Annals of Burns and Fire Disasters - vol. XX - n. 2 - June 2000

PIEDMONT FOUNDATION OF STUDIES AND RESEARCH ON BURNS: THE LAST THREE YEARS OF RESEARCH ACTIVITY

Teich Alasia S. Piedmont Foundation of Studies and Research on Burns, Turin, Italy

SUMMARY. An account is given of the main activities of the Piedmont Foundation of Studies and Research on Burns. The results of latest research are presented, together with important developments in the field of pathological scarring, new methods of skin cryoconservation, and peripheral sensory nerve regeneration after extensive burns. All the Foundation's efforts endeavour to promote close and productive links between researchers and clinicians in the field of burns in order to achieve significant advances in this important field of medical science.

The Piedmont Foundation of Studies and Research on Burns continued to support research in the field of burns related to clinical work in order to achieve its philosophy, which is to couple the needs of clinical science with a high level of basic science, know-how, and technology.
To achieve this goal the Foundation organized meetings and up-date courses, edited publications, and provided financial support for research.
Our efforts were , focused on certain main fields (pathological scarring, peripheral sensory nerve regeneration, tissue banking, and cryoconservation) together with a number of methodological projects (optimization of burn centre layouts, project for a hyperbaric chamber for burn patients, computer assisted management of a burn centre), linked by the common interest in burns.
We will briefly summarize the results of our research and in particular the development of studies in the field of pathological scarring, new methods of skin ciryoconservation, and peripheral sensory nerve regeneration after extensive burns.

Hypertrophic scarring

It is well known that wound healing is the result of a complex interaction of biochemical mediators, cells, and structure proteins that enables injured tissues to regain their original properties. When normal processes in the skin are altered, pathological scarring takes place and structural, aesthetic, and functional modifications become evident.
Post-burn pathological scarring is a very interesting field of speculation with regard both to its prevalence, which remains very high despite therapeutic advances, and to the complexity of its aetiopathogenesis, which remains obscure.
Recent studies have underlined the importance of the immune system in pathological scarring. Kelman Cohen and other researchers have demonstrated an increase of circulating immunoglobulins, the presence of anti-nucleus antibodies, and tissue immunoglobulins.
Some years ago, in collaboration with the Departments of Human Anatomy and Genetics in Turin, Italy, we initiated research that demonstrated an increase in number and morphological changes in Langerhans cells in the epithelium and dermis of pathological scar tissue, together with leukocyte infiltrates. The presence of these infiltrates has been observed in many skin diseases (i.e. lichen plarms, psoriasis, atopical dermatitis, etc.) that are characterized by the ectopical expression of HLA class II antigens on keratinocytes. The anomalous expression of HLA DR molecules and their modulation in the tissues involved are concomitant to other activation molecules such as intercellular adhesion molecule type 1 (ICAM-I), interleukin-2 receptor, and others. In the modulation of their expression a pivotal role is played by certain cytokines secreted by immunocompetent cells under different stimulation. Their controls are both a consequence and a causative agent of a cascade of reactions among cells involved in extracellular matrix remodelling.
On the basis of these considerations we decided to analyse the immunological factors of hypertrophic scarring through a study of the modulation of cells and molecules of the immune system directly in the affected tissue.
We then looked for different immunological activation markers in hypertrophic scars. We also investigated the production of certain regulatory cytokines, e.g. tumour necrosis factor (TNFa), IL-1, IL-6, interferon gamma (IFNg), and transforming growth factor (TGFb).
Briefly, our data underline that the ectopical expression of these different markers in hypertrophic scar tissue makes it possible to group it with the derimatosis caused by an altered immune response. In particular, the presence of these signals on keratinocytes further demonstrates their pivotal role, a role that hitherto has been overlooked, in the pathogenesis.
Our data also testify to the importance of fibroblasts in the evolution of scarring. The immune system thus plays a central role as regards both the presence of abundant leukocyte filtrates and their active status. In our opinion it is the altered co-operation between these cells that is responsible for the pathogenesis of hypertrophic scarring, in view of the fact that the production of cytokines, normally involved in extracellular matrix remodelling, shows a major impairment that leads to a disruption of the orchestration of messages regulating cell interactions.
It still has to be clarified whether an exogenous signal or a genetic predisposition can be considered the direct cause of hypertrophic scarring.

Analysis of cryoconservation techniques and quality controls

In recent years there has been an increasingly urgent need for cadaver skin in up-to-date treatment of extensive burns. For this reason the Foundation supported a programme for the analysis of different protocols for the cryoconservation of alloplastic skin. In particular, some studies were conducted for the assessment of tissue viability after freezing in order to standardize protocols for the harvesting, transport, and stocking of tissues in a regional tissue bank.

Peripheral nerve regeneration

The sensory recovery of transplanted skin is a fundamental feature that signifies the functional success of reconstruction. Sensory regeneration was discussed for many years: some researchers believed that clinical sensory recovery was due to structures surviving in the wound bed (like feeling through a glove), others suggested the regeneration of new structures in the transplants. One much discussed point was the survival of capsulated receptors and their reinnervation. Various conflicting data were obtained in different experimental conditions using unselective and repetitive histological techniques (heavy metal impregnation).
Research conducted in collaboration with the Department of Human Anatomy of the University of Turin led to the implementation of an immunchistochemical technique for the detection of a selective neural marker that repetitively stains all nerve structures in both normal and transplanted skin. This is a structural protein of all classes of nerve fibres, including autonomic fibres. Staining with this marker made it possible to demonstrate the presence of intra-epithelial fibres that are not yet clearly evident and to confirm the classification of capsular receptors (Meissner's corpuscles and simple coiled corpuscles, while all the others that are classically reported have to be regarded as artefacts). Our studies on scars, skin flaps, and skin grafts showed evidence of the regeneration of nerve structures as far as the superficial layers, but with a lower density than that of normal skin. Fibres spread from a deep dermal plexus into the dermal papillae, mainly terminating as dermal nerve endings or, less frequently, in contact with Merkel cells or as free intra-epithelial nerve endings. Skin flaps have been shown to contain capsulated receptors, although these were aberrantly reinnervated.
These findings conflict with the widely held opinion that scars and transplanted skin fail to regenerate and that sensory recovery depends uniquely on structures surviving in the wound bed.

Conclusions

Our efforts are thus aimed at establishing a close and productive relationship between researchers and clinicians in the field in order to achieve results of the greatest scientific significance in the stimulating field of burns.

RESUME. L'Auteur décrit les activités principales de la Fondation Piémontaise des Etudes et de Recherche sur les Brûlures. Les résultats des recherches les plus récentes sont présentées, comme aussi les développements importants dans le secteur de la cicatrisation pathologique, les nouvelles méthodes de la cryoconservation, et la régénération des nerfs sensoriels périphériques aprés les brolures étendues. Tous les efforts de la Fondation cherchent à promouvoir des rapports étroits et efficaces entre les chercheurs et les cliniciens dans le secteur des brûlures pour obtenir des progrés significatifs dans cet important secteur de la science médicale.

1. Stella M., Castagnoli C., Magliacani G., Richiardi P.: Fisiopatologia della cicatrizzazione patologica. (Revisione Bibliografica.) Riv. Ital. Chir. Plast., 21: 199-208, 1989.
2. Cohen LK., McCoy T., Mohanakumar T., Diegelmannn R.F.:Immunoglobulin, complement and histocompatibility antigen studies in keloid patients. Plast. Reconstr. Surg., 63: 689, 1979.
3. Cracco C., Stella M., Teich Alasia S., Filogamo G.: Comparative study of Langerhans cells in normal and pathological human scars. 11. Hypertrophic scar. Eur. J. Histochem., 36: 53, 1992.
4. Janssen de Limpens A.M.P., Cormane R.H.: Studies on the immunologic aspects of keloid and hypertrophic scars. Arch. Dermat. Res., 274: 259-64, 1982.
5. Castagnoli C., Stella M., Berthod C., Magliacani C., Momigliano Richiardi P.: TNF production in hypertrophic scarring. Cell Immunol., 147: 51-4, 1993.
6. Castagnoli C, Stella M., Magliacani G., Richiardi P.: The role of TNF alpha and beta cytokines in scar hypertrophy in burn patients: An inummohistochernical study. Arm. Medit. Burns Club, 8: 23-27, 1995.
7. Peruccio D., Castagnoli C., Stella M., D'Alfonso S., Momigliano Richiardi P., Magliacani C., Teich Alasia S.: Altered biosynthesis of TNF alpha is involved in post-burn hypertrophic scars. Burns, 20: 118-21, 1994.
8. Castagnoli C., Stella M., Magliacani G., Ferrone S., Momigliano Richiardi P.: Similar ectopic expression of ICAM-1 and HLA Class 11 molecules in hypertrophic scars following thermal injury. Burns, 20: 430-3, 1994.
9. Castagnoli C., Trombotto C., Stella M., Calcagni M., Magliacani G.: Funzione e organizzazione di una Banca della Cute. Atti 11 Congresso Associazione Italiana di Immunogenetica c Biologia dei Trapianti, June 1995.
10. Ranieri G., StelIa'M., Calcagni M., Teicb Alasia S., Cellino C., Panzica G.: Morphology of corpuscular receptors in hairy and nonhairy hurnan skin as visualized by antiserum to protein gene product 9.5 compared to anti-neuron-specific enolase and anti-S100 protein. Acta Anat., 144: 343, 1992.
11. Calcagni M., Stella M., Ranieri G., Cellino G, Panzica G.: Fibre nervose e recettori sensitivi nella cute urnana evidenziati con anticorpo antiprotein gene product (PGP) 9.5. Riv. Ital. Chir. Plast., 25:215,1993.