Alternative delivery of keratinocytes for epidermal replacement - Biomaterials for Treating Skin Loss - page 117

with immediate placement onto the wound, 31 once again, exploring the concept of
the wound as the 'tissue culture' environment. The development and clinical use
of three-dimensional tissue engineered skin constructs are discussed in Chapter 10.

8.2 Methods of keratinocyte delivery

A delivery system needs to ensure the even distribution of viable cells capable of
adhesion onto the surface of the wound. 32 The use of pre-confluent cells requires
that the surface is protected whilst the cells proliferate and migrate across the
wound and differentiate to a confluent, mature epidermal layer.
The keratinocytes are harvested from the dermal epidermal junction via a
process of enzymatic and physical dissociation. 2 Initially the cells are a mixed
population including melanocytes, papillary dermal fibroblasts and Langerhan's
giant cells with the keratinocytes. 33 The clinical use of the cells at this stage has
been practiced by our group since 1995 to augment wound healing in acute
injuries, donor sites 34 and to improve the surface in scar revision. 1,21 The cells are
delivered as a suspension in an aerosol using a nozzle fitted to a standard syringe.
Cells have been delivered to the wound bed immediately using a carrier dressing
to facilitate placement. 35 The cells have also been seeded onto carrier dressings for
later transfer to the wound bed as the cells develop into colonies. 36
In the process described by Green et al. 2 the cells are seeded onto a layer of
irradiated mouse 3T3 fibroblasts. As the primary colonies expand, the cells are
harvested and at this stage the cells may be used as a suspension delivered to the
wound directly 23 or using a carrier, or seeded onto a carrier for later use as the
colonies become established. 37,38
The delivery of cells to the wound can be at any stage in the expansion process
by direct application or by the use of a carrier. The seeding density is related to the
stage of the cells at the time of clinical use. 39 When used immediately with no prior
culture, the cell population will contain terminally differentiated keratinocytes and
the cell density will be expected to be greater than cells which are cultured and
colonies established prior to clinical application.
The need to understand the clinical indication is common to all techniques with
a clear understanding of what epidermal cells can achieve in the wound healing
spectrum. 32 There is a balance between the expansion ratio and the speed of
epithelial repair. 39 Understanding the influence of the wound bed on the speed and
area of epithelial repair for a given cell load will indicate the appropriate clinical
use. 1

8.3 Direct application

Direct application of the cells to the wound can be achieved by dripping, spraying
or air brushing techniques. The maintenance of viability of the cells in the system
is essential. 40