三氧化二砷对BXSB狼疮鼠生存时间及脾细胞分泌白细胞介素-4干扰素-γ的影响

目的探讨三氧化二砷（ATO）对BXSB狼疮小鼠生存时间、外周血抗dsDNA抗体水平及离体脾脏单个核细胞白细胞介素（IL）-4、干扰素（IFN）-γ分泌水平的影响。方法BXSB狼疮小鼠44只，随机分为2组，治疗组予腹腔内注射ATO，对照组注射等量生理盐水，酶联免疫吸附试验（ELISA）测其外周血抗dsDNA抗体水平，并将生存时间作比较；另外，通过体外培养脾脏单个核细胞比较C57和BXSB小鼠脾脏单个核细胞分泌IL-4、IFN-γ水平的差异，然后观察ATO对BXSB小鼠脾脏单个核细胞IL-4、IFN-γ分泌水平的影响，同时观察ATO对细胞活性的影响。结果ATO可延长BXSB狼疮小鼠的生存时间，显著降低其外周血中抗dsDNA抗体的水平；1μmol／L浓度的ATO对小鼠脾脏单个核细胞的活性无明显影响：与C57小鼠相比，BXSB狼疮小鼠脾脏单个核细胞在体外经有丝分裂原刺激之后分泌IL-4、IFN-γ水平较高，ATO能显著抑制其分泌。结论ATO可延长BXSB狼疮鼠的生存时间，降低其外周血抗dsDNA抗体的水平．提示ATO对BXSB狼疮鼠有一定的治疗作用；ATO抑制该小鼠脾脏单个核细胞IL-4和IFN-γ的分泌可能是其发挥治疗作用的重要机制之一。     

大黄和黄芪水提醇沉液注射治疗BXSB狼疮鼠的实验研究

目的 为了开发治疗系统性红斑狼疮（SLE）的新药，我们应用大黄、黄芪溶液注射治疗BXSB狼疮样小鼠，以观察其疗效。方法 应用水提醇沉法制取大黄黄芪溶液，15只BXSB雄性小鼠及6只雌鼠被随机分成三组，分别应用大黄黄芪溶液、地塞米松及生理盐水腹部皮下注射。考马斯亮蓝分光度法被用来检测小鼠尿蛋白含量，应用免疫荧光法检测小鼠血清抗核抗体（ANA）滴度及肾内免疫复合物沉积。结果 中药组及地塞米松组BXSB小鼠的尿蛋白含量及血清ANA滴度均明显低于生理盐水组（P＜0．01或P＜0．05），两组肾内免疫复合物沉积也轻于生理盐水组（P＜0．05）加药组BXSB小鼠的24h尿蛋白含量低于地塞米松组（P＜0．05）。结论 大黄、黄芪水提醇沉液可降低BXSB小鼠尿蛋白含量及血清ANA滴度，减少肾内免疫复合物沉积，对BXSB狼疮样小鼠有较为肯定的疗效。     

系统性红斑狼疮的基因表达谱及其免疫调控通路的研究

目的 通过基因表达谱勾画系统性红斑狼疮（SLE）发病机制中可能的免疫调控通路。方法 用寡核苷酸基因芯片研究了10例SLE（包括2例SLE同胞对）和10份正常对照的外周血白细胞基因表达谱。进行了基因表达谱和临床免疫表型的聚类分析和比较，筛选统计学显著性差异表达的SLE相关基因，并在转录和蛋白表达水平验证芯片表达谱结果。结果 聚类分析揭示SLE临床免疫表型与基因表达谱特征存在关联。获得25个显著性表达的SLE相关基因，其中大部分未经报道。转录水平直接验证了其中C／EBPD，LY6E，OAS2三个基因的表达，较大样本独立验证了C／EBPD，LY6E在SLE中的表达上调；蛋白质水平验证了C／EBPD的高表达。结论 SLE基因表达谱可能是其临床免疫表型的分子基础，探讨了通过基因表达谱进行SLE分型乃至鉴别诊断的可能性。通过显著性差异表达的SLE相关基因，推测干扰素及其免疫调控通路在SLE发病中可能扮演重要角色。     

应用基因芯片技术研究原发性胆汁性肝硬化患者外周血单个核细胞基因表达谱特征

目的 应用基因芯片技术分析原发性胆汁性肝硬化患者外周血单个核细胞的基因表达谱特征.方法 选取9例原发性胆汁性肝硬化患者、9名健康对照为研究对象,提取其外周血单个核细胞总RNA,进行人类全基因组寡核苷酸芯片(约22000个基因)检测,筛选出差异表达基因及相关信号通路.结果 原发性胆汁性肝硬化患者外周血单个核细胞中差异表达的基因共有79个,其中21个表达上调,58个表达下调.这些基因对应着27条信号通路,其中有6条通路参与了免疫调控及细胞凋亡过程,分别是自然杀伤细胞介导的细胞毒通路、Toll样受体信号通路、抗原加工提呈通路、细胞因子相互作用通路、T细胞受体信号通路、细胞凋亡通路.结论 原发性胆汁性肝硬化患者外周血单个核细胞中存在特有的差异表达基因,针对这些基因及相关信号通路的进一步研究,将为探索发病机制和寻找生物标志物提供新的方向.     

乙型肝炎病毒相关的肝硬化患者外周血单个核细胞基因表达谱的研究

目的分析乙型肝炎病毒（HBV）相关的肝硬化患者外周血单个核细胞的基因表达谱，探索慢性HBV相关的肝硬化形成的分子机制。方法应用含14 000条人体cDNA的微阵列芯片和来自外周血单个核细胞的标记cDNA，分析了10例慢性HBV相关肝硬化患者和10例健康人基因表达谱。通过应用GenePix 4000B扫描仪和ImaGene3．0分析软件比较Cy5标记的慢性HBV相关肝硬化来源cDNA与Cy3标记的健康人来源cDNA的杂交结果，获得个体基因的相对表达比值。结果和14 000条基因中，差异表达的基因有199条，占1．42％。其中108条基因表达水平上调，91条基因表达水平下调。这些差异表达的基因主要为细胞信号转导．细胞周期和代谢、凋亡及炎症相关类基因。结论HBV相关肝硬化发病过程中，发观众多基因的差异表达，为进一步阐明慢性HBV相关的肝硬化形成的分子机制提供基础。     

移植雄性骨髓的雌性小鼠肝组织肝再生相关基因的信号通路

目的:探讨骨髓形成肝细胞的分子机制.方法:采用移植♂骨髓的♀小鼠模型,♀Balb/c小鼠随机分为模型组和正常对照组,选用小鼠基因表达谱寡核苷酸芯片,利用反转录酶合成掺有荧光标记的cDNA探针,将探针与基因表达谱芯片杂交观察小鼠肝组织基因表达谱的变化,筛选样本之间杂交信号比值有差异表达的基因.采用生物信息学方法分析模型组中小鼠肝组织再生相关基因的信号通路变化规律.结果:♀小鼠在移植♂骨髓6 mo后肝组织的基因表达谱显著不同,对照组相对于正常组的差异表达基因有865条,已知功能基因有447条,其中上调基因92条,下调基因355条.与肝再生相关基因信号通路涉及HGF,TGF-β,Focal Adhesion,JAK-Stat,VEGF等信号通路,他们相关基因的上调表达促进肝细胞的增殖分化.TGF-β信号通路中相关基因下调,抑制信号通路的激活,减弱肝再生的负性作用,利于肝细胞的增殖分化.结论:♀小鼠移植♂骨髓后的肝组织基因表达谱显著变化,有可能通过其肝再生相关基因的信号通路激活或抑制调节肝再生.     

应用基因芯片检测系统性红斑狼疮患者外周血B淋巴细胞

目的 应用寡聚核苷酸基因表达芯片研究系统性红斑狼疮(SLE)患者与健康志愿者外周血B淋巴细胞基因表达谱的变化.方法 应用包含21522条70 mer长度Oligo DNA的寡聚核苷酸芯片(22K Human Genome Array)分别检测6例SLE患者和6名健康志愿者外周血B淋巴细胞基因表达谱,应用聚类分析法分析差异表达的基因.结果 SLE患者外周血B淋巴细胞和健康志愿者B淋巴细胞之间基因表达谱有显著差异,芯片中聚类表达显著上调的基因15个,主要包括Ⅰ型干扰素(IFN)通路中的相关分子;聚类表达显著下调的基因22个,主要包括神经内分泌相关的精氨酸甲基化转移酶家族等与雌激素代谢相关的基因.进一步对这些差异表达基因进行了初步的功能分类分析,分析结果表明这些基因主要与B淋巴细胞的JAK-STAT信号通路和激素代谢相关.结论 SLE患者外周血B淋巴细胞基因表达谱存在显著差异,芯片显示氨基酸代谢、激素代谢、细胞增殖和凋亡、炎症反应等相关生物学功能的改变可能参与SLE的发病.     

日本血吸虫感染过程中抑制性因子的表达特征

目的立足于基因组水平考察日本血吸虫慢性感染小鼠CD_4~+ T应答中抑制性因素的参与及可能的调控作用。方法采用高密度寡核苷酸芯片(Affymetrix芯片)对日本血吸虫感染0、3、6、13周小鼠脾脏中的CD_4~+ T细胞进行全基因组分析,获得相关抑制性因子的基因表达谱。结果基因芯片结果显示日本血吸虫感染从急性期至慢性期过程中抑制性因子的表达基因表达逐步上调,呈现基本一致的变化,包括抑制性细胞因子、阻断抗体、细胞凋亡、负调控分子的高表达等。结论从胞内信号转导通路至细胞凋亡的整体水平,多层次因素参与了日本血吸虫感染宿主免疫应答的负调控机制。     

采用cDNA芯片检测Krüppel样因子4过表达对C2C12细胞基因表达谱的影响

目的采用cDNA芯片检测Krüppel样因子4过表达对C2C12细胞中基因表达谱的影响。方法用cDNA芯片检测Krüppel样因子4过表达的C2C12细胞中基因表达谱的改变(以空载体细胞为对照);用MEDLINE、Genomatix等生物信息学数据库和分析软件对在Krüppel样因子4过表达的C2C12细胞中表达发生改变的已命名基因进行功能和细胞信号通路分析。结果采用cDNA芯片筛选到在Krüppel样因子4过表达的C2C12细胞中表达发生改变的基因605个;其中下调的基因为250个,已命名基因为155个;上调的基因为355个,已命名基因为255个;其中包含多个炎症相关基因和凋亡相关基因。上述炎症相关基因属于13条细胞信号通路,凋亡相关基因属于9条细胞信号通路。结论Krüppel样因子4过表达能够引起C2C12细胞中多个下游基因表达发生改变。     

MRL/lpr狼疮小鼠CD4~+ CD25~+调节性T细胞和相关基因的研究

目的 通过研究MRL/lpr系统性红斑狼疮(SLE)小鼠(模型组)胸腺、脾脏中CD4+CD25+调节性T细胞(Tr)数量和相关基因的变化,探讨Tr在SLE发病中的作用.方法 采用间接免疫荧光法检测模型组和对照组血清中抗核抗体水平,流式细胞术检测胸腺、脾脏中Tr数量,RT-PCR检测Tr功能基因表达水平.结果 抗核抗体模型组均为阳性,而对照组均为阴性;胸腺、脾脏中Tr数量模型组与对照组无明显差别;Foxp3的表达水平在模型组胸腺中与对照组无明显差别,而在模型组脾脏中却低于对照组;CTLA-4的表达水平在模型组胸腺和脾脏均高于对照组.结论 Tr数量的变化在MRL/lpr小鼠的发病中不起关键作用,而Tr功能的变化,尤其在外周的功能缺陷可能是其发病的原因之一,至于Tr的抑制功能是否真正发生变化,还有待于进一步通过体外抑制功能的检测来确定.     

Effects of FTY720 on BXSB lupus‐prone mice: a preliminary study

Aim: To examine the effects of a novel immunosuppressant, FTY720, on BXSB lupus‐prone mice, and to investigate its immune regulatory pathway by using a cDNA microarray. Methods: Male BXSB mice received oral administration of 10 mg/kg FTY720 twice a week from age 9 weeks. Levels of antinuclear antibodies in serum and histopathology of the kidney were evaluated. The gene expression profile of splenic lymphocyte was analysed by cDNA microarray. Results: FTY720 suppressed the production of antinuclear antibodies (P < 0.05) and reduced the deposition of IgG in glomeruli, compared to control animals. The microarray determined 247 differential expressed genes, which contained a cluster of genes related to lymphocyte trafficking and cytoskeletal remodeling. Conclusion: Our preliminary data suggests FTY720 has some immune suppressive effect on BXSB mice. The mechanism may be due to accelerate lymphocyte homing and redistribution which differs from traditional cytotoxic agents. Its clinical implication deserves further study.     

Effects of FTY720 in MRL-lpr/lpr mice: therapeutic potential in systemic lupus erythematosus

OBJECTIVE To examine the effects of a novel immunosuppressant, FTY720, on hematolymphoid cells and the clinical course of MRL-lpr/lpr (MRL/lpr) mice genetically predisposed to systemic lupus erythematosus (SLE). METHODS: Apoptosis of hematolymphoid cells was determined in vitro by FACScan after staining with propidium iodide or merocyanine 540. From 4 months of age, 15 female MRL/lpr mice received oral administration of 2 mg/kg each of FTY720, methylprednisolone (mPSL), or vehicle, 3 times per week. Therapeutic efficacy was evaluated by levels of anti-dsDNA antibodies in serum and the survival rate. In parallel, T cell proliferation and secretion of interleukin 2 (IL-2) induced by anti-CD3, phenotypes of the spleen, lymph node and bone marrow cells, as well as immunohistochemistry of the kidney, were examined in vitro. RESULTS: FTY720 at 2 microM induced apoptosis in more than 70% of double negative (CD4-/CD8-) T cells from the spleen of MRL/lpr mice in vitro. Oral FTY720 was tolerated well with no apparent side effects. FTY720 treated and control mice gained weight at an identical pace through to 9 months of age. FTY720 significantly suppressed the production of anti-dsDNA antibodies (FTY720 vs control: 1739 +/- 898 U/ml vs 410 +/- 356 U/ml at 8 months of age; p < 0.05) and reduced the deposition of IgG in glomeruli compared to control animals. At 9 months of age, the survival rate in the FTY720 treated mice was 86.9% compared to 33.0% in controls (p < 0.01). FTY720 decreased the number of double negative T cells from the spleen and lymph nodes in vivo, and increased T cell proliferation and IL-2 secretion induced by anti-CD3 stimulation in vitro. CONCLUSION: FTY720 suppressed the development of autoimmunity and prolonged the lifespan of female MRL/lpr mice. Suppression of autoimmunity, at least in part, may have resulted from an apoptogenic potential of FTY720. Hence, it could be useful for primary or adjunctive therapy of human SLE.     

The Sphingosine-1 Phosphate receptor agonist FTY720 dose dependently affected endothelial integrity in vitro and aggravated ventilator-induced lung injury in mice

Lung barrier protection by Sphingosine-1 Phosphate (S1P) has been demonstrated experimentally, but recent evidence suggests barrier disruptive properties of high systemic S1P concentrations. The S1P analog FTY720 recently gained an FDA approval for treatment of multiple sclerosis. In case of FTY720 treated patients experiencing multiple organ dysfunction syndrome the drug may accumulate due to liver failure, and the patients may receive ventilator therapy. Whereas low doses of FTY720 enhanced endothelial barrier function, data on effects of increased FTY720 concentrations are lacking. We measured transcellular electrical resistance (TER) of human umbilical vein endothelial cell (HUVEC) monolayers, performed morphologic analysis and measured apoptosis by TUNEL staining and procaspase-3 degradation in HUVECs stimulated with FTY720 (0.01-100 μM). Healthy C57BL/6 mice and mice ventilated with 17 ml/kg tidal volume and 100% oxygen for 2 h were treated with 0.1 or 2 mg/kg FTY720 or solvent, and lung permeability, oxygenation and leukocyte counts in BAL and blood were quantified. Further, electron microscopic analysis of lung tissue was performed. We observed barrier protective effects of FTY720 on HUVEC cell layers at concentrations up to 1 μM while higher concentrations induced irreversible barrier breakdown accompanied by induction of apoptosis. Low FTY720 concentrations (0.1 mg/kg) reduced lung permeability in mechanically ventilated mice, but 2 mg/kg FTY720 increased pulmonary vascular permeability in ventilated mice accompanied by endothelial apoptosis, while not affecting permeability in non-ventilated mice. Moreover, hyperoxic mechanical ventilation sensitized the pulmonary vasculature to a barrier disrupting effect of FTY720, resulting in worsening of ventilator induced lung injury. In conclusion, the current data suggest FTY720 induced endothelial barrier dysfunction, which was probably caused by proapoptotic effects and enhanced by mechanical ventilation.     

Effect of 2-amino-2-[2-（4-octylphenyl） ethyl] propane-1,3-diol hydrochloride （FTY 720） on immune liver injury in mice

AIM: To investigate the protective effect against two immune liver injury models in mice by 2-amino-2-[2-(4-octylphenyl) ethyl] propane-l,3-diol hydrochloride and its possible mechanisms in Con A-induced liver damage. METHODS: Liver tissue or hepatocyte injury was monitored biochemically by measuring alanine aminotransferase (sALT) and aspartate aminotransferase (sAST) activity. Hematoxylin & eosin (HE) staining was used for histopathological examination. To evaluate the role of IFN-γ and IL-4 in the liver injury, serum levels of IFN-γ and IL-4 were determined using commercially available ELISA kit at 12 h after Con A challenge. We also determined FTY 720-induced spleen cell apoptosis by flow cytometry analysis or spleen cell proliferation test. RESULTS: Different doses of FTY 720 treatment dramatically reduced circulating markers of hepatocyte injury in two kinds of immunological liver injury models. FTY 720 dramatically reduced the elevated serum IFN-γ and IL-4 levels after Con A injection. Effect of spleen cell supernatants treated with Con A or FTY 720 on hepatocytes showed that ALT activities in cultured hepatocyte supernatants in Con A treatment group increased markedly and FTY 720 could reduce this elevated ALT activities in FTY 720 treatment group. FTY 720 dose-dependently increased the percentage of apoptotic cells in T cells and inhibited splenocyte proliferation induced by Con A. CONCLUSION: Pretreatment with FTY 720 was shown to produce protective effect on the immune liver injury in mice. The possible mechanism of FTY 720 on Con A-induced liver damage is that it could inhibit lymphocyte proliferation and induce lymphocyte apoptosis, resulting in the reduction of IL-4 or IFN-γ release, and subsequently protecting liver from being damaged by Con A.     

Molecular profiling of LGL leukemia reveals role of sphingolipid signaling in survival of cytotoxic lymphocytes

T-cell large granular lymphocyte (LGL) leukemia is characterized by clonal expansion of CD3⁺CD8⁺ cells. Leukemic LGLs correspond to terminally differentiated effector-memory cytotoxic T lymphocytes (CTLs) that escape Fas-mediated activation-induced cell death (AICD) in vivo. The gene expression signature of peripheral blood mononuclear cells from 30 LGL leukemia patients showed profound dysregulation of expression of apoptotic genes and suggested uncoupling of activation and apoptotic pathways as a mechanism for failure of AICD in leukemic LGLs. Pathway-based microarray analysis indicated that balance of proapoptotic and antiapoptotic sphingolipid-mediated signaling was deregulated in leukemic LGLs. We further investigated sphingolipid pathways and found that acid ceramidase was constitutively overexpressed in leukemic LGLs and that its inhibition induced apoptosis of leukemic LGLs. We also showed that S1P₅ is the predominant S1P receptor in leukemic LGLs, whereas S1P₁ is down-regulated. FTY720, a functional antagonist of S1P-mediated signaling, induced apoptosis in leukemic LGLs and also sensitized leukemic LGLs to Fas-mediated death. Collectively, these results show a role for sphingolipid-mediated signaling as a mechanism for long-term survival of CTLs. Therapeutic targeting of this pathway, such as use of FTY720, may have efficacy in LGL leukemia.     

FTY720 suppresses humoral immunity by inhibiting germinal center reaction

FTY720 is a novel immunosuppressant that is highly effective in inhibiting rejection of allografts and autoimmunity in various animal models. It has been shown that the sphingosine 1 phosphate (S1P) receptors are the direct molecular targets of FTY720. However, the mechanisms responsible for inhibiting specific immune responses by FTY720 are not well resolved. In particular, there is no available information on whether or how this compound affects humoral immunity. We have investigated the effect of FTY720 treatment on B-cell response during the immune response to a well-defined T-dependent antigen. Our data demonstrated that germinal center reaction was significantly reduced in peripheral lymphoid tissues of mice treated with FTY720. In addition, FTY720 treatment inhibited the production of high-affinity, class-switched antibodies, but not the production of low-affinity, immunoglobulin M (IgM) antibody. Consistently, FTY720 did not have a significant effect on antibody response to a T-independent antigen. Our results may have important implications in application of FTY720 in immune regulation.     

FTY720 (fingolimod) efficacy in an animal model of multiple sclerosis requires astrocyte sphingosine 1-phosphate receptor 1 (S1P1) modulation

Sphingosine 1-phosphate (S1P), a lysophospholipid, has gained relevance to multiple sclerosis through the discovery of FTY720 (fingolimod), recently approved as an oral treatment for relapsing forms of multiple sclerosis. Its mechanism of action is thought to be immunological through an active phosphorylated metabolite, FTY720-P, that resembles S1P and alters lymphocyte trafficking through receptor subtype S1P(1). However, previously reported expression and in vitro studies of S1P receptors suggested that direct CNS effects of FTY720 might theoretically occur through receptor modulation on neurons and glia. To identify CNS cells functionally contributing to FTY720 activity, genetic approaches were combined with cellular and molecular analyses. These studies relied on the functional assessment, based on clinical score, of conditional null mouse mutants lacking S1P(1) in CNS cell lineages and challenged by experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis. All conditional null mutants displayed WT lymphocyte trafficking that responded normally to FTY720. In marked contrast, EAE was attenuated and FTY720 efficacy was lost in CNS mutants lacking S1P(1) on GFAP-expressing astrocytes but not on neurons. In situ hybridization studies confirmed that astrocyte loss of S1P(1) was the key alteration in functionally affected mutants. Reductions in EAE clinical scores were paralleled by reductions in demyelination, axonal loss, and astrogliosis. Receptor rescue and pharmacological experiments supported the loss of S1P(1) on astrocytes through functional antagonism by FTY720-P as a primary FTY720 mechanism. These data identify nonimmunological CNS mechanisms of FTY720 efficacy and implicate S1P signaling pathways within the CNS as targets for multiple sclerosis therapies.     

重组腺病毒程序性死亡配体-1治疗BXSB狼疮鼠的实验研究

目的 观察增强程序性死亡-1(PD-1)信号通路的功能对BXSB狼疮鼠病变的影响.方法 重组腺病毒程序性死亡配体-1(AdPD-L1,2.2×109 PFU/ml×0.5ml,一次量)尾静脉注射8周龄雄性BXSB狼疮鼠.结果 AdPD-L1经尾静脉注入BXSB狼疮鼠体内后,PD-L1可有效转染80%近端肾小管上皮细胞;12周时AdPD-L1组狼疮鼠的蛋白尿发生率为20%,对照组为100%,AdPD-L1组IgG型dsD-NA抗体光密度值为0.38,肾小球IgG荧光表达强度1.16.均较对照组明显下降(P<0.01),肾组织病理损伤也减轻.结论 增强PD-1信号通路的功能可减轻BXSB狼疮鼠的病变.     

Immunomodulator FTY720 Induces eNOS-dependent arterial vasodilatation via the lysophospholipid receptor S1P3

The novel immunomodulator FTY720 is effective in experimental models of transplantation and autoimmunity, and is currently undergoing Phase III clinical trials for prevention of kidney graft rejection. FTY720 is a structural analogue of sphingosine-1-phosphate (S1P) and activates several of the S1P receptors. We show that FTY720 induces endothelium-dependent arterial vasodilation in phenylephrine precontracted mouse aortae. Vasodilation did not occur in thoracic aortic rings from eNOS-deficient mice, implicating and effect dependent of activation of the eNOS/NO pathway. Accordingly, FTY720 induced NO release, Akt-dependent eNOS phosphorylation and activation in human endothelial cells. For biological efficacy, FTY720 required endogenous phosphorylation, since addition of the sphingosine kinase antagonist N',N-dimethylsphingosine (DMS) prevented activation of eNOS in vitro and inhibited vasodilation in isolated arteries. The endothelial phosphorylation of FTY720 was extremely rapid with almost complete conversion after 10 minutes as determined by mass spectrometry. Finally, we identified the lysophospholipid receptor S1P3 as the S1P receptor responsible for arterial vasodilation by FTY720, as the effect was completely abolished in arteries from S1P3-deficient mice. In summary, we have identified FTY720 as the first immunomodulator for prevention of organ graft rejection in clinical development that, in addition, positively affects the endothelium by stimulating NO production, and thus potentially displaying beneficial effects on transplant survival beyond classical T cell immunosuppression.     

Therapeutic approach for type 1 diabetes mellitus using the novel immunomodulator FTY720 (fingolimod) in combination with once‐daily injection of insulin glargine in non‐obese diabetic mice

Aims/Introduction: The therapeutic effectiveness against type 1 diabetes mellitus (DM) of the novel immunomodulator FTY720 (fingolimod), alone and in combination with insulin glargine, was examined in the non‐obese diabetic (NOD) mouse model. Materials and Methods: Female NOD mice that had developed DM spontaneously were divided into four groups: (i) an FTY720 (0.1 mg/kg, p.o., twice weekly)‐treated group; (ii) an insulin glargine (1.0 IU, s.c., once daily)‐treated group; (iii) a combination FTY720 + insulin glargine (0.1–1.0 IU, s.c., once daily)‐treated group; and (iv) a placebo (vehicle)‐treated group. Treatment was initiated at the time of onset of DM and continued for 70 days or until death. The therapeutic efficacy of FTY720, insulin glargine and FTY720 + insulin glargine was evaluated by measuring the ratio of insulin‐positive β‐cells/total islet area, the extent of islet inflammation (insulitis score), blood glucose levels, and serum C‐peptide levels. Results: Therapeutic administration of FTY720 to NOD mice with hyperglycemia (i.e. overt DM) significantly prolonged survival (P < 0.05 vs placebo). In the placebo group, all mice died within 63 days on the onset of DM; in contrast, 45% of FTY720‐treated mice survived during the observation period (up to 70 days after the onset of DM). Therapeutic administration of FTY720 in combination with insulin glargine to NOD mice with hyperglycemia further improved survival (P < 0.05) compared with either FTY720 or insulin glargine alone (i.e. 85% of FTY720 + insulin glargine‐treated mice survived to the end of the observation period). The efficacy of FTY720 in combination with insulin glargine was confirmed by histochemical, immunohistochemical and endocrinologic observations. Conclusions: Combination therapy with FTY720 plus insulin glargine is a promising candidate for the treatment of DM and may allow for a reduction in the frequency of insulin self‐injections. (J Diabetes Invest, doi: 10.1111/j.2040‐1124.2011.00160.x, 2011)