homologous pericardium as a biological dressing for treatment of distal limb wounds in horses: an Experimental study

HOMOLOGOUS PERICARDIUM AS A BIOLOGICAL DRESSING FOR TREATMENT OF DISTAL LIMB WOUNDS IN HORSES - AN EXPERIMENTAL STUDY¹

Maria Célia Ramos Bellenzani²

Júlia Maria Matera³

Márcia Regina Giacóia⁴

BELLENZANI, M.C.R.; MATERA, J.M.; GIACÓIA, M.R.– Homologous pericardium as a biological dressing for treatment of distal limb wounds in horses: an experimental study. Acta Cir. Bras., 13(4):238-42,1998.

SUMMARY: This study evaluates the effects of homologous pericardium as a biological dressing for treatment of distal limb wounds in horses. Five adult horses were put under general anesthesia and a circular wound was created over each metacarpus. The left limb wound was covered with a synthetic semi-occlusive dressing and the right limb wound was covered with pericardium stored in 98% glycerin. Fixation of the membrane to the wound margins was done with cyanoacrilate glue. Bandage changes and data collection were carried out every 72 hours, until spontaneous detachment of the pericardium occurred. Data evaluated were the aspect of the granulation tissue, the behaviour of the membrane and the wound area variation, assessed under digital image analysis. Anatomo-pathological study and non-parametric statistical analysis were also performed. Pericardium had a positive effect on the granulation tissue, but permanent occlusion of exsudative wounds cannot be recommended.

SUBJECT HEADINGS: Wound healing. Granulation tissue. Pericardium. Wounds. Horses.

INTRODUCTION

As commented by SILVER¹², the treatment of a wound should complement, rather than compete with the natural healing process.

In the search for the key to ideal wound healing, a wide range of synthetic and biological dressings has been tested. Bovine pericardium was used in guinea-pigs with promising results, which favoured its employment over xenologous implants, such as pigskin⁴.

In horses, the formation of exuberant granulation tissue following distal limb wounds in horses is very common and problematic, making such wounds a matter of great concern.

The purpose of this study was to evaluate homologous pericardium as a biological dressing in treatment of surgically created, full-thickness lower limb defects in horses, hoping to contribute with an additional alternative to control the formation of exuberant granulation tissue in this species.

METHOD

Five healthy adult horses of mixed breeds and different ages were used. They were kept confined and fed pellets, coast-cross hay and salt. Each horse was previously dewormed with Pirantel/Trichlorfon(Puriequi composto - Purina Nutrimentos Ltda.).

The day before surgery, both frontlimbs were clipped and antibiotic prophilaxis "Penicilin ⁽Pentabiótico veterinário - Fontoura-Wyeth)" was started. Horses were sedated with Romifidine (Sedivet - Boehringer - Ingelheim) and endotracheal intubation was obtained by IV infusion of Glyceril Guaiacol Ether (Henrifarma S. A.) and Midazolan (Dormonid - Produtos Roche Químicos e Farmacêuticos S.A.) in 500 ml of a 5% glucose solution, followed by a Ketamine (Vetarnacol - Laboratórios König S.A.) injection; anesthesia was mantained with Halotane (Halotano - Hoescht AG) and oxygen.

Wound demarcation was made in the mid third of the the dorso-lateral aspect of the metacarpus, with an x-ray film circular template of 2,5 cm diameter. The skin was incised with a scalpel blade and dissected away with curved scissors. Bleeding was controled by counterpressure. Wounds were covered with a three layers standard dressing consisting respectively of ADAPTIC (Adaptic - Johnson & Johnson), absorvent material (Intimu's Gel - Kimberly - Clark Kenko Ltda.) and elastic cotton wrap (Cremer) and were kept under pressure bandages.

Bandages were removed 24 hours after surgery and the wounds irrigated with saline solution, identified and photographed. Left limb wounds (A) were covered with the standard dressing and right limb wounds (B) were covered with pericardium and the standard dressing. Horses received Isopirine/Fenilbutazone (Tomanol - Boehringer Ingelheim) injections for three days and andantibiotic medication was continued to complete 72 hours.

The pericardium was collected from horses dead of non-infectuous causes, cleaned and stored in 98% glycerin at room temperature, for a minimum of 30 days. The fragment to be used was cut approximately 1cm broader than the wound, rinsed in saline and rehydrated in Penicilin antibiotic solution for 15 minutes. It was then dried between gauze swabbs and fixated to the wound margins with methyl-cyanoacrilate glue (Super Bonder - Loctite). A free distal 1,5 cm space was intentionally left for drainage.

Bandage changes were done at 72 hours intervals and photographs were taken at weekly intervals, until spontaneous detachment of the pericardium occurred. By this time, both wounds were cleansed and covered with a standard dressing. In the following day, bandages were removed and the wounds photographed. A biopsy specimen was then collected and the experiment was ended.

At each bandage change, the macroscopic aspect of the wounds was subjectively accessed, noting the evolution of the granulation tissue and the behaviour of the pericardium. The initial and final area of each wound were estimated by digital image analysis of the pictures taken. The percentage of increase or reduction in total area was then calculated. Digital statistic analysis (BIOSCAN 4.0 - Optimas) was performed by application of the Mann-Whitney non-parametric test (GRAPHPAD Instat tm. - Graphpad software V2.01).

RESULTS

Pericardium was inexpensive, easy to obtain, store, prepare and applicate to the wound margins. Animals showed no signs of disconfort or significant inflammation. ADAPTIC was satiafactory regarding non-adherence and permeability, but required fixation to the second layer of the dressing to prevent slippage from the wound surface. The sanitary napking was a practical and efficient alternative.

Data regarding time of evolution, behaviour of the membrane and granulation tissue are shown in Tables I and ^II. Alltohugh most wounds developed exuberant granulation tissue, pericardium showed some benificial effects, since granulation tissue was less exuberant and more uniform where the wounds were covered by the membrane. In wound 1B, the formetion of non-exuberant granulation tissue was coincident with prolongued integrity of the membrane (Figures 1 and 2).

TABLE I
– Behaviour of pericardium over wounds B. São Paulo, 1998.

Animal	Permanence over wound	Comments
1	31 days	Good covering of the wound throughout the experiment.
2	13 days	Rupture of the membrane + partial exposure of the superior half of the wound bed on day 7.
3	13 days	Rupture of the membrane + partial exposure of the superior half of the wound bed on day 7.
4	19 days	Rupture of the membrane + partial exposure of the superior half of the wound bed on day 7.
5	16 days	Rupture of the membrane + partial exposure of the superior half of the wound bed on day 13.

TABLE II – Final aspect of the granulation tissue in wounds A and B. São Paulo, 1998.

Animal

Granulation Tissue – Wound A

Granulation Tissue – Wound B

1

Exuberant, irregular, soft

Non-exuberant, uniform, firm

2

Exuberant, irregular, firm

Exuberant, irregular, firm

3

Exuberant, irregular, soft

Exuberant, uniform, firm (superior ½) Non-exuberant, uniform, firm (inferior ½)

4

Exuberant, irregular, soft

Exuberant, irregular, soft (superior ½) Less exuberant, uniform (inferior ½)

5

Exuberant, irregular, soft

Exuberant, uniform, firm (superior ½) Less exuberant (inferior ½)

Fig. 1
: Exuberant granulation tissue in wound 1A (standard dressing only).

Fig. 2
: Healthy granulation tissue in wound 1B (pericardium and standard dressing).

In animals 1 and 3, there was a reduction in the area of the wounds treated with pericardium. In all other cases, both wounds increased in area, allthough this was less marked in the ones treated with pericardium. Statistical data analysis, however, showed non-significant values.

The anatomo-pathological study showed a more intense inflammatory reaction in treated wounds (Figures 3 and 4).

Fig. 3
: Inflammatory reaction - wound 1A (standard dressing only). HE 40X.

Fig. 4
: More intense inflammatory reaction - wound 1B (pericardium and standard dressing). HE 40X.

DISCUSSION

Semi-occlusive non-adherent dressings are indicated for treatament of wounds in the reparative stage of healing¹³, their ideal composition consisting of a primariy non-adherent layer, a secundary absorbent and padded layer and a final protective layer. They are commonly employed as standard dressings and so have been selected for use this study.

The permanent occlusion of wounds B by pericardium minimized exposure to iatrogenic and environmental factors. It has been suggested that frequent dressing changes may impair wound healing¹⁵. In the light of this hypothesis, we believe the permanent covering by the membrane may have been responsible for our favourable results.

Abundant cleansing proved unnecessary, since the secretion accumulated under the biological dressing did not preclude the development of healthy granulation tissue. Also, the greater tendence to granulate documented in wounds A, as well as in exposed portion of wounds B, suggested that iatrogenic and environmental factors may stimulate granulation of distal limb wounds and that secretion may contain important factors to healing ^9,11,12.

As previously observed ⁴, pericardium could maintain its integrity for long periods of time. It acted as an occlusive dressing, retaining secretion to the wound surface, allthough its permeable structure and its effect on the granulation tissue suggest an incomplete occlusion.

Early membrane rupture precluded better evaluation of the dressing. Wound exsudation in the initial phase of healing rendered maintainance of a long permanence occlusive dressing difficult due to fluid accumulation. Similar complications were reported elsewhere² and were resolved by application of the dressing in previously granulated wounds. Thus, previous granulation or reapplication of the membrane may be required for best results.

The occurrence of exuberant granulation tissue in wounds treated with pericardium was not surprising, once it can be stimulated by wound occlusion^{3, 14}. Factors possibly involved would be the low local oxygen tension produced and the fluid retention, with consequent humoral factors and lactic acid producing cells accumulation^{7, 9}.

However, the less marked granulation observed under the membrane suggested an inverse relationship between granulation tissue exuberance and occlusion, confronting results reported by other authors⁸. We believe this can be related to the type of dressing, once pigskin, also an occlusive dressing, was said to stimulate granulation and at the same time impede its hypertrophy^5,10. Microenvironmental differences related to the permeability of each dressing could be the potencial factor involved¹¹.

The greater incidence of firm and uniform granulation tissue under the membrane suggests a relation between occlusion and healthy granulation. The fact that irregularity was coincident with the points of rupture of the membrane reinforces this hypotesis.

While covering the wound bed, pericardium might have imitated the beneficial effects of the scab, protecting the wound and providing a favourable environment to healing^6,15. This is supported by the behaviour of wound 1B, which was occluded for a longer period of time and coincidently had the best evolution. Poor oxygenation and dissecation of the wound surface are some disadvantages of the scab that could be reverted through its replacement by a permeable occlusive dressing¹³.

Allthough the area variation between wounds A and B was not statistically significant, results were interesting from a clinical point of view. The marked area reduction in wounds 1B and 3B and the smaller percentage of increase documented in the other wounds treated with pericardium suggest that contraction is not impaired by wound occlusion, as previously commented⁸.

It has been suggested that the application of adherent dressings over limb wounds in horses might prevent their expansion, resulting in less total area to heal¹. We believe the fixation of the membrane to the wound margins may have acted as an important mechanical factor, limiting the expansion process.

We found no definitive explanation for the greater inflammation observed in treated wounds. The membrane may have acted as a foreing body, inducing persistent inflammation, but further discussion would demand a more detailed anatomo-pathological analysis, which is beyond the scope of this study.

CONCLUSION

The pericardium can function as a biological dressing and tends to favour healthy granulation, but the application of a single long permanence dressing is not adequate for treatment of exsudative wounds. Fixation to the wound margins does not impede contraction and may hinder expansion.

Methyl – cyanoacrilate is a viable alternative for fixation of biological dressings.

BELLENZANI, M.C.R.; MATERA, J.M.; GIACÓIA, M.R.– Pericárdio homólogo como curativo biológico no tratamento de feridas na região distal dos membros em eqüinos: e studo experimental. Acta Cir. Bras., 13(4):238-42, 1998.

RESUMO: Este trabalho avalia o efeito do pericárdio homólogo como curativo biológico no tratamento de feridas cutâneas na região distal dos membros em eqüinos. Cinco animais foram operados sob anestesia geral, criando-se uma ferida circular em cada metacarpo. A ferida do membro esquerdo recebeu aplicação de um curativo sintético semi-oclusivo e a ferida contralateral foi coberta com um fragmento de pericárdio conservado em glicerina a 98%, fixado às margens da ferida com cianoacrilato. As trocas de curativo e coleta de dados foram realizadas a cada 72 horas, até o descolamento espontâneo do pericárdio. Avaliou-se as características do tecido de granulação, o comportamento do pericárdio e a variação de área das feridas, determinada por análise computadorizada de imagens. Realizou-se estudo anátomo-patológico e análise estatística não-paramétrica. Observou-se efeito benéfico do pericárdio sobre o tecido de granulação, porém concluiu-se que a oclusão permanente é inadequada para tratamento de feridas exsudativas.

DESCRITORES : Cicatrização. Granulação. Pericárdio. Feridas. Eqüinos.

Adress for correspondence

Rua Leonor de Quadros, 531

CEP 05691-020 São Paulo – SP.

Accepted for publication June 3, 1998

² Post-graduate Student, Surgery Department, FMVZ – USP.

³ Professor of Surgical Technique, Surgery Department, FMVZ – USP.

⁴ Post-graduate Student, Department of Pathology, FMVZ – USP.

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¹ Work performed at the Surgery Department of the Faculdade de Medicina Veterinária e Zootecnia da Universidade de São Paulo (FMVZ –USP).

Datas de Publicação

• Publicação nesta coleção
  14 Jan 1999
• Data do Fascículo
  Out 1998