Wound Healing as Well as Fibroblasts and Neutrophil Numbers in a Skin Exposed to Infrared and Electrical Stimulation

Objective: To investigate wound healing as well as fibroblasts and neutrophil numbers on a limited skin field which is previously exposed to combined Infrared and Electrical stimulation. Methods: 20 male rabbits were randomly assigned to two groups each one of 10 animals. The experimental group was exposed to combine Infrared (IR) and Electrical Stimulation (ES) at dorsal area for three days respectively, in day four an incision was made in the center of exposed skin then sutured. A biopsies were taken at 24,48,72 hours, week and two weeks after surgery for counting of fibroblasts and neutrophil numbers as well as for histological observation. The same procedure was made for control group without exposing to radiation and stimulation. Results: There was statistically significant difference in fibroblasts and neutrophil numbers between two groups (p<0.01). The number of these cells were higher in experimental group than control in all periods. Histological observation revealed good orientation of fibroblasts and collagen fibers in experimental group as well as excellent wound healing. Conclusion: This study suggested that application of IR combined with ES on limited skin field have beneficial effects on fibroblasts and neutrophil numbers of a wound created in same exposed field with good wound healing. Introduction The use of Infrared (IR) radiation in the treatment of variety of medical conditions has been studied for long time (Abou-Hala et al., 2007). IR radiation is that invisible portion of the electromagnetic spectrum adjacent to the long wavelengths, or red end, of the visible light range that extend up to the microwave range(Hideyoshi, 2003). This phototherapeutic phenomenon has been reported particularly by physicians performing phototreatment of poorly healing wounds. Regeneration and microcirculation processes are stimulated not only in the irradiated lesions, but are also seen in distant non-irradiated lesions in the same patient; simultaneously, improvement of patients’ immune, metabolic and hormonal status can be registered. Local changes are usually associated with the direct action of light on skin cells. Journal of Kirkuk University – Scientific Studies, vol 6, No2, 2011 51 The systemic mechanisms of photobiomodulation remain, however, non-elucidated.( Samoilova et al., 2004). Radiation therapy has profound effects, both acute and long-term, on skin and connective tissues. Radiation therapy also affects the time course and end result of wound healing, and the risk of postoperative complications(Wang et al., 2006). It has been reported that wounds combined with whole body irradiation heal slowly, but the mechanisms are not fully clarified (Qu et al., 2004). On the other hand, electrical stimulation of very low amplitude and frequency modulation has become an increasingly popular treatment modality. This form of stimulation has been referred to as microamperage electrical stimulation (MES). MES is defined as stimulations with a very low frequency (1 Hz or less) and low intensity or amplitude (1–1,000 μA) (Mohammad, 2006), in addition , numerous morphological and functional effects of electric stimulation have been identified, both at the cellular and at the tissue level(Sumano et al., 2002). Surface stimulation studies have shown that Electrical Stimulation (ES) can produce positive short-term changes in tissue health variables such as regional blood flow and pressure distribution(Bogie et al., 2000). Many aspects of treatment with ES have been studied. Several randomized controlled trials (RCTs) have evaluated ES with varying protocols using different currents and voltages for the healing of pressure sores, venous stasis ulcers, arterial ulcers, surgical wounds, and diabetic foot wounds(Hayes, 2003). Radiation, however, impairs wound healing (Schaffer et al., 2007). Many studies have showed negative effects of radiation such as gamma ray and external beam radiation of wound healing when body or limited part of body exposed to these radiations (Qu et al., 2004; Dubin et al., 2000). No one study to date however examined the beneficial or harmful effect of IR or ES or combination between them on healing of wound in field that previously exposed to these stimulators. The investigation into the effect of combined IR and ES on local skin field to the healing of wounds that are created in a same exposed field ,as well as on fibroblasts and neutrophil numbers was the main goal of this study. Material and methods We used 20 male rabbits, weighing mean of (1,500-1,900), with age 5-6 months. Animals were housed individually with access to food and water for a week before trial. All animals injected with penicilline streptomycine 0.75 ml\ day for three days as a protective dose. Exposure and surgical technique The dorsal skin area of 10 rabbits clipped and shaved ( about 10 cm2) the field was exposed to an infrared radiation (IR) for 5 minutes ,the distance between IR lamp and the field was approximately 35cm, and the Journal of Kirkuk University – Scientific Studies, vol 6, No2, 2011 52 exposure area was 5x5cm2.Immediatetly after that an electrodes were placed on sterile pads moisted with water and placed on shaved area, we applied electrical stimulation(ES) at frequency 0.5 Hz current intensity for 5 minutes, this procedure was applied for 3 days frequently*. After 24 hr from last application of combined IR and ES, the animals were anesthetized with 50 mg\kg kitamine hydrochloride mixed with 35 mg\kg xylazine intramuscularly. The area that have been receipted the stimulators was sterilized by a pad of cotton immersed in ethanol alcohol (70%) for 5 minutes, the pad removed and the area swapped with povidone iodine for further sterilization. An incision about 3cm was made in the centre of exposed skin area then sutured using interrupted suture pattern. The wound covered with sterile bandage for protection. The same surgical procedures were made in control group (10 rabbits) without application of the stimulators. Biopsy collection The biopsy collected at 24,48,72 hours, week and two weeks after the surgery both in experimental and control group (two animals for each period). The biopsies fixed in 10% formalin solution at least 24hr for tissue processing (clearing, infiltration and embedding in paraffin wax), the sections of block were done in microtome for cutting about 5 microns, the slides stained with heamatoxyline-eiosin for histological examination. Counting technique Counting of neutrophils and fibroblasts was expressed per microscopic field. The number of these cells were counted in 10 microscopic fields for each specimen using ALTAY microscope at magnification of 40x. Statistical analysis To make comparison between experimental and control group in cell numbers and also to observe the differences of cell numbers during different periods, we used F test with ANOVA table.