Biomedical Engineering Reference
In-Depth Information
7.3
In Vitro Versus In Vivo Conditions for Synthesis of Tissues
Why was an in vivo environment required for synthesis of rete ridges or a dermis?
Which indispensable reactants, apparently not available in vitro, were supplied by
the dermis-free skin defect? These questions can be answered, at least in part, by
considering the few irreducible reaction diagrams that were selected in the preced-
ing section. The requirement for a distinction between in vitro and in vivo protocols
(Fig.
7.1
) has important practical and theoretical implications (Yannas 2000).
In an effort to identify the components of a dermis-free defect that were required
in the syntheses we will limit the choices by elimination. First, we look for evidence
that a given reactant, exogenously supplied, participated in an irreducible reaction
diagram in vivo. Such participation in the irreducible diagram shows that addition
of the reactant was necessary to achieve the desired synthesis; if so, it follows that
the defect was not a supplier (at least, not an adequate supplier) of this required reac-
tant. For example, keratinocytes are a reactant that had to be exogenously supplied
in order to induce simultaneous synthesis of the epidermis and the dermis (Dg. S30)
but not of the dermis alone (Dg. S19). Another reactant that had to be exogenously
supplied to induce in vivo either synthesis of dermis (Dg. S19) or synthesis of skin
(Dg. S30) is the DRT. The data suggest, therefore, that keratinocytes and the DRT
were reactants that were required to be supplied by the investigator (exogenously);
if these reactants had been supplied by the defect (endogenously), there would have
been no requirement for their addition as reactants to the irreducible diagram.
Exogenous supply of fibroblasts was not required to synthesize any of the com-
ponents of skin, as seen by inspection of all four irreducible reaction diagrams, S2,
S13, S19, and S30. Since it is well-known that fibroblasts are critically involved in
synthesis of ECM (Clark et al. 1996b; Eckes et al. 1996), it appears that synthesis
of stroma resulted from fibroblasts that were spontaneously supplied entirely by
the defect (endogenous supply). A similar argument can be made about the absence
of microvascular endothelial cells from the irreducible diagrams; these cells are
responsible for angiogenesis (Madri et al. 1996). Since the irreducible diagrams de-
scribe the synthesis of vascularized stroma, it follows that the defect spontaneously
supplied microvascular endothelial cells; an exogenous supply was unnecessary.
These observations are consistent with the observation (Butler et al. 1998; Compton
et al. 1998) that DRT becomes spontaneously vascularized following grafting. Of
the tissue components in skin, the epidermis and the BM could be synthesized by
keratinocytes in vitro, in the absence of fibroblasts (Dgs. S2, S13). Synthesis of a
dermis, however, required the presence of the defect (Dg. S19). Since the synthesis
of tissues could not have proceeded in the absence of cells, it follows that all cells
required for synthesis of a dermis (Dg. S19) must have originated in the defect
(endogenous supply of cells). We arrive, therefore, at the hypothesis that synthesis
of the dermis, as in Dg. S19, required in vivo conditions primarily because of a re-
quirement for an endogenous supply of fibroblasts and endothelial cells.
If this simple analysis was sufficient to explain the data, synthesis of a vascu-
larized dermis should be possible in vitro by seeding the hypothetically required
fibroblasts and endothelial cells into an appropriate nondiffusible regulator, such
