34th Annual Scientific Meeting proceedings

1^o closure is always the 1^o aim

All wounds, either iatrogenic (the result of excision) or traumatic, should be primarily closed if at all possible. Secondary intention healing, particularly of the distal limb, is a very slow process. The resultant wound is covered by thin fragile epithelium that is then susceptible to further trauma (Figure 1).

Figure 1. Second intention healing of a large wound to the dorsal aspect of the hock. A. Wound at 3 weeks following initial closure and subsequent dehiscence; B. 9 months following dehiscence; C. Final result at 15 months

Reconstructive surgery can result in a tension-free closure and where applicable, is preferable to skin grafting since it will result in a more reliable result and produces a better cosmetic and functional outcome.

Reconstructive techniques used in horses include:

• Methods to reduce tension on the suture line
• Skin mobilization (Rotation flaps, pedicle advancement flaps, transposition flaps)
• Skin expansion

There has been little information in the literature regarding the outcome of primary intention healing. In a large retrospective study, complete primary healing after closure of traumatic wounds was successful in only 26% of horses and 41% of ponies (Wilmink 2002). Based on these disappointing results perhaps, wounds are often left to heal by secondary intention. However, in a recent study, using a novel tension relief system (Tension Tile System), primary intention healing was achieved in 69% of wounds overall (Comino 2023) (Figure 2). Moreover, this was irrespective of the time elapsed prior to surgery, refuting the so-called ‘golden period’, after which wounds should not be primarily closed.

Figure 2. Application of the Tension Tile System to a large wound on the cranial aspect of the stifle. A. Wound following debridement; B. Wound apposed and pre-placed horizontal mattress sutures tied; C. Wound at 2 weeks after injury at the time of suture removal

Why are we failing therefore to primarily close wounds successfully?

• Lack of knowledge
• Poor technique
• Inability to take into account both static and dynamic tension

Skin Grafting

If the skin defect is large or the location negates a skin mobilization technique, free skin grafting is eminently preferable over secondary intention healing. Techniques include island grafting, tunnel grafting, sheet grafting, and MEEK micrografting. Grafts can be categorized as full or partial thickness, based on the degree of dermis included.

A full-thickness skin graft includes all of the dermis, whereas a split-thickness skin graft is thinner and includes only parts of the dermis. In the latter, the skin appendages located in the deeper dermal layers remain at the site of removal and provide the resources necessary for the donor site to heal.

Full thickness grafts will lead to a superior functional and cosmetic effect, however there is a generally a lower acceptance rate due to the higher metabolic demand. In addition, the expansion ratio is limited and the donor site must be primarily closed. Therefore, the technique should be limited to smaller skin defects with a good vascular supply. The modified MEEK technique is a combination of split-thickness sheet grafting and island grafting, and has proven to be extremely successful in horses (Wilmink et al. 2006) (Figure 3). The technique has been recently adapted, enabling the use of full-thickness grafts in the standing horse (Wilmink 2024).

Figure 3. MEEK grafting of a dorsal MT3 wound A. Initial debridement with exposure of extensor tendon; B. Wound ready for grafting; C. Meek grafts applied; D: 12 weeks following grafting

The techniques used nowadays are not very different from the descriptions made

150 years ago. Some tools and devices have been improved, some refinements introduced, but the overall concepts remain.

Is There Anything New?

Despite few changes in the techniques themselves, several attempts have been made on the human side to address specific but frequent issues. Some of these techniques may be applicable to equine wound healing.

Obtain more time prior to skin grafting
Use of allografts or Xenografts (Nile tilapia) to temporarily cover large body surfaces after debridement, if split thickness grafts are not directly available.

Increase the supply of donor skin

Minced grafts
A new technique has been developed to increase skin graft expansion. Using a mincer with parallel cutting discs, 0.8 x 0.8 mm micrografts are created from a donor site of only 2.5 x 5cm (Figure 4). The grafts are mixed into a hydrogel and then applied to the recipient site with a spatula. In a prospective study of human chronic leg ulcers, the minced grafts were no less effective compared to a traditional 1:3 meshed graft, but with the advantage of being economical and allowing a 1:9 expansion ratio (Sanchez-Pinto 2023).

Figure 4. A. Head of the mincer with parallel cutting discs; (B) minced skin grafts applied over the recipient site with a spatula. C: Appearance of wound prior to application of a silicon dressing and hydrogel layer

Re-Cell
More recently, devices that permit autologous skin cell suspension and spraying (ReCell) have been developed and brought to the market, further expanding the tools available (Holmes 2023).

CEA (cultured epidermal autografts)
In this technique, cells isolated from small skin biopsies can be propagated in vitro and cultivated on biomaterials to cover skin defects. This has revolutionised the care of massively burned patients, allowing the coverage of burns that involved >90% of total body surface area. This has been further refined in cultured dermoepidermal autografts (CDEA), by adding fibroblasts to keratinocytes to improve skin laxity. CDEA have now been developed to the point where anatomical body parts such as a whole hand can be grown as a single unit (Pappalardo 2023) (Figure 5). The next generation of cultured autografts will include stem cells in order to recreate a full-thickness skin substitute that includes adipose, melanocytic, nervous, lymphatic, and vascular structures (Marino 2014). Attempts are being made to automatize the process of skin production through 3D-printing technologies, generating 100cm² of skin in 35 minutes (Cubo 2016).

Figure 5. Engineered edgeless human skin A: 3D printed hand scaffold; B: Hand-shaped dermis after 14 days of ECM remodelling; C: Ready for removal from scaffold

Deficiency in the wound bed

Almost all tissue can accept a free graft as long as it is well vascularized. Traditionally however, bone devoid of periosteum or tendon devoid of paratenon cannot be grafted directly. Dermal substitutes, such as Integra^® have been shown to overcome this limitation, serving as a scaffolding for the growth of neodermis. It consist of a bilaminate sheet of collagen I/GAG plus a temporary silicone sheet cover. Three weeks after application, the silicon sheet is removed and replaced with a split thickness sheet graft. In people, it is used increasingly for lower extremity wounds where coverage of bone and tendon is a common problem (Chang 2019).

Graft failure

Optimise wound bed preparation
Infection is one of the most important causes of skin graft failure. We are currently investigating the use of a fluorescence imaging device (Moleculight^TM) to detect elevated bacterial load prior to the grafting.

Secure grafts to skin (NPWT)
The use of negative wound pressure therapy both for wound bed preparation and to secure skin grafts in place has become widespread on the human side. In dogs, variables of graft acceptance were superior when NPWT was used in the 1^st week post-grafting. Fibroplasia was enhanced, open meshes closed more rapidly, and there was less graft necrosis (Stanley 2013). Although a promising technique in horses, currently the evidence is based on individual case reports (Rijkenhuizen 2005, Jordana 2011).

References

• Chang DK, Louise MR, Gimenez A, Reece E. The basics of integra dermal regeneration template and its expanding clinical applications. Semin Plast Surg 2019 33:185–189.
• Comino F, Pollock PJ, Fulton I, Hewitt-Dedman C, Handel I, Gorvy DA. A novel tension relief technique to aid the primary closure of traumatic equine wounds under excessive tension. Equine Vet J. 2024 56(3):514-521.
• Cubo, N.; Garcia, M.; del Cañizo, J.F.; Velasco, D.; Jorcano, J.L. 3D Bioprinting of Functional Human Skin: Production and In Vivo Analysis. Biofabrication 2016, 9, 015006.
• Holmes, J.H. A Brief History of RECELL® and Its Current Indications. J. Burn Care Res. Off. Publ. Am. Burn Assoc. 2023, 44, S48–S49.
• Jordana, M., Pint, E. & Martens, A., “The use of vacuum-assisted wound closure to enhance skin graft acceptance in a horse”, Vlaams Diergeneeskundig Tijdschrift 2011, 80(5), 343–350
• Marino, D.; Luginbühl, J.; Scola, S.; Meuli, M.; Reichmann, E. Bioengineering Dermo- Epidermal Skin Grafts with Blood and Lymphatic Capillaries. Sci. Transl. Med. 2014, 6, 221ra14.
• Pappalardo, A.; Alvarez Cespedes, D.; Fang, S.; Herschman, A.R.; Jeon, E.Y.; Myers, K.M.; Kysar, J.W.; Abaci, H.E. Engineering Edgeless Human Skin with Enhanced Biomechanical Properties. Sci. Adv. 2023, 9, eade2514.
• Rijkenhuizen, Astrid B. M. et al. “Can vacuum assisted wound management enhance graft acceptance.” Pferdeheilkunde Equine Medicine 21 2005: 413-418.
• Sanches-Pinto DC, Eriksson E, Gomez DS, Nunes MPT, Gemperli R, Soriano FG. Minced skin grafts for chronic wounds compared to conventional mesh grafts. Health Sci Rep. 2023 Jun 21;6(6):e1353
• Stanley BJ, Pitt KA, Weder CD, Fritz MC, Hauptman JG, Steficek BA. Effects of negative pressure wound therapy on healing of free full-thickness skin grafts in dogs. Vet Surg. 2013 Jun;42(5):511-22.
• Watfa,W.; di Summa, P.G.; Meuli, J.; Raffoul,W.; Bauquis, O. MatriDerm Decreases Donor Site Morbidity After Radial Forearm Free Flap Harvest in Transgender Surgery. J. Sex. Med. 2017, 14, 1277–1284.
• Wilmink JM, van Herten J, van Weeren PR, Barneveld A. Retrospective study of primary intention healing and sequestrum formation in horses compared to ponies under clinical circumstances. Equine Vet J. 2002; 34(3):270–3
• Wilmink JM, van den Boom R, van Weeren PR, Barneveld A. The modified Meek technique as a novel method for skin grafting in horses: evaluation of acceptance, wound contraction and closure in chronic wounds. Equine Vet J. 2006 Jul;38(4):324-9
• Wilmink JM, van Weeren PR. Skin grafting with the modified Meek technique in the standing horse using full thickness skin: Evaluation of acceptance, wound contraction and wound closure in chronic wounds. Equine Vet J. 2024 Jan 24. doi: 10.1111/evj.14064. Epub ahead of print. PMID: 38267820.