The effect of TGF-beta on keloid fibroblast proliferation and collagen synthesis.

Keloids are characterized by an overabundant deposition of collagen, and they recur frequently following excision. Fibroblasts isolated from keloid tissue and maintained in cell culture continue to express an increased capacity to produce collagen. In an effort to define the mechanisms responsible for keloid formation, the potential of exogenous transforming growth factor beta 1 (TGF-beta 1) to differentially affect DNA synthesis and collagen expression in cultured human fibroblasts derived from keloid or normal dermis was investigated. In this study, TGF-beta 1 at a concentration of 5.0 ng/ml was found to stimulate DNA synthesis of keloid-derived fibroblasts to a greater extent than fibroblasts derived from normal dermis. With a microassay to measure levels of collagenase-digestible radiolabeled proteins, TGF-beta 1 was found to elicit a greater increase in absolute collagen synthesis in keloid-derived fibroblasts compared with fibroblasts derived from normal dermis. Examination of tRNA(pro) pool-specific activities indicated that these observed differences in rates of collagen synthesis were not the result of unequal rates of proline transport or pool size. Likewise, TGF-beta 1 did not alter the uptake of vitamin C, an essential cofactor and mediator needed for maximal collagen expression. The increase in collagen synthesis by keloid-derived fibroblasts treated with TGF-beta 1 was accompanied by a corresponding increase in procollagen type I mRNA levels, indicating that the differential response of keloid and normal dermal fibroblasts to this growth factor is occurring primarily at a pretranslational level. These results suggest a unique sensitivity of keloid fibroblasts to TGF-beta 1 and thus a possible role for this mediator in keloid pathogenesis.

Hyaluronic acid impedes reepithelialization of skin graft donor sites.

Prior studies have suggested that hyaluronic acid (HA), a naturally occurring glycosaminoglycan, may accelerate the healing of superficial burn wounds. To objectively assess whether exogenous HA could accelerate the rate of healing and limit scar formation in superficial wounds, 11 patients with burn injuries who were undergoing skin grafting were given two separate skin graft donor sites of similar size and depth (1" x 1" x 0.016"). In a randomized, double-blind fashion, one of these partial-thickness wounds had repeated application with 1.5% HA; 100% glycerin, which served as a control of similar viscosity, was applied to the other wound. On alternating days, photographs were taken and then analyzed with computerized digital planimetry to objectively quantitate the rate of reepithelialization. In contrary to our hypothesis, HA significantly delayed wound healing compared to the glycerin control (time to > 95% reepithelialization: 10.3 +/- 2 days HA vs. 9.1 +/- 1.6 days control; mean +/- SD). Furthermore, no difference in the resultant scar at 6 weeks and at 3 months after wounding was evident. These results demonstrate that HA retards healing of partial thickness wounds in adults.

Comparison of the polyvinyl alcohol sponge and expanded polytetrafluoroethylene subcutaneous implants as models to evaluate wound healing potential in human beings.

Our current understanding of the complex processes involved in wound healing is based mainly on studies of animal models. Although this information has been useful, it may not totally reflect the response found in human beings. For example, human beings have a tendency to either "overheal," as seen in keloids and hypertrophic scar formation, or have deficient healing, as seen in chronic ulcer formation. No animal models are available to analyze these human clinical pathologic conditions. Therefore the objective of this study was to analyze the wound healing response in a large population (n = 40) of normal healthy human beings as a first step to begin studies of abnormal human wound healing. Simultaneously, a comparison was made between the polyvinyl alcohol implant and the expanded polytetrafluoroethylene implant model. Under sterile conditions with the use of local anesthesia, two preweighed polyvinyl alcohol implants and two standard 6 cm expanded polytetrafluoroethylene implants were placed subcutaneously in the upper arm of each subject. High-performance liquid chromatography was used to quantitate isoleucine and hydroxy-l-proline in acid hydrolysates of each implant. Isoleucine was used as an indicator of protein content in the tissue sample, whereas hydroxyproline reflected collagen content. No infectious or hemorrhagic complications were found in the 40 volunteers included in the study. No significant difference was found in isoleucine or hydroxy-l-proline content between postoperative day 7 polyvinyl alcohol implants and day 14 polyvinyl alcohol implants. In contrast, both isoleucine and hydroxy-l-proline content were significantly increased in day 14 expanded polytetrafluoroethylene implants compared with day 7 implants (p < 0.005 and p < 0.001, respectively). In addition, the ratio of hydroxy-l-proline to isoleucine was significantly increased in day 14 expanded polytetrafluoroethylene implants compared with day 7 expanded polytetrafluoroethylene and both day 7 and day 14 polyvinyl alcohol implants (p < 0.001). This observation suggests that by 14 days implantation of expanded polytetrafluoroethylene stimulated an increased deposition of collagen. No significant differences were found in the hydroxy-l-proline to isoleucine ratios among day 7 expanded polytetrafluoroethylene, day 7 polyvinyl alcohol, and day 14 polyvinyl alcohol implants. Histologic analyses correlated with the biochemical findings. These results suggest that expanded polytetrafluoroethylene may be the preferred implant for studies designed to examine pathologic processes associated with retarded wound healing. In contrast, the polyvinyl alcohol implant may be better suited for studies where a low background response is required. Moreover, the extreme variability in normal healthy volunteers seen in this study correlates clinically with the finding that, among the normal adult human population, there is a heterogeneous wound healing response.

The role of inflammatory cytokines in wound healing: accelerated healing in endotoxin-resistant mice.

UNLABELLED: The role of cytokines in normal wound healing remains poorly defined. In vitro, tumor necrosis factor alpha (TNF-alpha) decreases collagen accumulation, perhaps by increasing collagenase activity. The current study was undertaken to test the hypothesis that cytokines impair wound healing and collagen production. Wounds in C3H/HeJ (J) mice, which are characterized by a genetic defect in macrophage production of TNF and other cytokines in response to endotoxin, were compared with wounds in normal endotoxin-sensitive C3H/HeN (N) mice. METHODS: TNF (by murine ELISA) and collagenolytic activity (CA by in vivo labelled collagen fibril degradation assay) were measured in wound fluid from silicone reservoirs on post-wounding days 1, 3, and 5 (n = 5 per group). Hydroxyproline (HOP, nmol/mg sponge) incorporation (by HPLC) in polyvinylacohol sponges and breaking strength (BS, as determined by tensiometry) in linear wounds were assessed on days 5, 7, 10, and 14 (n = 5 per group). The data demonstrate significantly increased BS at 5 and 7 days in endotoxin-resistant J mice compared with that in endotoxin-sensitive N mice. An early, significant reduction in TNF production in J mice corresponded with a significant increase in CA on day 1, increased collagen production at day 7, and increased procollagen gene transcription at early time points. Conclusion. The reduced production of inflammatory cytokines including TNF in the wound fluid of J mice corresponded with an early improvement in wound tensile strength. An accelerated accumulation of collagen in the wounds of J mice, perhaps resulting from a significant decrease in collagenolytic activity or increased collagen production, are potential mechanisms. The data suggest that cytokines produced in normal healing of clean wounds may contribute to a delay in increased tensile strength.

Biology of fetal repair: the presence of bacteria in fetal wounds induces an adult-like healing response.

The minimal acute inflammatory response to tissue injury is one of the most dramatic differences between fetal and adult wound healing. Considering the prominent role of inflammation in adult tissue repair, this study tested the hypothesis that the minimal fetal inflammatory response to tissue injury plays a central role in the "scarless" fetal repair process. Sponge implants were treated with lethally irradiated or live bacteria and placed subcutaneously in fetal rabbits to test the ability of the fetus to mount an acute inflammatory response to bacterial antigens present at the wound site and to analyze the effects of this inflammatory response on fetal fibroplasia and neovascularization. After harvest, these implants were examined histologically for inflammation, fibroblast infiltration, collagen deposition, and neovascularization, and collagen deposition was measured using hydroxyproline quantitation by high-performance liquid chromatography. Bacteria-treated implants showed dose-dependent acute inflammatory responses and significant increases in collagen deposition compared with control sponges. Implants containing live bacteria demonstrated maximal fibroplasia and neovascularization. These findings suggest that, despite neutropenia and immaturity of the fetal immune system, the fetus is capable of mounting an acute inflammatory response to avirulent bacteria present at the wound site. Fetal inflammatory cells which respond to this bacterial stimulus appear capable of initiating an adult-like healing response. Thus, by failing to provide a bacterial stimulus for leukocyte recruitment at the site of tissue injury, the sterile fetal environment appears to play a role in effecting "scarless" fetal wound healing.