CN114470213A - Nampt抑制剂在制备预防、缓解和/或治疗组织缺血再灌注损伤中的药物中的应用 - Google Patents
Nampt抑制剂在制备预防、缓解和/或治疗组织缺血再灌注损伤中的药物中的应用 Download PDFInfo
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Abstract
本发明公开了NAMPT抑制剂和NAMPT基因在制备预防、缓解和/或治疗组织缺血再灌注损伤中的药物中的应用。本发明通过实验证实调控NAMPT活性能够切实影响巨噬细胞极化,NAMPT为影响巨噬细胞极化的关键基因,经动物实验验证调控NAMPT活性能够减轻小鼠肝缺血再灌注损伤;为开发新的预防、缓解和/或治疗组织缺血再灌注损伤的药物提供新的思路。
Description
技术领域
本发明涉及生物医药技术领域,具体涉及NAMPT抑制剂在制备预防、缓解和/或治疗组织缺血再灌注损伤中的药物中的应用。
背景技术
缺血再灌注损伤(ischemia-reperfusion injury,IRI)是肝移植术过程中伴随的病理生理过程,是影响肝脏功能,导致肝脏功能障碍甚至肝脏功能衰竭的关键因素。
线粒体作为能量代谢的核心场所在巨噬细胞的免疫代谢中发挥重要作用,其代谢途径决定了巨噬细胞极化后的状态。能否通过调控线粒体功能使巨噬细胞向抑制炎症方向极化具有潜在研究价值。
沉默信息调节转录因子3(Silencing information regulating transcriptionfactor 3,SIRT3)是主要定位于哺乳动物细胞线粒体基质,作为烟酰胺腺嘌呤二核苷酸(nicotinamide adenine dinucleotide,NAD)依赖的去乙酰化酶,它主要通过修饰线粒体内约70%的蛋白来调节其功能。其能激活肝脏激酶B1(liver kinase B1,LKB1)使其处于磷酸化状态,后者进而磷酸化激活腺苷酸活化蛋白激酶(AMP-activated protein kinase,AMPK),活化的AMPK又通过磷酸化活化环磷腺苷效应元件结合蛋白(cAMP-responseelement binding protein,CREB),最终上调了过氧化物酶体增殖物激活受体γ共激活因子-1α(peroxisome proliferators activated receptor γcoactivator-1α,PGC-1α)表达;同时PGC-1α又可介导***相关受体α(estrogen-related receptor,ERRα)与SIRT3启动子区域的序列基元(sequence motif)结合进一步促进SIRT3基因表达来形成正反馈弧。
烟酰胺磷酸核糖转移酶(nicotinamide phosphoribosyltransferase,NAMPT)作为补救合成途径(salvage pathway)的限速酶,是决定哺乳动物细胞内NAD水平的关键分子。NAMPT的表达丰度与细胞内NAD含量及SIRT3活性成正相关。
目前,已有大量的研究深入阐明IRI的发生机制,认为由肝脏巨噬细胞介导的过度炎症反应是IRI过程中的重要因素。肝脏中的巨噬细胞可依据功能不同分为M1和M2型巨噬细胞,可分别通过分泌促炎、抗炎因子双向调控无菌性炎症,在触发、维持及改善肝脏IRI中起关键作用。免疫代谢(Immunometabolism)主要研究免疫与代谢在生理及疾病中的相互影响。体内复杂的细胞微环境对免疫细胞的代谢反应会产生重要影响从而决定免疫细胞的命运。
发明内容
本发明的发明人在研究过程中发现NAMPT在缺血再灌注损伤中高表达,认为NAMPT可能是潜在的能够调控缺血再灌注损伤基因,发明人经过深入研究证实调控NAMPT活性能够减轻小鼠肝缺血再灌注损伤。基于此,本发明保护如下技术方案:
NAMPT抑制剂在制备预防、缓解和/或治疗组织缺血再灌注损伤中的药物中的应用。
在上述技术方案中,所述组织为肝。
在上述技术方案中,所述NAMPT抑制剂为FK866。
在上述技术方案中,NAMPT抑制剂通过调控巨噬细胞极化达到预防、缓解和/或治疗炎症的作用。
在上述技术方案中,NAMPT抑制剂通过PARP1通路调控巨噬细胞极化。
本发明还保护:
NAMPT基因在在制备预防、缓解和/或治疗组织缺血再灌注损伤中的药物中的应用,其特征在于:以NAMPT作为靶基因以设计并制备预防、缓解和/或治疗组织缺血再灌注损伤的药物或生物学制剂。
本发明在研究中,发现:
调控NAMPT影响巨噬细胞极化的WB(图4A)与PCR(图4B)实验结果显示:NAMPT的激动剂P7C3与抑制剂FK866能够分别使巨噬细胞M1极化指标TNF-α,IL-1β,iNOS升高和降低,表明调控NAMPT活性能够切实影响巨噬细胞极化。
在NAMPT抑制剂FK866处理中,添加NAMPT直接产物NMN作为回溯实验,结果显示(图5)NMN能够使降低的M1极化指标回升,表明NAMPT确实为影响巨噬细胞极化的关键基因。
对NAMPT的13个影响巨噬细胞极化的直接下游进行PCR检测,有且仅有PARP1表达具有差异(图6),表明PARP1可能为NAMPT调控巨噬细胞极化的关键通路。
抑制PARP1活性也会减轻巨噬细胞M1型极化程度,且在添加PARP1抑制剂时,NAMPT回溯实验无变化(图7),表明PARP1为NAMPT调控巨噬细胞极化的关键通路。
动物实验中假手术组、模型对照组、用药组的HE染色观察肝损伤情况的实验结果显示,NAMPT抑制剂FK866用药组的HE染色肝细胞胞质空泡化少于模型对照组,表明调控NAMPT活性能够减轻小鼠肝缺血再灌注损伤。
本发明的有益效果是:通过实验证实调控NAMPT活性能够切实影响巨噬细胞极化,NAMPT为影响巨噬细胞极化的关键基因,经动物实验验证调控NAMPT活性能够减轻小鼠肝缺血再灌注损伤;为开发新的预防、缓解和/或治疗组织缺血再灌注损伤的药物提供新的思路。
附图说明
图1是从GEO基因集中筛选肝移植再灌注损伤关键基因结果。
图2是在小鼠干缺血再灌注模型中加入NAMPT抑制剂的实验结果。
图3是探究巨噬细胞极化条件实验结果。
图4是用NAMPT激动剂与抑制剂调控NAMPT活性探究影响巨噬细胞极化的情况。
图5是NAMPT直接产物NMN的回溯实验以验证NAMPT对巨噬细胞极化的影响。
图6是采用PCR实验探索NAMPT影响巨噬细胞极化的下游通路、检测NAMPT直接下游基因的变化情况的实验结果。
图7是采用PARP1抑制剂AG14361验证PARP1是否能够调节巨噬细胞极化的实验结果。
具体实施方式
下面结合实施例对本发明作进一步说明,但并不因此而限制本发明。
下述实施例中的实验方法,如无特别说明,均为常规方法;所用生物化学试剂如无特殊说明,均为本领域常规试剂,均可商购获得。
主要试剂来源:
提取RNA:奕杉生物公司的RNA提取试剂盒。
PCR逆转录:MCE RT Master Mix for qPCR(gDNA digester plus)逆转录试剂盒。
q-PCR:MCE SYBR Green qPCR Master Mix(No ROX)。
ELISA:九邦生物公司的小鼠肿瘤坏死因子α(TNF-α)ELISA检测试剂盒。
NAMPT抑制剂FK866、NAMPT激动剂P7C3、NAMPT直接产物NMN、PARP1抑制剂AG14361:均为美国selleck公司产品。
脂多糖(LPS)购自美国Sigma公司,阿佛丁购自南京爱贝生物科技有限公司。
TNF-α、iNOS、NAMPT抗体均万英国abcam公司产品。
actin、tubulin抗体均购自武汉三鹰生物技术有限公司。
实施例中RT q-PCR中用到的PCR引物序列如表1中所示:
表1引物序列
引物名称 | SEQ ID NO | 引物序列(5'-3') |
Arg-1 F | SEQ ID NO.1 | CATATCTGCCAAAGACATCGTG |
Arg-1 R | SEQ ID NO.2 | GACATCAAAGCTCAGGTGAATC |
Tnf-α F | SEQ ID NO.3 | ATGTCTCAGCCTCTTCTCATTC |
Tnf-α R | SEQ ID NO.4 | GCTTGTCACTCGAATTTTGAGA |
Il-1β F | SEQ ID NO.5 | TCGCAGCAGCACATCAACAAGAG |
Il-1β R | SEQ ID NO.6 | AGGTCCACGGGAAAGACACAGG |
Actin F | SEQ ID NO.7 | CTACCTCATGAAGATCCTGACC |
Actin R | SEQ ID NO.8 | CACAGCTTCTCTTTGATGTCAC |
CD38 F | SEQ ID NO.9 | GCCATTTTACAAAAACAGCACC |
CD38 R | SEQ ID NO.10 | GCACAATCATCTTCAGCTCATT |
PARP1 F | SEQ ID NO.11 | CAAGAAATGCAGCGAGAGTATT |
PARP1 R | SEQ ID NO.12 | CAGAAGCAGGAGAAGTGGTAC |
PARP2 F | SEQ ID NO.13 | TGACACAGGAATTCTCAATCCA |
PARP2 R | SEQ ID NO.14 | TTAGAAGGTATCGCATACGGAC |
PARP3 F | SEQ ID NO.15 | ACCAGACCAACATCGGCAACAAC |
PARP3 R | SEQ ID NO.16 | TCCAGCAGAAGAAGCGACTACCC |
PARP4 F | SEQ ID NO.17 | CCAAGTTGCCACTCTGTCTCTGTC |
PARP4 R | SEQ ID NO.18 | AGGAATGAAGCCGTAGACCAGGAG |
PARP9 F | SEQ ID NO.19 | CTCCATTTCAGAGACTGTCAGT |
PARP9 R | SEQ ID NO.20 | CGTCACACAGCATACATTCAAT |
PARP10 F | SEQ ID NO.21 | AGGTGCTGACGGGTGACTATGG |
PARP10 R | SEQ ID NO.22 | ACAAAGATTCTGGGCTGCTGAAGG |
PARP14 F | SEQ ID NO.23 | GCTATGCTGGGAAGAACGCTACTG |
PARP14 R | SEQ ID NO.24 | ATTGGTGTCTGGTCTGGAATATGTGTC |
Sirt1 F | SEQ ID NO.25 | CGCTGTGGCAGATTGTTATTAA |
Sirt1 R | SEQ ID NO.26 | TTGATCTGAAGTCAGGAATCCC |
Sirt2 F | SEQ ID NO.27 | CAGCTACTTCAAGAAACATCCG |
Sirt2 R | SEQ ID NO.28 | TATTCTTTTCTGCAGGAGGTGT |
Sirt3 F | SEQ ID NO.29 | TCTATACACAGAACATCGACGG |
Sirt3 R | SEQ ID NO.30 | GCATGTAGCTGTTACAAAGGTC |
Sirt6 F | SEQ ID NO.31 | CCCAAGTGTAAGACGCAGTA |
Sirt6 R | SEQ ID NO.32 | GTCCAGAATGGTGTCTCTCAG |
Sirt7 F | SEQ ID NO.33 | CAACAGTGGGTGAGACAGAAGAGG |
Sirt7 R | SEQ ID NO.34 | TCTTAATCTGGGCTTTGGCTGGAAG |
实施例1、寻找影响肝移植再灌注损伤关键基因
从GEO基因集中筛选肝移植缺血再灌注基因集,最终得到GSE12720、GSE14951、GSE151648、GSE8748四个基因集。运用R软件“limma”包分别进行差异分析,并取交集,得到81个共同的差异基因。在MSigDB数据库中寻找免疫代谢相关基因,并与差异基因取交集,最终获得12个与免疫代谢相关的差异基因,分别是CDKN1A、EGR2、G0S2、GADD45B、HSPH1、IL1B、KLF6、NAMPT、PTGS2、SERPINE1、SLC2A3、SOCS3。运用Cytoscape软件的cytoHubba插件进行关键基因筛选,在12个基因中,NAMPT得分最高。运用R软件对NAMPT在四个基因集中的差异表达进行绘图。在四个基因集中验证,NAMPT均在缺血再灌注损伤中高表达。本发明涉及的NAMPT(人)在NCBI数据库中的编号:基因ID 10135,核苷酸序列编号NM_005746.3,蛋白序列NP_005737.1。NAMPT(鼠)在NCBI数据库中的编号:基因ID 59027,核苷酸序列编号NM_021524.2,蛋白序列NP_067499.2。
如图1所示,NAMPT在肝缺血再灌注损伤基因集中具有差异表达,且在缺血再灌注损伤中表达上调。结果显示:NAMPT是潜在的能够调控缺血再灌注损伤基因。
实施例2、动物实验
1、实验动物:C57BL/6小鼠由重庆医科大学动物中心提供。选取8-10周龄,雄性,体重25~30g,SPF级。
2、各组处理:
先将动物称重,编号,选择健康的小鼠9只,按照随机数字表法分为三组,每组三只,包括假手术组,模型对照组以及用药组。
假手术组:在室温条件下,用1.25%阿佛丁以0.2ml/10g剂量麻醉小鼠后,将小鼠固定。脱毛膏涂于小鼠胸前区,脱毛后75%酒精消毒胸前区备皮。用手术刀在小鼠剑突下划开一道约2cm口子破皮,再用手术剪剪开腹膜,用手在小鼠腹部左右两侧用力挤压,挤出肝脏,清晰暴露门静脉及其分支后,随即逐层缝合。局部消毒后,核实标记置于相应鼠笼。
模型对照组:麻醉、消毒方法同假手术组。处理同假手术组至清晰暴露门静脉及其分支。此时使用显微静脉血管夹夹闭小鼠门静脉右支,将其置于37℃温床缺血1.5h,随即逐层缝合。待其苏醒后,局部消毒,核实标记,置于相应鼠笼。
用药组:具体操作处理同模型对照组,并于术前24h、12h、0.5h腹腔注射FK866(10mg/kg)。
3、实验过程和结果:
小鼠缝合6h后,麻醉小鼠,浓度方法同前。在麻醉状态下用颈椎脱臼法处死各组小鼠,然后用手术剪剪开缝合线,并扩大切口,游离肝脏,取出后予以4%多聚甲醛固定。多聚甲醛固定后组织在经脱水、包埋、切片、制片后,予以HE染色观察肝损伤情况。
HE染色结果如图2所示,图2A、2B、2C分别是假手术组、模型对照组、用药组的结果,从图中看出,用药组的HE染色肝细胞胞质空泡化少于模型对照组。结论:调控NAMPT活性能够减轻小鼠肝缺血再灌注损伤。
实施例3、细胞实验
1.实验材料和方法
1.1骨髓来源巨噬细胞(BMDM)提取与培养:
动物准备:C57BL/6小鼠由重庆医科大学动物中心提供。选取8-10周龄,雄性,体重25~30g,SPF级。
提取细胞:用阿佛丁麻醉小鼠后(剂量方法同前),以颈椎脱臼法处死小鼠,并用75%酒精浸泡消毒。用手术剪在小鼠下腹部横向切开,用手扩大切口,并且使小鼠腿部皮肤完全剥离。
取胫骨:反向弯折小鼠膝关节,再将胫骨从肌肉中顶出,用手术剪将胫骨和脚钝性分离,得到较为干净的胫骨。
取股骨:用手大致钝性剥离小鼠股骨与肌肉,再用手术剪离断股骨与盆腔相连的肌肉,并将股骨与髋骨钝性分离,得到较为干净的股骨。
取下的骨头均浸泡于4℃无菌PBS缓冲液中暂时保存。
细胞培养:
超净台紫外消毒20min后,在超净台内,将取下的股骨与胫骨放入含无菌PBS缓冲液的10cm直径培养皿中。用手术剪去除胫骨与股骨两端,以5ml注射器吸取无菌PBS缓冲液冲洗骨髓腔,直至小鼠骨头变为半透明。
用100um细胞筛过滤冲出的液体,并收集至离心管中。
经1000rpm/10min离心后,去上清,再用完全培养基(DMEM高糖培养基,含10%FBS,1%青霉素-链霉素,M-CSF(巨噬细胞集落刺激因子,50ng/ml))重悬细胞,接种于六孔板中(每孔细胞量1.5*106)。于37℃,5%二氧化碳培养箱中静置培养。
骨髓来源巨噬细胞诱导需七天时间,第三天与第七天需要换液,重新添加完全培养基。1.2试剂应用方法:
LPS诱导M1型巨噬细胞极化,在BMDM(骨髓来源的巨噬细胞)诱导完成后,加入LPS(脂多糖,200ng/ml)处理24h。
FK866为NAMPT抑制剂,在加入LPS前0.5h添加FK866(100nM)处理24h。
P7C3为NAMPT激动剂:在加入LPS前2h添加P7C3(10uM)处理24h。
NMN为NAMPT直接产物:在加入LPS前0.5h添加NMN(500uM)处理24h。
AG14361为PARP1抑制剂:在加入LPS前1h添加AG14361(400nM)处理24h。
1.3实验技术路线:
提取原代小鼠骨髓来源巨噬细胞,以1.5*106种于六孔细胞培养板,经M-CSF诱导后加药处理,并用LPS刺激诱导M1型极化,24h后收取蛋白进行Western blot实验,提取RNA进行逆转录PCR实验,收集上清液进行ELISA实验。其中,
1、western blotting检测巨噬细胞极化相关标志物TNF-α、iNOS、CD163、Arg-1蛋白表达,以及通路中关键蛋白PGC-1α、PPAR-γ表达。
2、PT-PCR检测极化相巨噬细胞极化相关标志物TNF-α、iNOS、CD163、Arg-1的mRNA表达,以及通路中关键分子PGC-1α、PPAR-γ的mRNA表达。
3、流式细胞仪结合FITC-F4/80,PE-CD206和APC-CD11b检测M1/M2型巨噬细胞的比例。
4、应用ELISA检测上清液中巨噬细胞分泌的细胞因子。
5、线粒体功能检测:检测细胞氧耗率(OCR)、质子生成率(PPR)以及细胞外酸化率(ECAR)反应氧化磷酸化及糖酵解水平,测量NAD、ATP含量。
具体实验方法如下。
1.3.1 Western blot实验
蛋白提取:每孔BMDM中加入碧云天RIPA裂解液(强),冰上裂解15min,移入离心管内,经4℃离心机12000rpm/15min离心后,提取上清液,运用碧云天BCA Protein Assay Kit测定蛋白的浓度,再加入碧云天SDS-PAGE蛋白上样缓冲液混匀,沸水浴10min。
上样与电泳:配置好电泳凝胶,并在电泳槽内加入电泳液。将蛋白样品按计算出的浓度加入SDS-PAGE胶加样孔内,点样完成后开始电泳。
转膜:配置电转液,4℃预冷。PVDF膜浸泡甲醇10s活化。凝胶放于负极夹板上,PVDF膜放于凝胶上,夹上夹板。组装好电转槽开始电转。
封闭:将PVDF膜浸泡于碧云天QuickBlockTM封闭液15min。
一抗孵育:将PVDF膜放入含一抗iNOS(1:1000)、TNF-α(1;1000)、NAMPT(1:1000)、actin(1:2000)、tubulin(1:2000)的5ml离心管中,4℃摇床过夜。
二抗孵育:将PVDF膜用TBST洗涤3次,每次10min,再放入对应的二抗(1:8000)中,常温孵育1h。
蛋白检测:将PVDF膜用TBST洗涤3次,每次10min。利用Bio-Rad Chemi Doc XRS+凝胶成像***检测目的条带。采用Image Lab软件分析条带灰度值,用目的蛋白与内参蛋白比值来衡量蛋白的相对表达量。
1.3.2 PCR逆转录:
提取RNA:使用奕杉生物公司的RNA提取试剂盒。
吸干培养基,用适量PBS清洗一次。加入500ul的lysis buffer,用力吹吸10次,转移至EP管中,vortex10s充***解细胞。向裂解的细胞中加入等体积无水乙醇重复分混匀,再将液体移入离心柱。4000g/1min离心后,向离心柱中加入500ul wash buffer,再进行12000g/1min离心。将柱子放进无酶的1.5ml EP管上,开盖晾干两分钟。在RNA柱的膜中心加入20ul elution buffer,室温静置2min,再进行12000g/1min离心。使用Naro Photometer核酸浓度测定仪测定RNA浓度。
逆转录:使用MCE RT Master Mix for qPCR(gDNAdigesterplus)逆转录试剂盒。整个操作过程在冰上进行。在无酶EP管中加入5x gDNA digester Buffer 2ul、gDNAdigester 1ul、mRNA 1ug,用DEPC水将总体积补到10ul,42℃孵育2min。在上一步反应液中加入2×Super RT Mix 2ul,置于BIO RAD T100 Thermal Cycler中进行反应。具体程序为:25℃5min、42℃45min、85℃2min。
qPCR:使用MCE SYBR Green qPCR Master Mix(No ROX)。配置反应体系在冰上进行,依次加入SYBR Green Master Mix(2×)(No ROX)10ul、前引物0.4ul、后引物0.4ul、逆转录产物2ul、DEPC水7.2ul。
将反应体系放入BIO RAD CFX Connect Real-Time System中进行反应。具体程序为:95℃预热5min,再以95℃10s、60℃30s循环40次,最后用仪器默认程序测定溶解曲线。
1.3.3 ELISA
使用九邦生物公司的小鼠肿瘤坏死因子α(TNF-α)ELISA检测试剂盒。
上清液3000rpm/10min离心,去除颗粒和聚合物。先将试剂盒在室温平衡1h。待测样品每孔加入10ul,重复三次,再加入样本稀释液40ul,随后加入辣根过氧化物酶(HRP)检测抗体100ul,用封板膜封住反应孔,37℃恒温箱反应1h。弃去液体,吸水纸上拍干。每孔加入350ul洗涤液,静置1min,甩去洗涤液,吸水纸上拍干,重复五次。每孔加入底物A、B各50ul,37℃恒温箱孵育15min。再加入终止液50ul,于Thermo VARIOSKAN LUX酶标仪450nm处检测各孔OD值。
2实验结果和分析
图3显示了探究巨噬细胞极化条件。结果显示:诱导巨噬细胞M1型极化,LPS诱导最适浓度为200ng/ml,在ELISA(图3A)与PCR(图3B)中验证效果均显著。LPS能够使巨噬细胞M1型极化指标TNF-α,IL-1β升高,且NAMPT在M1型极化程度升高时也会升高,表明LPS能够成功诱导巨噬细胞M1型极化,且诱导NAMPT含量升高。
图4为调控NAMPT影响巨噬细胞极化的WB(图4A)与PCR(图4B)实验。结果显示:NAMPT的激动剂P7C3与抑制剂FK866能够分别使巨噬细胞M1极化指标TNF-α,IL-1β,iNOS升高和降低,表明调控NAMPT活性能够切实影响巨噬细胞极化。
图5是验证NAMPT对巨噬细胞极化的影响。结果显示:在NAMPT抑制剂FK866处理中,添加NAMPT直接产物NMN作为回溯实验,NMN能够使降低的M1极化指标回升,表明NAMPT确实为影响巨噬细胞极化的关键基因。
图6是探索NAMPT影响巨噬细胞极化的下游通路实验。对NAMPT的13个影响巨噬细胞极化的直接下游进行PCR检测,有且仅有PARP1表达具有差异,表明PARP1可能为NAMPT调控巨噬细胞极化的关键通路。
图7是验证PARP1是否能够调节巨噬细胞极化。实验结果显示:抑制PARP1活性也会减轻巨噬细胞M1型极化程度,且在添加PARP1抑制剂时,NAMPT回溯实验无变化,表明PARP1为NAMPT调控巨噬细胞极化的关键通路。
序列表
<110> 重庆医科大学
<120> NAMPT抑制剂在制备预防、缓解和/或治疗组织缺血再灌注损伤中的药物中的应用
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Claims (6)
1.NAMPT抑制剂在制备预防、缓解和/或治疗组织缺血再灌注损伤中的药物中的应用。
2.如权利要求1所述的应用,其特征在于:所述组织为肝。
3.如权利要求1所述的应用,其特征在于:所述NAMPT抑制剂为FK866。
4.如权利要求1所述的应用,其特征在于:NAMPT抑制剂通过调控巨噬细胞极化达到预防、缓解和/或治疗炎症的作用。
5.如权利要求4所述的应用,其特征在于:NAMPT抑制剂通过PARP1通路调控巨噬细胞极化。
6.NAMPT基因在在制备预防、缓解和/或治疗组织缺血再灌注损伤中的药物中的应用,其特征在于:以NAMPT作为靶基因以设计并制备预防、缓解和/或治疗组织缺血再灌注损伤的药物或生物学制剂。
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CA2401567A1 (en) * | 2002-08-05 | 2004-02-05 | University Health Network | Methods of preventing the inhibition of apoptosis and treating inflammatory conditions |
WO2009109610A1 (en) * | 2008-03-05 | 2009-09-11 | Topotarget Switzerland S.A. | Use of nad information inhibitors for the treatment of ischemia-reperfusion injury |
US20110020364A1 (en) * | 2009-07-23 | 2011-01-27 | Garcia Joe G N | Methods and compositions involving pbef inhibitors for lung inflammation conditions and diseases |
CN110996967A (zh) * | 2017-08-10 | 2020-04-10 | 华盛顿大学 | 使用烟酰胺单核苷酸进行治疗的组合物和方法 |
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CA2401567A1 (en) * | 2002-08-05 | 2004-02-05 | University Health Network | Methods of preventing the inhibition of apoptosis and treating inflammatory conditions |
WO2009109610A1 (en) * | 2008-03-05 | 2009-09-11 | Topotarget Switzerland S.A. | Use of nad information inhibitors for the treatment of ischemia-reperfusion injury |
US20110020364A1 (en) * | 2009-07-23 | 2011-01-27 | Garcia Joe G N | Methods and compositions involving pbef inhibitors for lung inflammation conditions and diseases |
CN110996967A (zh) * | 2017-08-10 | 2020-04-10 | 华盛顿大学 | 使用烟酰胺单核苷酸进行治疗的组合物和方法 |
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