Girish
K Srivastava 1, 2, David Rodriguez-Crespo 1, J. Carlos
Pastor 1, 2
1. Instituto
Universitario de Oftalmobiologia Aplicada (IOBA), Universidad de Valladolid,
Valladolid, Spain.
2. Centro en
Red de Medicina Regenerativa y Terapia Celular de Castilla y León, Spain.
Abstract
Phagocytosis
is involved in several functions including nutrient uptake, immune response,
inflammation, tissue homeostasis, cellular apoptotic bodies and debris
elimination. Its dysfunction has impacts on normal functioning of an organism.
It is involved in several diseases including the AMD pathogenesis, and
treatment approaches. Mostly an initial proof of concept is tested in a
laboratory conditions. Cell and tissue cultures need proper surfaces to grow
and maintain them in laboratory conditions. The study aims to evaluate effect
of two surfaces; tissue culture plate polystyrene and glass coverslip, on
phagocytosis performed by RPE cells growing on each surface. Fresh RPE cells
and ARPE-19 cell line were grown in corresponding cell culture conditions.
Phagocytosis in fresh RPE cell cultures was detected and recorded. ARPE-19
cells were incubated for 4 and 24 hours with 0.2 and 1 μl red fluorescent latex
beads, and then images were taken to measure the average numbers of cells,
cells performing no phagocytosis and phagocytosis and engulfed red fluorescent
latex beads. Results showed that RPE cell membrane was forming blebs releasing
black dots like particles as cell debris in cell culture medium. Subsequently
cytoplasmic membrane extensions were formed which had engulfed black dots like
particles. ARPE-19 cells are a good cellular model for performing initial in
vitro test in comparison to fresh RPE cells. There was no significant
difference between numbers of ARPE-19 cells growing on polystyrene and glass
surface performing latex bead phagocytosis. Both had engulfed red fluorescent
latex beads; however, numbers of engulfed beads was significantly higher for
glass surface than polystyrene surface. Results support the hypothesis that
surface of cell culture plates affects the cellular phagocytosis mechanism
which play a critical role in the functions in many cases such as in vivo
retina. These findings are very crucial when it comes to transfer a treatment
approach based on phagocytosis to the clinics and that the materials and the
conditions of the experiments can have very important
consequences.
Keywords: Retinal pigment epithelial cells, phagocytosis, cell growth supporting
surface, latex beads
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Ms. Ref. No.: A0201006 RA