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Parietal endodermal (PE) migration along rat trophectodermal (TE) cells coincides with the deposition of Reichert's membrane between these two cell layers. In this study, we compared the influences of fibronectin and laminin, two components of Reichert's membrane, on the migration and replication of PE-like cells from cultured rat inner cell masses (ICMs). We also explored the role of substrate nondeformability by comparing cell translocation on gels versus coatings of Matrigel (a tumor cell-derived basement membrane preparation) or of collagen. ICMs, isolated by immunosurgery from Day 5 blastocysts, were cultured on coatings of collagen IV, laminin, fibronectin, collagen I, or Matrigel, or on gels of the latter two substrates. Minimal laminin or fibronectin coating concentrations of 2.5 micrograms/ml were required for ICM attachment and cell migration. Migration was similar during the first 48 h of culture on fibronectin and on laminin; however, by 72 h, the extent of cell translocation on fibronectin was greater (1.5- to 2-fold) than that measured on laminin. Fibronectin-cultured ICM-derived cell clusters also contained 1.5- to 2-fold more cells than those on laminin. Migration did not occur on undiluted gels of Matrigel but was supported by diluted (1:10 and 1:20) Matrigel coatings. Similarly, cell migration on coatings of collagen IV reached almost 3-fold that measured on collagen I gels. Most of the cells migrating on fibronectin or collagen (I or IV) were flattened and elongated. In contrast, a high proportion of the cells migrating on laminin or Matrigel coatings were tall and rounded, with thin cytoplasmic extensions. Fibronectin- and collagen IV-cultured cells stained strongly for both collagen IV and laminin, but contained no fibronectin. In contrast, laminin-cultured cells contained fibronectin but were less immunoreactive for laminin and collagen IV. These findings indicate that substrate composition and resilience influence the in vitro migration and morphology of ICM-derived PE-like cells. A role for the TE cells in anchoring Reichert's membrane during development of the PE cell layer within the blastocyst is postulated. Furthermore, the sensitivity of cell morphology and differentiation to individual basement membrane components provides a potential key mechanism whereby an emerging basement membrane can regulate cell migration and differentiation, two fundamental processes that occur throughout embryonic development.

biology of reproduction

Extracellular Matrix Composition and Resilience: Two Parameters that Influence the in Vitro Migration and Morphology of Rat Inner Cell Mass-Derived Cells1