Studies of mitogenic and morphogenic signals during mouse limb development
Description
In an attempt to understand the patterning regulation during limb development, three approaches have been taken in current studies. First, the relationship between cellular position and growth has been studied in cultures of dissociated fragments of mouse limb bud cells. We have found that cells derived from posterior region of the limb proliferate at a much reduced rate than cells from anterior and central regions of the limb under the same culture conditions. Further, we have found that it is the fibroblastic cells in the limb that attribute to this position-specific growth. Since this position-specific growth in vitro unique to the posterior region of the limb bud complements the asymmetrical distribution of positional activity associated with the ZPA, it may represent an in vitro means by which positional identity can be evaluated. Furthermore, when anterior and posterior cells are co-cultured, a position-dependent stimulation of growth was observed, which is indicative of a mitogenic signal passing between positionally disparate cells Second, to test the idea that fibroblastic cells derived from ZPA region play an important role in pattern formation, both fibroblastic and myogenic cells were enriched from dissociated mouse ZPA and grafted into chick wing bud to test for polarizing activity. We found that the strength of polarizing activity can be increased by enriching for fibroblast population from mouse ZPA cells and decreased by enriching for myoblast population. This indicates that it is the fibroblastic not myogenic cells who have the positional signaling activity Third, to establish permanent cell lines that can signal, we have immortalized mouse limb posterior cells by transfecting with polyoma large T antigen. 5 cell lines have been established so far. Two lines have been tested for positional activity based on the induction of supernumerary limb following grafts into the anterior site of chick wing. Only one cell line (MPLB-1) displayed a weak level of polarizing activity. We also found that MPLB-1 line represents a precartilage mesenchymal cell line and have the potentials to become either chondrogenic cells or non-chondrogenic cells