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n my last column I described
the Ocular Research Center’s
efforts to examine glaucoma with a
model system that used a computer
machined plastic dish and the
dissected front of a human eye.
Growing and examining the key cells
in disease processes is another type of
model that allows scientists to gain
insight into the cellular mechanisms
behind disease processes as well as
the possibilities of using normal cells
to help alleviate the disease. In this
column, I would like to describe our
efforts to grow the key cells involved
in glaucoma, age related macular
degeneration and corneal endothelial
The ability of cells to grow in vitro
simply means that cells can be grown
in plastic dishes with a nutrient media
in incubators at 37°C. The cells have to
be removed from the tissue with great
care and under sterile conditions. If the
scientist does not dissect out just the cells
you want, the result is known as amixed
culture (several cell types growing in the
dish) and any experiments examining
characteristics of the cells would be
from all of the cell types in the dish. In
addition, if the tissue is not dissected
under sterile conditions the cultures
will be contaminated with bacteria. As
you can imagine, if the conditions are
favorable for the cells to grow, those
same conditions of a nutrient media, a
moist environment and warmth are also
ideal for bacteria to grow. Fortunately,
the ORC had a dissection hood donated
by the Lions for the initial dissection
which helps keep the procedure sterile
and a high quality dissectingmicroscope
to aid in dissecting just the cells we want
to grow.
Each cell type that the ORC
is growing has their own unique
characteristics that require specialized
procedures. Trabecularmeshwork tissue
is carefully dissected from the eye, cut
into pieces and placed in culture dishes
with a small amount of media. After
a few days the pieces are removed by
rinsing with media and the cells that
havemigrated from the tissue piece onto
the dish are allowed to grow. Once they
fill the small initial dish they have to
be transferred to a larger dish to allow
the cells to multiple. Unlike other cell
types, TMcells cannot be removed from
the dishes by using an enzyme to break
their attachments to the dishes. The
passage of these cells must begin by
placing beads on top of the cells and
letting the cells move onto the beads.
After several days the beads are rinsed
off of the original dish, placed into new
dishes where the cells move off of the
beads onto the new dish to multiply
and grow.
Retinal pigment epithelial (RPE)
cells are the most posterior layer of the
retina. These cells are pigmented and
interact with the photoreceptor cells
which are responsible for translating
light energy into an electrical signal to be
transmitted to the brain. It is believed that
the RPE cells may be the key to solving
the mystery of the disease entity known
as age related macular degeneration
(AMD). The ORC is doing something
rather unique in that we are growing
“old” RPE (from donors older than 50
yoa) rather than the RPE from donors
younger than 10 yoa which grow more
robustly. Many labs have shown that the
younger cells are easier to grow but our
older RPE cells are agematched controls
for the AMD cells. The challenge with
our RPE cells is to get them to grow as
well as not having mixed cultures. The
methods used to grow RPE cells often
also allow fibroblasts (connective tissue
cell type) to grow as well. In order to
insure only RPE cells are growing we
In the Realm of
6 | Lions Eye Institute Newsletter
Mitchell D. McCartney, Ph.D.
1,2,3,4,5 7,8,9,10,11,12
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