Elsevier

Burns

Volume 30, Issue 6, September 2004, Pages 531-538
Burns

The healing-promoting effect of saliva on skin burn is mediated by epidermal growth factor (EGF): role of the neutrophils

https://doi.org/10.1016/j.burns.2004.02.007Get rights and content

Abstract

Local skin trauma induces inflammatory responses resulting in local tissue and distant organ injury. EGF, a polypeptide hormone, mainly produced in saliva, is one of the major accelerators in wound healing. Wistar albino rats of both sexes received either bovine serum albumin or EGF (10 μg/kg) subcutaneously before a circular (18 mm diameter) partial thickness burn was induced. Afterwards, some rats were placed in separate cages to prevent licking, while the others were caged together to allow wound-licking. Treatments were continued for 5 more days and on the 5th day animals were decapitated. Histopathological analysis of skin damage and dermal myeloperoxidase (MPO) activity, as an index for neutrophil activity, were evaluated. Oxidant injury to the liver and intestines was determined by measuring glutathione (GSH) and malondialdehyde (MDA) levels, as well as MPO activity. The results demonstrate that healing of the burn wound on the skin is accelerated by both wound-licking and EGF administration, which also attenuated tissue neutrophil accumulation, suggesting the role of neutrophils as the source of mediators involved in delayed epithelial regeneration. Moreover, local dermal burn results in oxidant injury to the liver, concomitant with significant elevations in hepatic and intestinal GSH levels. Exogenous administration of EGF at physiological doses had no effect on inflammatory responses of the distant organs, while allowing the rats to lick the wound reduced the oxidant injury to the liver. Since saliva or EGF enhances skin wound healing, topical use of EGF-rich artificial saliva merits consideration for its use in burn patients.

Introduction

The local and systemic inflammatory response to severe cutaneous thermal trauma is extremely complex, resulting in both local burn tissue damage and subsequent edema as well as marked systemic effects, which demand urgent medical interventions to ensure the survival of the patient. Despite many advanced medical treatments, cutaneous thermal wound management is still a great problem. Animal studies suggest that saliva, providing a natural reservoir of growth factors along with its bactericidal effects, plays a vital role in enhancing wound healing [1]. It was demonstrated that epidermal regeneration is accelerated when surgical wounds were treated with saliva [2]. Hutson et al. [3] have shown that wound healing of the skin is enhanced by licking, i.e. transfer of saliva to the wound. On the other hand, rate of healing of experimentally induced cutaneous wounds was delayed in sialadenectomized animals [4], [5]. Among many peptide growth factors that are known to exist in the saliva, one of the most widely studied is epidermal growth factor (EGF). Several lines of evidence have suggested that salivary EGF significantly accelerates the rate of wound healing of the skin and gastric mucosa [6], [7], [8].

EGF, a 53-amino-acid polypeptide hormone of low molecular weight (∼6000 Da), was purified by Cohen [9], who first observed the ability of EGF to induce early eye opening and tooth eruption in neonatal mice. EGF is a potent mitogen, both in vitro and in vivo, not only for epithelial cells, but for mesothelial and endothelial cells as well. Likewise, EGF is implicated in the regulation of a wide variety of physiological and pathological processes, including embryogenesis, growth, tissue repair, regeneration and neoplasia [10]. The major source of EGF in the mouse is submaxillary gland, from which it was first isolated [9]. Since the extirpation of submaxillary glands reduces, but does not fully eliminate, EGF concentrations in the plasma or urine, it is apparent that additional EGF sources in the body are present. In the small intestine, it is secreted directly into the lumen by Brunner’s glands and it is also present in the bile, colostrum and mature breast milk [11]. EGF stimulates messenger RNA, DNA, and protein synthesis in many cell types [9], [12]. It has been also shown to stimulate keratinocyte division in vitro and epidermal regeneration in vivo [8], [13], [14]. Local daily application of EGF in rats enhances accumulation of granulation tissue cells, collagen and glycosaminoglycans in experimental wounds [15]. Similarly, topical EGF treatment improves the healing of alkali burned corneal wounds in mice [16]. Exogenous administration of EGF attenuates internal complications associated to thermal injuries [17]. Moreover, it is well known that animals demonstrate an instinctive wound-licking behavior and the beneficial effect of licking on wound healing may be, in part, due to the transfer of saliva containing EGF.

In the present study, in an attempt to clarify the impact of EGF on the healing-promoting effect of saliva on burn-wounds, rats with local thermal wounds were either allowed to lick their wounds, as well as to have communal licking, or they received exogenous EGF while licking was prevented. In addition, the effect of EGF on both local and distant organ inflammatory response to burn injury was studied using the extent of neutrophilic leukostasis.

Section snippets

Animals

Wistar Albino rats (n=38) of both sexes, weighing 250–300 g, were obtained from Marmara University School of Medicine Animal House. The rats were kept at a constant temperature (22±1° C) with 12 h:12 h light and dark cycles and were fed with standard rat pellets and tap water ad libitum. All experimental protocols were approved by Marmara University School of Medicine Animal Care and Use Committee.

Burn injury

Deep anesthesia was induced with ketamine (100 mg/kg, intraperitoneally, i.p.). Local thermal trauma

Results

In the vehicle-treated, licking-prevented burn grup, histopathological examination of the skin confirmed that the epidermis was totally disrupted or severely desquamated, with severe dermal edema and vasocongestion (Fig. 1). Subcutaneous tissues, along with hair follicles, were severely degenerated, while the collagen fibers showed massive separation, resulting in a high microscopic damage score (14.0±0.63) (Fig. 2A). In the licking-permitted group, epithelial disruption was observed in limited

Discussion

The results of the present study demonstrate that healing of burn wound on the skin is accelerated by both wound-licking and EGF administration, which also attenuate tissue neutrophil accumulation. Moreover, the results also show that a small and local dermal burn results in oxidant injury of the liver, which is still evident on the post-burn 5th day. At the same time, this local trauma appears to stimulate the replenishment of hepatic and intestinal GSH stores, resulting in significant

Acknowledgements

The authors are grateful to Mr. Salih Güzel for the design and construction of the steel plaque used to induce burn injury.

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