The role of perforin-mediated apoptosis in lichen planus lesions

Lichen planus is recognized as a T-cell-mediated disease. Histologically, it is characterized by the formation of colloid bodies representing apoptotic keratinocytes. The apoptotic process mediated by CD8⁺ cytotoxic T lymphocytes (CTLs) and NK cells mainly involves two distinct pathways: the perforin/granzyme pathway and the Fas/FasL pathway. So far, little is known regarding the role of perforin-mediated apoptosis in lichen planus. In the present study, the expression and distribution of perforin, T and NK cell subsets in the epidermis and dermis of lesional and nonlesional lichen planus skin were studied. Skin biopsy specimens from lesional and nonlesional skin of ten patients with lichen planus and eight healthy persons were analysed by immunohistochemistry. Significant accumulation of T cells, particularly of CD4⁺ and CD8⁺ subsets, was found in both epidermis and dermis of lichen planus lesions compared with nonlesional and healthy skin. There were no significant differences in the incidence of NK cells (CD16⁺ and CD56⁺) between lesional, nonlesional and healthy skin. Perforin expression was significantly upregulated in the epidermis of lichen planus lesions. In conclusion, accumulation of perforin⁺ cells in the epidermis of lichen planus lesions suggest a potential role of perforin in the apoptosis of basal keratinocytes.     

Parapsoriasis en plaques. Characterization of the cellular infiltrate using monoclonal antibodies

We report on a 65-year old woman suffering from parapsoriasis with large plaques. The cellular infiltrate was analysed by means of monoclonal antibodies and an immunoperoxidase technique. Our findings proved that the majority of the cellular infiltrate in the upper dermis is composed of helper-inducer T-cells (Leu 4+, Leu 3a+). A notable number of Leu-2a-reactive suppressor-cytotoxic T-cells associated with hydropic degeneration of the basal cell layer were found within the dermal infiltrate and the dermoepidermal interface. Large numbers of Leu-6-reactive dendritic Langerhans' cells were noted in the lower layers of the epidermis, in particular in areas where the epidermis showed focal exocytosis of lymphoid cells. Langerhans' cells, in contrast, were present in the upper portions of epidermis lacking exocytic T-cells. HLA-DR antigens were expressed on Langerhans' cells as well as on nearly all T-lymphocytes. Positive intercellular staining for HLA-DR antigens were only found in localized areas of the epidermis. Small numbers of both Leu-11B-reactive natural killer cells and cells expressing interleukin-2 receptors were seen within the basal cell layer and at the dermo-epidermal junction. Our findings suggest that a cell-mediated immune response which is directed against antigens expressed by keratinocytes may play an important role in the pathogenesis of parapsoriasis en plaques.     

Histologic characteristics of lichen planus transplanted onto nude mice and cultured in vitro

Summary Typical confluent lesions of lichen planus were transplanted onto nude mice and cultured in organ culture. The characteristic histologic appearance of lichen planus disappeared after grafting and became similar to normal skin within 6 weeks on nude mice: the dense lymphocyte infiltrate in dermis disappeared, the basal cell layer normalized, and the colloid bodies disappeared from epidermis, although some of them were found in dermis. The granular layer also normalized, but the stratum corneum remained hyperkeratotic 6 weeks after transplantation. In organ culture, characteristic histologic features of lichen planus disappeared in 3–5 days via a rapid necrosis of the upper part of the epidermis and formation of a new, normal-looking basal epidermis. These results suggest that lesions of lichen planus are primarily dependent on the influence of the host to maintain their typical histologic appearance.     

T cells and T-cell subsets in mycosis fungoides and parapsoriasis. A study of 18 cases with anti-human T-cell monoclonal antibodies and histochemical techniques

Skin lesions from 15 patients with mycosis fungoides (MF) and from three with parapsoriasis were studied immunohistochemically with monoclonal antibodies against T cells (Leu 1) and against T-cell subsets (Leu 2a, Leu 3a). Lymphoid cell reactivity was diverse among these sampled cases. In two cases of parapsoriasis and nine of MF, there was a predominance of helper/inducer (Leu-3a-reactive) cells over suppressor/cytotoxic (Leu-2a-reactive) cells. In one case of parapsoriasis and one (advanced tumor stage) of MF, there was suppressor/cytotoxic cell predominance. One case of MF showed strong reactivity for both T-cell subset markers. Four cases of MF (two plaque-stage and two tumor-stage) featured a predominant cell type in the dermis which was nonreactive for all three antibodies. The intraepidermal lymphoid cellularity was Leu-1-reactive in ten cases of MF and two of parapsoriasis. Among these 12 cases, the intraepidermal cellularity was Leu-2a-reactive in four and Leu-3a-reactive in three. The use of such studies of T-cell subsets on in situ cutaneous lymphoid infiltrates may demonstrate a correlation with cytomorphology, clinical stage, and disease prognosis.     

Plaque psoriasis vs. atopic dermatitis and lichen planus: a comparison for lesional T-cell subsets, epidermal proliferation and differentiation

BACKGROUND: T-cell infiltration in plaque psoriasis has recently been an important subject of investigation. Interestingly, comparative analyses of the disease-specific composition of the lesional T-cell infiltrate in plaque psoriasis and other inflammatory dermatoses have only sparsely been performed. OBJECTIVES: To compare plaque psoriasis vs. atopic dermatitis and lichen ruber planus with respect to T-cell subsets, epidermal proliferation and keratinization. PATIENTS AND METHODS: Biopsies were taken from untreated lesional skin of patients, six with psoriasis, six with atopic dermatitis and six with lichen planus. T-cell subsets (CD4+, CD8+, CD45RO+, CD45RA+, CD2+, CD25+), an epidermal proliferation (Ki-67) and a keratinization marker (K10) were stained immunohistochemically and quantified using image analysis. RESULTS: The high number of CD8+ T cells (52 +/- 13 cells mm(-1)) found in the psoriatic epidermis was not found in the epidermis of atopic dermatitis (9 +/- 4), nor in the epidermis of lichen planus (34 +/- 10). The other T-cell subsets in the epidermis and dermis showed no statistically significant differences between psoriasis and atopic dermatitis. In contrast to the limited presence of CD4+, CD8+ and CD2+ in the psoriatic dermis (110 +/- 19, 27 +/- 9, 127 +/- 41, cells mm(-1), respectively), more impressive numbers of these cells were observed in the dermis of lichen planus (300 +/- 53, 144 +/- 38, 272 +/- 48, respectively). CD45RO+ memory effector T-cell counts were significantly higher in the epidermis of lichen planus (39 +/- 10) than in psoriasis (19 +/- 5). Psoriatic epidermis proved to have major keratinocyte hyperproliferation (247 +/- 26 cells mm(-1) lamina basalis), as compared with atopic dermatitis (134 +/- 15) and lichen planus (128 +/- 20). Furthermore, a marked decreased expression of keratin 10 was observed in psoriasis (41% of epidermal area) contrary to atopic dermatitis (70%). CONCLUSIONS: Psoriatic epidermis exhibits a pronounced CD8+ epidermotropism with accompanying epidermal hyperproliferation and abnormal keratinization, which changes are only minimally expressed in atopic dermatitis and lichen planus. In plaque psoriasis, substantially fewer activated CD4+ and CD8+ T cells in the dermis and less CD45RO+ T cells in the epidermis are present in comparison with lichen ruber planus.     

Subpopulations of mononuclear cells in microscopic lesions of psoriatic patients. Selective accumulation of suppressor/cytotoxic T cells in epidermis during the evolution of the lesion

The age of microscopic lesions in psoriatic subjects was assessed from the stacking characteristics in the horny layer and related to type and density (cells/tissue volume) of mononuclear cells in the epidermis and the dermis determined by immunoperoxidase methods using monoclonal antibodies. Pan T cells (Lyt-2+, Lyt-3+, Leu-4+, OKT3+), T helper cells (Leu-3a+, OKT4+), T suppressor/cytotoxic cells (Leu-2a+, OKT8+), Ia+ cells and monocytes (OKM2+, BRL alpha mono+) were determined in epidermis and dermis. The psoriatic lesion was divided into regions underneath a parakeratotic and an orthohyperkeratotic/hypergranular portion of the horny layer and contrasted with perilesional and uninvolved psoriatic skin as well as with healthy skin. In the various regions and skin layers, the cell density was highest in parakeratosis and decreased toward normality with decreasing histologic abnormality. The relation between epidermal and dermal cell densities of the T-cell subsets was modified in the involved psoriatic skin with a selective preponderance of T suppressor/cytotoxic cells in the epidermis. The accumulation was present in the youngest lesion found (3 days) and cell densities were unchanged in older lesions. The findings suggests that the altered relationship in the subsets of T cells has an important role during the induction and progress of the psoriatic process in the skin.     

Sequential enumeration of peripheral blood T cell subsets in lichen planus

Peripheral blood T lymphocytes and T cell subsets were examined in fifteen patients with lichen planus prior to and for 4 months during treatment. The percentages of different T cell sub-populations were defined by indirect immunofluorescence using monoclonal antibodies OKT₃, OKT₄ and OKT₈. These are specific markers of total T cells, helper-inducer T cells and suppressor-cytotoxic T cells respectively. Decreased percentages of suppressor T cells and elevated helper suppressor ratios were observed before treatment and after 1 month of therapy. These changes had disappeared by the second month of treatment, by which time all the lesions had healed.     

Quantitative studies on nonlymphoid mononuclear cell subpopulations in cutaneous infiltrates. I. Stage-related changes of dendritic nonlymphoid mononuclear cells, Langerhans' cells, and macrophages in lichen planus lesions

In previous investigations on lichen planus, we suggested that in early lesions T4-positive cells might be antigen-specifically driven, whereas in late lesions T8-positive cells may be cytotoxic to keratinocytes. To verify this hypothesis, we investigated the following nonlymphoid mononuclear cell subpopulations in early versus late lichen planus lesions: interdigitating cells (phenotype: S100-positive, lysozyme-negative, T6-negative, M3-negative), Langerhans cells (phenotype: S100-positive, lysozyme-negative, T6-positive, M3-negative), macrophages (phenotype: S100-negative, lysozyme-positive, T6-negative, M3-positive). Interdigitating cells were moreover identified in semithin and ultrathin sections by distinctive morphological characteristics. The S100-positive/lysozyme-positive cell ratio was higher (p less than 0.01) in early lesions than late lesions. In dermis but not in epidermis (NS), of early lesions, T6-positive cells were less represented than S100 positive cells (p less than 0.025). Thus, Langerhans' cells largely predominated over interdigitating cells in epidermis, but the two populations were both represented in dermis. Lysozyme-positive and M3-positive cells, more abundant in late lesions than in early lesions (p less than 0.001), were often filled with pigment granules.     

A comparative immunohistochemical study of lichen planus and discoid lupus erythematosus

A comparative immunohistochemical study was performed on skin biopsies from 10 patients with lichen planus and 10 patients with discoid lupus erythematosus (DLE). A panel of antibodies against T lymphocytes (UCHL-1, OPD-4, CD8, CD45), B lymphocytes (L-26), granulocytes (Leu-M1), activation markers (Ki-1, LN-3), macrophages, fibroblasts and dendritic cells (FXIIIa, S-100, Mac-387, K.P-1, vimentin), endothelial cells (CD34), and epithelial cells (epithelial membrane antigen) was employed using a peroxidase-anti-peroxidase technique. The recently released CD8 antiserum required microwave antigen retrieval of formalin-fixed, paraffin-embedded tissue to label lymphocytes. The results showed many similarities in the lymphocyte subsets and macrophages between lichen planus and discoid lupus erythematosus. The most important differences between the two conditions were statistically significant increases in the number of S-100⁺ cells in the epidermis and dermis, FXIIIa⁺ cells in the dermis and CD34⁺ vessels within the inflammatory infiltrate in lichen planus.     

Detection of transferrin and C3d receptors in the skin of patients with various dermatoses

The localization of transferrin and C3d receptors in various skin lesions and normal appearing skin have been studied on sections with the PAP technique. The transferrin receptor was recognized in the lower epidermis from psoriatic plaques. Here it was more evident than in other inflammatory or hyperproliferative disorders where it was mainly detected on the basal cells. In healthy skin or lesions of lichen planus, scleroderma and ichthyosis the transferrin receptor was not detected in the epidermis. The C3d receptor was in normal skin found on the basement membrane and on elastic fibres in the papillary dermis. The basement membrane was strongly marked in pemphigoid but was not seen in lichen planus and Ehlers-Danlos syndrome. In patients with urticaria factitia, contact dematitis, psoriasis and Darier's disease the suprabasal cells also expressed C3d whereas in other dermatoses the epidermis was negative. Colloid bodies in lichen planus and GVH reactions expressed both the transferrin receptor and C3d.     

Epidermal DR+T6- dendritic cells in inflammatory skin diseases

T lymphocyte and dendritic cell subpopulations were counted in three biopsies each of endogenous eczema and pityriasis rosea and two of lichen planus and compared with previous findings in psoriatic lesions. In common with psoriasis, proportionately more CD4 T cells than CD8 T cells were DR+ in both epidermis and dermis of all lesions. In addition, total numbers of epidermal dendritic cells were significantly increased in endogenous eczema and pityriasis rosea, and variably in lichen planus lesions. Interestingly, a DR+T6- subpopulation of dendritic cells was present in varying proportions in all three skin lesion types. Electron microscopy of DR+T6- dendritic cells from psoriatic lesions, using an immunogold staining technique, showed the cells to be of the Langerhans' cell lineage. DR+T6- dendritic cells are a subpopulation of Langerhans' cells which are not specific to psoriasis, but present in the lesions of other benign, inflammatory skin conditions in which CD4 T cells are preferentially activated.     

LICHEN PLANUS: EVALUATION OF CELLS IN SKIN LESIONS AND OF T-LYMPHOCYTE SUBSETS IN BLOOD

The purpose of the present study was to examine the phenotype of cutaneous immunocompetent cells and to quantify Langerhans cells in Lichen planus by the use of monoclonal antibodies directed against T-cell populations. Helper cells (OKT4+) and Suppressor/cytotoxic cells (OKT8+) were observed in all cutaneous infiltrates, and numerous Langerhans cells were identified by OKT6, BL6, and BL2 (HLA-DR) in the epidermis and dermis. The quantification of Langerhans cells demonstrated that the number of dendritic cells in epidermis is greater in involved skin than in non-involved skin. In recent lesions, Langerhans cells are more abundant than in older lesions. The results in peripheral blood indicated a T Helper/Suppressor imbalance with a decreased T-Suppressor/cytotoxic subpopulation in patients with lichen planus diseases. In lichen planus, our results suggest an immunological reaction involving all the immunocompetent cell subpopulations with a first stage of information by Langerhans cells (OKT6+, BL6+, BL2+ (HLA-DR)) and Helper cells (OKT4+), and second stage mediated by Suppressor/cytotoxic cells (OKT8+).     

A histochemical profile of lichen simplex

Enzyme histochemical tests were performed on biopsies taken from patches of lichen simplex in eighteen patients and the results were contrasted with previous findings in psoriasis and lichen planus. The mitochondria! enzyme reactions were found to be increased within the epidermis, differing from those in lichen planus, in which there is a decreased density of reaction product for these tests. The non-specific esterase band was increased in width in lichen simplex contrasting with the absence of a discrete band in psoriasis. The DOPA reaction demonstrated the presence of positively reacting cells in the basal layer, and the ATPase reaction showed that there were many ATPase positive dendritic cells within the epidermis. It appears that in lichen simplex there is hyperplasia of all components of the epidermis in contrast to psoriasis where it seems the hyperplasia is almost restricted to keratinocytes.     

T cell subsets and Langerhans cells in lichen planus: in situ characterization using monoclonal antibodies

Skin biopsies from four patients with lichen planus were studied using monoclonal antibodies directed against T lymphocytes. Anti-T1 and anti-T3 antibodies, which react with all peripheral T cells, stained most cells in the dermal infiltrates. The majority of infiltrating cells also stained with anti-T4 and anti-T4b antibodies, which react with helper/inducer cells, whereas a minority of cells stained with anti-T8 antibody, which reacts with cytotoxic/suppres-sor cells. Surface IgM was not identified on any infiltrating cells, providing evidence against B cell participation. Intraepidermal and dermal cells with long cytoplasmic extensions stained with anti-T6 antibody in all cases, defining them as Langerhans cells or their precursors. T6-positive cells were seen in greater number than in normal control epidermis and dermis. The results indicate that well-developed lesions of lichen planus are characterized by an influx of helper/inducer T lymphocytes and increased numbers of Langerhans cells. These observations support the contention that cellular immunity is important in the pathogenesis of this disorder.     

Immunophenotyping of the eczematous flare-up reaction in a nickel-sensitive subject

An eczematous flare-up reaction, occurring at a previously involved site, which followed oral challenge with 5.6 mg of nickel in a 29-year-old nickel-sensitive woman, was biopsied and studied by immunohistochemistry. The cellular infiltrate in the dermis and epidermis at 8 days was predominantly of Leu 3a phenotype (helper/inducer T lymphocytes), with smaller numbers of Leu-2a-reactive (suppressor/cytotoxic) T lymphocytes. Many infiltrating cells were DR-positive. No increase in epidermal Leu-6-positive Langerhans cells was seen but Leu-6-reactive cells were noted in the dermal infiltrate. Keratinocytes showed some expression of class II antigen (mainly DR). In comparison with the 48-hour allergic patch test reaction, the eczematous flare-up site showed no increase in epidermal Langerhans cell numbers nor infiltration with macrophages, but the responses were similar since both showed a superficial T cell reaction in the skin.     

血管内皮生长因子在扁平苔藓及银屑病皮损中的表达

目的探讨扁平苔藓及银屑病患者皮损表皮血管内皮生长因子(VEGF)表达情况。方法用抗VEGF及CD34抗体行免疫组化染色,对扁平苔藓及银屑病皮损标本进行观察,计数表皮下方真皮的毛细血管密度。结果正常人表皮VEGF基本阴性,扁平苔藓及银屑病患者非病变部VEGF阴性,移行部表皮上层VEGF染色逐渐由弱到强,由间断到连续;病变部表皮VEGF阳性,以表皮上层细胞质的细颗粒状染色为主。CD34染色各部分真皮上层毛细血管密度为;扁平苔藓的非病变部(45.61±15.70)个/mm²、移行部(68.63±15.36)个/mm²、病变部(92.07±16.84)个/mm²;银屑病的非病变部(43.73±14.55)个/mm²、移行部(72.12±18.81)个/mm²、病变部(100.29±21.93)个/mm²,各病种三个部位之间比较差异均有统计学意义。结论扁平苔藓及银屑病皮损表皮分泌VEGF,并与真皮毛细血管增生扩张密切相关。     

Proliferative activity of CD8(+) T cells as an important clue to analyze T cell-mediated inflammatory dermatoses

Abstract To elucidate the pathogenesis of T cell-mediated inflammatory skin diseases, we examined the exact sites where CD8(+) T cells proliferate, correlating them with the localization of antigen-presenting dendritic cells. We performed CD8/Ki-67 double immunohistochemical staining and single staining for CD1a, CD68, and factor XIIIa on sections of paraffin-embedded tissue samples of inflammatory dermatoses in which T lymphocytes are thought to play a crucial role. The dermatoses were lichen planus (12 samples), acute graft-versus host disease (GVHD) (12 samples), chronic GVHD (10 samples), spongiotic dermatitis (8 samples) and psoriasis (7 samples). Labelling for Ki-67 among CD8(+) T cells was predominantly observed in the subepidermal lymphoid infiltrate, and was scanty in the epidermis. This suggested that proliferation of CD8(+) T cells occurred preferentially in the dermis. The labelling index for Ki-67 among dermal and epidermal CD8(+) cells was quite different among the different diseases studied (P < 0.05). They were rich in the subepidermal portion of the dermis of spongiotic dermatitis, acute GVHD and chronic GVHD, but rare in the dermis of psoriasis and lichen planus. A moderate infiltrate was also observed in lesional epidermis of spongiotic dermatitis, acute GVHD and chronic GVHD, whereas they was almost none in the epidermis of psoriasis and lichen planus. CD1a(+) dermal dendritic cells were densely distributed within the lymphoid infiltrate in the affected dermis of spongiotic dermatitis, psoriasis and lichen planus, whereas they were minimal in GVHD. These dermal dendritic cells are candidates as stimulators on T cells in the dermis. In conclusion, the proliferative status of T cells could be an important clue in the elucidation of the pathophysiology of T cell-mediated inflammatory dermatoses. Received: 13 December 2000 / Revised: 24 April 2001 / Accepted: 11 July 2001     

银屑病和扁平苔藓皮损ki-67及PCNA蛋白的表达

目的研究银屑病和扁平苔藓患者皮损表皮增殖状态，并比较ki-67和PCNA在增殖性皮肤疾病中表达的一致性。方法应用免疫组化法分别检测银屑病和扁平苔藓患者皮损处ki-67及PCNA的表达。结果正常对照组ki-67及PCNA弱阳性表达，仅在基底层有表达；扁平苔藓组ki-67及PCNA在基底层和棘层中均有阳性表达；银屑病组表皮基底层、棘层、颗粒层中角质形成细胞强阳性表达ki-67及PCNA。与正常对照组相比，扁平苔藓组、银屑病组ki-67及PCNA表达均增高（P〈0．05）；与扁平苔藓组相比，银屑病组ki-67表达增高（P〈0．05），PCNA无统计学意义。ki-67和PCNA在皮损中表达一致性差。结论ki-67和PCNA可能通过诱导角质形成细胞增殖参与了扁平苔藓和银屑病的发病，银屑病和扁平苔藓表现出不同的表皮增殖动力学状态。     

In situ identification of mononuclear cells in lichen planus

In this study, the in situ immunological typing of cell populations in lichen planus was attempted. T lymphocytes and suppressor/cytotoxic subsets, B lymphocytes, macrophages, immunocytes and Langerhans' cells were studied by one or more technical parameters and semiquantitative assessment of T cell populations were carried out. A critical evaluation of assays for T cell characterization was also attempted. T cells were found predominant in lichen planus infiltrate but macrophages were also many. Langerhans' cells were increased in the epidermis compared to normal skin and contact dermatitis.     

Lichen planus remission is associated with a decrease of human herpes virus type 7 protein expression in plasmacytoid dendritic cells

The cause of lichen planus is still unknown. Previously we showed human herpes virus 7 (HHV-7) DNA and proteins in lesional lichen planus skin, and significantly less in non-lesional lichen planus, psoriasis or healthy skin. Remarkably, lesional lichen planus skin was infiltrated with plasmacytoid dendritic cells. If HHV-7 is associated with lichen planus, then HHV-7 replication would reduce upon lichen planus remission. HHV-7 DNA detection was performed by nested PCR and HHV-7 protein by immunohistochemistry on lesional skin biopsies from lichen planus patients before treatment and after remission. Biopsies were obtained from lichen planus lesions before treatment (n = 18 patients) and after remission (n = 13). Before treatment 61% biopsies contained HHV-7 DNA versus 8% after remission (P = 0.01). HHV-7-protein positive cell numbers diminished significantly after remission in both dermis and epidermis. Expression of HHV-7 was mainly detected in BDCA-2 positive plasmacytoid dendritic cells rather than CD-3 positive lymphocytes. HHV-7 replicates in plasmacytoid dendritic cells in lesional lichen planus skin and diminishes after remission. This study further supports our hypothesis that HHV-7 is associated with lichen planus pathogenesis.