26 Oct 2016
Distal limb wound healing in horses is well known for being problematic, prolonged and expensive. Sarah Boys Smith considers two simple graft techniques achievable in standing, sedated patients.
Sarah Boys Smith
Job Title
Skin grafting should not be feared or considered as a last resort, but should be part of a well-formulated plan.
Grafting large wounds greatly reduces the healing and convalescence time, and improves the functional and cosmetic appearance of the healed wound.
Indications for grafting include full-thickness wounds that cannot be closed primarily or by using sliding flaps, and wounds that cannot easily heal by contraction and epithelialisation.
The healed skin graft should consist of epidermis, dermis and some adnexa, including some hair follicles.
This means the new skin will be much more resilient and elastic compared to the dense fibrogranuloma scar tissue that can occur after the formation of excessive granulation tissue.
The many types of grafts can be broadly classified into two types – pedicle and free skin.
Pedicle skin grafts are full-thickness and remain attached to the donor site by a vascular pedicle, so the graft does not depend on the vascularity of the recipient site to survive.
Free skin grafts are full-thickness or partial/split-thickness separated from their original blood supply that must establish a new vascular supply.
Pedicle skin grafts have a good cosmetic and functional result, but are rarely applicable in equine patients because it is often not practical to mobilise the graft sufficiently to reach the wound.
Free skin grafts are much more practical as grafts are most commonly taken and transferred to one patient (also termed an autograft). They can also be implanted into the wound as island grafts, or applied to the wound surface as sheet grafts.
Free skin grafts include pinch, punch, tunnel, meek, full-thickness sheet (meshed or unmeshed) and split-thickness sheet (meshed or unmeshed). This article focuses on pinch and punch.
The graft adheres to the recipient site by fibrin. Vessels and fibroblasts invade the fibrin clot within 48 to 72 hours and the graft should be firmly attached to the wound bed 10 days after implantation.
Plasmatic imbibition, during which the graft becomes oedematous until re-vascularisation occurs, nourishes the graft.
After 48 hours, capillaries cross the fibrin clot and anastomose with those in the graft before neovascularisation.
Re-vascularisation is established four to five days after implantation and lymphatic circulation is re-established at seven days.
The graft epidermis may die after implantation, exposing the pale dermis of the graft underneath. This is pale pink in colour, either similar or paler than the surrounding granulation tissue.
Epithelial cells migrate from the hair follicles in the graft and quickly cover it, and split-thickness grafts remain covered with flaky debris for several months until the eccrine glands regenerate.
Pigmentation is generally regained after about one month and hair is usually evident four to six weeks after implantation.
Reinnervation returns in humans after seven to nine weeks, but can be patchy with hyperaesthesia possible. This is also likely the case in horses.
The graft contracts immediately after it is harvested due to recoil of the elastin fibres within the graft.
Skin grafts in horses seem to stimulate contraction of the wound – the opposite effect seen in humans and other species.
The exact pathway for this mechanism is unclear, but is thought to be partly due to a reduction in the exposed granulation tissue – this results in a reduction in inflammation, which increases the rate of wound contraction and epithelialisation.
This “contraction effect” is reduced when:
Grafts will be accepted if the recipient site is free from infection and necrotic tissue, and is well vascularised. Fresh wounds, therefore, accept grafts most readily, although most equine wounds do not often lend themselves to grafting at this stage.
Non-infected granulation tissue remains the most common graft bed and has the advantage that a degree of contraction is likely to have occurred already, thus reducing the wound’s size.
Most non-infected tissue can accept grafts, except:
Tissues – such as adipose tissue, joint capsules and ligaments – are less likely to accept grafts. However, grafts can still be placed over tendons present within a wound as a process, as vascular bridging may occur.
This is where the vascular supply to the graft arises from the adjacent vascular granulation tissue.
Full-thickness grafts are more likely to survive in this situation compared to partial/split thickness grafts; however, the differential movement of a tendon or a ligament within a grafted granulation tissue bed will have a detrimental effect on overall graft survival.
The most common cause of graft failure is infection. However, inflammation, fluid accumulation underneath the graft and movement are also significant causes of failure.
Most bacteria (of particular relevance is β-haemolytic streptococci) produce proteolytic enzymes that catalyse the conversion of plasminogen to plasmin.
This causes fibrin to degrade, destroying the graft’s initial fibrinous attachment to the wound.
Pseudomonas species produce elastase, which damages the elastin in the dermis of the graft, to which the fibrin attaches.
Wound beds will never be completely free from bacteria, but the abundant blood vessels in the granulation tissue, and the phagocytic cells in the tissue, act as a barrier to bacterial invasion and colonisation.
Chronic inflammation causes the production of poor-quality granulation tissue, which, in turn, produces a moderate amount of purulent exudates – factors that have a negative effect on graft acceptance.
This is of more importance in adult horses compared to ponies, as the latter are less affected by this chronic inflammatory state.
Fluid accumulation beneath the graft – in the form of blood, serum or exudate – prevents the formation of the fibrinous attachment of the graft to the wound bed, obstructing vascularisation.
Grafts can survive for a few days through plasmatic imbibition. However, if capillaries between the wound bed and the graft are not established after a few days, the graft will die.
Because of this, it is important for haemorrhage to have stopped before the graft is embedded in the wound.
Any movement that disrupts the formation of the vascular supply and the stable fibrinous connection will reduce graft survival.
The most common form of motion is the shearing forces produced between the wound and the overlying bandage.
The use of splints and casts should, therefore, be carefully considered to reduce the movement of the distal limb, particularly if grafting has taken place over a highly mobile area, such as a joint.
The presence of mobile structures in the granulation tissue, such as exposed tendons and ligaments, will also affect graft survival. Horses should not be turned out during healing; they should remain on box rest.
Ensuring the recipient site is as well prepared as possible will optimise graft survival. As aforementioned, the area must be free of infection and necrotic tissue, and must be well vascularised.
Fresh tissue accepts grafts more readily than granulation tissue, but grafting is rarely feasible at this stage for equine wounds for many reasons, including the presence of too much necrotic or infected tissue.
New granulation tissue also accepts grafts more readily than mature granulation tissue, as vascularisation diminishes in the latter.
Many factors relating to the recipient site need to be considered before grafting is carried out.
A wound is assumed to be infected if it shows signs such as localised redness, swelling and the formation of purulent exudate.
However, even if the localising signs of infection are not present, some wounds still contain an infective concentration of bacteria.
Streptococcal infection is generally readily resolved using β-lactam antibiotics, although the addition of potassium clavulanate may need to be considered if the presence of β-lactamase-producing bacteria is isolated.
Pseudomonas-infected wounds typically produce a blue-green exudate with a strong odour of grape juice.
These bacteria are generally sensitive to aminoglycoside and silver sulfadiazine antibiotics.
Topical administration of these antibiotics is generally more successful than systemic administration, as the fibrin at the base of the granulation tissue prevents adequate penetration of the drug.
The most effective way to decrease infection or chronic inflammation in a wound is excision of the granulation tissue.
In addition to the wound surface, infection of the deeper structures may also need to be considered. This could involve the presence of infected bone (a sequestrum), tendon or ligament, or the presence of a foreign body.
A draining tract may be present, which could indicate one or more of these complications exists. Consideration to debriding these structures must be given before grafting is considered.
Exuberant granulation tissue, or granulation tissue that is infected or inflamed, should be excised to a level slightly below the margin of the surrounding skin.
This removes the more mature granulation tissue, bacteria and wound debris, and provides a granulation bed that is more vascular and readily acceptable of grafts.
This procedure can be performed in the standing horse, as granulation tissue does not have a nerve supply.
A bandage should be applied following granulation tissue excision to stop the associated haemorrhage and changed daily until the new granulation bed appears ready for grafting – this is typically at least 24 hours following excision, by which stage capillaries will have started to develop.
An alternative to excision, topical corticosteroids can be applied to the granulation tissue to reduce a chronic inflammatory state.
This is more applicable to smaller wounds or wounds where the amount of mature granulation tissue is not excessive.
Corticosteroids do have a detrimental effect on angiogenesis, but this downside is offset by the reduction in inflammation.
The ideal granulation tissue for accepting free grafts should be a flat, firm, vascular red, with minimal clefts. The wound should not bleed readily and, ideally, have a slight serosanguineous or slightly purulent discharge.
The donor site for obtaining both pinch and punch grafts is clipped, cleaned with a dilute skin cleanser or povidone-iodine solution and rinsed with physiological saline.
Local anaesthetic is placed subcutaneously proximal to the site to block the selected area of skin. Grafts are harvested at about 1cm intervals and, ideally, in a symmetrical pattern to improve the donor site’s cosmetic result during healing. Donor sites include:
Punch grafts are small, full-thickness grafts obtained directly from the horse or, less commonly, from an excised sheet of skin.
They are obtained and implanted into the granulation tissue using 5mm to 8mm skin biopsy punches. The wounds created by taking the grafts are generally left to heal by secondary intention, with small, inconspicuous scars being apparent.
Occasionally, the donor sites can be sutured or stapled closed to increase the speed of healing and decrease the risk of scarring.
The excess subcutaneous tissue should be removed from the graft before implantation so the dermal vascularisation is exposed, encouraging graft acceptance.
The recipient holes in the wound granulation tissue should be made using a punch biopsy slightly smaller than the one used to obtain the grafts, to account for primary graft contraction, and should be made to a depth corresponding to the depth of the graft – it is a common mistake to make the holes too deep.
They should be made about 6mm apart and from the distal aspect of the wound, working proximally, so the haemorrhage created does not prevent visualisation.
It is important the haemorrhage has stopped before the grafts are inserted into the recipient holes, so they should be made before the grafts are obtained.
In addition, cotton bud ends can be placed into the holes to help reduce haemorrhage and aid the location for graft implantation.
The recipient holes should be flushed with a small amount of saline to remove any blood clots before the grafts are inserted. Each graft is inserted using a small haemostat or tissue forceps, in the direction of the hair.
Pinch grafts are small, partial/split-thickness grafts harvested using a scalpel blade.
A small cone of skin is tented with tissue forceps or a hypodermic needle and excised with a sharp blade.
The discs should, ideally, be about 3mm in diameter, as this means they will be thin towards their periphery, but at nearly full thickness towards the centre.
The epidermis and only a portion of dermis are harvested, and the grafts are placed and covered on a gauze moistened with physiological saline until they are implanted.
To implant the grafts, a stab incision using a number 15 scalpel blade or hypodermic needle can be made into the granulation tissue, working from the distal aspect of the wound, to create a shallow pocket at an acute angle. The pockets are made 3mm to 5mm apart.
The graft is pushed into the pocket, epidermal side up, using a hypodermic/suture needle or small curved haemostat.
The cosmetic appearance is poorer for pinch grafts, so consideration to the direction of the hair growth is not made.
Punch grafts resist trauma better than pinch grafts and, because they contain all the properties of the surrounding skin, produce a better cosmetic and functional result.
They also have fewer exposed vessels, so their requirement for nourishment post-implantation is greater.
They are, therefore, not accepted as readily and the circumstances for graft retrieval (including removing any excessive subcutaneous tissue from the graft) and implantation must be ideal.
In ideal conditions, 60-75% of punch grafts survive, but pinch graft survival is more variable (50-75%). Punch grafts are, therefore, often preferred, despite the procedure being slightly more skilful to perform.
Pinch and punch grafting are easy techniques that do not require expensive equipment and can be achieved in standing, sedated horses. The key to its success is timing.
The recipient bed must consist of noninfected granulation (or fresh) tissue free from necrotic material and debris.
Excision of mature/excessive granulation tissue is paramount for graft acceptance to be maximised; this will encourage wound contraction to occur following graft implantation.
The significant advantages of wound grafting include reduced convalescence, improved cosmetic result and lower costs for the owner.
