Phenotypic characterization of skin lesions in urticaria pigmentosa and mastocytomas

In order to identify possible cellular abnormalities in human mastocytosis, sections from 13 urticaria pigmentosa lesions and 5 mastocytomas were compared with 5 normal skin specimens using histochemical, enzyme histochemical and immunohistochemical techniques. All toluidine blue-positive mast cells also reacted with FcRI and c-kit antibodies, almost all stained for tryptase, many for chymase and the myeloid workshop mast cell antibodies, few for FcRII and none for the proliferation marker Ki-67. Urticaria pigmentosa lesions contained fewer epidermal Langerhans cells and a lower percentage of avidin-positive mast cells than mastocytomas and normal skin. Mastocytomas exhibited generally weaker staining for mast cell markers and mostly lacked FcRI-bound IgE on mast cells and Langerhans cells, although the receptor was able to bind IgE in tissue sections. Most of the mast cell antibodies also reacted with other cell types. Only toluidine blue, avidin, tryptase and chymase stains were mast cell specific. Mast cells in mastocytosis thus differed only to a minor degree from normal mast cells, although distinct pathomechanisms may play a role in urticaria pigmentosa and mastocytosis.     

Evidence for altered mast cell proliferation and apoptosis in cutaneous mastocytosis

BACKGROUND: Mastocytosis presents as a focal or generalized increase of mast cells, particularly in the skin, but also in other organs. Activating mutations of KIT (formerly c-kit), the receptor of the mast cell growth factor stem cell factor (SCF), appear to play a key role in the pathogenesis of sporadic adult onset mastocytosis. However, these mutations are not present in childhood-onset and familial mastocytosis and also fail to explain the heterogeneity of adult-onset disease. Other factors such as prolonged survival of mast cells may therefore participate in causing and modulating the pathological increase of mast cells in mastocytosis. OBJECTIVES: To examine the expression of proliferation and apoptosis markers in the mast cells of cutaneous mastocytosis lesions in order to gain further insight into the pathogenesis of mastocytosis. METHODS: Lesional cutaneous biopsies from eight infants with solitary mastocytomas, five children with multiple mastocytomas, 11 children with generalized urticaria pigmentosa, 12 adults with urticaria pigmentosa, and skin from seven normal controls were used in this study. Serial sections were stained with toluidine blue to quantify mast cell numbers and with antibodies against the proliferation marker Ki67 protein, the tumour suppressor protein p53, and the inhibitor of cyclins and cyclin-dependent kinases p21WAF1/CIP1, using the alkaline phosphatase antialkaline phosphatase technique. The terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end labelling (TUNEL) method was used to assess apoptosis. RESULTS: Cutaneous mast cell counts were significantly increased in all patient sections, particularly in childhood lesions, and similarly, a small but significant increase of proliferation was found in the lesional mast cells of all patients. Enhanced mast cell numbers and proliferation was associated with a significant decrease of TUNEL staining, particularly in mastocytomas. p53 expression was highly variable, with an overall significant increase in all patient skin mast cells, whereas p21 expression was barely observed at all. CONCLUSIONS: These findings further support the concept that an imbalance of mast cell proliferation and apoptosis is prevalent in mastocytosis lesions that may account in part for the increased focal mast cell accumulation in this condition.     

Two novel mast cell phenotypic markers, monoclonal antibodies Ki-MC1 and Ki-M1P, identify distinct mast cell subtypes

Summary In order to identify more specific or selective mast cell markers, the reactivity of two monoclonal antibodies, Ki-MC1 and Ki-M1P, was studied by immunohistochemistry in two human cell lines (mast cell line HMC-1, basophilic cell line KU812), in mast cells cultured from blood precursors, in adherent mononuclear cells from peripheral blood, and in mast cells of tissue sections from 1 3 urticaria pigmentosa lesions, live mastocytomas and live normal skin specimens. Toluidine blue staining, fluorescence staining with FITC-conjugated avidin, and immunohistochemical staining (APAAP) with other mast cell reactive monoclonal antibodies, was performed for comparison. Double staining with the APAAP method, using the Ki-antibodies and toluidine blue, was also carried out. Both Ki-antibodies showed reactivity for skin mast cells, but with a different staining pattern. In addition, the Ki-MC1 antibody did not react with the cell lines, and reacted only with a few peripheral blood mononuclear cells and cultured mast cells. In contrast, the Ki-M1P antibody reacted with almost all cultured mast cells and blood mononuclear cells, but stained only about one-half of lesional and one-fifth of normal skin mast cells. Ki-M1P also reacted with many toluidine blue-negative dermal cells, particularly in urticaria pigmentosa. Ki-MC1 antibody can thus be considered as a useful additional marker for normal skin mast cells. In contrast, the Ki-M1 P antibody primarily identifies immature mast cells and monocytes/macrophages, suggesting that these cell types probably originate from the same bone marrow precursor.     

Demonstration of the high-affinity IgE receptor on human Langerhans cells in normal and diseased skin

Summary Epidermal dendritic cells of normal adult foreskin, and of lesional skin from patients with atopic eczema, stasis eczema and urticaria pigmentosa are shown to be highly reactive with two different monoclonal antibodies (29C6 and 6F7) specific for extracellular domains of the α-chain of the high-affinity IgE receptor. By their distribution pattern, the reactive cells are Langerhans cells. This is confirmed by immunoelectron microscopic demonstration of Birbeck granules in the labelled epidermal cells. Very weak staining is observed on the same cells with an antibody (TÜl) against the low-affinity IgE receptor. Pre-incubation of the sections with IgE partially blocks binding of 6F7 antibody. Langerhans cells, together with dermal mast cells, can therefore bind IgE with high efficiency, and may in this way participate in IgE-mediated cutaneous diseases.     

The bone marrow in urticaria pigmentosa and systemic mastocytosis. Cell composition and mast cell density in relation to urinary excretion of tele-methylimidazoleacetic acid

The bone marrow sections from five normal subjects and 18 patients with mastocytosis were examined to establish criteria to distinguish urticaria pigmentosa from systemic mastocytosis. Nine patients had increased numbers of mast cells in bone marrow sections stained with a long toluidine blue staining technique specific for mast cells, whereas five patients exhibited increased numbers of mast cells on May-Grünwald-Giemsa-stained smears of bone marrow. A positive correlation between the number of mast cells in sections of the bone marrow and the urinary excretion of the main histamine metabolite tele-methylimidazoleacetic acid was found. In ten of the examined bone marrow specimens, focal lesions containing mast cells, lymphocytes, and eosinophils appeared. The presence of these focal lesions together with either an increased number of mast cells in bone marrow sections and/or increased urinary excretion of telemethylimidazoleacetic acid is considered diagnostic of systemic mastocytosis. No patient exhibited myeloproliferative condition or other major hematologic abnormality.     

Absence of MHC class II antigen on mast cells at sites of inflammation in human skin

Abstract Since the presence of major histocompatibility complex (MHC) antigens has recently been reported on murine and human mast cells under various conditions, we have investigated their expression on mast cells in different types of cutaneous inflammation. Cryostat sections from lesional biopsies of patients with psoriasis, atopic eczema, chronic urticaria, lichen planus, bullous pemphigoid and urticaria pigmentosa were immunohistochemically stained with monoclonal antibodies against MHC class I and class II antigens using a double staining APAAP/toluidine blue methodology. While strongly positive staining with the antibody directed against MHC class I antigens was found on nearly all mast cells in normal skin and in inflammatory dermatoses, reactivity for HLA-DR and HLA-DQ antigens on mast cells could not be detected, except for less than 2% of cells with doubtful staining. Human mast cells therefore probably play no significant rôle as antigen-presenting cells in the conditions studied.     

C5a receptors are detectable on mast cells in normal human skin and in psoriatic plaques but not in weal and flare reactions or in urticaria pigmentosa by immunohistochemistry

The expression of the receptor for the anaphylatoxin C5a on mast cells was studied with three monoclonal antibodies directed to the N-terminal domain of the C5a receptor. Human skin was investigated by immunohistology applied to sequential 2 μm sections of acrylate-embedded tissues. All anti-C5a receptor antibodies stained c-kit ⁺ or tryptase ⁺ cells which were metachromatic after toluidine blue staining in normal human skin. The binding of anti-C5a receptor antibodies was inhibitable by a peptide representing the first 31 amino acids of the C5a receptor. A similar expression of C5a receptors was found on mast cells in chronic psoriatic plaques. However, C5a receptors were not detectable on mast cells in weal and flare reactions or in lesional skin of urticaria pigmentosa. These findings suggest that (1) anti-C5a receptor antibodies directed to the N-terminal domain of the receptor are suitable tools for the identification of mast cells in acrylate-embedded sections of human skin, (2) mast cell activation in weal and flare reactions results in C5a receptor downregulation or receptor blockade and (3) mast cells in urticaria pigmentosa lack a typical marker of normal human skin mast cells. Received: 29 November 1995     

Systemic mastocytosis

Systemic mastocytosis associated with urticaria pigmentosa seems to be strikingly more common than previously assumed. The diagnosis can be established by the investigation of bone marrow sections, whereas bone marrow smears are less reliable. Some mediators are produced by the enhanced number of mast cells; telemethyl imidazole acetic acid is the most suitable mediator to calculate the size of the mast cell pool. Investigations like this might offer an alternative to the examination of bone marrow sections in future.     

Dermatoscopic findings of urticaria pigmentosa

Mastocytosis is a rare disease characterized by proliferation and accumulation of mast cells in various organs. The maculopapular cutaneus mastocytosis is divided into three subtypes: papular/plaque variant, urticaria pigmentosa and eruptive macular telangiectasia perstans. Dermoscopic may help to better characterize the different forms of cutaneus mastocytosis. We report a 55 year-old female with urticaria pigmentosa and its dermoscopy.     

Mast cells with bilobed or multilobed nuclei in a nodular lesion of a patient with urticaria pigmentosa

Mast cells with bilobed or multilobed nuclei have only rarely been observed in the bone marrow of patients with systemic mastocytosis and in a case of subdural mast cell sarcoma. To our knowledge, they have not been reported in cutaneous mast cell disease. We report a rare occurrence of mast cells with bilobed or multilobed nuclei (atypical mast cell type II) in a nodular lesion of a 24-year-old woman with urticaria pigmentosa. The typical and atypical mast cells were confirmed by Giemsa and Leder's naphthol-AS-D-chloroacetate esterase stains and by immunohistochemical staining for tryptase and KIT protein (CD117). Although the nodular lesion with atypical mast cells did not appear to be cytologically malignant, the occurrence of atypical mast cells in a nodular lesion but not in a papular lesion might denote progression of the disease as suggested by the emergence of cells positive for p53.     

Intestinal mucosal mast cells: enumeration in urticaria pigmentosa and systemic mastocytosis

Endoscopic gastrointestinal mucosal biopsy specimens from one patient with systemic mastocytosis and five with urticaria pigmentosa (UP) were fixed with Carnoy's reagent and then stained for chloracetate esterase. The mast cell population densities were enumerated in the mucosa using cursor planimetry. Compared with controls, mast cell counts were increased in gastric and duodenal but not sigmoid mucosae. On a histological basis, systemic involvement would appear commoner in urticaria pigmentosa than is generally expected. Gastrointestinal symptoms did not relate to elevated mucosal mast cell counts.     

Cutaneous mastocytosis exacerbated by pinworms in a young boy

Cutaneous mastocytosis in children has an indolent course and undergoes spontaneous regression. Many triggering factors may cause mast cell degranulation and clinical manifestations. Knowledge of these factors is important for patients and their families. We report a case of exacerbation of urticaria pigmentosa due to mast cell degranulation caused by Enterobius vermicularis, which has not been reported before as a triggering factor.     

Adult mastocytosis: an immunophenotypic and flow-cytometric investigation

Skin mast cells of five patients with mastocytosis/urticaria pigmentosa and from four normal controls were immunophenotyped with a panel of 17 monoclonal antibodies; peripheral blood mononuclear cells from the patients were similarly labelled and analysed by flow cytometry. Cutaneous mast cells expressed leucocyte common antigen and markers of macrophage/monocyte lineage suggesting a common progenitor in the ontogeny of the two cell types. The mast cells from the patients differed from the controls, expressing HLA-DR in four cases, and this may be a feature of proliferating cells. The patients' peripheral blood T lymphocytes showed a marked increase in the activation markers HLA-DR and CD25 suggesting a contributory role, possibly via cytokines, in driving mast-cell proliferation.     

Immunohistochemical demonstration of migration inhibitory factor in different types of urticaria

Because urticarial lesions can persist for extended periods of time, we have investigated the histochemical expression of an antibody against the cytokine macrophage inhibitory factor in 23 patients with different types of urticaria. Positive staining of upper and middermal dendritic cells was noted in sections from all three biopsy specimens of acute urticaria, eight of chronic urticaria, and all six of urticaria pigmentosa lesions. In all but one biopsy specimen, endothelial cells reacted as well. In three sections (two chronic urticaria, one urticaria pigmentosa), luminal lining cells of sweat glands were also noted to stain positively. In contrast, lesional skin from all eight patients with pressure urticaria was negative, as was the clinically normal skin of all patients, with the exception of one patient with urticaria pigmentosa. The data suggest that cytokines may be involved in lesions of acute type immunologic processes and that they need not be expressed in delayed type reactions.     

Perivascular mast cells in urticaria pigmentosa

We have quantified perivascular mast cells in cases of urticaria pigmentosa, urticaria, and dermal hypersensitivity reactions. To facilitate reproducibility, the mast cells were counted for a precisely defined vessel unit. These vessel units were divided arbitrarily into those ≤55μm and > 55μm in largest diameters. Urticaria pigmentosa showed an average of 6.4 ± 1.9 and 22.8 ± 13.2 masts cells for vessel unit ≤ 55μm and ≤ 55μm, respectively. Urticaria yielded a lower number of mast cells: 1.5 ± 0.2 and 2.9 ± 0.9 for the same respective vessel units. Dermal hypersensitivity reactions revealed an average of 1.6 ± 0.4 and 2.2 ± 0.7 mast cells, and the normal skin showed 1.5 ± 0.3 and 2.4 ± 0.6 mast cells for each of the vessel units of ≤ 55μm and > 55μm. The perivascular mast cell distributions of urticaria pigmentosa are statistically different from those of urticaria and dermal hypersensitivity reactions with p < 0.0001. No statistical difference was noted between urticaria, dermal hypersensitivity reactions, and normal skin. The percentages of vessel units ≤ 55μm with ≥ 5 mast cells and vessel units < 55μm with ≥ 10 mast cells were determined for each case. The average percentage of vessel units for the former and latter in urticaria pigmentosa was 82.6% and 58.9%, respectively. Urticaria yielded 0% and 0.2%, respectively. None of vessel units in the dermal hypersensitivity reactions or normal skin contained more than 5 mast cells in vessel units ≤ 55μm and more than 10 mast cells in vessel units > 55μm. Cases of urticaria pigmentosa can be distinguished from other cutaneous eruptions containing mast cells using a simple counting technique on Giemsa stained sections.     

Serum tryptase measured with B12 and G5 antibody-based immunoassays in mastocytosis patients and its relation to histamine turnover

Serum tryptase was measured with the B12 and G5 antibody-based immunoassays in 25 adult patients with mastocytosis and in 18 controls. Twelve patients had uncomplicated cutaneous mastocytosis (urticaria pigmentosa) and 13 had urticaria pigmentosa with systemic symptoms. Tryptase levels were compared with histamine turnover estimated as urinary excretion of the main histamine metabolite tele-methylimidazoleacetic acid. Elevated B12 tryptase levels (> 20 μg/L) were found in most mastocytosis patients, including five of eight patients with only cutaneous manifestations who had a low urinary histamine metabolite excretion. This indicated a higher sensitivity for diagnosing mild mastocytosis on the basis of levels of serum tryptase as opposed to urinary methylimidazoleacetic acid. However, the serum B12 tryptase assay could not differentiate between urticaria pigmentosa patients with and without systemic disease: the measurement of histamine metabolite excretion probably reflects the mast cell burden more accurately. Serum G5 tryptase levels were generally low in both controls and mastocytosis patients.     

Dermal mast cell granules bind interstitial procollagenase and collagenase.

In order to identify structures in human skin that bind collagenase, sections from frozen or paraffin-embedded skin were incubated with either procollagenase or activated collagenase. After washing, bound procollagenase or collagenase was detected by immunofluorescence microscopy. In normal skin, procollagenase bound only to isolated granular dermal cells that were identified as mast cells on the basis of staining with fluoresceinated avidin and pinacyanol erythrosinate. When mast cells were degranulated by exposure to the ionophore A23187, extracellular granules bound procollagenase. Of various pathologic conditions examined, the highest binding of procollagenase occurred in specimens of urticaria pigmentosa. Procollagenase bound to granular cells and to abundant granules scattered throughout the dermis. Binding could be abolished by pre-treatment of tissue sections with heparinase or by pre-incubation of procollagenase with soluble heparin, suggesting that heparin is the binding agent in the granules. Activated collagenase also bound to dermal mast cells but in addition bound strongly to the dermal collagen. Enzymatic activity of activated collagenase was not inhibited by heparin in concentrations up to 10 mg/ml. There is evidence that mast cell tryptase can contribute to procollagenase activation. This study further supports a role for mast cells in collagenolysis by demonstrating that heparin from mast cells binds procollagenase and possibly serves as a reservoir for procollagenase, which may then subsequently be activated.