Douglas L. Helm

Microdeformation of three-dimensional cultured fibroblasts induces gene expression and morphological changes.

Background:Vacuum-assisted closure induces microdeformations of the wound surface and accelerates healing of complex wounds; however, a thorough understanding of the biology of cellular mechanotransduction is lacking. We hypothesized that fibroblast shape and function can be altered in an in vitro vacuum-assisted closure device. Methods:A 3-dimensional fibrin matrix with cultured murine fibroblasts and an intervening polyurethane foam was exposed to 125 mm Hg suction and compared with similar wells without suction. We measured fibroblast proliferation and morphology using fluorescence microscopy and gene expression change using real-time reverse-transcriptase polymerase chain reaction at 24, 48, and 72 hours. Results:Wells exposed to suction induced significant proliferation of fibroblasts and morphologic changes visible by larger, rounder, and notable dendrite-like branching and process extensions. Type 1 collagen alpha 1 (COL1A1), fibroblast growth factor 2 (FGF2, bFGF), and transforming growth factor beta 1 (TGF&bgr;1) were all up-regulated after 48 hours of exposure to suction. Smooth muscle actin alpha 2 (Acta2, &agr;-SMA) was up-regulated after 72 hours. Conclusions:Microdeformations produced by the combination of polyurethane foam and suction are associated with increased fibroblast proliferation and up-regulation of gene expressions in fibroblasts.

Foam pore size is a critical interface parameter of suction-based wound healing devices.

Background: Suction-based wound healing devices with open-pore foam interfaces are widely used to treat complex tissue defects. The impact of changes in physicochemical parameters of the wound interfaces has not been investigated. Methods: Full-thickness wounds in diabetic mice were treated with occlusive dressing or a suction device with a polyurethane foam interface varying in mean pore size diameter. Wound surface deformation on day 2 was measured on fixed tissues. Histologic cross-sections were analyzed for granulation tissue thickness (hematoxylin and eosin), myofibroblast density (&agr;-smooth muscle actin), blood vessel density (platelet endothelial cell adhesion molecule-1), and cell proliferation (Ki67) on day 7. Results: Polyurethane foam–induced wound surface deformation increased with polyurethane foam pore diameter: 15 percent (small pore size), 60 percent (medium pore size), and 150 percent (large pore size). The extent of wound strain correlated with granulation tissue thickness that increased 1.7-fold in small pore size foam–treated wounds, 2.5-fold in medium pore size foam–treated wounds, and 4.9-fold in large pore size foam–treated wounds (p < 0.05) compared with wounds treated with an occlusive dressing. All polyurethane foams increased the number of myofibroblasts over occlusive dressing, with maximal presence in large pore size foam–treated wounds compared with all other groups (p < 0.05). Conclusions: The pore size of the interface material of suction devices has a significant impact on the wound healing response. Larger pores increased wound surface strain, tissue growth, and transformation of contractile cells. Modification of the pore size is a powerful approach for meeting biological needs of specific wounds.

Analysis of Nerve and Neuropeptide Patterns in Vacuum-Assisted Closure-Treated Diabetic Murine Wounds

Background: Reestablishment of the peripheral nervous system occurs in parallel with wound healing. With accelerated wound healing seen with the vacuum-assisted closure device, the authors studied its effects on nerve fiber regeneration, nerve sprouting, and the stimulation of neuropeptides and neurotrophins. Methods: A vacuum-assisted closure device was applied to a full-thickness diabetic mouse wound using continuous or cyclical modes and compared with foam dressing or occlusive dressing controls, using 10 mice per group. Nerve fibers, substance P, calcitonin gene-related peptide, and nerve growth factor were analyzed using two-dimensional immunohistochemistry and real-time reverse-transcriptase polymerase chain reaction. Results: A significant increase in dermal and epidermal nerve fiber densities and in substance P, calcitonin gene-related peptide, and nerve growth factor expression was seen in vacuum-assisted closure–treated wounds. Cyclical treatment mode correlated with the largest increase in granulation tissue production, wound surface microdeformations, and a slightly faster wound closure rate. Conclusions: This study suggests that vacuum-assisted closure therapy can modulate nerve fiber and neuropeptide production in the wound. Optimized kinetics of vacuum-assisted closure application may provide an opportunity for clinicians to further improve wound healing in denervated wounds such as pressure sores and diabetic foot ulcerations.

Waveform modulation of negative-pressure wound therapy in the murine model

Background: Negative-pressure wound therapy applied with a porous foam interface has been shown to accelerate granulation-tissue formation when a cyclic application mode of suction is applied, but the optimal waveform has not been determined. The authors hypothesized that changes in the suction waveform applied to wounds would modulate the biological response of granulation tissue formation. Methods: A vacuum-assisted closure device (Kinetic Concepts, Inc., San Antonio, Texas) was applied to full-thickness wounds in 48 male diabetic mice (C57BL/KsJ-Lepr db), which were treated with six different waveforms: square waveforms of 125 mmHg of suction for 2 minutes, alternating with 50 mmHg of suction for 2 minutes, 5 minutes, or 10 minutes; triangular waveform with a 7-minute period oscillating between 50 and 125 mmHg; and static suction at 125 mmHg or static suction at 0 mmHg (occlusive dressing). Wounds were quantitatively evaluated for granulation tissue thickness as well as the number of proliferating cells and the number of blood vessels of the newly formed granulation tissue. Results: At 7 days, the continuous and triangular waveforms induced the thickest granulation tissue, with high rates of cellular proliferation and blood vessel counts compared with square wave and occlusive dressing control wounds. Decreasing square waveform frequency significantly increased granulation tissue thickness, cellular proliferation, and blood vessel counts. Conclusions: Waveform modulation has a significant effect on granulation tissue formation, angiogenesis, and cellular proliferation in excisional wounds in diabetic mice. The rapid change in pressure seen in our square wave model may be detrimental to granulation tissue formation.

Controlled induction of distributed microdeformation in wounded tissue via a microchamber array dressing

Mechanical stimuli are known to play an important role in determining the structure and function of living cells and tissues. Recent studies have highlighted the role of mechanical signals in mammalian dermal wound healing. However, the biological link between mechanical stimulation of wounded tissue and the subsequent cellular response has not been fully determined. The capacity for researchers to study this link is partially limited by the lack of instrumentation capable of applying controlled mechanical stimuli to wounded tissue. The studies outlined here tested the hypothesis that it was possible to control the magnitude of induced wound tissue deformation using a microfabricated dressing composed of an array of open-faced, hexagonally shaped microchambers rendered in a patch of silicone rubber. By connecting the dressing to a single vacuum source, the underlying wounded tissue was drawn up into each of the microchambers, thereby inducing tissue deformation. For these studies, the dressings were applied to full-thickness murine dermal wounds with 200 mmHg vacuum for 12 h. These studies demonstrated that the dressing was capable of inducing wound tissue deformation with values ranging from 11 to 29%. Through statistical analysis, the magnitude of the induced deformation was shown to be a function of both microchamber height and width. These results demonstrated that the dressing was capable of controlling the amount of deformation imparted in the underlying tissue. By allowing the application of mechanical stimulation with varying intensities, such a dressing will enable the performance of sophisticated mechanobiology studies in dermal wound healing.

SPLIT rectus abdominis myocutaneous double free flap for extremity reconstruction

A Mathes and Nahai type III muscle, such as the rectus abdominis muscle, can be utilized to cover two separate wounds simultaneously utilizing its dual blood supply thereby minimizing donor site morbidity and operative time. We report a case for treatment of bilateral Gustillo type IIIB lower extremity injuries treated with a single rectus abdominis muscle split into two free flaps, with one based on the deep inferior epigastric vessels and one on the superior epigastric vessels to cover the contralateral wound. In our patient, both lower extremity wounds were covered with muscle flaps from the same donor site in a single operation, salvaging both limbs with progression to unassisted ambulatory status. We show in this case report that the utilization of the vascular anatomy of the rectus muscle allows for division of the flap into two flaps, permitting preservation of the contralateral abdominal wall integrity and coverage of two wounds with a single muscle.

Chapter II.5.14 – Burn Dressings and Skin Substitutes

Cost does play a role in the development of burn dressings and skin substitutes. Physicians must determine the optimal treatment they can provide to a patient considering the economic resources available. Also, although the development of dressings and skin substitutes for the treatment of burn injury is rewarding, our current marketplace has not proven to be supportive of this industry. One of the problems is that burn injury correlates to a very small market, and in order for a dressing or skin substitute to be truly successful it must also be able to be applied to a wider market such as that for general wound care, or the company must sell other products. Skin protects the body from microorganisms and external forces, integrates complex sensory nervous and immune systems, controls fluid loss, and serves important aesthetic functions. Large skin deficits have motivated the development of improved technologies to replace and restore skin. Although many of these technologies were originally developed to treat burns, these technologies have been extended to other areas, including chronic wounds and acute surgical wounds. This chapter explores the factors that need to be considered when developing wound dressings and skin substitutes, and also reviews the technologies currently available to treat patients.

Extended V-Y Advancement Flap Reconstruction of a Large Posterior Upper Midline Trunk Defect.

Summary: Large posterior upper trunk defects can be challenging to reconstruct. Trapezius or latissimus dorsi myocutaneous flaps are principally utilized for reconstruction; however, some defects may not be amenable to this standard approach. Here, we describe a patient with a full-thickness skin and subcutaneous tissue loss of the upper back and inferior cervical region after dermatofibrosarcoma protuberans resection. A large, extended V-Y flap was used for closure of this wound secondary to its location, size, and orientation. This approach preserves shoulder function, allows for readvancement of the flap as needed, and is a reconstructive option for patients with large upper back defects.