Differences between Solution and Membrane Forms of Chitosan on the In Vitro Activity of Fibroblasts

Year 2015, Volume: 32 Issue: 1, 69 - 78, 01.01.2015

Bahar Uslu Burcu Biltekin Seçnur Denir Ayhan Bilir Suna Özbaş Turan Jülide Akbuğa Serap Arbak

Abstract

Background: Chitosan, a linear polysaccharide, has
been recently used in biomedical applications. In vitro
studies have demonstrated its effect on cellular growth
and its stimulatory action on cellular layer formation.
Aims: The present study aims to compare the proliferative
effects of chitosan in two forms, membranous
and solution forms, on Swiss 3T3 mouse embryonic
fibroblasts.
Study Design: In vitro study.
Methods: Three experimental groups were formed:
cells were cultured in a normal medium without
chitosan (Control Group); cells were cultured either
in a medium containing 2.0% chitosan in membranous
form (Membrane Group) or chitosan solution
at a concentration of 2.0% (Solution Group).
Two different methods were used in the experiments:
cells cultured on the medium containing
chitosan in solution or membranous forms
(method 1); and chitosan solution or membranous
forms were added into the medium containing previously
cultured cells (method 2).
Results: Scanning electron microscopic investigations
of the experimental groups revealed cells with welldefined
cellular projections, intact cellular membranes
and tight intercellular junctions. They were especially
prominent in the membrane group of method 1 and in
the membrane and solution groups of method 2. Mouse
monoclonal anti-collagen 1 primary antibody was used
to indicate collagen synthesis. Prominent collagen synthesis
was detected in the membrane groups on the 10th
day of culture for both methods. Bromodeoxyuridine
(BrdU) and MTT assays were performed in order to
assess cellular proliferation and viability, respectively.
BrdU labelling tests indicated a higher proliferation
index in the membrane group of method 1 on the 5th
and 10th days. For the second method, the membranous
form on the 10th day and solution form on the 5th
day were the most effective groups in terms of cellular
proliferation. MTT results reflected a high cellular viability
in method 1 on the 5th day of treatment with the
membranous form, whereas cellular viability was highest
in the solution form of method 2 on the 5th day.
Conclusion: The membranous form of chitosan induced
a significant proliferative effect and increased
the ratio of cell-to-cell junctions of Swiss 3T3 mouse
embryonic fibroblasts. Conveniently, the solution form
also resulted in enhanced cell proliferation and viability
compared to the control group. As the solution form
is easy to prepare and apply to cells compared to the
membrane form, the application of Chitosan directly to
media appears to be a convenient alternative for tissue
engineering approaches

Keywords

Cell culture, cell viability, chitosa, collagen immunohistochemistry, scanning electron microscopy, Swiss 3T3

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