α-谷胱甘肽-S-转移酶监测肝移植术后肝脏功能的研究

目的 探讨肝移植术后监测α－谷胱甘肽－Ｓ－转移酶（α－ＧＳＴ）的价值。方法 建立大鼠原位肝移植模型，并将模型分为同种异体移植组、同基因移植组和单纯开腹３组，动态监测血清α－ＧＳＴ，谷丙转氨酶（ＡＬＴ）、谷草转氨酶（ＡＳＴ）活性变化。结果 非排斥组α－ＧＳＴ在术后２ｄ内恢复正常，ＡＬＴ、ＡＳＴ需要３ｄ后才恢复正常。排斥组α－ＧＳＴ每天平均变化率与ＡＬＴ、ＡＳＴ相比差异有显著性（Ｐ〈０．０５），自术后     

奥替普拉在阻断细胞恶性转化时对某些基因表达的影响

奥替普拉（ｏｌｔｉｐｒａｚ）来源于十字花科（Ｃｒｕｃｉｆｅｒａｓｅ）蔬菜，全称５（２吡嗪基）４甲１，２２羟基３硫酮，文献报道它为一种防癌抑癌剂［１］。我们用致癌素香烟凝聚物（ｃｉｇａｒｅｔｅｓｍｏｋｉｎｇｃｏｎｄｅｎｓａｔｅＣＳＣ）诱发...     

细胞凋亡过程中核糖体蛋白质S3a水平变化的研究（三）：——检测细胞凋亡过程中RPS3a水平的变化

目的：检测细胞凋亡过程中RPS3a水平的变化，探讨RPS3a在细胞凋亡中的作用。方法：用放射菌素D（Act D）诱导HL－60细胞发生凋亡。应用WesternBlot法检测细胞凋亡过程中RPS3a水平的变化，结果：HL－60细胞在诱发凋亡1小时后，RPS3a的水平与对照组比较显著升高，之后迅速降低，在诱导2h时其水平已低于正常对照组的水平，并随时间推移逐渐降低，在细胞凋亡的晚期，即诱导6h时，RPS3a已降至不可检测的水平，结论：RPRS3a水平出现的一过性增高随后迅速降低，这可能是发挥其核糖体外的功能，作为信号诱导细胞凋亡；在细胞凋亡的早期RPRS3a的不可逆性降致使核糖体的完整性丧失，从而抑制蛋白质的合成，最终导致细胞凋亡。     

谷胱甘肽S—转移酶异质性研究

本文以不同状态的大鼠肝细胞为对象,采用生化和病理学方法研究了谷胱甘肽S-转移酶(GST)的异质性。根据GST同工酶酶谱分忻、GST活性改变状况和GST同工酶表达活性的改变等结果,表明GST无论在正常状态或病理状态下均具有显著的异质性。     

Sodium 2-propenyl thiosulfate derived from garlic induces phase II detoxification enzymes in rat hepatoma H4IIE cells

There is evidence that onions and garlic protect against cancer in humans. It has been suggested that this effect is partly due to the organosulfur compounds in Allium vegetables and that these substances act through induction of phase II detoxification enzymes. Here, we hypothesized that alk(en)yl thiosulfates, sodium n-propyl thiosulfate (NPTS), and sodium 2-propenyl thiosulfate (2PTS), which were identified in onions and garlic, respectively, may induce phase II enzymes. Therefore, rat hepatoma cells (H4IIE) were cultured with 1 to 100 μmol/L of NPTS or 2PTS for 48 hours at 37°C; and the activities and messenger RNA (mRNA) expression levels of phase II enzymes in H4IIE cells were investigated. The effects of diallyl trisulfide and tert-butylhydroquinone, known as phase II inducers, were also examined as positive controls and compared with the responses of NPTS and 2PTS. Quinone reductase (QR) activity and mRNA expression levels of QR and epoxide hydrolase 1 were significantly increased by 2PTS (P < .05-.005). In particular, QR activity was increased at a relatively low concentration of 2PTS (10 μmol/L). However, glutathione S-transferase activity and mRNA expression levels of glutathione S-transferase A5 and uridine diphosphate glucuronosyl transferase 1A1 were not changed by 2PTS. In contrast, NPTS did not affect the activities and mRNA expression levels of these phase II enzymes. These results show that 2PTS can induce phase II enzymes, and its inductive effect is comparable or superior to that of diallyl trisulfide and tert-butylhydroquinone.     

The identification of potential alternative biomarkers of nitrofurazone abuse in animal derived food products

Semicarbazide (SEM) was considered to be a characteristic protein-bound side-chain metabolite of the banned veterinary drug nitrofurazone and used as a marker of nitrofurazone abuse. It was recently discovered that SEM can arise in food from sources other than nitrofurazone. This uncertainty over the source of SEM may be overcome if alternative markers specific to tissue-bound nitrofurazone residues can be determined. The structure of nitrofurazone metabolites in vivo and particular proteins to which they are bound are not known. These proteins with altered structure due to the presence of the drug metabolites can be considered as potential alternative biomarkers of nitrofurazone abuse. The proteins implicated in the in vivo binding of nitrofurazone were separated and identified. A crude mixture of proteins extracted from the liver of a rat treated with the drug was separated using a series of different techniques such as preparative isoelectric focusing and size exclusion HPLC. Multiple fractions were assayed by LC-MS/MS to detect the presence of SEM. The proteins containing SEM residues were identified by peptide mass mapping using trypsin digestion and MALDI-TOF. The first protein identified as containing high concentration of SEM was albumin. It was also shown that low molecular weight species within a protein mixture whose main constituent was glutathione S-transferase contained a high concentration of SEM. The chemical composition of these components is under investigation. Preliminary data suggest the SEM forms part of a nitrofurazone metabolite conjugated to glutathione.     

Identification of aspartic acid-203 in human thymidine phosphorylase as an important residue for both catalysis and non-competitive inhibition by the small molecule “crystallization chaperone” 5′-O-tritylinosine (KIN59)

Thymidine phosphorylase (TP) is a catabolic enzyme in thymidine metabolism that is frequently upregulated in many solid tumors. Elevated TP levels are associated with tumor angiogenesis, metastasis and poor prognosis. Therefore, the use of TP inhibitors might offer a promising strategy for cancer treatment. The tritylated inosine derivative 5′-O-tritylinosine (previously designated KIN59) is a non-competitive inhibitor of TP which was previously found to be instrumental for the crystallization of human TP. A combination of computational studies including normal mode analysis, automated ligand docking and molecular dynamics simulations were performed to define a plausible binding site for 5′-O-tritylinosine on human TP. A cavity in which 5′-O-tritylinosine could fit was identified in the vicinity of the Gly405-Val419 loop at a distance of about 11 Å from the substrate-binding site. In the X-ray crystal structure, this pocket is characterized by an intricate hydrogen-bonding network in which Asp203 was found to play an important role to afford the loop stabilization that is required for efficient enzyme catalysis. Site-directed mutagenesis of this amino acid residue afforded a mutant enzyme with a severely compromised catalytic efficiency (V_max/K_m of mutant enzyme ∼50-fold lower than for wild-type TP) and pronounced resistance to the inhibitory effect of 5′-O-tritylinosine. In contrast, the D203A mutant enzyme kept full sensitivity to the competitive inhibitors 6-aminothymine and 6-amino-5-bromouracil, which is in line with the kinetic properties of these inhibitors. Our findings reveal the existence of a previously unrecognized site in TP that can be targeted by small molecules to inhibit the catalytic activity of TP.     

耐药癫痫患者和难治性癫痫大鼠脑组织与外周血GST-π蛋白表达

目的考察耐药癫痫患者和难治性癫痫大鼠脑组织与外周血谷胱甘肽S-转移酶π(glutathione S-transferaseπ,GST-π)的表达。方法临床收集2010年1月至2014年3月在我院神经外科行手术治疗的耐药性癫痫患者32例,选择同期因脑血管畸形手术切除的10例正常脑组织为对照;动物试验:收集15只对照组和20只难治性癫痫大鼠组,难治性癫痫依据锂-匹罗卡品癫痫模型制备。检测人与大鼠外周血和脑的GST-π水平表达,并进行组间比较。结果耐药癫痫患者脑组织的颞叶20例,额叶6例,枕叶6例。GST-π在耐药癫痫患者和难治性癫痫大鼠脑的颞叶、额叶、枕叶中均有表达,存在于细胞胞浆,染成浅黄色至棕黄色颗粒,与对照组比,耐药癫痫患者和难治性癫痫大鼠脑和外周血GST-π表达明显增加(P0.05)。结论 GST-π可能参与耐药癫痫的发病,检测外周血GST-π为耐药性癫痫的治疗提供参考。