonstrate an elevated apoptosis in the Tg ovaries. (c) Altered expression patterns of key ovary-specific genes in the ovaries of pW224 Tg mice at 4 months of age. In situ hybridization using³⁵S-UTP labeled antisense probes was performed. Panels in columns 1 and 3 are the bright field views of tissue sections, while panels in columns 2 and 4 are photographs showing corresponding dark field views to visualize the hybridization signals. Umodl1 genotype is indicated on the top. The molecular markers examined are listed on the left. Scale bar, 100 μm