841. Collagen- and Elastin-Rich Biomaterial Isolated from the Skin



841. Collagen- and Elastin-Rich Biomaterial Isolated from the Skin
Collagen- and Elastin-Rich Biomaterial Isolated from the Skin of Aplysia californica
Victoria A. Webster, Phillip McClellan, Ozan Akkus, Umut Gurkan, Hillel J. Chiel, Roger D. Quinn
Case Western Reserve University
Introduction: We have previously reported on the
isolation of collagen-rich material from the skin of
Aplysia californica for use in biohybrid robotics and
scaffold fabrication. However, mechanical analysis of
electrocompacted collagen sheets found that those
fabricated from bovine collagen were an order of
magnitude stiffer than those from Aplysia collagen[1].
Aplysia lives at temperatures lower than that of the
mammalian physiology and Aplysia collagen gels at room
temperature unlike the mammalian collagens. Therefore,
there may be interesting biomaterials based application,
Figure 2. Aplysia collagen sheet with Verhoeff
such as on demand thermal release of drugs. The aim of
(Left) and Van Gieson (Right) stain. Dark grey
this study was to assess and characterize the collagen
indicates Elastin while red indicates Collagen.
isolated from Aplysia.
birefringence under polarized light microscopy (Figure 1).
Additionally, staining slices using Verhoeff-Van Gieson
Methods: Animals were obtained from Marinus
stain resulted in staining suggesting the presence of
Scientific and South Coast Biomarine. They were
elastin, in addition to collagen (Figure 2).
subsequently maintained in 16 C artificial seawater
SDS-PAGE gel electrophoresis indicated while that
(Instant Ocean, Quarium Systems) in aerated aquariums.
distribution is similar to that of rat or bovine
Skin was harvested and stored at -20 oC for processing.
the molecular weights of Aplysia collagen
The collagen-rich material was isolated using a previously
be greater than the rest. Furthermore, well
reported procedure [1]. The molecular weights of isolated
species was subsequently characterized using SDS-PAGE
beta, and alpha
analysis (Bio-Rad). Electrophoresis of the gels was
bands developed in
performed at 150V for 2.5 hrs, after which gels were
the Aplysia
removed from their holders and stained with Coomassie
collagen, indicating
Blue for 30 min. Gels were subsequently rinsed in
that the triple helical
distilled water and destained. The molecular weights of
structure known in
the bands were compared to those of commercially
purchased bovine collagen (Collagen Solutions) and
collagen applies to
previously isolated rat tail
Figure 3. SDS-PAGE Analysis. 1:
the material from
tendon collagen.
Ladder, 2: Rat Tail Tendon Collagen,
Aplysia as well.
The isolated material
3: Aplysia collagen-rich material, 4:
(Figure 3).
was compacted into
Bovine Collagen
Figure 1. Picrosirius red stained
histological section of
compacted Aplysia collagenrich material imaged using
polarized light microscopy.
randomly aligned
scaffolds using a
previously reported
technique [2] modified to
use plate electrodes.
Scaffolds were fixed and
sections of 5 micron
thickness were prepared
and stained histologically
using picrosirius red or
Verhoeff-Van Gieson
staining protocols. They
were then imaged under
bright field and polarized
lighting conditions.
Results: Histological analysis of scaffolds fabricated
from the isolated material confirmed the presence of
collagen-rich and elastin-like molecules. Picrosirius red
staining revealed both orange (indicative of Type I
collagen) and blue (indicative of Type III collagen)
Characterization of collagen-rich material isolated from
the skin of Aplysia californica indicates that while the
material does contain collagen, it is also contaminated
with non-collagenous proteins such as elastin. For use in
biohybrid robotics, the ability to fabricate such compliant
scaffolds may be advantageous, as it limits the cellular
force needed to deflect the scaffold.
Webster, V. Biomimetic and Biohybrid Systems
2016, pp. 365–374.
Cheng, X. Biomat. 28(22), pp. 3278–88, 2008.
Acknowledgements: This material is based upon work
supported by the NSFGRFP under Grant No. DGE0951783 and a GAANN Fellowship (Grant No.
P200A150316). This study was also funded in part by
grants from the NSF (Grant No. DMR-1306665), and the
NIH (Grant No. R01 AR063701).

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