Unknown,Transcriptomics,Genomics,Proteomics

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Time course assay in Mouse Pre-Somitic Mesoderm

ABSTRACT: Stem cell-derived tissues have wide potential for modelling developmental and pathological processes as well as cell-based therapy. However, it has proven difficult to generate several key cell types in vitro, including skeletal muscle. In vertebrates, skeletal muscles derive during embryogenesis from the presomitic mesoderm (PSM). Treatment of mouse ES cells with a combination of the secreted Wnt activator R-Spondin3 and the BMP inhibitor Noggin generated cells expressing the early PSM marker Mesogenin1 (Msgn1) with high efficiency. To confirm their identity, we mapped gene expression profiles at successive stages of PSM differentiation in vivo and showed that the differentiated ES cells closely corresponded to the posterior PSM domain that expresses Msgn1 and then with time in culture matured to acquire the profile of the anterior Pax3 domain. When grafted into injured adult muscle in vivo these Pax3-expressing-cells generated large numbers of muscle fibers. Our system therefore efficiently produces myogenic precursors in vitro by recapitulating stepwise early myogenic differentiation in vivo. These findings should advance the development of cellular therapies for muscle degenerative diseases. Pre Somitic Mesoderm (PSM) were dissected into 7 pieces in Stage E8.5 Mouse embryo. The experiment was designed to have biological triplicate in each pieces. The PSM is a symmetric structure, left and right area were obtained from one embryo, another set of dissection was obtained from another embryo.

ORGANISM(S): Mus musculus

SUBMITTER: philippe moncuquet

PROVIDER: E-GEOD-39613 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Within a given vertebrate species, the total number of vertebrae in each anatomical domain is precisely defined and shows little variation among individuals. In contrast, this number can vary tremendously between different species, ranging from as few as six vertebrae in frogs to as many as several hundred in some snakes and fish. Segmental precursors of the vertebrae, called somites are produced sequentially in the embryo from the presomitic mesoderm (PSM), until a final number characteristic of the species, is reached. Here, we show in the chicken embryo that, by controlling the rate of axis elongation, Hox genes control the total number of somites generated by the embryo. We observed that activation of the most posterior Hox genes in somite precursors of the tail bud correlates with an abrupt slowing-down of the speed of axis elongation. We show that progressively more posterior Hox genes, which are collinearly activated in somitic precursors of the epiblast, repress Wnt activity with increasing strength. This leads to a graded repression of the Brachyury/T transcription factor, reducing mesoderm ingression and slowing down the elongation process. Due to the continuation of somite formation, the PSM, which is not fed with sufficient supply of new cells posteriorly, becomes progressively exhausted, ultimately leading to an arrest of segment formation. Our data provide a conceptual framework to explain how the cross-talk between the segmentation clock and the Hox clock accounts for the diversity of vertebral formulae across animal species. Primitive streak aera corresponding to PSM precursors were dissected in Stage 9 somites chicken embryo overexpressing HoxA13 or a control H2B-Venus. The experiment was designed to have biological duplicate in each conditions. The gain-of-function was obtained by electroporating the embryo at Stage 5HH with a vector containing HoxA13 under the CAGGS promoter with an H2B venus reporter. In order to have enough material for the microarray, 7 embryos were pooled in each sample before the hybridization.

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Time course assay in Mouse Pre-Somitic Mesoderm

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