Biomedical Engineering Reference
In-Depth Information
tive” to the “bottom-up approach” of the in vitro reconstitution of ECM
biopolymers which consists of the isolation of assembled matrix structures
from living tissues without fragmentation, e.g. the collection of decellular-
ized tissue as medically applied in the form of xenogenic (porcine) transplant
heart valves.
Obviously, any “bottom up” reconstitution depends on the availability of
isolated, dissociated ECM biopolymers. Enzymatically driven fragmentation
of ECM forms like connective tissue often results in structurally altered bio-
polymers which may exhibit limited capability of reconstitution.
Collagen I-based fibrillar assemblies do certainly represent the best-
studied example of in vitro reconstituted ECM structures [55]. Collagen is
used as a generic term for proteins forming a characteristic triple helix of
three polypeptide chains. So far, 27 genetically distinct collagen types have
been described. Type I collagen forms more than 90%oftheorganicmassof
bone and is the major collagen of tendons, skin, ligaments, cornea, and many
interstitial connective tissues. The collagen type I triple helix is formed as
a heterotrimer by two identical
2(I)-chain. The fibrils
are indeterminate in length, insoluble, and form elaborate three-dimensional
arrays that extend over numerous cell lengths. Studies of the molecular basis
of collagen fibrillogenesis have provided insight into the trafficking of procol-
lagen (the precursor of collagen) through the cellular secretory pathway, the
conversion of procollagen to collagen by the procollagen metalloproteinases,
and the directional deposition of fibrils involving the plasma membrane and
late secretory pathway [56]. Fibrils arranged in elaborate three-dimensional
arrays, such as parallel bundles, orthogonal lattices and concentric weaves
provide the base for dedicated matrix structures in tendons and ligaments,
in the cornea, and in bone. The fibrils are synthesized and secreted by fibro-
blasts and might self assemble thereafter but accumulating evidence suggests
that fibril assembly can begin in the secretory pathway and at the plasma
membrane.
The reconstitution of collagen I is attractive both for the relevance due
to the abundance and dominating role in various tissues and for the rela-
tively easy access and reconstitution. Enzymatically fragmented tropocolla-
gen, collected from various kinds of connective tissue, can be dissolved in
acidic solutions of sufficiently high ionic strength and precipitated into the
fibril form upon neutralizing the solution. Variation of temperature and elec-
trolyte concentration was demonstrated to influence the fibrillogenesis with
respect to the dynamics of the process, however, the resulting fibrils show
a very similar structure almost independent of the “history” of their for-
mation and so do the rheological characteristics of the resulting gels. The
spontaneous self-assembly of monomeric collagen was attributed to the en-
tropy gain upon binding of collagen molecules implying that hydrophobic
interactions between collagen monomers are the major driving force for fib-
ril formation [1, 57]. In addition to hydrophobic interactions the contribution
α
1(I)-chains and one
α
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