Biology Reference
In-Depth Information
20
Using Organotypic Tissue Slices as Substrata
for the Culture of Dissociated Cells
Daniel E. Emerling and Arthur D. Lander
1. Introduction
Dissociated cell culture has proved extremely valuable for studying the func-
tional relationship between specific cell surface and extracellular matrix (ECM)
molecules and a variety of cellular behaviors such as proliferation, differentia-
tion, migration, and neuronal process outgrowth. When cells are cultured on
substrata made from purified cell surface or ECM components, the direct
actions of these molecules on cells can be examined easily. However, for some
in vitro studies, it is desirable to examine the functions of such molecules in an
environment that more closely approximates that which exists in vivo. Such
studies might include, for example, investigations in which the endogenous
distribution and heterogeneity of such molecules is important, screens for
uncharacterized factors that effect a particular cell behavior, or studies exam-
ining the combinatorial effects of the many cell surface and ECM components
that occur in intact tissue.
We devised an assay in which dissociated cells are plated onto living slices
of tissue, much as they can be plated onto tissue culture plastic. This procedure
allowed us to examine cell behaviors within the organotypic environment of a
tissue substratum. Primary neurons were plated onto slices of living mouse
forebrain, cocultured at 37
C, fixed, and then examined. The degree of attach-
ment of the cells and the orientation of process outgrowth from them was found
to be highly dependent upon the anatomical region of the forebrain slice onto
which the cells settled when plated (
Fig. 1
and
ref.
1
). When the same cells
were plated onto cryostat sections of the same tissue, neither the degree of
overall attachment and outgrowth nor the region-dependent effects were as
dramatic
(1)
.
°
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