Annals of Burns and Fire Disasters - vol. IX - n. 3 - September 1996

USE OF CULTURED HOMOLOGOUS KERATINOCYTES IN THE LOCAL TREATMENT OF LVELUS SYNDROME

Napoli B., D'Arpa N., Masellis M., D'Amelio L., Genovese M.

Divisione di Chirurgia Plastica e Terapia delle Ustioni, Ospedale Civico, Palermo, Italy

SUMMARY. The authors describe two cases of toxic epidermal necrolysis (Lyell's syndrome). The first case did not have a clear pharmacological aetiology, occurring in a patient with a lymphoproliferative condition (non-Hodgkin's lymphoma). The second patient had taken a number of drugs and the aetiology was probably attributable to anticonvulsants (phenobarbital, phenytoin and carbarnazepinc). The cases did not present any complications and were treated with vitamins, gastroprotective and rehydrating therapy, appropriate nutritional support, without the use of steroids, and with local application of cryopreserved cultured homologous keratinocytes which promoted extremely rapid re-epithelialization of the lesions (48 h). The specific literature regarding both Lyell's syndrome and the problerns involved in the cultivation of human epithelium are reviewed and the Authors find confirmation of the biological foundations and the validity of the local treatment which they applied.

Introduction

Lyell's syndrome resembles a partial-thickness burn because of the presence of erythema and the formation of bullae, the surface of which consists of the epidermis, with subsequent detachment exposing the dermal layer of the cutis.
The modifications of the skin covering that occur in Lyell's syndrome require general and local treatment analogous to that required in burns. For this reason toxic epidermal necrolysis (as Lyell's syndrome is also known) is a pathology that is often treated in Burn Centres.
The purposes of the local treatment of Lyell's syndrome are:

• to reduce the intensely painful burning feeling
• to prevent heat and fluid loss to prevent bacterial contamination and septic processes to prevent the lesions from becoming deeper
• to promote re-epithelialization

The methods of local treatment described in the literature (air-fluidized bed,2 hyperbaric oxygen therapy,' antibiotic and antiseptic treatment,'-' skin substitutes` 14) are not always sufficient to achieve these purposes. Barring complications, re-epithelialization generally occurs within two weeks (as in partial-thickness burns) and never in less than a week.

Clinical cases and methods
We describe here two recent cases of Lyell's syndrome in which we successfully used cultured homologous keratinocytes for topical treatment.
The homotransplant donors were tested and found negative for Toxo IgG and IgM, CNIV IgG and IgM, HbsAg and HCV, HIV 1+2, and TPHA.
The patients were subjected to the removal of cutis in negative Nikolsky's sign areas for laboratory culture and conservation of autologous keratinocytes to be used if necessary as an autologous dressing in the post-acute phase of the disease.
Patient with non-Hodgkin's lymphoma diagnosed histologically in 1994. In December 1995, following the appearance of large submandibular swellings, the patient was given radiation therapy (15 sessions) and pharmacological treatment (dexamethasone 4 mg per day for 45 days). On 24 March 1996, two days after the last radiation session, the appearance was observed of rubescence and oedema with subsequent de-epithelialization in the face and neck, progressively involving the trunk and upper limbs. The patient was admitted to the Dermatology Department and diagnosed as suffering from Lyell's syndrome. On 31 March 1996 he was transferred to our division.

Objective examination on admission
The face presented lesions in the healing phase; zones of erythema and de-epithelialization were present in the neck, upper limbs, trunk, and lower limbs (Fig. 1). Nikolsky's sign was positive. The visible and explorable mucosae were not affected.

Fig. 1 Fig. 2 Fig. 3 Fig. 4

Diagnosis
The skin biopsy performed on I April 1996 provided histological confirmation of the clinical diagnosis. The lymphocyte typing was also compatible with the diagnosis (reduced T lymphocytes with reduced helper- suppressor ratio). Activated lymphocytes present. Increased NK lymphocytes (CD 16).

Clinical course and treatment
During the course of the disease the patient did not present any blood gas analysis modifications and chest radiography was normal. Electrolytes remained normal, as also kidney function (urea and creatininaemia) and liver function (transaminase, bilirubin, alkali phosphatase).
On admission the patient presented leucopenia (WBC x 10'/ml 2.53) with neutropenia (62.9%) and thrombocytopenia (P1t x 101/tul 83). During the course of the disease there was a progressive increase in these values towards normalization when the patient was discharged (WBC x 101/ml 5.86; neutrophils 72.5%; P1t x 101/mI 123).

Haemocoagulation remained within the normal range.
Swabs from the right hand and neck developed germs which generally contaminate Lyell's syndrome lesions, i.e. strains of Staphylococcus aureus and epidermidis B-1actamase-producing and non-producing.
The patient was given vitaminic, antibiotic and gastroprotective treatment together with heparin prophylaxis and rehydrating therapy.

Local treatment
On 1 April 1996 cultured homologous keratinocytes were applied on some de-epithelialized zones.
The zones presenting only macular erythema were dressed with dry petrolatum gauze and Desogen tincture. On the same day skin was removed from a zone with negative Nikolsky's sign and sent to the laboratory for culturing.
On 3 April the zones dressed with keratinocyte cultures were found to be completely re-epithelialized.The patient was dismissed on 10 April 1996. Nikolsky's sign was negative in all body areas.

Case 2 C.M., female, aged 57 yr
The patient, admitted on 31 March 1996 to the Neurology Department and subjected to CT scan, presented a roundish formation in the right posterior parietal cortical area surrounded by a clearly visible perilesional oedematous condition. The patient was given anti-oedema therapy (mannitol, furosemide and dexamethasone) together with gastroprotective therapy (ranitidine) and anticonvulsants (phenobarbital).
On 1.0 April 1996 the patient was transferred to the Neurosurgery Department, where the same therapy was continued, with the exception of steroids, but with double the dosage of phenobarbital (Gardenal 100 mg, two tablets daily).
Four days later, following the appearance of a diffuse erythematous reaction thought to be of allergic origin, therapy was continued with steroids (betamethasone, Bentelan, 4 mg, 1 phial twice daily) and antihistamines (dexchlorpheniramine-polaramin AR, one pill twice daily).
As phenobarbital was considered to be the cause of the reaction, it was suspended and replaced by phenytoin (Dintoina 100 mg, two tablets daily), with the addition of carbamazepine (Tegretol 200 rng, two tablets daily).
On 30 April the patient presented a new diffuse erythematous reaction which persisted for several days, despite the resumption of steroid and antihistaminic treatment and the interruption of anticonvulsant therapy.
On 11 May the patient was transferred to the Dermatology Department, where the skin rash became more complicated with the appearance of diffuse bullae and the characteristic features of Lyell's syndrome.
Four days later (15 May 1996), the patient was transferred to our department.

Objective examination on admission
The patient presented an intense and diffuse macular erythema in the lower limbs, with a lesser degree of erythema in the abdomen; there was disintegration of the epidermis in the glutei, arms, and limited areas of the legs (Fig. 5). Nikolsky's sign was positive in parts affected by macular erythema. The visible and explorable mucosae were not affected.

Fig. 5 Fig. 6 Fig. 7 Fig. 8

Diagnosis
The clinical diagnosis was confirmed by biopsy on 16 May. The results of lymphocyte typing were also consistent with the clinical and histological diagnosis (reduced T lymphocytes with normal helper- suppressor ratio; increased NK lymphocytes [CD 16]).

Clinical course and treatment
This case, like the first, presented an uncomplicated clinical course. Leucocytes and platelets remained within normal limits. Swabs from the gluteal regions developed strains of Acinetobacter and Staphylococcus epidermidis not producing B-1actarnase.
The patient was given vitaminic and gastroprotective treatment, heparin prophylaxis and rehydrating therapy. No antibiotics were administered and steroid treatment was gradually reduced and in the end suspended.

The patient fed regularly per os.
On 18 May 1996 infusion therapy was suspended and the patient was encouraged to leave her bed.

Local treatment
On 15 May 1996 (the day of admission to our Department) some de-epithelialized zones were dressed with cultured homologous keratinocytes.
The zones with macular erythema and initial formation of small bullae were dressed with dry petrolatum gauze and Desogen tincture. On 16 May skin was removed for culture from a zone with negative Nikolsky's sign. On 17 May the zones treated with keratinocytes were found to be completely reepithelialized.
The patient was discharged on 27 May 1996. Nikolsky's sign was negative in all parts of the body.

Discussion

The local treatment of Lyell's syndrome varies considerably. Apart from hyperbaric oxygen therapy, which is preferred by reanimators, the choice of treatment ranges from antiseptics and antibiotics to biological or synthetic skin substitutes.
Antiseptics and antibiotics applied on extensive areas may cause problems of toxicity due to absorption and some, e.g. silver sulphadiazine, may even induce Lyell's syndrome; while skin substitutes present problems of application and require perfect adherence, without which they are ineffective.
The literature relative to Lyell's syndrome does not mention the use of cryopreserved cultured homologous keratinocytes.
However, bibliographical research into the applications of cultured skin provides evidence for the biological confirmation of the validity of its use.
De Luca and Cancedda.` found that when burned areas were covered with homologous keratinocytes the take is only apparent, and that what in fact occurs is the proliferation and migration of residual keratinocytes in the lesion. Instead of "take" they use the term "epidermal regeneration", which better describes the real clinical effect of the application of homologous keratinocytes (a biological dressing). The stimulus for epithelial regeneration comes from the intense secretory activity (cytokines, hormones and numerous growth factors) performed by the homograft sheets; this is the reason for the rapidity of re-epithelialization.
This is the basis found in the literature of the clinical application of cultured homologous keratinocytes in pathological forms such as ulcers in the lower limbs, dermo-epidermal graft donor areas, sandwich technique with widemesh net graft, and superficial and deep dermal burns.`
It was therefore concluded that, like superficial burns, Lyell's syndrome, with its superficial epithelial necrosis leaving the dermis intact, could benefit from the use of cultured homologous keratinocytes.

Conclusion

Previously we had treated cases of Lyell's syndrome in various ways (using antiseptics, antibiotics, dry petrolatum gauze, and synthetic skin substitutes) with results comparable to those in the literature.
In the two cases we describe here, both without complications and not serious, apart from the problem of the advanced age of the first patient, local treatment with cultured homologous keratinocytes determined re-epithelialization of the affected areas in only 48 hours and led to complete healing. Both patients were placed on air-beds (Mediscus).
We would make the following conclusions:

1. Cultured homologous keratinocytes should be taken into consideration in the local treatment of Lyell's syndrome (which we believe should be conservative treatment) in a burns centre with a laboratory for cell culture.

2. Skin should be removed immediately after admission of the patient from areas with negative Nikolsky's sign for the culturing of autologous keratinocytes. These can then be applied in patients with complications in the chronicizing phase, in slow-healing areas (e.g. gluteal regions or other pressure areas) or in cases of recidivation of the disease.

SUMMARY. Les Auteurs décrivent deux cas de nécrolyse épidermique toxique (syndrome de Lyell). Le premier cas s'agissait d'un patient sans étiologie pharmacologique évidente qui présentait une condition lymphoproliférative (lymphome non hodgkinien). L'autre patient avait pris divers médicaments et la cause probable de la maladie était les anticonvulsivants (phénobarbital, phénytoïne et carbamazépine). Pour le traitement des deux cas, non compliqués, les Auteurs ont pratiqué une thérapie vitaminique, gastroprotective, réhydratante, avec un support nutritionnel approprié, sans l'emploi de stéroïdes et, localement, l'application d'une couverture de kératinocytes homologues cultivés et congelés qui a déterminé la réépithélialisation des lésions avec une rapidité impressionante (48 h). Les Auteurs, après avoir examiné la littérature spécifique sur le syndrome de Lyell et celle qui s'occupe des problèmes de la culture de l'épithélium humain, ont trouvé la confirmation des bases biologiques et de la validité du traitement local qu'ils ont employé.

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