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Fig. 6. Tbx5 misexpression leads to changes in cardiac proliferation and morphology. The overall morphology of stage 40 (A) uninjected embryos or (B,C) embryos injected with increasing amounts of Tbx5 RNA, as indicated. Arrows denote the location of the heart. (D-G) Whole-mount in-situ hybridization showing expression of (D,E) Nkx2.5 and (F,G) myosin light chain (MLC) in (D,F) uninjected stage 37 embryos and (E,G) stage-matched embryos injected with 1 ng of Tbx5 RNA. (H) Tbx5 misexpression leads to an increase in the cardiac mitotic index at stage 37. Mitotic index was calculated as the percentage of cardiac cells labeled with pH3. The data represents the mean of at least three different embryos. Error bars denote the standard deviation, and * denotes a statistically significant difference between Tbx5-injected and control embryos (at P<0.05). (I) Mitotic index for sections of the neural tube corresponding to the same position as the heart along the anterior-posterior axis. The data represents the mean mitotic index of four different embryos per condition, with four sections analyzed per embryo. Error bars denote the standard deviation. (J) Tbx5 misexpression leads to an alteration in the timing and order of the cardiac differentiation program. RT-PCR analysis of the expression of Nkx2.5 as well as heart-specific isoforms of MHC, troponin and tropomyosin, and skeletal-muscle-specific genes, MyoD and muscle actin, throughout early and mid-gestation stages of development in control (`C') or Tbx5-injected (`T') stage-matched embryos. All samples are derived from a single batch of eggs and identical results were achieved in at least two independent sets of experiments for each marker. EF1-alpha was used as a control for all RT-PCR reactions. |
