Donald P Hill

Sequential Cytokine Therapy for Pressure Ulcers: Clinical and Mechanistic Response

OBJECTIVE To compare the healing response of sequential topically applied cytokines to that of each cytokine alone and to a placebo in pressure ulcers, and to evaluate the molecular and cellular responses. SUMMARY BACKGROUND DATA Because of a deficiency of cytokine growth factors in chronic wounds and the reversal of impaired healing in animal models, pressure ulcer trials have been performed with several exogenously applied growth factors. Because single-factor therapy has not been uniformly successful, combination or sequential cytokine therapy has been proposed. Laboratory data have suggested that sequential treatment with granulocyte-macrophage/colony-stimulating factor (GM-CSF)/basic fibroblast growth factor (bFGF) might augment the previously reported effect of bFGF alone. METHODS A masked, randomized pressure ulcer trial was performed comparing sequential GM-CSF/bFGF therapy with that of each cytokine alone and with placebo during a 35-day period. The primary measure was wound volume decrease over time. Cytokine wound levels and mRNA levels were serially determined. Fibroblast-populated collagen lattices (FPCLs) were constructed from serial fibroblast biopsies. Cellular ultrastructure was evaluated by electron microscopy. Changes in ease of surgical closure and its relative cost were determined. RESULTS Ulcers treated with cytokines had greater closure than those in placebo-treated patients. Patients treated with bFGF alone did the best, followed by the GM-CSF/bFGF group. Patients treated with GM-CSF or bFGF had higher levels of their respective cytokine after treatment. Patients with the greatest amount of healing showed higher levels of platelet-derived growth factor (PDGF) on day 10 and transforming growth factor beta (TGFbeta1) on day 36. Message for the bFGF gene was upregulated after treatment with exogenous bFGF, suggesting autoinduction of the cytokine. FPCLs did not mimic the wound responses. Ultrastructure of wound biopsies showed response to bFGF. Treatment with any of the cytokines improved the wound by allowing easier wound closure. This was most marked for the bFGF-alone treatment, with a cost savings of

A prospective randomized evaluator-blinded trial of two potential wound healing agents for the treatment of venous stasis ulcers

9,000 to

Long-term outcome study of growth factor-treated pressure ulcers

9,200. CONCLUSIONS Treatment with bFGF resulted in significantly greater healing than the other treatments in this trial. The clinical response appeared to be related to upregulation of the bFGF message and to increased levels of PDGF-AB, bFGF, and TGFbeta1 in the wounds and changes in ultrastructure. The resultant improvements could be correlated with cost savings.

Wound-healing trajectories as outcome measures of venous stasis ulcer treatment.

Chronic wounds such as venous stasis ulcers have become a socioeconomic problem. Even with successful initial management, the recurrence rate approaches 70%. With the advent of new wound healing agents, nonoperative attempts to heal these wounds appear indicated. This study reports a prospective randomized evaluator-blinded trial comparing two potential wound healing agents to an inert vehicle placebo. Eighty-six evaluable patients completed the trial. Silver sulfadiazine 1% in a cream proved to statistically reduce the ulcer size compared with a biologically active tripeptide copper complex 0.4% cream formulation or the placebo. There was no difference between the latter two treatments. Silver sulfadiazine has been shown to allow keratinocyte replication and to have antiinflammatory properties. In this trial its antibacterial action was not used since all ulcers had comparable bacterial levels (less than or equal to 10(5)/gm of tissue) before treatment. These results suggest that the silver sulfadiazine cream used in this study may facilitate healing in wounds healing largely by the process of epithelialization.

In vivo characterization of keratinocyte growth factor-2 as a potential wound healing agent.

BACKGROUND Exogenous application of growth factors have been reported in an attempt to accelerate healing of chronic wounds. Most of the trials were of brief duration with short to no follow-up periods. Long-term outcome studies are sparse for pressure ulcer therapies with success rates around 30% for both operative and nonoperative treatments. METHODS Follow-up evaluations were performed serially up to 12 months for patients completing a 35 day blinded, placebo-controlled cytokine clinical trial of pressure ulcers. RESULTS Fifty-four of 61 patients completed the follow-up period with 68.5% of the patients (37 of 54) being healed after 1 year. Of patients healing > or =85% during the active treatment phase, 84.6% were healed after 1 year compared with 61% of those that healed <85% during treatment (P <0.05). CONCLUSION Long-term outcome was better in this growth factor trial than with surgical or standard nonoperative treatment of pressure ulcers. Since only patients receiving exogenously applied cytokines achieved >85% closure during the treatment phase of the trial, the excellent long-term outcome appears attributable to the cytokine therapy.

In vitro fibroblast populated collagen lattices are not good models of in vivo clinical wound healing

Outcome measures of venous ulcer healing are not uniformly accepted. Stringent criteria of 100% closure fail to provide information of healing over the entire span of repair. Wound‐healing trajectories (plot of percentage of wound closure versus time of wound treatment) were constructed for 232 patients treated in eight clinical trials at two independent wound care/research centres. Trajectories were constructed for ulcers that totally healed (100% closure) and those that did not (<100% closure) over a 20‐week period. Kaplan–Meier survival plots determined the percentage of patients achieving total healing versus time of treatment. The wound‐healing trajectories were almost identical for patients achieving complete ulcer healing, as were the trajectories for patients with less than 100% closure. The trajectories for the ulcers healing completely were significantly different from those with <100% closure. Only 60% of all patients achieved 100% closure by 20 weeks. Using linear regression, it was predicted that it would take 31 weeks for all patients to achieve total healing. Total healing is an inadequate outcome measure for healing of venous stasis ulcers. Clinical trials using this measure would require excessive time periods. As wound‐healing trajectories for patients treated at two centres mimic one another, shifting the wound‐healing trajectory from one of impaired healing to one of a more ideal healing course may be considered a better outcome measure for determining healing of venous stasis ulcers.

Minoxidil and Wound Contraction

Human keratinocyte growth factor‐2 exerts a proliferative effect on epithelial cells and mediates keratinocyte migration. It has also been shown to increase both deposition of granulation tissue and collagen and maturation of collagen. Because these properties should affect the healing trajectory of wounds, this study set out to investigate the effects of keratinocyte growth factor‐2 on the healing of three different types of wounds. Human meshed skin grafts explanted to athymic “nude” rats, surgical incisions in Sprague‐Dawley rats, and acute excisional rat wounds inoculated with Escherichia coli were used. Two concentrations of recombinant human keratinocyte growth factor‐2 were compared to a vehicle control and keratinocyte growth factor‐1. Keratinocyte growth factor‐2 significantly accelerated the rate of epithelialization in the meshed skin graft model and effected a modestly more rapid gain in breaking strength of surgical incisions than keratinocyte growth factor‐1 or the vehicle control treatment. Neither keratinocyte growth factors accelerated wound closure by contraction of the excisional wounds. Based on these data, keratinocyte growth factor‐2 may be useful in accelerating healing in wounds healing mainly by the process of epithelialization such as venous stasis ulcers, partial thickness burn wounds, and skin graft donor sites. It might also accelerate the gain in incisional wound strength in acute surgical or traumatic wounds.

Microfluidic and transducer technologies for lab on a chip applications

In chronic wounds, the healing process is prolonged and incomplete, proceeding in an uncoordinated manner, and resulting in poor anatomical and functional outcome. There have been numerous attempts to discover models that mimic human wound healing processes. The fibroblast populated collagen lattice is one such model that has been proposed. This study evaluated whether the fibroblast populated collagen lattice can be a model of chronic wound healing using the pressure ulcer as a paradigm. Fibroblast cultures of wound biopsies and wound volume measurements were obtained serially during a four arm blinded, placebo‐controlled sequential cytokine clinical trial of pressure ulcers. Fibroblasts obtained from study patients were added to collagen lattices and contraction was determined daily for 10 days. Collagen gel‐area measurements were converted to reflect percentage of gel contraction. These data of both edge and base wound biopsies on days 0, 10, and 36 were categorized into treatment groups and one‐way analysis of variance showed no significant differences in contraction among these groups. When considering all fibroblast populated collagen lattices, there was significantly greater contraction at days 10 and 36 for cells from both edge and base biopsies compared to day 0 (p < 0.05). The Spearman Rank Correlation test comparing all patients with fibroblast populated collagen lattice results from fibroblasts obtained at the edge or base of the wound at days 0, 10, and 36 and clinical pressure ulcer healing on day 36 showed no correlation. This lack of correlation not only persisted for each of the four treatment arms but also for responder status based on decrease in wound volume over the 35 day trial period. In conclusion, chronic wound healing is a complex process that is not modeled by in vitro fibroblast populated collagen lattices.

Advances towards reliable identification and concentration determination of rare cells in peripheral blood

Minoxidil has been proposed as a potential topical inhibitor of wound contraction and proliferative scarring. Suggestions for this application are derived from in vitro investigations demonstrating inhibition of various fibroblastic functions. The purpose of this study was to attempt to establish in vivo support of these effects using an established animal model of wound contraction. Standardized cutaneous wounds were created on the dorsum of Sprague-Dawley rats, which were divided equally into six treatment groups. Wounds were treated daily after tracing their unhealed areas. On complete closure of the wounds, analyses of the contraction rates and tensile strength were performed for comparison among groups. Minoxidil did not demonstrate significant inhibition of wound contraction rates relative to either an inert vehicle, an active vehicle, or no treatment. Contrarily, as previously demonstrated in this animal model, silver sulfadiazine did demonstrate significant inhibition of wound contraction rates relative to both vehicles. No significant difference in tensile strength was demonstrated among groups. These observations do not support the proposed use of minoxidil as an “antifibrotic” agent.

Wound Healing Trajectories as Predictors of Effectiveness of Therapeutic Agents

Point-of-care diagnostic devices typically require six distinct qualities: they must deliver at least the same sensitivity and selectivity, and for a cost per assay no greater than that of todays central lab technologies, deliver results in a short period of time (<15 min at GP; <2h in hospital), be portable or at least small in scale, and require no or extremely little sample preparation. State-of-the-art devices deliver information of several markers in the same measurement.