The Use of a Silk Fibroin-Based Dressing on Full Thickness Facial Skin Wounds: A Randomised Controlled Clinical Trial

Introduction

Wound healing is a natural, orderly, and timely reparative process that results in sustained restoration of anatomic and functional integrity. Facial trauma can cause abrasions on skin surfaces, and various surgical procedures often result in open wounds. These wounds are covered by dressings or grafts to prevent microbial infection, excessive fluid loss, foreign material contamination, wound contracture and optimize the rate of healing.¹ Research has proven the concept of an optimum environment for wound repair and the involvement of the dressing material in establishing and maintaining such an environment. This has led to the development of wound dressings from the traditionally passive to the more functionally active dressings.²

Desirable properties of a wound dressing include being protective, antimicrobial, absorbent, non-adherent, preventive of heat and fluid loss, and being aesthetically acceptable. Silk Fibroin is a material that promotes collagen synthesis and re-epithelialization, which is considered to be proper for the generation of biomedical products such as blended materials because of its minimal adverse effects on the immune system.³ Asiaticoside, derived from Centella asiatica at low doses also promotes wound re-epithelialization by stimulation of factors essential for angiogenesis.⁴

The present study aimed to evaluate the effectiveness of Silk Fibroin (SF)-Asiaticoside membrane in full-thickness skin wounds, to assess its acceptance as a dressing material and its effects on raw wounds in the face.

Materials and Methods

This study was conducted in the Oral and Maxillo-facial Surgery Department of a tertiary referral research institute in Karnataka, India, after obtaining ethical committee approval and informed consent from the patients. All procedures performed in this study conformed to the principles of the Helsinki Declaration.

Twenty four adult patients with facial skin defects were randomly allocated to the study and control groups. Patients with full-thickness facial wounds caused by trauma or surgical procedures (Post excision, Flap donor sites), subjects aged 18 years and above were included in the study. Pregnancy, uncontrolled systemic diseases, and any known allergy to silk or petrolatum were considered as exclusion criteria. Patients who underwent skin grafts for facial wounds with reporting time more than 72 hours post-trauma and those who had infected wounds were also excluded from the study.

Materials

We used silk protein-based wound dressing sheets containing Silk Fibroin Matrix (46%) and Asiaticoside extracted from Centella asiatica (0.6%W/W) based on a sterile bi-laminate silk film having hydrophobic and hydrophilic ends. The hydrophobic side comes in contact with the wound, which allows the absorption of exudates produced as a part of the natural wound-healing process. The hydrophobic end does not allow atmospheric moisture to enter but allows free passage of air through the micro pores present on the film.

Petrolatum gauze (PG) is a non-adherent dressing that can be used as a contact layer on open wounds. Due to the absence of any bioactive properties, PG was chosen as control to evaluate the performance of Silk Fibroin-based membrane as a wound dressing.

Methodology

All surgical procedures were performed under strict aseptic conditions. Thorough debridement was done in all abrasions caused by trauma. In the first group of patients, the SF membrane was taken out of its sterile pouch and cut in slight excess of the wound size using sterile scissors and was applied directly over the wound. In the second group of patients, PG dressing material was taken out of its sterile container and placed over the wound. In both groups of patients, sterile gauze was placed over the dressing membrane and was secured in position using surgical tape or roller gauze. The dressing was evaluated by removal of the overlying sterile gauze dressing and was re-assessed periodically.

Scoring criteria

Weekly scoring was done on the first day of the dressing and the 1st, 2nd, 3rd, and 4th week after the dressing was first placed. Evaluation of the results was based on the following parameters.

Wound surface area⁵

The wound surface area was calculated on the day of first dressing and on the four subsequent weeks by the greatest length and width method.

Greatest length of the wound (L) x Greatest width of the wound (W) = Wound Surface Area

The criteria for evaluation of granulation, pain relief, exudation, adherence and number of dressing changes are presented in Table 1.

Total Score

The total score was calculated at the end of each week by adding the scores obtained in the parameters of pain relief, granulation, epithelialization, adherence, wound exudation and frequency of dressing change. Each subject was awarded a score on a scale of 0 to 12 to give an indication of the overall effectiveness of the dressing materials.

Statistical analysis

Data were entered into Microsoft Excel (Windows 7; Version 2007), and analyses were done using the Statistical Package for Social Sciences (SPSS) for Windows software (version 22.0; SPSS Inc, Chicago). The mean values of subjective variables were calculated using Freidman's test. A comparison of values obtained for both groups was made using the Mann-Whitney U test. Comparison of mean of quantitative variables was analyzed using one way ANOVA test. Paired t-test was used to compare the variation of the mean of each time period, variable compared to pre-treatment. The level of significance was set at 0.05.

Results

Twenty-four patients between the age of 18 and 75 years (Mean 34.8±14.69) were included in the study. The mean age of the patients included in the study group was 31.5 years (±12.9), and the control group was 38.1 years (±16.1). Nineteen male and six female patients were included in the study. Three subjects had tissue defects caused by surgical procedures (flap donor sites), and 21 were caused due to trauma.

Wound surface area

A statistically significant change in the mean wound surface area was found in both the groups (P=0.03 study, P=0.02 control) during the first week of the study.

However, no significant difference in the change in wound surface area was found between the two groups over four-week period, and both groups followed a similar trend (Figure 1).

The scores awarded for wound surface area, granulation, exudation, pain relief, epithelialization, dressing adherence, frequency of change and the total scores over four weeks are presented in Table 2.

SF membrane was found to induce faster granulation at the end of the first week (P <0.05), and faster epithelialization during the second week (P=0.01). SF showed significantly lower exudation during the initial two weeks. Adherence to PG was significantly less during the entire study period. SF required significantly fewer dressings throughout the four week follow-up period (P <0.05). Significantly higher (P <0.05) total scores were recorded in the SF membrane group during the entire four weeks of the study. 

Discussion

Winter in 1962 established that occluded wounds epithelialize faster than wounds left open without dressing.6 It is now recognized that wound dressings are not just passive materials that occlude open wounds, but can actively contribute to the wound healing process.¹

Silk Fibroin has been used because of its unique set of properties which include biocompatibility, low immunogenicity, biodegradability, high water and oxygen uptake, robust mechanical properties, and cost-effectiveness. Silk Fibroin is used in various forms like films, sponges, hydrogels, electrospun mats and tubes.³

It was observed that the SF membrane had good conformability to the wound beds i.e., it was supple and adapted well to the wound bed irrespective of the contour and shape of the wound.

Hasatsri et al. ⁷ reported reduced healing time in wounds treated with Silk Fibroin dressings (11±6 days) when compared to wounds treated with Paraffin Gauze (14±6 days). Similar results were reported by Zhang et al. ⁸ in their study, with a mean healing time of 9.86±1.79 days for partial thickness wounds treated with Silk Fibroin films. Both studies reported faster rates of wound healing when compared to control groups, which is comparable to the findings of our study.

The mean wound surface area and the rate of shrinkage of the wound were found to be comparable in both the groups. P values obtained indicated no significant difference in the rate of wound healing between the study and control groups. Hasatsri et al.⁷ and Zhang et al., ⁸ used Silk Fibroin films containing no additional bioactive components in their studies. We used Silk Fibroin membranes containing 46% Silk Fibroin protein and Asiaticoside, which could account for the longer healing times. This can also be attributed to the formation of hypertrophic granulation tissue in one patient of SF group, which increased the duration of wound healing.

Zhang et al. ⁸ reported that Silk Fibroin membrane provided coverage of the sensitive nerve endings present in the wound surface area, thus contributing to the pain relief reported by patients. The lack of an inflammatory response to Silk Fibroin has also been cited as a factor influencing pain relief. We found pain relief to be similar in the SF membrane group when compared to the PG during the first two weeks of the study, with the average pain scores being lesser in the SF group as was found during the subsequent two weeks. Schulz et al. ⁹ found no significant difference in the levels of pain reported by patients treated with Silk Fibroin dressings and other dressings during their study period. Their findings are in accordance with the findings of this study.

Wounds treated with SF showed early and faster granulation tissue formation compared to the control group. 10 out of 12 patients received a score of 2 by the end of first week, and all patients in the study group (n=12) scored 2 by the end of second week, in contrast to the PG group, which took the subjects three weeks to achieve granulation more than 2/3rd of the wound bed. This correlates with the findings of Zhang et al.,⁸ who reported the formation of healthy granulation tissue in all patients treated with a Silk Fibroin film during a 10- day period. The early formation of granulation tissue may also be attributed to the presence of Asiaticoside in the dressing membrane. However, no histological study was carried out to correlate the clinical findings.

Hasatsri et al. ⁷ found early epithelialization and wound healing in patients treated with Silk Fibroin dressings as compared to those wounds treated with paraffin gauze. They concluded that it was because silk fibroin could induce migration, adhesion, and proliferation of epidermal cells. We found the rate of epithelialization of wounds in the SF group was higher during the first two weeks as compared to the PG group, but the results for the third and final weeks were similar. Significantly higher (P=0.015) scores for epithelialization were recorded at the end of second week in SF membrane group. Although the rate of epithelial proliferation was found to be higher during the initial two weeks, the overall rate of wound shrinkage was found to be similar in both groups.

Exudation was significantly lower (P <0.05) in wounds treated by the SF membrane during the four weeks study period. This can be attributed to the semi-permeability of Silk Fibroin membrane to moisture and also to the high water and oxygen uptake exhibited by Silk Fibroin. This is in accordance with the finding of minimal exudation of wounds treated with a Silk fibroin film reported by Schulz et al. ⁹ and Zhang et al. ⁸ The low inflammatory potential of Silk Fibroin is one of the contributing factors for minimal exudation.

Both the SF membrane and PG were found to be easily adaptable to irregular wound surfaces. The mean adherence scores achieved by PG during the initial two weeks were higher than those attained by SF. PG was easier to remove from the wound bed. This was due to the lack of an emollient base in the SF membrane.

The frequency of dressing change was significantly lesser in the SF group throughout the study period (P <0.001).The PG dressing required multiple dressing changes during the initial three weeks of study period, primarily due to the larger amounts of exudate produced from the wounds. In healing wounds, the SF membrane was easily removed from the epithelial portions of the wound, resulting in lesser pain during dressing removal and better patient compliance during dressing changes. Less frequent dressing changes cause lesser disruption of healing wounds and enhance rates of wound healing.

Zhang et al.⁸ also reported a similar finding of nonadherence. Upon wound healing, the SF membrane became spontaneously detached from the regenerated skin areas on the wound bed and thus caused less trauma on removal.

The total scores obtained by wounds treated with SF membrane during the first week (9.91) and second week (11) were significantly higher (P <0.001) than those obtained by PG (5.66 & 7.75). Once the initial healing phase was completed, both dressings obtained similar scores during the last two weeks. The higher scores obtained by SF membrane point towards the overall effectiveness of the membrane as a dressing material.

In the clinical trial by Shulz et al.,⁹ only minor inflammatory reactions to Silk dressings were reported, while other studies^7.8 described no adverse reactions. In this study, no local inflammatory or allergic reactions to SF were noted. This finding reinforces the fact that Silk Fibroin is a clinically safe biomaterial with minimal antigenic potential.

Limitation of this study was that significant results could not be obtained in all parameters due to smaller sample size. In the absence of published clinical trials with larger sample size, conclusions about the effectiveness of Silk Fibroin-based dressing materials cannot be arrived at. Further studies regarding the use of Silk Fibroin-based biomaterials are required to fully realize its potential in wound healing applications.

Conclusion

SF membrane was found to be more effective in inducing the formation of granulation tissue and rate of epithelialization was found to be faster in wounds dressed with SF membrane. Wound exudation in the SF membrane group was found to be significantly lesser than the PG group and hence fewer dressing changes were required. The SF membrane did not evoke any antigenic or inflammatory responses and was found to be suitable for wound dressing purposes. Considering the above findings, the Silk Fibroin Asiaticoside membrane was found to be a more suitable dressing for extraoral wounds than petrolatum gauze when used judiciously in controlled clinical settings. However, more studies are required to explore the full potential of Silk Fibroin biomaterials in wound healing .

Conflicts of interest

Nil

Acknowledgements

The authors would like to acknowledge Mr. Vivek Mishra of Fibroheal Woundcare Ltd., Bangalore for providing the Silk Fibroin based dressings used in this study.