Th 细胞因子IL-17、IFN-γ、IL-4 在狼疮性肾炎中的表达及意义

目的:探讨T辅助细胞(Th)相关细胞因子在狼疮性肾炎发病中的免疫机制作用。方法:64例系统性红斑狼疮患者和28例健康体检者作为对照,采用酶联免疫吸附测定法(ELISA法)检测所有受试者血清IL-17、IFN-γ、IL-4水平,并对其与SLEDAI、SDI、24小时尿蛋白量相关性进行研究。结果:狼疮性肾炎组血清IL-17水平显著高于狼疮无肾炎组和健康对照组(P<0.001),狼疮性肾炎组血清IFN-γ水平显著高于狼疮无肾炎组(P<0.05)和健康对照组(P<0.01),血清IL-4水平在狼疮性肾炎组、狼疮无肾炎组均显著高于健康对照组(P<0.01)。狼疮性肾炎组IFN-γ/IL-4比值显著高于狼疮无肾炎组(P<0.01)和健康对照组(P<0.05);狼疮无肾炎组IFN-γ/IL-4比值显著低于健康对照组(P<0.01)。SLE患者血清IFN-γ表达水平与SLEDAI积分呈正相关(r=0.402,P<0.05),血清IL-17、IL-4表达水平与SLEDAI、SDI、抗ds-DNA抗体、C3、24小时尿蛋白量均无相关性。结论:狼疮性肾炎患者外周血中IL-17、IFN-γ、IL-4等促炎细胞因子均有不同程度升高促起炎症发生及组织损伤,参与了狼疮性肾炎的免疫发病过程。     

Increased serum interleukin 17 in patients with systemic lupus erythematosus

Interleukin 17 (IL-17) is a Th17 cytokine associated with inflammation, autoimmunity and defense against some bacteria, it has been implicated in many chronic autoimmune diseases including psoriasis, multiple sclerosis and systemic sclerosis. However, whether IL-17 plays a role in the pathogenesis of systemic lupus erythematosus (SLE) remains unclear. In the present study, we aimed to investigate the serum IL-17 level in patients with SLE and it’s associations with disease manifestations and activity. Fifty-seven patients with SLE and 30 healthy volunteers were recruited. Serum IL-17 levels were examined by enzyme linked immunosorbent assay (ELISA). Statistic analyzes were performed by SPSS 10.01. Results show that serum IL-17 levels were significantly elevated in SLE patients as compared with normal controls. Nevertheless, no associations of serum IL-17 level with clinical and laboratory parameters were found; no significant difference regarding serum IL-17 level between SLE patients with nephritis and those without nephritis was found; no significant difference was found between Less active SLE and More active SLE; Correlation analysis between serum IL-17 levels and SLEDAI showed no association. Taken together, our results indicate increased serum IL-17 levels in SLE patients, suggesting that this cytokine may trigger the inflammatory process in SLE. However, no associations of serum IL-17 level with disease manifestations were found. Therefore, further studies are required to confirm this preliminary data.     

Infection of Female BWF1 Lupus Mice with Malaria Parasite Attenuates B Cell Autoreactivity by Modulating the CXCL12/CXCR4 Axis and Its Downstream Signals PI3K/AKT,NFκB and ERK

Systemic lupus erythematosus (SLE) is a prototypic autoimmune disease characterized by abnormal autoreactivity in B cells. Lymphocytes and their soluble mediators contribute to the disease pathogenesis. We recently demonstrated that infecting lupus mice with malaria confers protection against lupus nephritis by attenuating oxidative stress in both liver and kidney tissues. In the current study, we further investigated B cell autoreactivity in female BWF1 lupus mice after infection with either live or gamma-irradiated malaria, using ELISA, flow cytometry and Western blot analysis. The lupus mice exhibited a significant elevation in plasma levels of IL-4, IL-6, IL-7, IL-12, IL-17, IFN-α, IFN-γ, TGF-β, BAFF and APRIL and a marked elevation of IgG2a, IgG3 and ant-dsDNA autoantibodies compared with normal healthy mice. Infecting lupus mice with live but not gamma-irradiated malaria parasite partially and significantly restored the levels of the soluble mediators that contribute to the progression of lupus. Furthermore, the B cells of lupus mice exhibited an increased proliferative capacity; aberrant overexpression of the chemokine receptor CXCR4; and a marked elevation in responsiveness to their cognate ligand (CXCL12) via aberrant activation of the PI3K/AKT, NFκB and ERK signaling pathways. Interestingly, infecting lupus mice with live but not gamma-irradiated malaria parasite restored a normal proliferative capacity, surface expression of CXCR4 and B cell response to CXCL-12. Taken together, our data present interesting findings that clarify, for the first time, the molecular mechanisms of how infection of lupus mice with malaria parasite controls B cell autoreactivity and thus confers protection against lupus severity.     

Increased expression of costimulatory markers CD134 and CD80 on interleukin-17 producing T cells in patients with systemic lupus erythematosus

Introduction  

There is growing evidence that interleukin 17 (IL-17) producing T cells are involved in the pathogenesis of systemic lupus erythematosus (SLE). Previous studies showed that increased percentages of T-cell subsets expressing the costimulatory molecules CD80 and CD134 are associated with disease activity and renal involvement in SLE. The aim of this study was to investigate the distribution and phenotypical characteristics of IL-17 producing T-cells in SLE, in particular in patients with lupus nephritis, with emphasis on the expression of CD80 and CD134.     

IL-17 与狼疮性肾炎关系的研究新进展

系统性红斑狼疮是一种涉及多系统损害的自身免疫性疾病,其主要并发症及死亡原因之一是狼疮性肾炎。研究表明,IL-17和产IL-17细胞可以浸润肾脏,与其他细胞因子协同作用,引起肾脏局部炎症反应。拮抗IL-17的生物制剂已应用于银屑病、强制性脊柱炎等免疫介导炎症性疾病,但在系统性红斑狼疮及狼疮性肾炎的研究尚少。本文对IL-17与狼疮性肾炎的关系进行综述,探讨IL-17及其抑制剂在狼疮性肾炎治疗的作用及发展前景。     

Renal Dnase1 Enzyme Activity and Protein Expression Is Selectively Shut Down in Murine and Human Membranoproliferative Lupus Nephritis

Background

Deposition of chromatin-IgG complexes within glomerular membranes is a key event in the pathogenesis of lupus nephritis. We recently reported an acquired loss of renal Dnase1 expression linked to transformation from mild to severe membranoproliferative lupus nephritis in (NZBxNZW)F1 mice. As this may represent a basic mechanism in the progression of lupus nephritis, several aspects of Dnase1 expression in lupus nephritis were analyzed.

Methodology/Principal Findings

Total nuclease activity and Dnase1 expression and activity was evaluated using in situ and in vitro analyses of kidneys and sera from (NZBxNZW)F1 mice of different ages, and from age-matched healthy controls. Immunofluorescence staining for Dnase1 was performed on kidney biopsies from (NZBxNZW)F1 mice as well as from human SLE patients and controls. Reduced serum Dnase1 activity was observed in both mesangial and end-stage lupus nephritis. A selective reduction in renal Dnase1 activity was seen in mice with massive deposition of chromatin-containing immune complexes in glomerular capillary walls. Mice with mild mesangial nephritis showed normal renal Dnase1 activity. Similar differences were seen when comparing human kidneys with severe and mild lupus nephritis. Dnase1 was diffusely expressed within the kidney in normal and mildly affected kidneys, whereas upon progression towards end-stage renal disease, Dnase1 was down-regulated in all renal compartments. This demonstrates that the changes associated with development of severe nephritis in the murine model are also relevant to human lupus nephritis.

Conclusions/Significance

Reduction in renal Dnase1 expression and activity is limited to mice and SLE patients with signs of membranoproliferative nephritis, and may be a critical event in the development of severe forms of lupus nephritis. Reduced Dnase1 activity reflects loss in the expression of the protein and not inhibition of enzyme activity.     

血β2微球蛋白水平在系统性红斑狼疮中的临床意义

目的：探讨系统性红斑狼疮（systemic lupus erythematosus,SLE）患者血β2-MG水平与疾病活动的相关性及其临床意义。方法：随机抽取2012年2月-2012年7月我科收治的62例SLE患者（SLE组）和同期在我院门诊体检的健康体检者40例（对照组）,检测和比较两组血清β2-MG、自身抗体、补体水平,并对SLE患者进行SLEDAI评分,分析SLE患者血清β2-MG水平与自身抗体、补体水平和SLEDAI评分的相关性。结果：SLE组血β2-MG水平（3．11±0．40μg／mL）显著高于对照组（2．23±0．23μg/mL）,差异有统计学意义（P〈0．05）;其中发生口腔溃疡、浆膜炎及蛋白尿的SLE患者的血β2-MG水平与无此三种表现的患者相比显著升高,差异有统计学意义（P〈0．05）。SLE患者的血β2-MG水平与抗ds—DAN抗体、SLEDAI均呈显著正相关（分别为r=0．289,r=0．361,P〈0．01）,与C3呈负相关（r=-0．271,P〈0．05）。结论：SLE患者血β2-MG水平高于正常,可作为SLE疾病活动指标用于监测疾病活动。     

Therapeutic potential of DCB-SLE1, an extract of a mixture of Chinese medicinal herbs, for severe lupus nephritis

The pathogenesis of lupus nephritis is mainly attributable to a complex interaction between the innate and adaptive immune systems, including T and B cell function abnormalities. In addition to autoantibody production and immune complex deposition, Th1 and Th17 cytokines may play key roles in the development and progression of lupus nephritis. Acute onset of severe lupus nephritis remains a challenge in terms of prevention and treatment. In the present study, we evaluated the therapeutic effects of DCB-SLE1, an extract of a mixture of four traditional Chinese medicinal herbs (Atractylodis macrocephalae Rhizoma, Eucommiae cortex, Lonicerae caulis, and Hedyotidis diffusae Herba), on an accelerated severe lupus nephritis model, characterized by acute onset of proteinuria, azotemia, autoantibody production, and development of severe nephritis, induced by twice weekly injection of New Zealand black/white F1 mice with Salmonella-type lipopolysaccharide. DCB-SLE1 was administered daily by gavage starting 2 days after the first dose of induction of lipopolysaccharide, and the mice were euthanized at week 1 or week 5. The results showed that DCB-SLE1 significantly ameliorated the hematuria, proteinuria, renal dysfunction, and severe renal lesions by 1) suppression of B cell activation and decreased autoantibody production; 2) negative regulation of T cell activation/proliferation and natural killer cell activity; 3) suppression of IL-18, IL-6, and IL-17 production and blocking of NF-κB activation in the kidney; and 4) prevention of lymphoid and renal apoptosis. These results show that DCB-SLE1 can protect the kidney from autoimmune response-mediated acute and severe damage through systemic immune modulation and anti-inflammation pathways.     

Physiologic role for enhanced renal thromboxane production in murine lupus nephritis.

To investigate the physiologic significance of enhanced renal thromboxane production in murine lupus nephritis, we measured renal hemodynamics and eicosanoid production in MRL-lpr/lpr mice from 8 to 20 weeks of age. Over this age range, MRL-lpr/lpr mice develop an autoimmune disease with nephritis similar to human systemic lupus erythematosus (SLE). In these studies, glomerular filtration rate (GFR) and PAH clearance (CPAH) decreased progressively with age in MRL-lpr/lpr mice, but not in controls. This impairment of renal hemodynamics was associated with increased renal thromboxane production, as well as increased excretion of both thromboxane B2 (TxB2) and 2,3-dinor TxB2 in urine. There was an inverse correlation between renal thromboxane production in MRL-lpr/lpr mice and both GFR and CPAH. Furthermore, there were positive correlations between thromboxane production by the kidney and both the severity of renal histopathology and serum anti-DNA antibody levels measured in individual animals. Enhanced urinary excretion of TxB2 and the development of renal dysfunction also coincided temporally with the appearance of increased levels of interleukin 1 beta (IL-1 beta) mRNA in renal cortex. Acute administration of the specific thromboxane receptor antagonist GR32191 to MRL-lpr/lpr mice restored GFR to normal in early stages of the autoimmune disease. However, in animals with more advanced nephritis, the effect of acute thromboxane receptor blockade on renal hemodynamics was less marked. We conclude that thromboxane A2 is an important mediator of reversible renal hemodynamic impairment in murine lupus, especially in the early phase of disease.     

Cutting edge: protein phosphatase 2A confers susceptibility to autoimmune disease through an IL-17-dependent mechanism

The contribution of individual molecular aberrations to the pathogenesis of systemic lupus erythematosus (SLE), an autoimmune disease that affects multiple organs, is often difficult to evaluate because of the presence of abundant confounding factors. To assess the effect of increased expression of the phosphatase protein phosphatase 2A (PP2A) in T cells, as recorded in SLE patients, we generated a transgenic mouse that overexpresses the PP2Ac subunit in T cells. The transgenic mouse displays a heightened susceptibility to immune-mediated glomerulonephritis in the absence of other immune defects. CD4(+) T cells produce increased amounts of IL-17 while the number of neutrophils in the peripheral blood is increased. IL-17 neutralization abrogated the development of glomerulonephritis. We conclude that increased PP2Ac expression participates in SLE pathogenesis by promoting inflammation through unchecked IL-17 production and facilitating the development of end-organ damage.     

Amelioration of lupus nephritis by serum amyloid P component gene therapy with distinct mechanisms varied from different stage of the disease

Background

Our previous study revealed that administration of syngeneic female BALB/c mice with excessive self activated lymphocyte-derived DNA (ALD-DNA) could induce systemic lupus erythematosus (SLE) disease, indicating that overload of self-DNA might exceed normal clearance ability and comprise the major source of autoantigens in lupus mice. Serum amyloid P component (SAP), an acute-phase serum protein with binding reactivity to DNA in mice, was proved to promote the clearance of free DNA and prevent mice against self-antigen induced autoimmune response. It is reasonable to hypothesize that SAP treatment might contribute to alleviation of SLE disease, whereas its role in ALD-DNA-induced lupus nephritis is not fully understood.

Methodology/Principal Findings

The ratios of SAP to DNA significantly decreased and were negatively correlated with the titers of anti-dsDNA antibodies in ALD-DNA-induced lupus mice, indicating SAP was relatively insufficient in lupus mice. Herein a pcDNA3-SAP plasmid (pSAP) was genetically constructed and intramuscularly injected into BALB/c mice. It was found that SAP protein purified from the serum of pSAP-treated mice bound efficiently to ALD-DNA and inhibited ALD-DNA-mediated innate immune response in vitro. Treatment of ALD-DNA-induced lupus mice with pSAP in the early stage of SLE disease with the onset of proteinuria reversed lupus nephritis via decreasing anti-dsDNA autoantibody production and immune complex (IC) deposition. Further administration of pSAP in the late stage of SLE disease that had established lupus nephritis alleviated proteinuria and ameliorated lupus nephritis. This therapeutic effect of SAP was not only attributable to the decreased levels of anti-dsDNA autoantibodies, but also associated with the decreased infiltration of lymphocytes and the reduced production of inflammatory markers.

Conclusion/Significance

These results suggest that SAP administration could effectively alleviated lupus nephritis via modulating anti-dsDNA antibody production and the inflammation followed IC deposition, and SAP-based intervening strategy may provide new approaches for treating SLE disease.     

Kallikrein Transduced Mesenchymal Stem Cells Protect against Anti-GBM Disease and Lupus Nephritis by Ameliorating Inflammation and Oxidative Stress

Previously we have shown that kallikreins (klks) play a renoprotective role in nephrotoxic serum induced nephritis. In this study, we have used mesenchymal stem cells (MSCs) as vehicles to deliver klks into the injured kidneys and have measured their therapeutic effect on experimental antibody induced nephritis and lupus nephritis. Human KLK-1 (hKLK1) gene was transduced into murine MSCs using a retroviral vector to generate a stable cell line, hKLK1-MSC, expressing high levels of hKLK1. 129/svj mice subjected to anti-GBM induced nephritis were transplanted with 10⁶ hKLK1-MSCs and hKLK1 expression was confirmed in the kidneys. Compared with vector-MSCs injected mice, the hKLK1-MSCs treated mice showed significantly reduced proteinuria, blood urea nitrogen (BUN) and ameliorated renal pathology. Using the same strategy, we treated lupus-prone B6.Sle1.Sle3 bicongenic mice with hKLK1-MSCs and demonstrated that hKLK1-MSCs delivery also attenuated lupus nephritis. Mechanistically, hKLK1-MSCs reduced macrophage and T-lymphocyte infiltration into the kidney by suppressing the expression of inflammation cytokines. Moreover, hKLK1 transduced MSCs were more resistant to oxidative stress-induced apoptosis. These findings advance genetically modified MSCs as potential gene delivery tools for targeting therapeutic agents to the kidneys in order to modulate inflammation and oxidative stress in lupus nephritis.     

Increased interleukin-23 receptor<Superscript>+</Superscript>T cells in peripheral blood mononuclear cells of patients with systemic lupus erythematosus

Introduction  

Systemic lupus erythematosus (SLE) is an autoimmune disorder characterized by production of autoantibodies and immune complex deposition in various organs. Aberrations in the T lymphocyte compartment and dysregulated cytokine production are key features of SLE pathogenesis and disease progression. Recently, the role of the interleukin (IL)-17/IL-23 axis in the pathogenesis of SLE has been reported. IL-23 and IL-23R are essential for expansion of pathogenic IL-17-producing T lymphocytes and have been shown to be important in the pathogenesis of lupus in animal models.     

系统性红斑狼疮患者血清中IL-17 的检测及临床意义

目的:通过检测系统性红斑狼疮(SLE)患者外周血中IL-17的水平,探讨其临床意义。方法:用酶联免疫吸附法(enzymelinked immunosorbent assay,ELISA)检测61例SLE患者及30例健康人血清IL-17水平,并收集整理SLE患者的临床资料及实验室数据,分析其与临床的关系。结果:活动期SLE患者血清IL-17水平明显高于正常对照组(P<0.01),与SLE非活动组相比,差异亦有统计学意义(P<0.01)。但SLE非活动组与正常对照组间无统计学意义。结论:IL-17水平在活动期SLE患者血清中表达明显增高,且与SLEDAI评分呈正相关,提示IL-17可能参与了SLE疾病的病理过程,可能与疾病活动的关系密切。     

Activated protein C attenuates systemic lupus erythematosus and lupus nephritis in MRL-Fas(lpr) mice

Systemic lupus erythematosus (SLE) is a chronic autoimmune disease leading to inflammatory tissue damage in multiple organs (e.g., lupus nephritis). Current treatments including steroids, antimalarials, and immunosuppressive drugs have significant side effects. Activated protein C is a natural protein with anticoagulant and immunomodulatory effects, and its recombinant version has been approved by the U.S. Food and Drug Administration to treat severe sepsis. Given the similarities between overshooting immune activation in sepsis and autoimmunity, we hypothesized that recombinant activated protein C would also suppress SLE and lupus nephritis. To test this concept, autoimmune female MRL-Fas(lpr) mice were injected with either vehicle or recombinant human activated protein C from week 14-18 of age. Activated protein C treatment significantly suppressed lupus nephritis as evidenced by decrease in activity index, glomerular IgG and complement C3 deposits, macrophage counts, as well as intrarenal IL-12 expression. Further, activated protein C attenuated cutaneous lupus and lung disease as compared with vehicle-treated MRL-Fas(lpr) mice. In addition, parameters of systemic autoimmunity, such as plasma cytokine levels of IL-12p40, IL-6, and CCL2/MCP-1, and numbers of B cells and plasma cells in spleen were suppressed by activated protein C. The latter was associated with lower total plasma IgM and IgG levels as well as lower titers of anti-dsDNA IgG and rheumatoid factor. Together, recombinant activated protein C suppresses the abnormal systemic immune activation in SLE of MRL-Fas(lpr) mice, which prevents subsequent kidney, lung, and skin disease. These results implicate that recombinant activated protein C might be useful for the treatment of human SLE.     

Granulin Exacerbates Lupus Nephritis via Enhancing Macrophage M2b Polarization

Background and Aims

Lupus nephritis (LN), with considerable morbidity and mortality, is one of the most severe manifestations of systemic lupus erythematosus (SLE). Yet, the pathogenic mechanisms of LN have not been clearly elucidated, and efficient therapies are still in great need. Granulin (GRN), a multifunctional protein linked to inflammatory diseases, has recently been reported to correlate with the disease activity of autoimmune diseases. However, the role of GRN in the pathogenic process of LN still remains obscure. In this study, we explored its potential role and underlying mechanism in the pathogenesis of LN.

Methodology/Principal Findings

We found that serum GRN levels were significantly up-regulated and were positively correlated with the severity of LN. Overexpression of GRN in vivo by transgenic injection remarkably exacerbated LN, whereas down-regulation of GRN with shRNA ameliorated LN, firmly demonstrating the critical role of GRN in the pathogenesis of LN. Notably, macrophage phenotype analysis revealed that overexpression of GRN could enhance macrophage polarization to M2b, a key mediator of the initiation and progression of LN. On the contrary, down-regulation of GRN resulted in impaired M2b differentiation, thus ameliorating LN. Moreover, we found that MAPK signals were necessary for the effect of GRN on macrophage M2b polarization.

Conclusion/Significance

We first demonstrated that GRN could aggravate lupus nephritis (LN) via promoting macrophage M2b polarization, which might provide insights into the pathogenesis of LN as well as potential therapeutic strategies against LN.     

Mannose-Binding Lectin Blunts Macrophage Polarization and Ameliorates Lupus Nephritis

Background

Deficiency in clearance of self nuclear antigens, including DNA, is the hallmark of systemic lupus erythematosus (SLE), a chronic autoimmnue disease characterized by the production of various autoantibodies, immune complex deposition and severe organ damage. Our previous studies revealed that administration of syngeneic BALB/c mice with activated lymphocyte-derived DNA (ALD-DNA) could induce SLE disease. Mannose-binding lectin (MBL), a secreted pattern recognition receptor with binding activity to DNA, has been proved to be a modulator of inflammation, but whether MBL takes responsibility for DNA clearance, modulates the DNA-mediated immune responses, and is involved in the development of DNA-induced SLE disease remain poorly understood.

Methodology/Principal Findings

The levels of serum MBL significantly decreased in lupus mice induced by ALD-DNA and were negatively correlated with SLE disease. MBL blunted macrophage M2b polarization by inhibiting the MAPK and NF-κB signaling while enhancing the activation of CREB. Furthermore, MBL suppressed the ability of ALD-DNA–stimulated macrophages to polarize T cells toward Th1 cells and Th17 cells. Importantly, MBL supplement in vivo could ameliorate lupus nephritis.

Conclusion/Significance

These results suggest MBL supplement could alleviate SLE disease and might imply a potential therapeutic strategy for DNA-induced SLE, which would further our understanding of the protective role of MBL in SLE disease.