Surgical Therapies

Abstract

Cellular grafting for vitiligo has evolved since its inception. Non-cultured cellular grafting has superseded cultured techniques as the preferred modus operandi in clinical practice. The reasons are straightforward: it can be completed in a few hours, does not require the use of xenobiotics (except trypsin) or culture media and is of significantly lower costs. Although it does not offer the possibility of cell expansion and cryopreservation for future transplant, this can be supplanted by harvesting a new piece ultrathin split skin graft. Under proper training, harvesting an ultrathin split skin graft (not beyond the papillary dermis) is fast and results in no or minimal scarring. Innovative methods to simplify non-cultured cellular grafting have also been successfully achieved.

Melanocytes can now be extracted not only from the epidermis for transplantation but from outer root sheath of hair follicles.

Non-cultured cellular grafting has proven to be a safe and effective surgical treatment for stable vitiligo, substantiated by various long-term studies over the last two decades. The imposition of strict legislative regulation on this technique is unwarranted and will only drive up cost and deprive patients’ access to this innovative procedure.

Annex A: Surgical Protocol for NCES (Adopted and Modified from [27])

• 1. The first step is always the most important. It is crucial to harvest an ultrathin split skin graft. We usually choose the left lateral hip as the donor site, and the size is one-third to one-fifth the vitiligo area to be transplanted. The donor site is cleansed with chlorhexidine 4% and outlined with skin marking ink, before performing an anaesthetic field block using 2% lignocaine. A silver dermatome is used to harvest a split skin graft not beyond the papillary dermis, which can be gauged by the translucency of the graft (Fig. 36.3a). Too thick a graft will impede trypsinization and can lead to scarring of the donor site. This is a skill to be acquired and a learning curve. It requires the help from an assistant in stretching the skin taut (using, e.g. a sterilized wooden spatula), so that the surface is as flat as possible. Sterile liquid paraffin provides good lubrication for the dermatome to glide on the donor skin surface.

• 2. The split skin graft is placed in a sterile tray and washed in normal saline, before placing it in a petri dish containing 10 ml of trypsin-EDTA 0.25%, pre-warmed to 37 °C (Fig. 36.3b). The graft is then cut into smaller pieces, and the petri dish is covered and placed in an incubator, set at 37 °C, for 30 min (Fig. 36.3c).

• 3. After incubation, the petri dish containing the donor tissue is placed in a laminar flow chamber, wherein the extraction of epidermal cells takes place. The lid of the petri dish is lifted, inverted and placed on the benchtop of the chamber. The tissue pieces are removed from trypsin and placed on the inverted lid. Here, using fine forceps, the epidermis is peeled away from the dermis and their surfaces scraped with the forceps to dislodge the epidermal cells including melanocytes.

• 4. The tissue-cell mixture is then collected and placed into a 50 ml Falcon tube containing 10 ml of PBS. The tube is capped and the suspension mixed by shaking and twirling tube for 30 s. In so doing, the cells will be further dislodged and suspended in the PBS.

• 5. The suspension is decanted into a petri dish, and 10 ml of soybean trypsin inhibitor is added to neutralize trypsin. The tissue debris are physically removed and discarded from the mixture. The neutralized suspension is then pipetted into a 50-ml Falcon tube, through a 40-μm filter, to remove the residual tissue debris.

• 6. The filtrate, containing the cell suspension, is then centrifuged at 120 × g for 5–8 min. Epidermal cells will settle to the bottom of the tube as a pellet (Fig. 36.3d). The supernatant is discarded, and the cell pellet is resuspended in 2–5 ml of PBS (depending on the size of the pellet and the area of recipient site).

• 7. The final suspension is then applied onto laser-ablated or dermabraded vitiliginous recipient sites. The depth of ablation of the recipient area is gauged by pinpoint bleeding. The application of the cell suspension is carried out using a pipetter which offers good control of the amount dispensed. A small amount is applied to avoid run-off, controlled by surface tension of the fluid (Fig. 36.3e). Collagen dressing (Neuskin-F, Eucare, India), cut into small pieces, is then applied to hold the suspension in place (Fig. 36.3f), and the area is further secured with Hypafix dressings (Smith & Nephew, UK) (Fig. 36.3g).

• 8. The dressings are kept in place for 5–7 days before removal. Post-grafting phototherapy is not required in our experience. Patients can expose the recipient sites to sunlight daily for 5–10 min.

• 9. If the grafting is successful, signs of repigmentation are usually evident by 4 weeks in Indian, 8 weeks in Chinese (Fig. 36.4a, b) and 12 weeks in Caucasian patients.