系统性红斑狼疮小鼠模型的研究近况

系统性红斑狼疮(SLE)小鼠模型的研究对揭示SLE发病原因、发病机制及探索治疗方法具有重要价值。本文介绍了几种SLE小鼠模型的研究近况,以BXSB小鼠为主,同时介绍(NZBxNZW)F1、MRL/lpr及诱导的SLE样小鼠,从小鼠的品系来源、发病特点、免疫学特征、相关基因以及治疗状况等几个方面进行了综述。     

空肠弯曲菌慢性感染诱致自身免疫反应的机理初探

用空肠弯曲菌CJ—S131株感染昆明种小鼠3个月,观察到:(1)感染鼠血清中,出现高滴度的抗ds—DNA和ss—DNA抗体;(2)Con A诱导的抑制细胞功能降低和TH/Ts细胞比值增加;(3)PFC形成和LPS诱致的淋巴细胞转化作用增强:(4)DTH和Con A及PHA诱致的淋巴细胞转化作用增强。提示,CJ—S131感染小鼠使其T_S细胞功能降低,导致T_H细胞功能偏亢,B细胞功能亢进,这可能与自身免疫反应的产生有关。     

T细胞接种对SLE样小鼠防治作用的探讨

本文在空肠弯曲菌（CJ-S131）免疫诱导的小鬼SLE样综合征模型上,观察了T细胞接种（T CellVaccination,TCV）的预防和治疗作用。用于 TCV的细胞为 CJ-S131预致敏的同系小鼠 T细胞。结果显示TCV可明显地抑制CJ-S131诱导的特异性迟发型超敏反应,相反却显著地增强抗dsDNA抗体和抗外膜蛋白抗体生成。因此TCV对SLE样小鼠没有预防及治疗作用。本文对此现象进行了讨论。     

系统性红斑狼疮模型小鼠脾脏细胞凋亡及可能机制

目的:研究系统性红斑狼疮(SLE)模型小鼠的脾脏细胞凋亡及其相关调控机制.方法:采用ConA活化的同系脾脏细胞诱导BALB/c小鼠SLE,通过检测其外周血自身抗体、肾组织病理学改变确定SLE小鼠模型诱导成功.脾脏细胞凋亡的检测采用末端脱氧核苷酸转移酶介导的缺口末端标记法,免疫细胞化学法检测脾脏细胞Bcl-2和NF-кB的表达.结果:ConA活化的同系脾脏细胞成功诱导BALB/c小鼠发生SLE,与盐水对照组和未活化脾脏细胞组比较,SLE模型小鼠脾脏细胞凋亡率明显降低,Bcl-2和NF-кB的表达均增加(P<0.01).结论:SLE模型小鼠脾脏细胞凋亡率降低,可能是通过Bcl-2和NF-кB的表达增加所致.     

ICAM-1 参与系统性红斑狼疮中免疫球蛋白产生的实验研究

目的:探讨ICAM-1在系统性红斑狼疮(SLE)免疫球蛋白产生中的作用。方法:流式细胞术检测分析SLE患者、健康者、anti-CD3 Ab/IFN-α刺激健康者CD4~+T细胞及Pristane诱导的狼疮样小鼠及对照鼠中CD4~+T细胞的活化表型;分选获得Pristane处理小鼠来源的CD4~+T细胞和B细胞,并于体外交叉共培养,加入ICAM-1阻断抗体或同型对照抗体,ELISA检测培养上清中IgG和IgM水平。结果:SLE患者及anti-CD3 Ab/IFN-α刺激健康者CD4~+T细胞表达HLA-DR,CD69和ICOS等分子的细胞比例明显高于对照组。Pristane诱导的狼疮样小鼠CD4~+T细胞的活化程度也显著高于对照;Pristane诱导的狼疮样小鼠血清中抗snRNP和抗dsDNA自身抗体水平显著高于对照组;Pristane处理小鼠来源的活化CD4~+T细胞与B细胞共培养可以促进B细胞产生IgM和IgG。而经抗ICAM-1抗体阻断处理的培养上清中IgM含量低于对照组,而IgG含量显著高于对照组。结论:Pristane来源的活化CD4~+T细胞可促进B细胞分化,ICAM-1分子可能在B细胞分化早期发挥作用,但不利于IgM型向IgG型的类别转换,为ICAM-1在SLE中抗体类别转化的分子机制提供新的理论依据。     

褪黑素通过下调ERK和AKT活化来抑制Toll样受体9触发的巨噬细胞促炎症因子产生

<正>目的:研究褪黑素(melatonin,MT)对Toll样受体9(TLR9)配体CpG诱导巨噬细胞产生促炎症因子的影响及其机制。方法:先采用不同浓度MT处理小鼠腹腔巨噬细胞或小鼠巨噬细胞系Raw264.7,然后采用CpG刺激细胞;通过定量PCR检测MT受体MT1和MT2表达,通过ELISA和定量PCR检测细胞因子表达,通过Western blot检测信号转导蛋白的活化。结果:     

活化淋巴细胞与慢性GVHR诱导的SLE样小鼠模型的比较

目的 :将本室建立的用活化淋巴细胞诱导的系统性红斑狼疮 (SLE)样小鼠模型与国际上公认的用慢性GVHR诱导的SLE样小鼠模型进行比较 ,进一步探索SLE的发病机理。方法 :分别将亲代Balb c小鼠淋巴细胞经静脉和用ConA活化的(Balb c×C5 7BL 6 )F1代小鼠淋巴细胞经皮下途径输入F1代小鼠 ,用ELISA测定IgG类抗dsDNA抗体和抗组蛋白抗体 ,用免疫荧光法检测抗核抗体 (ANA)荧光核型和肾小球内免疫复合物沉积 ,用免疫印迹法检测抗可溶性核抗原 (ENA)抗体。结果 :亲代淋巴细胞免疫F1代小鼠所致的慢性GVHR和活化F1代小鼠淋巴细胞均可诱导F1代小鼠产生高滴度的抗dsDNA抗体、抗组蛋白抗体等ANA ,并且肾脏都有明显的IgG类免疫复合物沉积。但亲代淋巴细胞免疫组ANA核型以颗粒型、核仁型为主 ,ENA多肽谱多在 32、47、6 7kD处显色 ;而活化淋巴细胞免疫组以胞浆型、周边型、均质型为主 ,ENA多肽谱在 2 8、47、6 7kD处显色。结论 :这 2种方法均可诱导出SLE样综合征 ,但其抗核抗体谱有所不同。     

1-磷酸鞘氨醇受体2抑制脂多糖诱导的急性肺损伤

 目的: 探讨1-磷酸鞘氨醇受体2(S1P2R)对脂多糖(LPS)诱导的急性肺损伤(ALI)中的作用及机制。方法: 野生小鼠和S1pr2^-/-小鼠经气管滴注LPS,建立急性肺损伤动物模型。LPS注射24 h时观察肺组织的病理改变,测定支气管肺泡灌洗液(BALF)中的蛋白浓度、总细胞数、中性粒细胞的比值及TNF-α、IL-6细胞因子的表达。为了观察S1P2R在肺损伤中的作用机制,LPS注射10 min前野生小鼠和S1pr2^-/-小鼠经尾静脉注射一氧化氮合酶抑制剂L-NAME,LPS注射12 h时,再观察肺的病理组织学变化以及BALF中的蛋白浓度,总细胞数及TNF-α、IL-6细胞因子表达的变化。结果: 与野生小鼠比较,S1pr2^-/-小鼠恶化LPS诱导的急性肺损伤,BALF中的蛋白浓度、总细胞数,中性粒细胞比值及炎症细胞因子表达显著增加。而L-NAME的预处理显著抑制在S1pr2^-/-小鼠LPS诱导加重的急性肺损伤。结论: S1P2R通过抑制NO合成,维持血管屏障,从而抑制急性肺损伤。     

系统性红斑狼疮合并动脉粥样硬化的小鼠模型建立和评价

目的动脉粥样硬化(AS)相关心血管病变是系统性红斑狼疮(SLE)患者最主要的致病和死亡因素,但是其具体机制不明。本研究旨在建立SLE合并AS的小鼠模型并进行评估,为研究SLE合并AS的机制和治疗奠定基础。方法将SLE模型(Fasl~(-/-))和动脉粥样硬化模型(apoE~(-/-))小鼠进行杂交纯化;采用PCR方法鉴定小鼠的基因型;检测小鼠血清中肾功能和血脂等生化指标;分析肾脏和心脏等病理改变。结果 PCR结果显示SLE和AS杂合小鼠为Fasl和apo E基因纯合突变;小鼠血清肌酐增高,血清中出现抗ds-DNA抗体。血清中总胆固醇、甘油三酯和低密度脂蛋白增加,高密度脂蛋白降低;小鼠肾脏出现狼疮性肾炎病理改变,心脏有明显动脉粥样斑块沉积。结论 SLE和AS模型小鼠杂合纯化的小鼠,同时表现出SLE和AS样病变,成功建立了SLE合并AS的小鼠模型,为后续研究奠定了基础。     

C3d对基因免疫诱导的HBV特异性体液免疫应答的调节作用

观察补体C3d对HBV基因免疫诱导的特异性体液免疫应答的调节作用 ,为增强HBV基因疫苗免疫效果寻求新途径。将HBV preS2 /S编码基因分别插入真核表达载体TR4 2 1和含有 3拷贝C3d编码基因的TR4 2 1 C3d3质粒 ,构建重组质粒TR4 2 1 preS2 /S和TR4 2 1 preS2 /S C3d3。采用肌肉注射法对BALB/c小鼠实施基因免疫 (10 0 μg/ 10 0 μl/只小鼠 ) ,以空质粒为对照 ,定期采集血清。ELISA法检测免疫小鼠血清特异性抗 HBs IgG及其亚型 ,并采用NaSCN竞争ELISA法检测其亲合力。结果表明 ,TR4 2 1 preS2 /S C3d3重组质粒免疫组诱导的特异性抗 HBs IgG水平明显高于TR4 2 1 preS2 /S重组质粒免疫组 (P <0 0 5 ) ;而且TR4 2 1 preS2 /S C3d3重组质粒免疫组诱导的抗 HBs IgG抗体的亲合力 (ED50 :1 375 )显著高于TR4 2 1 preS2 /S重组质粒组 (ED50 :0 875 ) ,但C3d并不改变基因免疫诱导的特异性抗HBs IgG各亚型水平的格局 ,仍以IgG2b和IgG2a为主。提示C3d可增强基因免疫诱导的HBV特异性体液免疫应答 ,并促进特异性抗体亲和力的成熟 ,这为提高HBV基因疫苗的免疫效果提供了新的途径。     

Beneficial Effect of Eucommia Polysaccharides on Systemic Lupus Erythematosus-like Syndrome Induced by Campylobacter jejuni in BALB/c Mice

The stem bark of Eucommia ulmoides Oliv. is commonly used for the treatment of hypertension, rheumatoid arthritis, lumbago, and ischialgia in traditional Chinese medicine. This study was to determine whether the crude polysaccharides (EUPs) isolated from the stem bark of E. ulmoides had beneficial effects on lupus-like syndrome in mice. BALB/c mice were immunized with CJ-S₁₃₁ in Freund’s complete adjuvant on day 0, and then boosted on day 14. EUPs 15 or 30 mg kg⁻¹·day⁻¹, or prednisone 5 mg kg⁻¹·day⁻¹ was given to BALB/c mice intragastrically from day 0 to 34. Treatment with EUPs 15 or 30 mg kg⁻¹·day⁻¹ for 35 days protected kidney from glomerular injury with reduced immunoglobulin deposition and lowered proteinuria. The increased production of serum autoantibodies and total immunoglobulin G (IgG) was also inhibited. These findings suggested that Eucommia polysaccharides had a beneficial effect on systemic lupus erythematosus-like syndrome induced by CJ-S₁₃₁ in BALB/c mice.     

CR1/CR2 deficiency alters IgG3 autoantibody production and IgA glomerular deposition in the MRL/lpr model of SLE

CR1 and CR2 expression is decreased by approximately 50% on B cells of patients with systemic lupus erythematosus (SLE). Expression is also decreased in the MRL/lpr murine model of SLE prior to the development of clinical disease, suggesting that this alteration may play a role in pathogenesis. To determine whether the decrease in receptor levels affects the development of SLE, we analyzed MRL/lpr mice in which CR1/CR2 expression was altered by gene targeting. Mice from each cohort (Cr2+/+, Cr2+/-, and Cr2-/-) were analyzed biweekly for the development of proteinuria and autoantibodies. Kidneys were examined at 12 and 16 weeks for evidence of immune complex deposition and renal disease. Deficiency of CR1/CR2 did not affect survival or development of renal disease as measured by proteinuria. Mice deficient in CR1/CR2 had significantly lower levels of IgG3 rheumatoid factor (RF) and total serum IgG3, suggesting a specific defect in production of IgG3 in response to endogenous autoantigens. Since IgG3 RF has been associated with the development of vasculitis in this model, we examined the mice for alterations in development of this clinical manifestation. Although there was no difference in the development of ear necrosis among the three groups, renal arteritis was not identified in any of the Cr2+/- mice, whereas it was present in 20% of the Cr2+/- and 40% of the Cr2+/+ mice. Finally, significantly higher levels of IgA were seen in the glomeruli of Cr2+/- mice compared to Cr2+/- or Cr2+/+ mice, suggesting that CR1/CR2 are involved in either the regulation of IgA production or the clearance of IgA immune complexes. Together these data support the concept that alterations in CR1/CR2 expression or function affect the regulation of autoantibody production and/or clearance and may have clinical consequences.     

小鼠脾淋巴细胞可能存在5HT_3受体

本文用3H TdR掺入法和MTT法观察 5HT3受体激动剂 1 phenylbiguanide(PBG )、 5HT3受体拮抗剂格拉司琼 (granisetron )和托烷司琼 (tropisetron )对体外培养ICR小鼠脾淋巴细胞ConA ,LPS刺激的增殖和NK细胞活性的影响。结果表明PBG ( 10 6～ 10 4mol/L )抑制ConA刺激的脾细胞增殖反应和脾细胞IL 2的生成 (P <0 0 5 ) ;增强LPS剌激的脾细胞增殖反应 (P <0 0 5 ) ;格拉司琼和托烷司琼双相影响 ,即在低浓度 ( 10 7～ 10 6 mol/L )促进、在较高浓度 ( 10 5～ 10 4mol/L )抑制ConA刺激的增殖反应 (P <0 0 5 ) ;二药均浓度依赖抑制LPS刺激的脾细胞增殖效应。PBG对脾淋巴细胞增殖的影响被同时加入格拉司琼或托烷司琼拮抗 ,格拉司琼和托烷司琼减弱PBG 10 5mol/L对脾细胞增殖反应的影响。本实验 5HT3受体激动剂或拮抗剂浓度不明显影响脾NK细胞活性和无丝裂原刺激的增殖反应。结果提示小鼠脾T细胞和B细胞表面可能存在对淋巴细胞增殖有不同影响的 5HT3受体     

空肠弯曲菌与可提取核抗原的分子交叉反应

取空肠弯曲菌（ＣＪ－Ｓ１３１）免疫的Ｂａｌｂ／ｃ小鼠脾细胞与骨髓瘤细胞ＳＰ２／Ｏ融合，制备出４种能与空肠弯曲菌和可提取核抗原（ＥＮＡ）起反应的单抗。以Ｈｅｐ－２细胞作基质的免疫荧光染色表明，４种单抗均呈阳性结果，证实为抗核抗体。当单抗腹水被空肠弯曲菌菌体吸收后，其抗ＥＮＡ的活性明显降低，证明这４种单抗与空肠弯曲菌ＣＪ－Ｓ１３１和ＥＮＡ存在交叉反应性。免疫印渍试验表明，４种单抗均与空肠弯曲菌ＣＪ－Ｓ１３１的４３ｋＤ外膜蛋白反应。这一结果提示：空肠弯曲菌ＣＪ－Ｓ１３１的４３ｋＤ外膜蛋白与ＥＮＡ之间可能存在相似的分子构象，这为自身免疫病发机制的分子模拟假说提供了实验的依据。     

Differential effects of oestrogen in murine lupus: acceleration of glomerulonephritis and amelioration of T cell-mediated lesions.

Oestrogen is known to accelerate glomerulonephritis and autoantibody production in human and murine systemic lupus erythematosus (SLE). In this study we demonstrate that treatment of castrated autoimmune MRL +/+ mice with physiological doses of oestrogen results in enhanced immunoglobulin and autoantibody production as well as increased deposition of IgG in renal glomeruli. Accelerated development of glomerulonephritis was also evident from the increase of albuminuria. Interestingly, in contrast to these deteriorative effects of oestrogen on immune complex-mediated disease we now show that the lymphocytic infiltrations in the submandibular glands and perivascular lesions in the kidneys were significantly diminished after exposure to oestrogen. This remarkable impact of physiological oestrogen levels on the outcome of SLE in MRL +/+ mice is postulated to be the result of a differential effect on T and B cell-mediated immune responses.     

Delivering PD-1 inhibitory signal concomitant with blocking ICOS co-stimulation suppresses lupus-like syndrome in autoimmune BXSB mice

BXSB mice spontaneously develop an autoimmune syndrome characterized by hypergammaglobulinemia, autoantibody production, and the development of fatal glomerulonephritis that closely resembles systemic lupus erythematosus (SLE) in humans. While blocking positive T cell co-stimulation has shown effectiveness in preventing the onset of murine lupus, deliberate delivering negative co-stimulation to halt unwanted T and B cell activation has not been tested. We developed a recombinant adenovirus containing the full-length mouse PD-L1 gene (Ad.PD-L1) to engage the immunoinhibitory receptor PD-1 on activated lymphocytes to prevent lupus nephritis in BXSB mice. This strategy was further reinforced by concomitant injection of anti-ICOSL(B7h) mAb to block ICOS-mediated co-stimulation. The combined therapy dramatically delayed the onset of proteinuria, effectively inhibited IgG autoantibody production, and significantly reduced hypercellularity and deposition of IgG in glomeruli, resulting in almost complete amelioration of lupus nephritis in these animals. Our results indicate the therapeutic potential of simultaneous stimulation of PD-1-mediated pathway and blockade of ICOS-B7h co-stimulation in the prevention of human lupus nephritis.     

甲醛化空肠弯曲菌(CJ-S_(131))与佐剂诱导的自身免疫动物模型

甲醛化CJ-S_(131)辅以佐剂免疫小鼠,可诱导自身免疫反应。模型鼠出现的T、B细胞功能改变,血清出现高水平的自身抗体,在肝、肾、肠等组织出现的炎症性病理损伤,均类似于CJ-S_(121)慢性感染的小鼠。本模型优点在于实验周期短,用菌体抗原免疫小鼠代替复杂的感染过程,而诱导自身免疫反应。本模型可用于诸如免疫调节剂的筛选,自身免疫的分子机理等自身免疫反应的实验研究。     

Alpha‐melanocyte stimulating hormone ameliorates disease activity in an induced murine lupus‐like model

Alpha‐melanocyte stimulating hormone (α‐MSH) is a neuropeptide exhibiting anti‐inflammatory activity in experimental models of autoimmune diseases. However, no studies thus far have examined the effects of α‐MSH on systemic lupus erythematosus (SLE). This study aimed to determine the effects of an α‐MSH agonist in induced murine lupus. Here we employed female Balb/cAn mice in which lupus was induced by pristane. Groups of lupus animals were treated daily with the α‐MSH analogue [Nle4, DPhe7]‐α‐MSH (NDP–MSH) (1·25 mg/kg) injected intraperitoneally or saline for 180 days. Normal animals comprised the control group. Arthritis incidence, plasma immunoglobulin (Ig)G isotypes, anti‐nuclear antibodies (ANA) and plasma cytokines were evaluated. Renal function was assessed by proteinuria and histopathological lesion. Glomerular levels of IgG, α‐smooth muscle actin (α‐SMA), inducible nitric oxide synthase (iNOS), C3, CD3, melanocortin receptors (MCR)1, corticotrophin‐releasing factor (CRF) and α‐MSH was estimated by immunohistochemistry. When compared with normal controls, lupus animals exhibited increased arthritis, IgG levels, ANA, interleukin (IL)‐6, IL‐10, proteinuria and mesangial cell proliferation together with glomerular expression of α‐SMA and iNOS. Glomerular expression of MCR1 was reduced in lupus animals. NDP‐MSH treatment reduced arthritis scores by 70% and also diminished IgG1 and IgG2a levels and ANA incidence. In the glomerulus, NDP–MSH treatment reduced cellularity by 50% together with reducing IgG deposits, and expression levels of α‐SMA, iNOS and CRF were also all decreased. Taken together, our results suggest for the first time that α‐MSH treatment improves several parameters of SLE disease activity in mice, and indicate that this hormone is an interesting potential future treatment option.     

Immunopathology of early and clinically silent lupus nephropathy.

Detailed immunopathologic studies of early or silent renal alterations in systemic lupus erythematosus have been sparse. The renal biopsies of 16 lupus patients with normal renal function, including 8 with hematuria and/or proteinuria of recent onset, and 8 without clinically detectable renal disease were investigated by light, immunofluorescence, and electron microscopy. Immunoglobulins, complement components, and electron-dense deposits were detected in glomeruli of all patients, regardless of morphologic appearance or lack of clinical evidence of renal involvement. Features of membranous glomerulonepritis were observed in 4 patients with substantial proteinuria. In the remaining 12 patients, including 3 with hematuria and 4 with slight proteinuria, either minimal glomerular alterations or features of mesangial proliferative glomerulonephritis were seen. Transformation of the original disease was demonstrated in 3 of 3 patients rebiopsied within 2 years. The significance of these findings is discussed in relation to a) the spectrum of clinical and immunopathologic alterations in lupus nephritis and b) transformation of the original disease.