Cross-Reactions of Human Lupus Autoantibodies with 8-Methoxypsoralen Photomodified DNA Fragments

DNA fragments of around 200 base pair (average size) have been covalently crosslinked with 8-methoxypsoralen under 365 nm UV light. The photoadduct, induced antibodies in rabbits with a titer of > 1:12,800 by direct bindng ELISA. Binding data showed that the induced antibodies are conformation-specific recognizing restricted conformational change at site of crosslinking. Human autoantibodies against DNA, bound not only to native DNA but to the photomodified DNA fragment as well. In addition, binding patterns of SLE sera obtained from different patients were remarkably similar, indicating the recognition of altered conformation of the modified polymer by naturally occurring SLE anti-DNA autoantibodies.     

IL-21单克隆抗体对MRL/lpr狼疮小鼠的治疗作用*

目的: 探讨IL-21单克隆抗体对MRL/lpr狼疮小鼠的免疫治疗作用。方法: 将20只MRL/lpr狼疮小鼠随机分为模型组和治疗组,每组10只;同年龄同性别C57BL/6小鼠10只作为正常组。治疗组小鼠每周腹腔注射IL-21单克隆抗体(100 μg),正常组及模型组小鼠每周腹腔注射等量生理盐水(100 μg),连续干预8周。干预结束后观察小鼠皮毛、活动等一般性状及浅表淋巴结大小,并采集小鼠血液、尿液及肾脏标本。采用Western blot法检测三组小鼠肾组织中IL-21蛋白表达情况;采用ELISA法比较三组小鼠血清抗ds-DNA抗体、ANA抗体、血尿素氮、肌酐和炎症因子IL-17A、TGF-β1水平;采用生物化学法比较三组小鼠24 h尿蛋白水平。结果: 与正常组比较,模型组小鼠肾组织IL-21、血清抗ds-DNA、ANA抗体水平、尿素氮、肌酐、IL-17A浓度及24 h尿蛋白水平均升高(P＜0.05),TGF-β1浓度降低(P＜0.01);与模型组比较,治疗组小鼠肾组织IL-21、血清抗ds-DNA、ANA抗体水平、尿素氮、肌酐、IL-17A浓度、24 h尿蛋白水平均降低(P＜0.05),TGF-β1浓度升高(P＜0.01)。结论: 腹腔注射IL-21单克隆抗体可改善MRL/lpr狼疮小鼠免疫功能与肾损害,提示治疗机制可能与重塑Th17/Treg相关细胞因子平衡有关。     

EDTA/gelatin zymography method to identify C1s versus activated MMP‐9 in plasma and immune complexes of patients with systemic lupus erythematosus

Gelatin zymography analysis is a sensitive method and commonly used to characterize and quantify the presence of the gelatinases (MMP‐2 and MMP‐9) in biological samples. In human plasma samples from healthy controls and systemic lupus erythematosus (SLE) patients, we observed a gelatinolytic molecule at 80 kDa, suggestive for activated human MMP‐9. However, by developing and using the EDTA/gelatin zymography method and after purification of the 80 kDa entity, we proved that this molecule was the C1s subunit of the complement system. The zymolytic capacity of C1s was validated and found to be enhanced, in the absence of calcium and in the presence of EDTA. Our findings indicate that for correct identification of gelatinolytic proteins in complex biological samples the use of EDTA/gelatin zymography for enzyme development is advised. In addition, by quantification of EDTA/gelatin zymography analysis and ELISA, we observed that the levels of C1s were higher in plasma and immune complexes of SLE patients than of healthy individuals. Therefore, our data imply that C1s may become a marker for the diagnosis of SLE.     

Potential roles and targeted therapy of the CXCLs/CXCR2 axis in cancer and inflammatory diseases

The chemokine receptor CXCR2 and its ligands are implicated in the progression of tumours and various inflammatory diseases. Activation of the CXCLs/CXCR2 axis activates multiple signalling pathways, including the PI3K, p38/ERK, and JAK pathways, and regulates cell survival and migration. The CXCLs/CXCR2 axis plays a vital role in the tumour microenvironment and in recruiting neutrophils to inflammatory sites. Extensive infiltration of neutrophils during chronic inflammation is one of the most important pathogenic factors in various inflammatory diseases. Chronic inflammation is considered to be closely correlated with initiation of cancer. In addition, immunosuppressive effects of myeloid-derived suppressor cells (MDSCs) against T cells attenuate the anti-tumour effects of T cells and promote tumour invasion and metastasis. Over the last several decades, many therapeutic strategies targeting CXCR2 have shown promising results and entered clinical trials. In this review, we focus on the features and functions of the CXCLs/CXCR2 axis and highlight its role in cancer and inflammatory diseases. We also discuss its potential use in targeted therapies.     

Immunogenicity of superoxide radical modified-DNA: studies on induced antibodies and SLE anti-DNA autoantibodies

Superoxide anion radical (SAR) is formed in almost all aerobic cells and it is the most abundant species generated by several enzymatic and non-enzymatic pathways in mammalian tissues, leading to unfavorable alteration of biomolecules including DNA. The SAR-modified macromolecules have been implicated in several disease states including disorders of inflammation. The SAR-induced damage to DNA showed hyperchromicity, single strand breaks, decrease in melting temperature, and modification of bases. Superoxide modified-DNA in rabbits elicited high titer antibodies and showed diverse antigens binding characteristics. The induced antibodies recognized native DNA and other nucleic acid polymers. Anti-DNA IgG from SLE sera, purified on Protein-A-Sepharose matrix, exhibited increased recognition of superoxide anion radical modified-DNA than native DNA in competitive immunoassay. The visual formation of immune complex between induced antibodies and native DNA, and between SLE anti-DNA IgG and superoxide modified-DNA, is a clear indication of property sharing between SLE autoantibodies and experimentally induced antibodies against superoxide modified-DNA.