The Clinical Effect of Phenytoin on Oral Wound

Aims: To study the effect of oral phenytoin on healing of oral wound in buccal mucosa of rabbits.Materials and Methods: This study was carried out on twenty healthy male rabbits weighing between 1.0 – 1.5 Kg, they were divided into 2 groups; first group consisted of 10 untreated rabbits (control) and second group consisted of 10 rabbits treated by phenytoin at dose of 60 mg/Kg orally along with 3 ml/Kg of sterile water using cavage needle for 10 days. All animals were anesthetized with amixture of xylazine hydrochloride and ketamine hydrochloride at 0.5, 50 mg/Kg intramuscular respectively, then a standard wound was made on buccal mucosa of each rabbit, all animals were kept under observation, and their wounds was measured every day with respect to surface area (length x width) in cm 2 , type of wound tissue and duration of healing. Results: t – test analysis was performed to test the differences in wound characteristics of both groups, it was found that there was significant differences between control and treatment groups (p< 0.001). Conclusions: Systemic use of phenytoin can delay oral wound healing of buccal mucosa.


INTRODUCTION
Delayed healing or chronic wounds are a significant health care problem today; the quest for better woundhealing agent is one of the oldest challenges for medical and dental practice. One agent that has been tried in wound healing is phenytoin (1) . Oral phenytoin (diphenyl hydantoin) is used widely for the treatment of convulsive disorders. It served as an effective anticonvulsant against tonicclonic and partial seizures. Although the mechanism of action responsible for this effect of phenytoin is not established, prolonging Na + channel inactivation is the most compelling explanation for its anticonvulsant effect. (2) A common side effect with phenytoin treatment of epilepsy is the development of fibrous overgrowth of gingiva, apparent stimulatory effect of phenytoin on connective tissue suggested an exciting possibility for its use in wound healing. (3) Researchers suggested that phenytoin has a powerful effect in skin wounds healing and it may be useful in clinical practice. (4)(5)(6) The aim of this study was to evaluate the effectiveness of oral phenytoin in oral wound healing in rabbits.

MATERIALS AND METHODS
The study was carried out on twenty healthy male rabbits weighing between 1.0 -1.5 Kg. The animals were kept in standard animal housing condition with the room temperature of 25±2°C. All rabbits were anesthetized by intramuscular administration of xylazine hydrochloride (Holland, Castenray, interchemra) and ketamine hydrochloride (Aleppo -Syria, El-Saad) at dose of 0.5 , 50 mg/Kg respectivly. A standard surgical incision was applied to the buccal oral mucosa of the upper jaw of all rabbits (about 1 cm length) horizontally using surgical blade (no. 12). The procedure was carried out at the pharmacology Lab. / College of Dentistry / University of Mosul. After oral mucosal incision, rabbits were divided into 2 groups as follows: first group consist of 10 untreated rabbits (control), second group consist of 10 rabbits treated with phenytoin sodium (Germany, Gödecke AG), each rabbit was given phenytoin at dose of 60 mg/Kg orally diluted with 3 ml/Kg of sterile water using cavage needle. The dose of drug were calculated based on body weight of rabbits in comparison to human dose. (2) Phenytoin treatment was given 2 days before surgical incision application to all rabbits in the treatment group and continue for 8 days later. (7)(8)(9) After doing the surgical incisions there was an every day observation and measurement of wounds size, they are categorized with respect to surface area that is calculated by measuring the greatest length and the greatest width (side to side) using a centimeter ruler then multiply these two measurements (length × width) to obtain an estimate of surface area in cm 2 ,the subscores of wound surface area were as follow: The type of wound tissue that is present in wound bed was determined by clinical observation and the subscores were as follows: 4 -Necrotic Tissue (Eschar): black, brown, or tan tissue that adheres firmly to the wound bed 3 -Slough: yellow or white tissue that adheres to the wound bed 2 -Granulation Tissue: pink or beefy red tissue with a shiny, moist, granular appearance.
1 -Epithelial Tissue: for superficial ulcers, new pink or shiny tissue that grows in from the edges or as islands on the wound surface. 0 -Closed/Resurfaced: the wound is completely covered with epithelium. Then these parameters were added to obtain total score. Acomparison of total scores measured over time and the duration of healing in days provide an indication of the improvement or deterioration in wound healing. (10) The wounds were measured by the same clinician with the animals in the same position at regular intervals (every day), digital camera was also used to had an image of wounds for both groups of rabbits. (11 -15)

RESULTS
Descriptive statistics showed that the complete healing of oral wound in rabbits was seen after 7 days in control group, while in treatment group it was achieved after 10 days (Table 1).
Ttest analysis was performed to test the differences in parameter of wound characteristics including surface area (length × width in cm 2 ), tissue type and total scores for both control and treatment groups in all study days (Table 2).  It was found that there were significant differences between control and treatment groups (p < 0.001) in relation to wound size (length × width in cm 2 ) during all study days, while for tissue type there were significant differences between control and treatment groups during study days except in second and 3 rd days of study that there was a non significant differences between two groups (p = 1.000, p = 0.177) respectively.Pictures of control and treatment groups were showed in (Figures 1, 2) respectively.

DISCUSSION
The wound healing process involves many complex factors. These may be classified as local factors, systemic factors and organ and species variability in response to injury. (7) Chronic wounds are a significant health care problem. Phenytoin (diphenyl hydantoin) which was introduced into therapy in 1937 for the effective control of convulsive disorders has been tried in wound healing, (1) a common side effect with chronic phenytoin treatment is the development of fibrous over growth of gingiva. This apparent stimulatory effect of phenytoin on connective tissue suggested an exciting possibility for its use in wound healing. (16) Both topical and oral uses of phenytoin for wound healing are within the guidelines set forth by the FDA. (17) In this study oral phenytoin has been tried in healing of wounds in buccal mucosa of rabbit mouth, mean time to complete healing was 7 days in the control group of rabbits compared with 10 days in the phenytoin group. Significant differences can be seen between control and phenytoin treatment groups. According to the relation to wound size and tissue type, they were found in the results of this study and provide evidence that phenytoin can cause delayed healing in the wounds of buccal mucosa and this was in agreement. Some studies indicated that phenytoin can increase the risk of oral infections and the intensity of bleeding and exudates of wounds which contribute to delay in wound healing. (4,18) While in other studies, topical phenytoin on skin may have the potential to alter the dynamics of wound healing, and to promote it mainly by increasing granulation tissue formation. (19,20) Favorable therapeutic responses has been reported with oral phenytoin in patients with epidermolysis bullosa, however, oral administration of this drug is not widely accepted as effective. (1) Also topical use of phenytoin as mouth wash has been found to accelerate wound healing and resolution of mucositis. (5) This overview provided the evidence that phenytoin can accelerate wound healing mainly when used topically, while oral administration of this drug is not so effective, this can be due to the differences between the pharmacokinetics of topical phenytoin compared to oral one. (18) Studies indicated that topical phenytoin accelerated wound healing by increasing the number of wound macrophages and improving the macrophage function, also this topical agent act locally and do not undergo classic systemic metabolism modifications which can not be achieved when phenytoin was used orally. (21,22) Another hand phenytoin mainly accelerate wound healing on skin which differ from oral wounds in relation to healing capacities glucose transporter protein-1 (GLUT-1) which is membrane protein act as carrier for glucose uptake into cell thus its expression can be a parameter of cell proliferation activity and wound healing. In the skin, glucose transporter protein-1 (GLUT -1) observed in the epidermal basal cell layer while the buccal mucosa showed no GLUT-1 protein expression in its basal cell layer, and this can affect the healing rate of wounds in skin and oral buccal mucosa. (23) So, different formulations are required for oral mucosal wound healing.

CONCLUSIONS
An interesting proposal is to use phenytoin (topical and\or systemic) to enhance wound healing, considerable evidence exists that this procedure holds promise. There is an interest in applying modern knowledge of growth factor to aid in the healing of oral wounds because phenytoin is widely available and relatively inexpensive, it might be helpful in some cases, research in this area is clearly needed.