The presence of tryptase-positive and bikunin-negative mast cells in psoriatic skin lesions

Human mast cells are well known to produce a serine protease, tryptase, which appears to play a pathogenic role in various skin inflammations. It was previously reported that a rat homologue of bikunin may inhibit tryptase activity. Various type of cells (i.e. keratinocytes) are able to produce this protein inhibitor, it still remains unclear if bikunin is present in dermal inflammatory milieu, in which mast cells, through secretion of tryptase, play an inflammatory role. Therefore, the purpose of the present study was to exploit expression and production of bikunin in dermis and dermal constituents. We first compared the dermal mast cells in psoriatic lesions with those in lesional skin of atopic dermatitis or of chronic eczema by use of immunoelectron microscopy and immunohistochemical analyses using antibodies to bikunin and tryptase. Then, we tested what kinds of cytokines may regulate the de novo synthesis of bikunin. To do so, RNA was extracted from a human mastocytic cell line, HMC-1, reverse-transcribed, and semiquantitative RT-PCR was performed using primers specific for bikunin. With immunoelectron microscopy, bikunin was found to localize on the cell membrane, while tryptase was in the secretary granules of the mast cells. In psoriatic lesions, around 70% of dermal mast cells were positive for both tryptase and bikunin, and the remaining was mostly positive for tryptase, but the expression of bikunin was under the detection limit of the experimental setting. This observation was seen in only psoriatic lesions, even in almost cured lesions, while in atopic dermatitis or chronic eczema only mast cells doubly positive for bikuin and tryptase were seen. In HMC-1, bikunin was constitutively expressed at an mRNA level, which was upregulated by stimulation with interleukine-4, but was suppressed by interferon-γ. Bearing in mind the concept that in psoriasis local cytokine milieu is shifted toward a Th1 pattern (predominant secretion of interferon-γ), tryptase-positive, bikunin-negative mast cells may be induced.     

Fc gamma-receptors and HLA-DR antigens on endothelial cells in psoriatic skin lesions

Receptors for the Fc-part of IgG (FcR) and HLA-DR antigens on endothelial cells in normal and lesional skin from patients with psoriasis were studied in cryostat sections, using soluble immune complexes and monoclonal antibodies. FcR and HLA-DR antigens were detected on endothelial cells of dermal vessels both in sections of normal and lesional skin. The expression of FcR varied from one vessel to another and on endothelial cells within one and the same vessel. The expression of FcR and HLA-DR antigens was enhanced in sections of lesional skin compared with normal skin and most pronounced in lesional skin from active psoriasis. The enhanced expression may be mediated by interferon produced in psoriatic lesions. The presence of FcR and HLA-DR antigens on endothelial cells adds further evidence of he involvement of these cells in immune processes in the skin.     

银屑病患者皮损间充质干细胞环状RNA表达异常研究

目的 探讨环状RNA（circRNA）在银屑病发病中的作用。方法 分离、培养15例银屑病患者皮损及15例健康对照皮肤间充质干细胞（MSC）,用流式细胞仪及多向分化法进行鉴定。用RNA测序法检测circRNA表达,并进行详细的生物信息学分析。挑选7个差异表达的circRNA构建circRNA-microRNA相互作用网络,从中挑选3个与其相关的microRNA进行qRT-PCR验证。银屑病患者组和对照组qRT-PCR验证结果采用两独立样本t检验。结果 RNA测序结果显示,共检测到6 323个circRNA,其中3 227个为本实验首次发现。与对照组相比,患者组中有129个circRNA呈差异表达,其中123个表达上调,6个表达下调。挑选7个差异circRNA进行circRNA-microRNA相互作用预测,显示与这7个circRNA关系密切的银屑病相关microRNA,包括miR-17-5p、miR-30e-5p、miR-142-3p/5p、miR-369-3p、miR-184、miR-4490、miR-654-3p、miR-423-5p等。qRT-PCR也证实,这7个差异circRNA在患者组也表达上调。与对照组相比,挑选的3个microRNA在银屑病皮损中低表达（t值分别为3.993、3.217、2.918,均P < 0.05）。结论 银屑病皮损MSC circRNA表达异常,并可能参与了银屑病发病。     

银屑病患者皮损和外周血中CD123+BDCA-2+ 浆细胞样树突细胞的表达

目的 探讨CD123+BDCA-2+浆细胞样树突细胞（PDC）在寻常性银屑病患者皮损和外周血中的表达及意义。方法 收集22例寻常性银屑病患者和15例健康人皮肤组织和外周血,免疫组化法检测患者皮损和健康人皮肤组织中PDC的表达水平,流式细胞仪检测外周血中PDC的比例。结果 免疫组化结果显示,银屑病皮损中PDC的表达水平为（10.1 ± 2.1）/mm2,健康对照组为（0.4 ± 0.6） /mm2,银屑病组显著高于健康对照组（t = 17.34,P < 0.01）。银屑病患者外周血中CD123+BDCA-2+PDC占外周血单一核细胞的比例为（0.17 ± 0.07）%,健康对照组为（0.33 ± 0.20）%,银屑病组显著低于健康对照组（t = 4.48,P < 0.01）。结论 PDC通过在皮损中重新分布,在银屑病外周血中下降,可能与银屑病的发生、发展相关。     

Enkephalin-like immunoreactivity in human skin is found selectively in a fraction of CD68 – positive dermal cells: increase in enkephalin-positive cells in lesional psoriasis

Opioid peptides are synthesized in neurons, endocrine cells, monocytes/macrophages and B and T lymphocytes. They interact with opioid receptors located on immune cells and nociceptive nerve terminals. Because opioid peptides might be of importance in inflammatory skin diseases, for example psoriasis, sections of skin from psoriatic patients were immunohistochemically stained with antisera against methionine and leucine enkephalin, CD68 (KP1, PG-M1), calprotectin (M747), M130 (Ber-MAC3), CD1a and CD3. Enkephalin-like activity was detected selectively in dermal CD68-positive macrophages/monocytes. The activity showed no association with the activation markers M747 and Ber-MAC3. There was a statistically significant increase in enkephalin-positive cells in involved psoriatic skin compared with uninvolved and normal skin. These results were confirmed by radioimmunoassay which showed elevated levels in extracts from involved psoriatic skin compared with uninvolved skin (81%) and normal skin (204%). Furthermore, preproenkephalin mRNA of an expected size was detected in involved psoriatic skin. If the increased levels of enkephalins present in monocytes/macrophages in psoriatic skin lesions reach the threshold for biological activity, they may play a role in the regulation of the inflammatory processes seen in this skin disease. Received: 3 April 1996     

Immunohistochemical analysis of sensory nerves and neuropeptides,and their contacts with mast cells in developing and mature psoriatic lesions

The distribution of the neuropeptides substance P (SP), vasoactive intestinal polypeptide (VIP) and calcitonin gene related peptide (CGRP) was studied immunohistochemically in psoriatic skin during the Koebner response (6 h, 2 days, 7 days, 14 days, 21 days), and in mature psoriatic plaques, of 37 psoriatic patients. The morphological association of sensory nerves, SP and VIP with papillary mast cells was also monitored. The nerves containing SP, VIP or CGRP were very scanty in control skin, and in non-lesional and Koebner-negative psoriatic skin. The first psoriatic lesions were seen 7 days after tape stripping the symptomless psoriatic skin. SP- and VIP-containing nerves were slightly increased in Koebner-positive specimens, but the increase was very prominent in dermal papillae of mature psoriatic plaques. In the plaques, nerve-mast cell contacts were significantly increased (p<0.001) compared with non-lesional psoriatic skin. Only SP-positive fibres were detected in the epidermis and in contact with papillary mast cells. VIP was mainly located around capillaries where SP was also found. No change was noted in CGRP-positive fibres between lesional and non-lesional specimens. The appearance of SP and VIP in the capillary walls is morphological evidence for their function as vasodilators in psoriatic lesion. A slight increase in SP- and VIP-positive fibres in Koebner-positive specimens suggests that these neuropeptides may participate in the inflammatory reaction at an early stage. Their prominence in mature psoriatic plaques in turn indicates a role for them in the maintenance of psoriatic lesions. Morphological contacts between mast cells and SP-containing nerves give further evidence to the view that SP is capable of amplifying the inflammatory reaction also through the axon-reflex mechanism.Part of this work was presented at the meeting of the European Society for Dermatological Research, London, UK, 4–7 April 1992     

In situ expression of messenger RNA of interleukin-1 and interleukin-6 in psoriasis: interleukin-6 involved in formation of psoriatic lesions

Summary Psoriasis is a disease of abnormal proliferation and differentiation of epidermal cells. Several cytokines released by keratinocytes are implicated as factors responsible for this pathological condition of the epidermis. In order to elucidate the role of these cytokines in psoriasis, messenger RNA (mRNA) expression of interleukin-1 (IL-1) and IL-6 in psoriatic epidermis was investigated using biotin-labelled complementary DNA (cDNA) of the cytokines. Messenger RNA of IL-1 was weakly detected in some normal healthy epidermis specimens and more strongly in all the perilesional uninvolved psoriatic epidermis specimens. It was also expressed in the transitional zone between uninvolved and fully developed psoriatic skin, but was not expressed in lesional skin. In contrast, IL-6 mRNA was rarely expressed in normal healthy epidermis, but was expressed in perilesional uninvolved psoriatic epidermis, in the transitional zone and in the fully developed lesional epidermis, with the maximum intensity in the transitional zone. Expression of mRNA of IL-6 receptor showed a similar tendency to that of IL-6. It was expressed in psoriatic epidermis, most strongly in the transitional zone, but not in normal healthy epidermis. IL-6 was demonstrated immunohistochemically in psoriatic epidermis, but IL-6 receptor was demonstrated only in the transitional zone. Thus IL-6 and its receptor expression correlated well with the formation of psoriatic lesions where IL-1 may initiate their expression. IL-6 may play an important role in the pathogenesis of psoriasis.     

银屑病患者皮损间充质干细胞的培养鉴定及其表达HES1和CXCL6的研究

【摘要】 目的 培养鉴定银屑病患者皮损处真皮间充质干细胞（DMSC）,并研究DMSC的发状分裂相关增强子1（HES1）和趋化因子配体6（CXCL6）表达情况。方法 分离培养15例银屑病患者皮损及18例正常人皮肤的DMSC,利用流式细胞仪进行表型鉴定,实时荧光定量聚合酶链反应（RT-PCR）及免疫印迹（Western blot）检测HES1和CXCL6的mRNA及蛋白表达水平,两组间比较采用t检验。结果 银屑病组DMSC形态与正常对照组无差异,但HES1 和CXCL6基因的mRNA水平分别是对照组的3.56和3.44倍,且差异有统计学意义（P < 0.05）。在蛋白水平,银屑病组DMSC中HES1和CXCL6的表达明显高于对照组（P < 0.05）。结论 银屑病患者皮肤DMSC HES1及CXCL6表达升高可能参与银屑病的发病。     

Skewed distribution of natural killer cells in psoriasis skin lesions

Psoriasis is a hyper‐proliferative disease of the skin in which immunological mechanisms play a direct pathogenetic role. There have been limited studies of natural killer (NK) cells in psoriasis. The aim of this study was to examine the phenotype of NK cells in skin biopsies and peripheral blood mononuclear cells from patients with psoriasis and healthy controls. CD56⁺CD16^? and CD56⁺CD16⁺ NK cells were isolated from lesional skin, unaffected skin and PBMC of psoriasis patients, and normal skin and PBMC from healthy controls. The expression of CD57, NKG2A and NKG2C was assessed by flow cytometry. NK cells in psoriasis skin lesions were skewed in their expression of CD57, a marker of NK cell maturity, with CD57 expression significantly reduced and NKG2A expression increased on NK cells in lesional and unaffected skin compared to controls. These data suggest that in this patient cohort, NK cells could be isolated from psoriasis lesions and exhibit an immature phenotype.     

Mast cell density in psoriatic skin. The effect of PUVA and corticosteroid therapy

Summary Numbers and volume fractions of mast cells in nonlesional and chronic lesional skin of psoriatic patients were compared with those of normal control skin. Mast cell densities were similar in psoriatic nonlesional and normal control skin. The superficial dermis of lesional psoriatic skin contained more mast cells than either normal or nonlesional psoriatic skin. Neither PUVA nor corticosteroid treatment for 3–4 weeks significantly reduced mast cell numbers or volume fractions in lesional skin, although both treatments clinically and histologically markedly improved the lesions. The results indicate that the initiation of the healing process in psoriatic plaques is not correlated with the mast cell density. The remaining high mast cell density may be normalized later, or after a longer therapy.     

Expression of HLA-DR and OKT6 antigens on keratinocytes and dendritic cells in pemphigus

Summary The expression of HLA-DR and OKT6 antigens by epidermal cells in pemphigus was examined by immunohistochemical methods using skin biopsy specimens from human patients with pemphigus vulgaris. Changes in the distribution and subpopulations of Langerhans cells were observed in lesional (involved) skin. HLA-DR-positive, OKT6-negative dendritic cells were numerous and generally confined to the margins of epidermal clefts. OKT6-positive dendritic cells were rare or absent in lesional skin. The HLA-DR-positive, OKT6-negative dendritic cells associated with intraepidermal clefts are most likely functionally active antigen-presenting cells, which may play a role in lesion development. Aberrant expression of HLA-DR molecules by keratinocytes was observed in both involved and uninvolved skin, and may function to facilitate the recognition of surface-bound pemphigus antigen(s) by immunocompetent cells.     

Stimulation of T cells by autologous mononuclear leukocytes and epidermal cells in psoriasis

Summary Based on reports suggesting aberrant cell-mediated immunity and altered infiltration of immunocompetent cells into the skin in psoriasis, we studied the stimulation of T cells by autologous non-T mononuclear leukocytes (autologous mixed lymphocyte reaction, AMLR) and by epidermal cells isolated from lesional and clinically uninvolved skin in psoriasis (autologous mixed epidermal cell lymphocyte reaction, AMECLR). Age- and sex-matched individuals served as controls. We found that the AMLR in psoriasis (n=11) was similar to that in healthy controls (n=16); furthermore, cell proliferation was alike in the presence of either 5% AB-serum or autologous serum. By contrast, while the AMECLR in healthy controls (n=9) resembled that in psoriatics employing epidermal cells from univolved skin, epidermal cells from lesional sites (n=10) induced a significantly higher proliferation of autologous T cells in the AMECLR (P<0.01). We conclude that the in vitro stimulation of T cells by non-T mononuclear leukocytes is normal in psoriasis and is not regulated by autologous serum. Lesional psoriatic epidermal cells, however, are more active in stimulating autologous T cell proliferation than cells from univolved psoriatic or normal epidermis.     

Involvement of insulin-like growth factor-I in psoriasis as a paracrine growth factor: dermal fibroblasts play a regulatory role in developing psoriatic lesions

Abstract To investigate the contribution of dermal fibroblasts to the development of psoriasis, we examined the expression of mRNA for insulin-like growth factor-I (IGF-I) and its regulator IGF-I binding proteins (IGFBPs) in psoriatic fibroblasts by RT-PCR. We also studied the effect of inflammatory cytokines including interferon gamma (IFN-γ), tumor necrosis factor alfa (TNF-α), and IFN-α on the expression of IGF-I and IGFBPs in the fibroblasts. Semiquantitative analysis revealed that IGF-I mRNA expression in psoriatic fibroblasts (PF) was significantly higher than in control fibroblasts (CF). However, no significant difference in IGF-I mRNA was shown between nonlesional psoriatic fibroblasts (NF) and CF. Treatment with IFN-α in vitro upregulated IGF-I mRNA in PF and in CF. TNF-α appeared to downregulate IGF-I mRNA in PF but had no effect on CF. IFN-γ did not show a significant effect on IGF-I mRNA levels in any type of fibroblast. IGFBP-3 mRNA was expressed equally in PF and CF, and was not affected by cytokines. The expression of IGFBP-5 mRNA in PF was downregulated by IFN-γ and TNF-α. Taken together, these results indicate that dermal fibroblasts may contribute to the epidermal hyperplasia of psoriasis by promoting keratinocyte proliferation through IGF-I, whose secretion could be modulated by inflammatory cytokines. Received: 8 May 2000 / Revised: 25 August 2000 / Accepted: 24 October 2000     

Quantitative enzyme-histochemical analysis of tryptase- and chymase-containing mast cells in psoriatic skin

Summary Tryptase-containing mast cells have recently been found to be increased in the upper dermis of psoriatic lesions. In the present study, the distribution of chymaseand tryptase-containing mast cells was morphometrically analysed at different dermal levels of lesional and non-lesional psoriatic skin (12 patients) as well as normal human skin. Mast cell tryptase was identified enzyme-histochemically, using Z-Gly-Pro-Arg-MNA as the substrate. For demonstrating mast cell chymase, a simple and specific enzyme-histochemical staining method was developed, using Suc-Val-Pro-Phe-MNA as the substrate. All mast cells positive for chymase were also positive for tryptase and Giemsa stain. Although the number of tryptase-positive mast cells was slightly increased throughout the dermis of lesional psoriatic skin, this increase was most pronounced in the upper dermis immediately beneath, and in close contact with, the epidermis. In contrast, the number of chymase-positive mast cells was clearly decreased in the upper dermis of psoriatic lesions, but not in the deeper dermis, as compared with non-lesional psoriatic skin. In addition, all chymase-positive mast cells observed in the upper dermis were very weakly stained when compared with those in the deeper dermis. No differences were found between non-lesional psoriatic skin and normal skin in which the number of mast cells containing chymase was 72–73% of the number containing tryptase. The present results suggest that T mast cells particularly, containing tryptase but no chymase, proliferate in psoriatic lesions, and that the increase in tryptase activity and the decrease in chymase activitiy in the upper dermis may lead to an imbalance in the biochemical regulatory systems.     

尖锐湿疣皮损活化抗原CD69,CD38,HLA-DR的表达及其意义

目的 探讨活化抗原CD69，CD38，HLA—DR在尖锐湿疣（CA）皮损角质形成细胞（KC）和真皮浸润单核细胞（MNC）表达的变化。方法 免疫组化方法检测CA30例初发患者、20例复发患者皮损和15例正常人包皮KC和真皮浸润MNC上CD69，CD38，HLA—DR的表达并进行对比。结果 CA患者初发组和复发组皮损KC的CD69，CD38，HLA—DR阳性表达率分别为93.3％和90％、93．3％和90％、100％和100％，真皮浸润MNC的CD69，CD38，HLA—DR阳性表达率分别为0和0、66．7％和40％、76．7％和70％，正常人对照组均不表达。初发组和复发组皮损KC和真皮浸润MNC的CD69，CD38，HLA—DR表达强度比较显示，初发组明显高于复发组（P〈0.05）。结论 CA患者局部细胞免疫处于高水平的激活状态，这种免疫激活状态在抗病毒感染中有着一定的作用。