TXNIP基因与肿瘤发生及转移

硫氧还蛋白结合蛋白（thioredoxin interacting protein, TXNIP）在细胞增殖、凋亡、分化的过程以及肿瘤、应激性疾病的发生中具有重要功能. 作为一个氧还反应的调节子,TXNIP能与硫氧还蛋白(thioredoxin, Trx)相结合,下调Trx的表达,而Trx则在DNA的损伤及细胞凋亡机制中有着关键的作用. 本文阐述了TXNIP基因的特征及其蛋白的生物学功能, 并简要总结TXNIP在人类肿瘤中低表达的研究成果. TXNIP基因是一个新的抑癌基因,它在人类乳腺癌、肝癌、肺癌等癌组织细胞中均表达下降, 并且与肿瘤的转移相关. TXNIP的缺失可以促使肿瘤细胞的增殖和抑制细胞凋亡的进程. 而在抗肿瘤机制中, TXNIP可通过参与细胞周期阻滞、低氧调节、影响NK细胞(natural killer cell, NK cell)发育等过程介导肿瘤的发生发展.     

使用腺病毒过表达TXNIP对H9C2心肌细胞损伤和凋亡的影响北大核心CSCD

本文旨在使用病毒转染技术,观察正常培养条件下的心肌细胞单纯过表达硫氧还蛋白相互作用蛋白(thioredoxin inter-acting protein,TXNIP)是否可以引起细胞损伤和凋亡的发生,并分析其作用途径。对数生长期的H9C2心肌细胞分为三组:正常培养组、空病毒(Ad-eGFP)组、TXNIP过表达(Ad-TXNIP-eGFP)组,使用正常糖脂浓度(5mmo1/L葡萄糖)的DMEM培养基,均于转染72h后收集细胞和培养基进行指标测定。结果显示,细胞转染成功,72h转染效率达到高峰。与Ad-eGFP组相比,Ad-TXNIP-eGFP转染组TXNIPmRNA(P<0.05)和蛋白表达(P<0.01)均明显升高;心肌caspase-3(P<0.05)和LDH活性明显升高(P<0.01);使用流式细胞仪测得细胞凋亡明显增加(P<0.01)。Trx活性、与Trx相关的自由基损伤以及介导氧化应激损伤凋亡途径的p38激酶的活性检测显示,与Ad-eGFP组相比,TXNIP过表达组的Trx活性明显降低(P<0.01),反映膜脂质过氧化损伤的指标丙二醛(malondial dehyde,MDA)、3-硝基酪氨酸明显升高(P<0.01),p38激酶活性明显升高(P<0.01)。这些结果提示,通过腺病毒转染单纯过表达TXNIP可以引起正常糖脂浓度培养条件下的心肌细胞发生损伤和凋亡,其具体机制与抑制Trx活性、增加自由基损伤和p38激酶介导的凋亡有关。     

Trx/TXNIP在脑卒中的研究进展

脑卒中是导致中老年人群死亡最主要原因之一,其具有较高的致死率和致残率,且每年的发病率呈逐年上升的趋势,严重危害人类的生命和健康,因此寻找有效的诊断及治疗脑卒中的靶点具有重要意义。硫氧还蛋白(Trx)是细胞内主要的硫醇还原剂,通过调节细胞内氧化还原状态,参与细胞内多种信号通路转导过程,具有二硫化物还原酶活性,通过抗氧化效应,减轻脑卒中后神经元氧化应激损伤。硫氧还原蛋白相互作蛋白(TXNIP)是Trx的内源性抑制剂,通过绑定/抑制Trx的活性,破坏细胞内氧化还原平衡,促进氧化应激,而抑制或敲除TXNIP具有明显的神经保护作用。最新研究表明Trx/TXNIP可经多种途径参与脑卒中病理生理过程。本文通过分析Trx和TXNIP的研究现状,以及探讨Trx系统在中枢神经系统中的定位和Trx系统在缺血性脑卒中的研究进展,展望Trx/TXNIP参与脑卒中的病理生理过程的信号途径,拟对Trx/TXNIP在脑卒中的作用机制进行综述,为脑卒中的治疗提供新思路。     

硫氧还蛋白在凋亡途径中的作用机制

硫氧还蛋白（Trx）是体内广泛存在的氧化还原蛋白,其家族中两种重要的硫氧还蛋白：硫氧还蛋白1（thioredoxin1,Trx1）和硫氧还蛋白2（thioredoxin2,Trx2）都含有保守的-Cys-Gly-Pro-Cys-还原序列。由于Trx具有调节细胞生长增殖和抗凋亡的作用,因此Trx在凋亡途径中的作用机制就成为了对抗肿瘤的研究热点。     

硫氧还蛋白结合蛋白-2在糖和脂代谢中的作用

硫氧还蛋白结合蛋白-2(thioredoxin binding protein-2,TBP-2)属于硫氧还蛋白结合蛋白家族成员,与还原型硫氧还蛋白结合,抑制其还原活性。在生物体内,TBP-2不但参与细胞内的氧化还原调节,还具有调节细胞生长繁殖和促进细胞凋亡的作用。TBP-2在葡萄糖和脂肪代谢中的作用被广泛研究。TBP-2高表达时,胰岛细胞凋亡,与糖尿病发生相关;而TBP-2缺失,与高脂血症发病有关。本文综述了TBP-2在糖脂代谢中的作用。     

硫氧还蛋白研究进展

硫氧还蛋白(Trx)是一类广泛存在于真核及原核生物体内的小分子多功能蛋白质。Trx具有调节细胞的生长、抑制细胞凋亡及调节基因转录等功能,并且它与硫氧还蛋白还原酶(TrxR)、烟酰腺嘌呤二核苷磷酸(NADPH)共同构成了生物体内重要的硫氧还蛋白系统,对维持体内稳定的氧化还原状态具有重要的作用。以Trx为对象,综述了其结构特点、分类分布及其生物学活性等方面的研究现状,以期为相关研究提供参考。     

转反义硫氧还蛋白基因小麦萌发种子中蛋白质的变化

硫氧还蛋白h（thioredoxin h,Trx h）是一类广泛存在于生物体内的多功能活性蛋白,分子量约为12kD,它通过还原靶蛋白中的二硫键参与酶活性调节、抗胁迫、信号传导等许多重要的生命活动。硫氧还蛋白h能促进谷物类种子萌发过程,主要表现在以下2个方面：（1）在籽粒萌发期间,硫氧还蛋白可通过还原储存蛋白的分子内二硫键使其更易于被降解;（2）硫氧还蛋白也可以直接地通过将酶还原或者间接地通过使酶抑制蛋白失活而激活酶。源于Phalaris coerulescens的trxs基因（thioredoxin s,trxs）与小麦硫氧还蛋白h基因（thioredoxin h,trx h）同属于硫氧还蛋白基因家族,它们的cDNA有94％的同源性,表达产物也有相似的生物功能。我们采用基因枪法将反义trxs基因导入小麦,获得了可稳定遗传的小麦,并检测出转基因种子中硫氧还蛋白h表达量、水溶蛋白和醇溶蛋白的还原状态以及α-淀粉酶活性均低于对照小麦;另外,通过模拟降雨抗穗发芽试验证实转基因株系具有很强的抗穗发芽能力。以转反义trxs基因抗穗发芽小麦为材料,检测反义trxs基因小麦籽粒萌发过程中蛋白质的变化,探讨转反义trxs基因小麦的抗穗发芽机理。研究表明反义trxs基因能够减缓KCl可溶性蛋白中Chloroform-methanol（CM）蛋白向代谢类蛋白的转化进程,在萌发初期降低籽粒代谢类蛋白的含量,使籽粒代谢速度下降,而CM蛋白主要包含一些分子量小于20kD的蛋白质。在籽粒成熟过程中,硫氧还蛋白能够阻止麦谷蛋白亚基形成谷蛋白聚合体的过程,在转基因小麦中麦谷蛋白更易于形成大分子量的谷蛋白大聚合体,使得转基因小麦中的谷蛋白在萌发初期更难于被水解,因此转基因小麦籽粒会因谷蛋白难于降解而萌发较慢。另外,反义trxs基因减慢了麦胚中10kD蛋白的降解过程。     

硫氧还蛋白系统在COPD抗氧化中的作用研究进展

硫氧还蛋白系统是由硫氧还蛋白(thioredoxin,Trx)、硫氧还蛋白还原酶(thioredoxin reductase,TrxR)和还原型辅酶Ⅱ(NADPH)组成的多功能小分子蛋白系统,广泛表达的硫氧还蛋白作为蛋白质二硫键的还原酶,它参与很多生理过程,并发挥重要生物学功能,包括调节机体的氧化还原反应、抑制细胞凋亡、调节转录因子DNA结合活性以及免疫应答等,其中一重要作用是参与调节细胞氧化还原状态以对抗氧化应激。因此在一些炎症性疾病如慢性阻塞性肺疾病、急性呼吸窘迫综合征、肺间质疾病、哮喘、肺结节病等的发生发展中扮演重要角色,本文对硫氧还蛋白系统在慢性阻塞性肺疾病中的抗氧化作用作一综述。     

硫氧还蛋白系统在COPD抗氧化中的作用研究进展

硫氧还蛋白系统是由硫氧还蛋白（thioredoxin,Trx）,硫氧还蛋白还原酶（thioredoxinreductase,TrxR）和还原型辅酶Ⅱ（NADPH）组成的多功能小分子蛋白系统,广泛表达的硫氧还蛋白作为蛋白质二硫键的还原酶,它参与很多生理过程,并发挥重要生物学功能,包括调节机体的氧化还原反应、抑制细胞凋亡、调节转录因子DNA结合活性以及免疫应答等,其中一重要作用是参与调节细胞氧化还原状态以对抗氧化应激。因此在一些炎症性疾病如慢性阻塞性肺疾病、急性呼吸窘迫综合征、肺间质疾病、哮喘、肺结节病等的发生发展中扮演重要角色,本文对硫氧还蛋白系统在慢性阻塞性肺疾病中的抗氧化作用作一综述。     

硫氧还蛋白抗体柱的制备

目的:探索硫氧还蛋白(Trx)抗体柱对Trx融合蛋白纯化的可行性。方法与结果:对含有Trx基因的质粒表达载体pTrxFus进行改造,在Trx读框之后加入6×His序列,并在大肠杆菌中表达C端带有6×His标签的Trx,经Ni2+柱亲和纯化后制备多克隆抗体;把经蛋白A纯化后的抗体偶联在溴化氰活化的琼脂糖凝胶上,制成Trx抗体柱;用此抗体柱纯化与Trx融合表达的豇豆胰蛋白酶抑制剂(CpTI),SDS-PAGE结果显示获得了纯度较高的Trx-CpTI。结论:用Trx抗体制成的免疫亲和层析柱可以有效纯化Trx融合蛋白。     

糖尿病诱导硫氧还蛋白相互作用蛋白(TXNIP)对小鼠胰岛β细胞衰老的影响*

目的: 本研究旨在观察糖尿病中硫氧还蛋白相互作用蛋白(TXNIP)的表达是否影响胰岛β细胞衰老。方法: 正常小鼠(db/m)、糖尿病小鼠(db/db)随机各取6只,血糖仪检测其空腹血糖值,Western blot检测胰腺组织TXNIP蛋白表达、免疫化学染色检测胰腺组织中衰老相关β-半乳糖苷酶活性,Western blot检测胰腺组织衰老相关指标p16、p21、Rb表达的变化。INS-1胰岛β细胞随机分7组(n=6),用各组慢病毒(30 μl)转染4~6 h后,嘌呤霉素(PM, 3 μg/m)筛选7 d,构建Normal组(正常组)、Scramble ShRNA组(干扰空病毒组)、TXNIP-ShRNA-1(TXNIP沉默一组)组、TXNIP-ShRNA-2(TXNIP沉默二组)组、TXNIP-ShRNA-3组(TXNIP沉默三组)、Ad-GFP组(过表达空病毒组)、Ad-TXNIP-GFP组(TXNIP过表达组)稳转INS-1胰岛β细胞株,检测其TXNIP蛋白表达、衰老相关β -半乳糖苷酶活性、衰老相关指标。结果: 与正常小鼠相比,db/db组小鼠空腹血糖显著上升(P＜0.01),胰腺组织TXNIP蛋白表达显著升高(P＜0.05),胰腺组织β -半乳糖苷酶阳性染色率增加,p16、p21、Rb蛋白表达显著升高(P＜ 0.05)。与Ad-GFP组相比,Ad-TXNIP-GFP组β -半乳糖苷酶阳性染色率增加,p16、p21、Rb蛋白表达均显著增加(P＜0.01)。与Scramble ShRNA组相比,TXNIP-ShRNA组β -半乳糖苷酶阳性染色率降低,p16、p21以及Rb蛋白表达均降低(P＜0.05)。结论: 糖尿病可通过上调TXNIP表达,诱导胰岛β细胞衰老。     

Vitamin D receptor activation in a diabetic-like environment: potential role in the activity of the endothelial pro-inflammatory and thioredoxin pathways

High blood and tissue concentrations of glucose and advanced glycation end products (AGEs) are thought to play an important role in the development of diabetic vascular complications. Thioredoxin interacting protein (TXNIP) is up-regulated in response to high levels of glucose and is an endogenous inhibitor of thioredoxin (TRX), and may play a contributory role in the occurrence of diabetic-related vascular diseases. Vitamin D inhibits endothelial proliferation and is a cardiovascular protective agent. The present study evaluated the impact of paricalcitol and calcitriol on the endothelial inflammatory and TXNIP pathways in cultured endothelial cells exposed to a diabetic-like environment. Fresh human umbilical vein cord endothelial cells (HUVEC) were treated for 24h with 200 μg/ml AGE-HSA and 250 mg/dl glucose concentrations, with paricalcitol or calcitriol. IL6, IL8, NFκB (p50/p65), receptor of AGE (RAGE), TXNIP, and TRX expressions were evaluated at the levels of mRNA, protein, and TRX activity. Calcitriol and paricalcitol significantly down-regulated the markers involved in the inflammatory responses. Only paricalcitol induced a significant decrease in TXNIP mRNA and protein expressions. Neither paricalcitol nor calcitriol affected TRX reductase activity or TRX mRNA and protein expressions. Our findings indicate that in an endothelial diabetic-like environment, paricalcitol and calcitriol significantly decreased the expression of genes involved in the inflammatory pathway. In this in vitro study, it seems that the TRX antioxidant system was not involved. The different effects found between paricalcitol and calcitriol might reflect the selectivity of vitamin D receptor (VDR) activation.     

p38 MAPK pathway is involved in high glucose-induced thioredoxin interacting protein induction in mouse mesangial cells

Excessive reactive oxygen species (ROS) play a key role in the pathogenesis of diabetic nephropathy. The thioredoxin (TRX) system, a major thiol antioxidant system, regulates the reduction of intracellular ROS. Here we show that high glucose (HG) inhibits TRX ROS-scavenging function through p38 mitogen-activated protein kinase (MAPK)-mediated induction of thioredoxin interacting protein (TXNIP) in mouse mesangial cells (MMCs). Knockdown of TXNIP in MMCs reversed HG-induced reduction of TRX activity and inhibited HG-induced activation of p38 MAPK and increased synthesis of TGF-β1 and fibronectin. These data suggest that HG-induced overexpression of TXNIP in MMCs, which may be via the p38 MAPK pathway.     

Thioredoxin interacting protein (TXNIP) induces inflammation through chromatin modification in retinal capillary endothelial cells under diabetic conditions

Chronic hyperglycemia and activation of receptor for advanced glycation end products (RAGE) are known risk factors for microvascular disease development in diabetic retinopathy. Thioredoxin‐interacting protein (TXNIP), an endogenous inhibitor of antioxidant thioredoxin (TRX), plays a causative role in diabetes and its vascular complications. Herein we investigate whether HG and RAGE induce inflammation in rat retinal endothelial cells (EC) under diabetic conditions in culture through TXNIP activation and whether epigenetic mechanisms play a role in inflammatory gene expression. We show that RAGE activation by its ligand S100B or HG treatment of retinal EC induces the expression of TXNIP and inflammatory genes such as Cox2, VEGF‐A, and ICAM1. TXNIP silencing by siRNA impedes RAGE and HG effects while stable over‐expression of a cDNA for human TXNIP in EC elevates inflammation. p38 MAPK‐NF‐κB signaling pathway and histone H3 lysine (K) nine modifications are involved in TXNIP‐induced inflammation. Chromatin immunoprecipitation (ChIP) assays reveal that TXNIP over‐expression in EC abolishes H3K9 tri‐methylation, a marker for gene inactivation, and increases H3K9 acetylation, an indicator of gene induction, at proximal Cox2 promoter bearing the NF‐κB‐binding site. These findings have important implications toward understanding the molecular mechanisms of ocular inflammation and endothelial dysfunction in diabetic retinopathy. J. Cell. Physiol. 221: 262–272, 2009. © 2009 Wiley‐Liss, Inc     

糖、脂代谢及氧化应激与糖尿病肾病的相关性

糖尿病肾病是糖尿病的主要并发症之一。近年来,糖尿病肾病的发病率呈逐渐上升趋势。研究发现,糖、脂代谢与糖尿病肾病的发生密切相关,而氧化应激也在糖、脂代谢异常中起重要作用。本文综述了有关糖、脂代谢与糖尿病肾病相关性研究,以及硫氧还蛋白及硫氧还蛋白结合蛋白2在糖尿病肾病发生中的作用。     

All‐trans retinoic acid ameliorates hepatic stellate cell activation via suppression of thioredoxin interacting protein expression

Activation of hepatic stellate cells (HSCs) is the effector factor of hepatic fibrosis and hepatocellular carcinoma (HCC) development. Accumulating evidence suggests that retinoic acids (RAs), derivatives of vitamin A, contribute to prevention of liver fibrosis and carcinogenesis, however, regulatory mechanisms of RAs still remain exclusive. To elucidate RA signaling pathway, we previously performed a genome‐wide screening of RA‐responsive genes by in silico analysis of RA‐response elements, and identified 26 RA‐responsive genes. We found that thioredoxin interacting protein (TXNIP), which inhibits antioxidant activity of thioredoxin (TRX), was downregulated by all‐trans retinoic acid (ATRA). In the present study, we demonstrate that ATRA ameliorates activation of HSCs through TXNIP suppression. HSC activation was attenuated by TXNIP downregulation, whereas potentiated by TXNIP upregulation, indicating that TXNIP plays a crucial role in activation of HSCs. Notably, we showed that TXNIP‐mediated HSC activation was suppressed by antioxidant N‐acetylcysteine. In addition, ATRA treatment or downregulation of TXNIP clearly declined oxidative stress levels in activated HSCs. These data suggest that ATRA plays a key role in inhibition of HSC activation via suppressing TXNIP expression, which reduces oxidative stress levels.     

Molecular changes in hepatic metabolism in ZDSD rats–A new polygenic rodent model of obesity,metabolic syndrome,and diabetes

In recent years, the prevalence of obesity, metabolic syndrome and type 2 diabetes is increasing dramatically. They share pathophysiological mechanisms and often lead to cardiovascular diseases. The ZDSD rat was suggested as a new animal model to study diabetes and the metabolic syndrome. In the current study, we have further characterized metabolic and hepatic gene expression changes in ZDSD rats. Immuno-histochemical staining of insulin and glucagon on pancreas sections of ZDSD and control SD rats revealed that ZDSD rats have severe damage to their islet structures as early as 15 weeks of age. Animals were followed till they were 26 weeks old, where they exhibited obesity, hypertension, hyperglycemia, dyslipidemia, insulin resistance and diabetes. We found that gene expressions involved in glucose metabolism, lipid metabolism and amino acid metabolism were changed significantly in ZDSD rats. Elevated levels of ER stress markers correlated with the dysregulation of hepatic lipid metabolism in ZDSD rats. Key proteins participating in unfolded protein response pathways were also upregulated and likely contribute to the pathogenesis of dyslipidemia and insulin resistance. Based on its intact leptin system, its insulin deficiency, as well as its timeline of disease development without diet manipulation, this insulin resistant, dyslipidemic, hypertensive, and diabetic rat represents an additional, unique polygenic animal model that could be very useful to study human diabetes.     

Thioredoxin inhibits NMDA-induced neurotoxicity in the rat retina

Thioredoxin (TRX) plays a variety of redox-related roles in organisms. To investigate its function as an endogenous redox regulator in NMDA-induced retinal neurotoxicity, we injected NMDA with TRX, mutant TRX or saline into the vitreous cavity of rat eyes. Retinal ganglion cells were rescued by TRX, compared with saline, when evaluated by retrograde labeling analysis at 7 days after NMDA injection. TRX, but not its mutant form, prevented NMDA-induced apoptosis in the retina, as measured by terminal deoxynucleotidyl transferase-mediated UTP nick-end labeling. The induction of caspase 3 and 9, but not caspase 8, by NMDA was significantly lower in TRX-treated eyes than in saline-treated eyes. NMDA-induced activation of the MAPKs, p38 kinase and c-Jun N-terminal kinase after 6 h and of the MAPK kinases (MKKs) MKK3/6 and MKK4 after 3 h was markedly suppressed in retinal ganglion cells by TRX but not by the mutant form. NMDA-induced increases in protein carbonylation, nitrosylation and lipid peroxidation were also suppressed in TRX-treated eyes. We concluded that the intravitreous injection of TRX effectively attenuated NMDA-induced retinal cell damage and that suppression of oxidative stress and inhibition of apoptotic signaling pathways were involved in this neuroprotection.