Th17细胞和调节性T细胞在系统性红斑狼疮患者中的研究

目的 研究系统性红斑狼疮(SLE)患者外周血Th17细胞、调节性T细胞及联系两者的细胞因子白细胞介素(IL)-6的变化及其与病情活动的相关性,探讨Th17细胞、调节性T细胞在SLE发病机制中的作用.方法 收集103例SLE患者及28名健康者,SLE活动性的判断采用SLE疾病活动指数(SLEDAI)评分方法,其中非活动组(SLEDAI≤9分)患者37例,活动组(SLEDAI＞9分)患者66例.用四色分选流式细胞仪检测外周血Th17细胞、调节性T细胞百分数,用流式细胞仪微球捕获芯片技术(CBA)检测血清IL-6浓度,并分析其与病情活动的相关性.统计学处理采用t检验、秩和检验和Spearman相关分析.结果 ①SLE患者组Th17细胞[(1.2±1.1)%]、IL-6[(35±92)pg/ml]显著高于健康对照组[(0.6±0.4)%、(6±3)pg/ml],而调节性T细胞[(1.6±1.2)%]显著低于健康对照组[(2.6±1.8)%].②SLE活动组Th17细胞[(1.6±1.7)%]显著高于SLE非活动组[(1.0±0.7)%]与健康对照组(P均＜0.05),SLE活动组调节性T细胞[(1.5±1.3)%]显著低于SLE非活动组[(2.1±2.0)%]与健康对照组(P均＜0.05),SLE活动组Th17/调节性T细胞(0.68±0.34)显著高于SLE非活动组(0.24±0.20)与健康对照组(0.13±0.09,P均＜0.05),SLE活动组IL-6[(41±22) pg/ml]显著高于SLE非活动组[(32±28) pg/ml]与健康对照组(P均＜0.05);SLE非活动组与健康对照组之间Th17细胞、调节性T细胞、Th17/调节性T细胞、IL-6差异均无统计学意义(P＞0.05).③Th17细胞百分数与SLEDAI呈正相关,调节性T细胞百分数与SLEDAI呈负相关,Th17/调节性T细胞与SLEDAI呈正相关,IL-6浓度与SLEDAI评分之间呈正相关,Th17细胞与调节性T细胞呈负相关.结论 SLE患者外周血Th17细胞、调节性T细胞及IL-6浓度较健康对照组显著增高,并与疾病活动性密切相关,说明Th17细胞、调节性T细胞可能在SLE的发生、发展中起重要作用.

Abstract：

Objective To study the changes of Th17 cell, regulatory T cell (Treg) and interleukin (IL)-6 in the peripheral blood of patients with systemic lupus erythematosus (SLE) and their relationship with disease activity. Methods Percentage of Th17 and Treg in the peripheral blood of 103 patients with SLE and 28 healthy volunteers were detected by flow cytometry. The concentration of IL-6 in SLE patients and healthy volunteers was detected by cytometric bead array (CBA). The disease activity of SLE was measured by SLEDAI. SLE patients were divided into two groups: stable SLE (SLEDAI≤ 9, n=37) and active SLE (SLEDAI＞9, n= 66). The change of Th17, Treg, IL -6 and their relationship with disease activity were analyzed. Nonparamentric tests, t -test and spearman correlation were used for statistical analysis. Results The percentage of Th17 cells and the concentration of IL-6 in the peripheral blood in patients with SLE was higher than that in normal controls [respectively for (1.2±1.1)%, (35±92) pg/ml and (0.6±0.4)%, (6±3) pg/ml, P＜0.05]. However, the percentage of Treg in patients with SLE was lower than that in normal controls [respectively for (1.6±1.2)%,(2.6±1.8)%, P＜0.05]. The percentage of Th17, Th17/Treg IL-6 level in active SLE patients was higher than those in inactive SLE and those in normal controls (P＜0.05). However, the percentage of Treg in active SLE was lower than that in stable SLE patients and that in normal controls (P＜ 0.05). The percentage of Th17, Th17/Treg and concentration of IL-6 was positively correlated to disease activity(P＜0.05). But the percentage of Treg had negative correlation with the percentage of Th17 and disease activity (P＜0.05). Conclusion Th17, Treg and serum IL-6 in SLE patients are abnormal and they maybe contribute to the pathogenesis of SLE.     

Systemic lupus erythematosus and regulatory T cells

A global depletion of FoxP3+ CD25(bright) CD4+ regulatory T cell is observed during lupus flares. This phenomenon is not the consequence of the relocalization of Tregs in diseased organs but could be related to their specific sensitivy to Fas mediated apoptosis. Several therapeutic perspectives can be drawn taking into account these pathophysiological insights.     

Quantification of regulatory T cells in peripheral blood of patients with systemic lupus erythematosus

Regulatory T cells (Tregs) are supposed to stop immune responses in the course of immune activation. However, chronic activation of immune system in systemic lupus erythematosus (SLE) and many other autoreactive disorders are evidence of malfunction of this system. Therefore, it is plausible to quantify presence of these cells in different diseases. Forty-one patients with diagnosis of SLE were enrolled in this study. Patients were divided into two groups of patients with active and inactive disease based on the disease activity score. Flow cytometry analysis was used to determine the frequency of regulatory T cells in peripheral blood according to high expression of CD25 and intracellular Forkhead/winged-helix (Foxp3). Further 30 healthy individuals considered as control group. Significantly less CD4+CD25hi regulatory T cells were detected in active patients (P < 0.001) compared to healthy individuals. The percentage of CD4+CD25hi cells were inversely correlated with the SLEDAI disease score in patients with active disease (r = −0.837, P < 0.0001). Patients with active disease had lower frequencies of CD4+Foxp3+ cells. However, increased frequencies of CD4+Foxp3+ T cells were observed in peripheral blood of patients with inactive disease compared with active patients or healthy individuals (P < 0.010). Moreover, a significant difference between the proportion of CD4+CD25-Foxp3+ population in healthy controls and patients with active disease was shown (P < 0.0005). Presence of lower frequencies of Tregs in patients with SLE could be evaluated as an immune turbulence and could be employed as a target for immunotherapeutic manipulation. However, controversies need to be resolved.     

Tumour necrosis factor alpha is expressed in refractory skin lesions from patients with subacute cutaneous lupus erythematosus

OBJECTIVES: To investigate whether tumour necrosis factor alpha (TNFalpha) is expressed in subacute cutaneous lupus erythematosus (SCLE) skin lesions. METHODS: The in situ expression of TNFalpha in refractory lesional and non-lesional skin biopsy specimens from patients with SCLE was analysed using an immunohistochemical approach. At the time of biopsy these patients were receiving treatment with systemic medications such as antimalarial agents, immunosuppressive drugs, and thalidomide. Expression of TNFalpha was also evaluated in cutaneous lesions of patients with other inflammatory and neoplastic skin diseases as controls. RESULTS: The data showed that refractory lesional skin tissue from patients with SCLE displays a strongly positive distribution of TNFalpha, particularly within the epidermis. No prominent staining was seen in non-lesional skin from the same group of patients or in cutaneous lesions from the control group. CONCLUSIONS: These findings suggest that TNFalpha is localised and produced by epidermal cells within SCLE skin lesions and support its potential role in the pathogenesis of SCLE. The tissue localisation of TNFalpha may represent a potential therapeutic target providing a new perspective in the treatment of refractory skin lesions in patients with SCLE.     

Deep skin lesions of lupus erythematosus

The authors report 2 cases of lupus erythematosus (LE) with deep cutaneous lesions. In both cases the lupus panniculitis presented as hard, infiltrated subcutaneous plaques situated symmetrically on the external aspect of both arms and on the upper parts of the buttocks. The diagnosis was confirmed on clinical, histological, immunohistological and therapeutic data. These 2 reports and the 43 cases in the literature dating from 1967, illustrate the main features of lupus panniculitis. The clinical appearances and sites are very stereotyped and immediately suggest the diagnosis even in the absence of other clinical and/or biological signs of lupus. The evolution is very slow and is characterised by dramatic regression of the inflammatory signs with synthetic antimalarial drugs. These deep skin lesions are observed in both purely cutaneous chronic lupus and in systemic lupus which is usually relatively inactive. This prevents the identification of a special form of this disease with a specific evolution and prognosis.     

Leflunomide-induced subacute cutaneous lupus erythematosus with erythema multiforme-like lesions

Leflunomide is an immunosuppressive agent that acts by inhibiting pyrimidine synthesis in lymphocytes and other rapidly proliferating cells, as well as by suppressing tumor necrosis factor-alpha-induced cellular responses. A number of leflunomide-related adverse events have been reported. Among cutaneous side effects, a few cases of subacute cutaneous lupus erythematosus have been described. We report a previously undocumented reaction to leflunomide, manifesting as subacute cutaneous lupus erythematosus and erythema multiforme-like lesions, in a young woman treated with this drug for ankylosing spondylitis. Withdrawal of leflunomide combined with a short cycle of systemic corticosteroid led to the resolution of the patient's rash, indicating this drug as being responsible for the development of the disease. We conclude that leflunomide might have triggered the occurrence of both subacute cutaneous lupus erythematosus and erythema multiforme in a patient with pre-existing autoimmune diathesis. The suppressive effect of this drug on tumor necrosis factor-alpha-related mechanisms might have played a role in the induction of such a unique reaction to leflunomide.     

Bullous skin lesions in systemic lupus erythematosus

Bullous lesions are an unusual finding in systemic lupus erythematosus (SLE). A number of other bullous disorders can coexist with SLE; therefore it is often uncertain whether the bullous eruption represents a manifestation of SLE. Here we present a case of bullous lupus erythematosus (BL) in which the skin lesions preceded other clinical manifestations of SLE. Using a review of 27 other cases of BL in the literature. the differential diagnosis, clinical and histopathologic presentation, and treatment of BL is discussed. Based on this review a set of criteria for the diagnosis of BL is proposed.     

Presence of cutaneous interferon-alpha producing cells in patients with systemic lupus erythematosus

Systemic lupus erythematosus (SLE) patients have increased levels of interferon-alfa (IFN-alpha) in the circulation but a reduced number of functionally intact natural IFN-alpha producing cells (IPC) in peripheral blood. In search for tissue localisation of activated IPC, we investigated skin biopsies from SLE patients for the occurrence of such cells. Eleven SLE patients with inflammatory skin lesions and six healthy controls were biopsied. An immunohistochemical technique (IH) and in situ hybridisation (ISH) were used to detect intracellular IFN-alpha protein and IFN-alpha mRNA, respectively. In all 11 biopsies from SLE lesions, a high number of IPC were detected by IH. In the nonlesional SLE biopsies we could also demonstrate IPC in 10/11 patients. In 6/11 SLE patients, IFN-alpha mRNA containing cells could be detected in the specimens. A low number of IPC were detected in 1/6 healthy controls by IH, but no ISH positive cells were seen. Our results demonstrate that SLE patients have active IPC in both dermal lesions and in noninflammatory skin. A recruitment of IPC from blood to peripheral tissues may explain the low number of circulating natural IPC in SLE patients. Because the type I IFN system is involved in the SLE disease process, these results are of interest for the understanding of the pathogenesis in SLE.     

CD4+CD25+ T cells in skin lesions of patients with cutaneous leishmaniasis exhibit phenotypic and functional characteristics of natural regulatory T cells

Endogenous regulatory T (Treg) cells are involved in the control of infections, including Leishmania infection in mice. Leishmania viannia braziliensis is the main etiologic agent of cutaneous leishmaniasis (CL) in Brazil, and it is also responsible for the more severe mucocutaneous form. Here, we investigated the possible involvement of Treg cells in the control of the immune response in human skin lesions caused by L. viannia braziliensis infection. We show that functional Treg cells can be found in skin lesions of patients with CL. These cells express phenotypic markers of Treg cells--such as CD25, cytotoxic T lymphocyte-associated antigen 4, Foxp3, and glucocorticoid-induced tumor necrosis factor receptor--and are able to produce large amounts of interleukin-10 and transforming growth factor- beta . Furthermore, CD4+CD25+ T cells derived from the skin lesions of 4 of 6 patients with CL significantly suppressed in vitro the phytohemagglutinin-induced proliferative T cell responses of allogeneic peripheral-blood mononuclear cells (PBMCs) from healthy control subjects at a ratio of 1 Treg cell to 10 allogeneic PBMCs. These findings suggest that functional Treg cells accumulate at sites of Leishmania infection in humans and possibly contribute to the local control of effector T cell functions.     

初发系统性红斑狼疮患者外周血CD4+调节性T细胞CD73表达

目的 研究CD73在初发活动性系统性红斑狼疮(SLE)患者外周血CD4+调节性T细胞的表达情况,探讨其在SLE发病中的作用.方法 采用流式细胞术检测29例初发未经治疗的活动期SLE患者(SLE组)和22例健康人(健康对照组)外周血CD4+CD25+CD73+T细胞百分率及CD4+CD73+、CD4+CDhi25、CD4+CD25+T细胞中叉头状转录因子3(FOXP3)蛋白表达,同时对CD73表达水平与SLE活动指标进行相关性分析.结果 SLE组患者外周血CD4+ CD25+ CD73+T细胞百分率低于健康对照组[(1.25±1. 32)%vs(2.35±1.09)%,P＜0.01].SLE组和健康正常对照组,CD73在C4+ CDhi25T细胞的表达水平[(29.05±12.53)%、(43.35±10.09)%]高于CD4+ CD25+T细胞[(17.48±6.92)%、(29.98±10.39)%,P＜0.001];FOXP3蛋白在CD4+ CD73+ T细胞[(65.36±14.40)%、(63.80±14.05)%]、CD4+ CDhi25 T细胞的表达水平[(67.30±13.04)%、(56.30±9.21)%]明显高于CD4+ CD25+ T细胞[(45.70±12.74)%、(43.98±5.17)%,P＜0.001],在CD4+ CD73-T细胞几乎不表达,而在CD4+ CD73+ T细胞、CD4+ CDhi25 T细胞中的表达差异无统计学意义(P值均大于0.05).CD73在CD4+ CD25+ T细胞的表达水平与SLE疾病活动指数、ESR、C反应蛋白、抗补体C1q、抗核小体抗体均无相关性(P＞0.05).结论 CD73可作为调节性T细胞新的表面标记,其在调节性T细胞中的异常表达可能参与SLE的发病机制.     

Neonatal lupus erythematosus with multisystem organ involvement preceding cutaneous lesions.

Here we report a case of neonatal lupus erythematosus syndrome presenting with multisystem organ involvement, including anemia, thrombocytopenia, purpura, bloody diarrhea, enzymatic liver abnormalities, splenomegaly and pneumonitis. These findings preceded the cutaneous rash that was the clue for the diagnosis. The patient's mother had an undiagnosed subacute cutaneous lupus erythematosus. The various forms of onset of neonatal lupus erythematosus syndrome are emphasized.     

Synchronous skin lesions in mother and baby with neonatal lupus erythematosus

Clinical Rheumatology -     

Management of skin disease in patients with lupus erythematosus

Skin disease in patients with lupus erythematosus may be subdivided into two broad categories - those represented by a 'specific' histopathology, the interface dermatitis, and those with changes that are not specific to lupus erythematosus, for example, vasculitis, mucin infiltration, etc. The specific skin lesions that are most common are discoid lupus erythematosus (DLE) and subacute cutaneous lupus erythematosus (SCLE). Evaluation will allow the treating physician to assign a prognosis. Cutaneous lesions can generally be managed with standard therapies. Patients with discoid LE and subacute cutaneous LE are generally photosensitive, and therefore sunscreens, protective clothing and behavioural alteration should be discussed with all patients. Topical corticosteroids are a standard form of therapy, but 'newer' agents such as retinoids, calcipotriene and tacrolimus might be effective. Antimalarial agents are generally effective. Attempts to reduce or stop smoking may aid in the control of cutaneous LE. The choice of alternative therapy is personal, and discussions of the risks and benefits should be carefully documented.     

Chloroquine treatment reduces the number of cutaneous HLA-DR+ and CD1a+ cells in patients with systemic lupus erythematosus

Systemic lupus erythematosus (SLE) is a chronic autoimmune disease that can be exacerbated by exposure to ultraviolet radiation (UVR). The number and phenotype of antigen presenting cells in the skin play a role in cutaneous immune response generation. Although antimalarials are widely used in SLE treatment, their mode of action is not completely elucidated. The aim of our study was to determine the effect of chloroquine treatment on HLA-DR+ and CD1a+ cell number in locally irradiated (three minimal erythema doses of UVB) and normal appearing skin in SLE patients and healthy subjects. A significantly higher number of HLA-DR+ and CD1a+ cells were found in both locations in SLE patients compared with controls. Following three months of daily chloroquine treatment (250 mg), the HLA-DR+ and CD1a+ cell counts were significantly reduced in both irradiated and unirradiated sites of SLE patients, although still higher than in controls. Chloroquine treatment reduces the number of antigen presenting cells in the skin of SLE patients, and this effect may explain the antimalarials beneficial immunoregulatory and anti-inflammatory properties.