Gerald S. Lazarus

Transplantation of Psoriatic Skin onto Nude Mice

Involved psoriatic epidermis maintains its histologic appearance, increased labeling index, and increased level of plasminogen activator after grafting onto athymic nude mice. Epidermis from clinically uninvolved psoriatic skin develops an increase in plasminogen activator activity after grafting for 6 weeks on nude mice and demonstrates an increased labeling index. Normal control skin maintains its low level of plasminogen activator and labeling index after grafting. These results indicate that psoriatic skin can maintain and develop markers of psoriatic skin independent of the host.

Characterization of a chemotactic and cytotoxic proteinase from human skin

A proteinase (EC 3.4.-.-) active at physiological pH has been isolated from human skin utilizing gel filtration and affinity chromatography techniques. The proteinase has a molecular weight of approx. 28 000 and it is inhibited by alpha 2-macroglobulin, alpha 1-antitrypsin, C-1 inactivatory, soybean trypsin inhibitor and diisopropyl fluorophosphate. 2njection of 1 ng of purified proteinase into rabbit skin induces polymorphonuclear leukocyte infiltration of the cutis. Inhibition of enzyme activity with diisopropyl fluorophosphate inhibits the chemotactic effect. Addition of 0.2 microgram/ml of purified proteinase to fibroblast cultures kills the cells within minutes. This proteinase may play a significant role in modulating the inflammatory response after cellular injury.

Proteinases of human epidermis; a possible mechanism for polymorphonuclear leukocyte chemotaxis

Three neutral proteinases (EC 3.4.--.--) and cathepsin D have been identified in human epidermis utilizing a highly sensitive radioactive method. The proteinases were extracted in 1.0 M KC1 and 0.1% Triton X-100 and separated by Sephadex G-75 chromatography. The neutral proteinase peaks were all inhibited by diisopropyl fluorophosphate and thus were serine proteinases. Incubation of the enzyme fractions with [3H] diisopropyl fluorophosphate followed by sodium dodecyl sulfate polyacrylamide gel electrophoresis demonstrated that the two larger molecular weight proteinases were enzyme mixtures. The small molecular weight [3H] diisopropyl fluorophosphate proteinase migrated as a single band. Injection of the small molecular weight neutral proteinase into rabbit skin produced a polymorphonuclear leukocyte infiltration and edema. The reaction was not observed with the diisopropyl fluorophosphate-inhibited enzyme fraction. The release of neutral proteinases may be one of the signal events in the epidermal inflammatory response.

Minidose heparin therapy for vasculitis of atrophie blanche

Atrophie blanche vasculitis is due to thromboocclusion of dermal blood vessels. A 28-year-old white man with a very severe case of this disease had a complete remission when he was treated with minidose heparin sodium injections. As little as 5,000 U of heparin every 3 days controlled his vasculitis. Further reduction of heparin dose resulted in an exacerbation of disease which was controlled with more vigorous heparin therapy. The role of minidose heparin in cutaneous disease is discussed.

Cellular serine proteinase induces chemotaxis by complement activation.

INFILTRATION of inflamed or injured tissues by polymorphonuclear leukocytes is a fundamental pathophysiological response. Undoubtedly, there are numerous mechanisms by which leukocytes are attracted to an area of damage. One possibility is that cellular injury could release or activate a proteolytic enzyme which could generate chemotactic factors. Extracts of parenchymatous tissues1 and cells2 are chemotactic when they are incubated with complement sufficient serum. Macrophages3 and leukocytes4 contain a proteinase which generates chemotactic peptides, and recently human polymorphonuclear leukocytes have been shown to secrete an enzyme which activates complement5. Lazarus and Barrett extracted and characterised a serine proteinase which induced polymorphonuclear leukocyte infiltration of the skin when injected intradermally6. Subsequently, this proteinase, which is both cytotoxic and phlogistic, has been purified to homogeneity from whole human skin7, human epidermis8, human lymphocytes9 and cultures of newborn mouse epithelium (G.S.L., unpublished). This report demonstrates that this proteinase is significantly more active than trypsin, plasmin or Pronase in generating polymorphonuclear leukocyte accumulation and that chemotactic activity is largely dependent on complement activation.

Neutral proteinase of rabbit skin: An enzyme capable of degrading skin protein and inducing an inflammatory response

Abstract 1. 1. A neutral proteinase has been extracted from rabbit skin in the presence of 1 M KCl and purified 86-fold. 2. 2. The enzyme was active at neutral pH; it was not inhibited by diisopropylphosphorofluoridate, thiol-blocking reagents or EDTA. 3. 3. About 85% of the activity in skin was located in the dermis, the remainder in the epidermis. 4. 4. Incubation of skin with enzyme resulted in the release of trichloroacetic acid-soluble peptides from the tissue. 5. 5. Intradermal injection of the purified neutral proteinase into rabbits produced whealing within 15 min and acute leukocytic infiltration within 18 h. It is suggested that this enzyme may play a part in the initiation of an inflammatory reaction in skin.

Penicillamine-induced pemphigus. Immunoglobulin from this patient induces plasminogen activator synthesis by human epidermal cells in culture: mechanism for acantholysis in pemphigus

The incubation of cultured epidermal cells with IgG obtained from a 56-year-old man with penicillamine-induced pemphigus resulted in an increase in extracellular and intracellular plasminogen activator. This suggests that penicillamine-induced pemphigus and spontaneously occurring pemphigus share common pathophysiologic processes in the induction of blister formation.

Antibody-Induced Proteinase Activation: A Proposed Mechanism for Pemphigus

SummaryThe current state of understanding of pemphigus includes the following:1.Pemphigus is an autoimmune disease. In all variants a circulating autoantibody is found which binds to epidermal cells. In vivo antibody may be found deposited in the epidermis of patients.2.The autoantibody levels generally correlate with disease activity indicating a relationship between antibody and clinical disease.3.Although complement components are found in lesional skin, complement does not appear to be necessary for dissolution of the epidermal cement substance.4.The treatment of pemphigus with corticosteroids has drastically reduced mortality rates.5.Three different groups have presented results in two different experimental systems which indicate that subsequent to binding of pemphigus antibody to epidermal cells a proteinase is activated. This proteinase(s) degrades the in-tercellular cement substance of epidermis which results in loss of cellular adhesion and acantholysis. There are numerous questions still remaining. What is the nature of the proteinase(s) and the surface protein(s) it cleaves? Does the binding of pemphigus antibody to the cell surface induce enzyme synthesis, specific enzyme activation, or generalized lysosomal secretion? The answers to these questions will have broad biologic relevance since they may elucidate the role of anticell surface antibodies in disease states.

Molecular mechanisms of pathogenesis

Abstract The first clue to the etiology of pemphigus was established in 1964, when Beutner and Jordon 1 demonstrated that serum from pemphigus patients contained autoantibodies that bound to an intercellular substance (ICS) of skin and mucosa. Subsequently, skin biopsies revealed in vivo deposition of auto-antibodies in the epidermis of pemphigus patients. 2 While these results were intriguing, they did not prove that antibody played a role in the acantholytic process. The following three lines of evidence do support a role for antibody in the induction of acantholysis in pemphigus: correlation of antibody titers with disease activity 3,4 ; induction of acantholytic lesions in the skin of neonatal mice 5 or in grafts of human oral mucosa on nude mice 6 following passive transfer of patient IgG; and induction of acantholytic lesions in expiants of normal human skin following incubation with patient IgG. 7,8 The first two lines of evidence have been discussed in other chapters of this volume. It is our intention to describe experimental results from several laboratories confirming the pathologic events induced in normal human skin by serum or IgG preparations from pemphigus patients and to discuss the molecular events leading to the pathophysiology of pemphigus.

Immunocytochemical localization of chemoattractant proteinase in whole human skin, fibroblasts, lymphocytes, and granulocytes.

A monospecific antibody to purified human skin chemotactic proteinase was raised in rabbits and shown to produce a single line of identity against crude and purified human proteinase. The antibody was used to localize the proteinase in human fibroblasts, polymorphonuclear leukocytes, lymphocytes, and sections of whole skin. All cells demonstrated particulate extranuclear staining. Whole human skin demonstrated staining of the epidermis, especially in the granular layer; particulate staining was also found in the dermis. Staining could be blocked by preincubation of the rabbit anti-human neutral proteinase with purified proteinase antigen. These data indicate that neutral proteinase is located in a number of different human cells in a particulate distribution.