somatic-genome-engineered-mouse-models-using-vivo-microinjection

Somatic Genome-Engineered Mouse Models Using <em>In Vivo</em> Microinjection and Electroporation

Germline genetically engineered mouse models (G-GEMMs) have provided valuable insight into in vivo gene function in development, homeostasis, and disease. ...

McGill University

The oviduct/fallopian tube is a tube-like structure that extends from the uterus to the ovary. It is an essential reproductive organ that provides an environment for internal fertilization and preimplantation development. However, our knowledge of its regional and cellular heterogeneity is still limited. Here, we examined the anatomical complexity of mouse oviducts using modern imaging techniques and fluorescence reporter lines. We found that there are consistent coiling patterns and turning points in the coiled mouse oviduct that serve as reliable landmarks for luminal morphological regionalities. We also found previously unrecognized anatomical structures in the isthmus and uterotubal junction, which likely play roles in reproduction. Furthermore, we demarcated the ampulla-isthmus junction as a distinct region. Taken together, the oviduct mucosal epithelium has highly diverse structures with distinct epithelial cell populations, reflecting its complex functions in reproduction.

Anatomical and cellular heterogeneity in the mouse oviduct-its potential roles in reproduction and preimplantation development†.

Ovarian cancer is the most lethal gynecologic cancer to date. High-grade serous ovarian carcinoma (HGSOC) accounts for most ovarian cancer cases, and it is most frequently diagnosed at advanced stages. Here, we developed a novel strategy to generate somatic ovarian cancer mouse models using a combination of  electroporation and CRISPR-Cas9-mediated genome editing. Mutation of tumor suppressor genes associated with HGSOC in two different combinations ( with and without  resulted in successfully generation of HGSOC, albeit with different latencies and pathophysiology. Implementing Cre lineage tracing in this system enabled visualization of peritoneal micrometastases in an immune-competent environment. In addition, these models displayed copy number alterations and phenotypes similar to human HGSOC. Because this strategy is flexible in selecting mutation combinations and targeting areas, it could prove highly useful for generating mouse models to advance the understanding and treatment of ovarian cancer. SIGNIFICANCE: This study unveils a new strategy to generate genetic mouse models of ovarian cancer with high flexibility in selecting mutation combinations and targeting areas.

Modeling High-Grade Serous Ovarian Carcinoma Using a Combination of  Fallopian Tube Electroporation and CRISPR-Cas9-Mediated Genome Editing.

Female fertility gets cilia(r) and cilia(r): ciliary defects in the oviduct compromises female fertility†.

Owing to technical advances in single-cell biology, the appreciation of cellular heterogeneity has increased, which has aided our understanding of organ function, homeostasis, and disease progression. The oviduct (also known as the fallopian tube) is the distalmost portion of the female reproductive tract. It is essential for reproduction and the proposed origin of high-grade serous ovarian carcinoma (HGSOC). In mammals, the oviduct is morphologically segmented along the ovary-uterus axis into four evolutionally conserved regions. It is unclear, however, if there is a diversification of epithelial cell characteristics between these regions. In this study, we identify transcriptionally distinct populations of secretory and multiciliated cells restricted to the distal and proximal regions of the oviduct. We demonstrate that distal and proximal populations are distinct lineages specified early in Müllerian duct development and are maintained separately. These results aid our understanding of epithelial development, homeostasis, and initiation of disease from the oviduct.

Oviduct epithelial cells constitute two developmentally distinct lineages that are spatially separated along the distal-proximal axis.

Epithelial cells lining the oviduct/fallopian tube are essential in reproduction and have been identified as the cell-of-origin in high-grade serous ovarian carcinoma (HGSOC). This protocol describes the generation of organoids from mouse oviduct epithelial cells, providing a powerful  tool to study epithelial homeostasis and malignant transformation. We also outline a protocol for whole-mount immunofluorescence and 3D confocal imaging. In addition, we describe approaches of viral transduction to investigate gene function in organoid development and epithelial cell behavior. For complete details on the use and execution of this profile, please refer to Ford et al. (2021).

Article	Year
Anatomical and cellular heterogeneity in the mouse oviduct-its potential roles in reproduction and preimplantation development†. Biology of reproduction\| PubMed ID: 33693543	2021
Modeling High-Grade Serous Ovarian Carcinoma Using a Combination of Fallopian Tube Electroporation and CRISPR-Cas9-Mediated Genome Editing. Cancer research\| PubMed ID: 34301761	2021
Female fertility gets cilia(r) and cilia(r): ciliary defects in the oviduct compromises female fertility†. Biology of reproduction\| PubMed ID: 34427303	2021
Oviduct epithelial cells constitute two developmentally distinct lineages that are spatially separated along the distal-proximal axis. Cell reports\| PubMed ID: 34496237	2021
Protocol to generate mouse oviduct epithelial organoids for viral transduction and whole-mount 3D imaging. STAR protocols\| PubMed ID: 35199031	2022

Keerthana Harwalkar

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