Fat Prospects for Healing Cutaneous Wounds: New Activities Under the Sheets

In this issue of Diabetes, Kato et al. (1) describe new results for dermal wound healing in full-thickness skin defects in Zucker diabetic fatty (ZDF) rats. The team fabricated and applied transplanted living cell sheets comprising allogeneic adipose-derived stem cells (ASCs) harvested from the inguinal fat pads of normal rats. They attempt to address the current challenges with abnormal wound healing in increasing numbers of patients with diabetes who present with difficult cutaneous wound healing problems (2,3). This deficiency is often clinically characterized by persistent inflammation, hypergranulation, excessive tissue bed exudates, and chronic inability to resolve, resulting in enhanced bacterial burdens, infections, ulcerations, and more serious complications. Delayed cutaneous healing in these patients is attributed to combinations of predisposing factors associated with chronic diabetes pathology, including oxidative imbalance, vascular abnormalities, neuropathy, infection, and local metabolic deficiencies intrinsic to diabetes. Pathological disruption of normal regenerative and degenerative extracellular matrix processing alters the dermal matrix composition, resulting in reduced amounts and quality of tissue matrix (2,3). Natural balances between proinflammatory cytokines (e.g., interleukin [IL]-1α and -1β and tumor necrosis factor-α) stimulating endogenous protease activities and prohealing cytokines (e.g., IL-4, IL-8, IL-10, IL-11, IL-13, granulocyte macrophage colony-stimulating factor, MCP-1, and macrophage inflammatory protein 2) apparently do not exist in diabetic tissues. Additionally, reduced levels of potent growth factors platelet-derived growth factor, basic fibroblast growth factor, epidermal growth factor, and transforming growth factor-β are found in chronic diabetic wounds (4).

Importantly, contributions of various progenitor cell populations in diabetic pathophysiological consequences—for restoring homeostatic tissue balance and for stimulating wound healing in diabetic wounds—are still not well understood. Cutaneous wound site multicellular competence and engagement is essential for normal cytokine/chemokine/growth factor dynamics that produce dermal tissue homeostasis. However, cellular deficiencies and disruption in diabetic wounds compromise this process and also elicit impaired angiogenesis and neovascularization associated with abnormal endothelial cell functions and vascular endothelial growth factor and stromal cell-derived factor 1 bioactivities, contributing to the diabetic wound healing problem (5,6). Recent evidence demonstrates that bone marrow–derived progenitor cells are correlated with new blood vessel formation in wounds (7,8). Cutaneous injury mobilizes these progenitor cells from marrow, and resulting circulating progenitor cells then transport to injury sites, transmigrate into wounded tissue, and initiate repair functions, among them new vessel formation (7,9). While numbers of marrow-derived progenitors in diabetic and wild-type mice are relatively similar, reduced mobilized circulating progenitor cells are present in diabetic mice both before and after wounding (5). Harvesting marrow cells and adoptively transferring them, either topically or subcutaneously, into diabetic wounds overcomes impaired progenitor cell mobilization, injury homing, and migration. This approach significantly improved diabetic wound healing (10,11). Nonetheless, while this adoptive progenitor cell strategy for diabetic wounds is effective in rodents, its translation to clinical practice will be difficult.

Kato et al. (1) overcame these translational issues to address long-standing wound healing challenges in full-thickness wounds in the ZDF rat model. They used a mature technology that this group has developed for fabricating living cell sheets from a large variety of tissue types (12). Their technique exploits a novel cell culture approach, amenable to many biopsied cell types, that yields intact confluent cell sheets without trypsin or dispase digests. The cell sheets then can be used with their intact endogenous extracellular matrix for direct application to tissues (13,14). Cell sheets are readily stacked into thick-sheet multilayers, patterned into complex shapes, and mixed with multiple cell types to yield tissue-like constructs (14,15). Fabricated sheets exhibit numerous advantages over digested single-cell suspensions often produced for cell therapies, including cutaneous healing. Recognized sheet benefits include sutureless tissue adherence in transplantation, rapid integration and growth, instantaneous structural integrity, mechanical robustness and epithelial sealing, and spontaneous neovascularization. Additionally, these sheets have demonstrated abilities to spontaneously differentiate, produce endogenous growth factors and signaling cues, and become tissue in the host (16). These capabilities represent the portfolio of dynamic physiological enhancements required to overcome refractory wound healing processes in the population with diabetes, and these living cell sheets are distinct from the current array of wound coverings generally clinically applied in nonhealing diabetic wounds. Kato et al. (1) harvested ASCs, a derivative mesenchymal stromal progenitor cell type, from wild-type laboratory rats and fabricated the isolated ASCs into cell sheets and then harvested and applied them as sheets to cutaneous wounds in ZDF rats (Fig. 1).

ASCs have known regenerative pluripotencies. The authors show that, unlike single-cell progenitor cutaneous local injections, cell sheets retain high densities of ASCs at the wound bed. While recent studies have used similar ASC sheets in nondiabetic rodent cutaneous wound healing models (17,18), Kato et al. used a diabetic rodent model more relevant to the human diabetic foot ulcer condition under clinical debridement. Their larger full-thickness skin defect deliberately exposed underlying bone, removing the local periosteum on exposed bone along with its progenitor cells and healing potential. Additionally, recognized disparities between rodent contractile epidermal wound healing and human reepithelializing healing were addressed by using the ZDF rat known to exhibit compromised contractile healing and skin-sutured contractile healing constraints. The authors show accelerated wound healing from allogeneic transplanted ASC cell sheets, increased wound site vascular density, rapid production of important arrays of growth factors, retention of most progenitor cells in the sheet after weeks of transplantation, and no obvious clinical signs of immune rejection from allogeneic sourcing.

Importantly, beyond preclinical proof of concept in many diverse tissue regeneration models, analogous autologous human-derived cell sheets using this same approach are in human clinical trials currently—in corneal resurfacing, cartilage replacement, esophageal stricture prevention, and periodontal ligament reconstruction. Translation of this technology to other preclinical models is currently very active, aptly demonstrating the attractive and versatile capabilities of autologous cell harvest, sheet fabrication, and transplantation to regenerate functional tissue. Given the demonstrated ability of allogeneic progenitor cells to modulate host immunity and perhaps inflammation in transplant scenarios (19), the strategy demonstrated by Kato et al. (1) might readily also be extended to human use the same way the results were achieved in rodents—that is, by transplanting competent adipose-derived progenitors from readily harvested adipose sources (e.g., liposuction) in donors without diabetes to patients with diabetes requiring assistance with healing troublesome cutaneous wounds.