尿微量白蛋白与β_2微球蛋白在糖尿病肾病早期的临床意义

目的探讨尿微量白蛋白(mALB)和尿β2微球蛋白(β2MG)对糖尿病肾早期诊断的临床意义。方法将研究对象分为糖尿病组与对照组。免疫透射比浊法检测尿微量白蛋白和尿β2微球蛋白(β2MG),酶法检测血清肌酐和尿素氮。结果糖尿病组患者尿微量白蛋白和尿β2微球蛋白(β2MG)水平均显示高于正常对照组(P<0.01),具有统计学意义;两组血清尿素氮、肌酐数值增高不明显,无统计学意义(P>0.05);mALB和β2MG联合检测的阳性率明显高于单一检测阳性率,组间比较差异具有(高度)统计学意义(P<0.05,P<0.01)。结论尿微量白蛋白和β2微球蛋白水平对糖尿病肾病的早期诊断具有重要的意义,联合检测可提高糖尿病早期肾功能损害的检出率。     

多项指标联合检测在糖尿病肾病早期诊断价值

目的：探讨尿α1-微球蛋白（α1-m）、β2-微球蛋白（β2-m）、Tamm-Horsfall蛋白（TH蛋白）、尿微量自蛋白（MAIb）及尿微量白蛋白／肌酐（MAIb／Cr）的检测在糖尿病肾病（DN）的早期诊断及病情进展中的应用价值。方法：100例糖尿病患者按照尿蛋白阴性、＋、＋＋、＋＋＋、＋＋＋＋分为5组，用放射免疫分析检测α1-m、β2-m、TH蛋白，用免疫透射比浊法测定患者尿中的MAIb），用酶法测定尿肌酐的含量，并与正常对照组作比较。结果：糖尿病患者尿蛋白阴性时，各项指标与对照组差异无显著性，当尿蛋白出现阳性时，各项指标与对照组差异有显著性．随病变进展异常改变更趋显著。结论：5项指标联合检测更有助于提高DN的早期检出率以及肾脏受损程度的鉴别．对DN的防治有重要价值。     

β2-微球蛋白在糖尿病肾病早期诊断中的临床意义

目的探讨糖尿病肾病（DN）早期诊断中β2-微球蛋白（β2-MG）检测的临床价值。方法选取2012年1-8月确诊为DN患者80例为观察组,选取同期进行健康体检合格者80例为对照组,两组受试者均进行β2-MG检测,同时检测与肾功能评价密切相关的血清生化指标尿素（Urea）和肌酐（Cr）水平,并对结果进行综合分析。结果观察组β2-MG、Urea、Cr水平明显高于对照组,两组比较,差异有统计学意义（P〈0．05）;不同病期DN患者β2-MG呈递增性升高,Ⅱ、Ⅲ、Ⅳ期DN患者β2-MG水平明显高于I期患者,差异有统计学意义（P〈0．05）,I期DN患者B2．MG检测阳性率为88．5％,与同期Urea、Cr阳性栓出率（53．8％、46．2％）比较,差异有统计学意义（P〈0．05）。结论β2-MG是早期诊断DN的一项良好的血清生化指标。其水平变化可为临床诊治DN提供依据。     

依那普利治疗早期糖尿病肾病31例

目的观察依那普利治疗早期糖尿病肾病的疗效。方法将65例2型糖尿病并早期糖尿病肾病患者随机分为观察组(33例)和对照组(32例),两组均给予常规综合治疗,包括降血糖、利尿、给予必需氨基酸及低盐、优质低蛋白饮食等,观察组在常规治疗基础上加用马来酸依那普利片口服,两组均连续治疗12周。对两组患者监测血压、测定空腹血糖(FPG)、餐后2 h血糖(2 h PBG)、血尿素氮(BUN)、肌酐(Scr)、内生肌酐清除率(Ccr);采用免疫透射比浊法检测尿微量白蛋白(u-ALB),采用微粒子酶免分析法检测β2微球蛋白(β2-MG)。结果治疗后观察组FPG,2 h PBG,Scr,Ccr改善程度明显优于对照组(P<0.05),观察组u-ALB和β2-MG水平显著降低(P<0.01)。结论依那普利能明显降低早期糖尿病肾病患者尿微量白蛋白和β2微球蛋白水平,有助于保护肾功能,延缓糖尿病肾病的进展。     

2型糖尿病血清β2-微球蛋白的临床意义

目的探讨2型糖尿病与血清β2-微球蛋白的关系。方法采用放射免疫分析法对30例正常人,72例2型糖尿病患者（其中早期糖尿病肾病35例,简写为DN,糖尿病未出现肾损害37例,简写为NDN）进行血清β2-微球蛋白测定,并分组进行分析。结果DN组血清β2-微球蛋白含量明显高于对照组,P〈0．01,差异有统计学意义。对照组、NDN组及DN组血清β2-微球蛋白的含量逐渐递增,NDN组与对照组相比,P〉0．05,差异无统计学意义。结论血清β2-微球蛋白在DN组明显升高,提示该指标对预测早期糖尿病肾病有重要意义     

血清β2-微球蛋白及尿白蛋白与早期糖尿病肾病相关性研究

目的血清β2-微球蛋白、尿微量白蛋白的测定在早期糖尿病肾病诊断中的意义。方法采用散射比浊法,对常规检测尿蛋白阴性的糖尿病患者进行血清β2-微球蛋白、尿白蛋白的含量检测。结果糖尿病尿蛋白阴性患者的血清β2-微球蛋白及尿白蛋白的含量均显著高于正常对照组。结论联检血β2-微球蛋白、尿白蛋白是预测糖尿病肾损害的简便可行的方法,对糖尿病肾病的早期诊断具有重要的临床价值。     

尿微量白蛋白和α1-微球蛋白在糖尿病肾病早期筛查和治疗中的作用

目的 探讨尿微量白蛋白(U-Alb)和α1-微球蛋白(α1-MG)在糖尿病肾病早期筛查和治疗中的作用.方法 测定96例糖尿病患者和93例健康体检者血尿素氮(BuN)、肌酐(Cr)、尿蛋白定性、尿微量白蛋白和α1-微球蛋白并进行分析.结果 糖尿病组尿微量白蛋白和α1-微球蛋白的测定结果明显高于健康对照组(P<0.01),具有统计学意义.结论 定期进行尿微量白蛋白和α1-微球蛋白的联合检测对糖尿病肾病早期诊断有重要意义.     

尿视黄醇结合蛋白在糖尿病肾病早期的诊断意义

目的 探讨尿视黄醇结合蛋白(RBP)在糖尿病肾病早期的诊断意义。方法 采用酶联免疫法(ELISA)测定78例非胰岛素依赖型糖尿病(NIDDM)患者和30例健康人的尿RBP,同时测定尿微量白蛋白(Alb)和尿β_2-微球蛋白(β_2-MG)。结果 NIDDM患者RBP较正常对照组增高,尿RBP与尿Alb、尿β_2-MG呈正相关。结论 尿RBP可作为诊断早期糖尿病肾病的敏感指标。     

DM2肾病患者血清胱抑素C和尿β2-m,mAlb水平检测的临床意义

目的 探讨DM2肾病患者血清胱抑素C和尿β2-m,mAlb水平的变化及临床意义.方法 应用放射免疫分析法和酶联法对36例DM2肾病患者进行了血清胱抑素C和尿β2-m,mAlb水平检测,并与35名正常健康人作比较.结果 DM2肾病患者血清胱抑素C和尿β2-m,mAlb水平均非常显著地高于正常人组（P＜0.01）,且血清胱抑素C和尿β2-m,mAlb水平呈显著正相关（r值分别为0.5984、0.6012,P值均小于0.01）.结论 检测血清胱抑素C和尿β2-m,mAlb水平的变化对了解病情和指导临床治疗均具有一定的临床实用价值.     

血、尿α_1-微球蛋白、β_2-微球蛋白在糖尿病肾病早期诊断中的意义

采用放射免疫测定方法（ＲＩＡ）,观察４３例糖尿病患者血、尿α１微球蛋白（α１ＭＧ）、β２微球蛋白（β２ＭＧ）及血肌酐（ＳＣｒ）变化。结果显示：常规尿蛋白阴性,ＳＣｒ含量异常者占２５５％,尿中α１ＭＧ／Ｃｒ、β２ＭＧ／Ｃｒ、血α１ＭＧ、β２ＭＧ异常者分别占７９０７％、６７４４％、３７２１％、５５８１％。各种微球蛋白测定的阳性率与ＳＣｒ比较,差异有显著性（Ｐ＜００５）。结果说明血、尿α１ＭＧ、β２ＭＧ在诊断糖尿病早期肾损伤中有一定的临床意义。并提示血、尿α１ＭＧ、β２ＭＧ测定在反映肾脏损伤中比ＳＣｒ敏感。     

Further observations on in vitro and in vivo effects of 2,5-hexanedione on glyceraldehyde-3-phosphate dehydrogenase

The in vitro and in vivo effect of aliphatic diketones has been studied on glyceraldehyde-3-phosphate dehydrogenase (GAPDH) d,l-glyceraldehyde-3-phosphate: NAD oxidoreductase (phosphorylating EC 1.2.1.12 activity). Neurotoxic diketone, 2,5-hexanedione (2,5-HD), but not 2,4-hexanedione (2,4-HD), a non-neurotoxic diketone, inhibited GAPDH in rat brain homogenate preincubated with 25 mM diketones for 20 min. If the preincubation period was increased to 2 h, approximately 25% and 55% inhibition of GAPDH activity was observed with 1 mM and 5 mM 2,5-HD respectively. The inhibiton of GAPDH activity was also seen in sciatic nerves but not in the brain or liver homogenates of rats chronically intoxicated with 2,5-HD for 12 weeks. The inhibition of GAPDH by 2,5-HD appears to be selective, and thus confirms earlier data from this laboratory.     

Deregulation of glycolysis in cancer: glyceraldehyde-3-phosphate dehydrogenase as a therapeutic target

Introduction: Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a key glycolytic enzyme, but recent studies have shown its non-glycolytic role in cell death, survival mechanisms and diseases. Increase in glycolysis, in particular overexpression of GAPDH, has been considered an important feature of many types of cancer cells. This review focuses on the role of GAPDH in carcinogenesis and the possibility of using this target for anticancer therapy.

Areas covered: In this review, the studies targeting GAPDH in human cancer as well as its functions in normal and cancer cells are described and discussed.

Expert opinion: GAPDH is an essential component of the glycolysis energy system, which is actively employed in cancer cells. Analysis of the so-called bioenergetics signature (the ratio of beta-F1-ATPase and GAPDH proteins) of different cancer types can be used for estimation of the cell metabolic activity, cancer aggressiveness and response to chemotherapy. Recent studies suggest GAPDH as a promising target for therapy of some carcinomas. Incidentally, limitations of this approach may come from the versatility of the GAPDH enzyme, since it combines glycolytic, pro-apoptotic and other activities. Hence, targeting GAPDH may lead to unexpected results concerning normal cells and therefore requires further research.     

A prolyl oligopeptidase inhibitor, Z-Pro-Prolinal, inhibits glyceraldehyde-3-phosphate dehydrogenase translocation and production of reactive oxygen species in CV1-P cells exposed to 6-hydroxydopamine

We studied the ability of prolyl oligopeptidase (POP) inhibitors, Z-Pro-Prolinal and JTP-4819, to prevent translocation of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and formation of reactive oxygen species (ROS), in 6-hydroxydopamine (6-OHDA) and cytosine arabinoside (Ara-C) treated monkey fibroblast (CV1-P) and human neuroblastoma (SH-SY5Y) cells. The cells were pretreated with POP inhibitors (30 min) before addition of toxicants. GAPDH was analyzed by Western hybridization, ROS by fluorescent 2′7′-dichlorodihydro-fluorescein diacetate, and viability by the MTT method. Both toxicants induced GAPDH translocation to the particulate fraction (mitochondria and nuclei). Z-Pro-Prolinal was able to inhibit the translocation in 6-OHDA-exposed CV1-P cells. In SH-SY5Y cells and in JTP-4819 pretreated cells, no prevention of translocation was seen. However, the intensity of GAPDH in cytosolic fraction increased. Both inhibitors blocked 6-OHDA-induced ROS-production to the control level in CV1-P but, not in SH-SY5Y cells, without affecting their viability. In conclusion, POP inhibitors are able to prevent certain cell stress related factors such as ROS production or GAPDH translocation.     

Antioxidant and prooxidant effects of quercetin on glyceraldehyde-3-phosphate dehydrogenase.

Anti- and prooxidant properties of quercetin under different conditions were investigated using glyceraldehyde-3-phosphate dehydrogenase, a glycolytic enzyme containing essential cysteine residues. Quercetin was shown to produce hydrogen peroxide in aqueous solutions at pH 7.5, this resulting in the oxidation of the cysteine residues of the enzyme. Quercetin significantly increased oxidation of GAPDH observed in the presence of ferrous ions, particularly when FeSO(4) was added to the solution containing GAPDH and quercetin. The results suggest the formation of hydroxyl radical in the case of the addition of FeSO(4) to a quercetin solution. At the same time, quercetin protects GAPDH from oxidation in the presence of ascorbate and Fe(3+). In the absence of metals, quercetin protects SH-groups of GAPDH from oxidation by the superoxide anion generated by the system containing xanthine/xanthine oxidase.     

Effector-induced dissociation of glyceraldehyde-3-phosphate dehydrogenase discriminated by urea solvation

The dissociation of glyceraldehyde-3-phosphate dehydrogenase (GAPD) from pig muscle in water solutions (0.1 M phosphate, pH 7) at increased urea concentrations was studied by means of frontal-gel chromatography, intrinsic (TRP) fluorescence, differential absorption spectroscopy and selective chemical modification at TRP-193. The results are in agreement with a consecutive two-step model of dissociation of the tetramer and the dimer (C*_T= 0.42 M urea < C*_D= 1.39 M urea). The binding of effector(s) destabilizes the oligomeric structures (ΔG^T changes from -1.00 to -0.54 kcal/mol; ΔG^D from -2.30 to -1.22 kcal/mol). The introduction of the bulky Koshland-reagent group to TRP-193 at the subunit-subunit interface leads to a decrease of the stability with δΔG ? 1 kcal/mol, owing to TRP-193…TYR-39…TYR-92 cluster destruction. By using lobster GAPD atomic coordinates (PDB file 1GPD) and pig muscle GAPD amino-acid sequence, a tentative molecular model was constructed and the subunit contacts in terms of the Lee-Richard static accessibilities were described. A detailed analysis of the dissociation as a transfer of the buried residues from the molecular interface to the urea solutions was performed.     

Structural basis for the thermal stability of glyceraldehyde-3-phosphate dehydrogenases

The amino acid sequences of two thermophilic and five mesophilic glyceralde-hyde-3-phosphate dehydrogenases have been compared with the known three-dimensional structure of this enzyme to determine the factors responsible for thermal stability. The changes are greatest in the S-loop regions at the center of the tetramer, which show a quantitative increase in hydrophobicity and polarity that can strengthen subunit interactions in a complementary manner. The S-loops also show increases in residue volume and bulk that may indicate a tighter packing at the molecular center. In addition, there are changes in the secondary structural parameters indicating that the helices, in particular, may be more stable in the thermophilic proteins. Increases in the hydrophobicity of domain and subunit contacts for the Thermus aquaticus glyceraldehyde-3-phosphate dehydrogenase may explain why it is the most thermostable protein in this series.     

Degradation of glyceraldehyde-3-phosphate dehydrogenase induced by acetylleucine chloromethyl ketone in U937 cells

We examined whether any changes were induced in cellular proteins by an inhibitor of acylpeptide hydrolase (ACPH) (EC 3.4.19.1), acetylleucine chloromethyl ketone (ALCK), which was shown in our previous report to induce apoptosis of human U937 cells. Extract prepared from U937 cells in 0.05% Triton X-100-PBS was incubated with ALCK at 37 degrees, and then analyzed using SDS-PAGE. A 36kDa protein in the cell extract was decreased markedly during the incubation period. This protein was purified and identified as glyceraldehyde-3-phosphate dehydrogenase (GAPDH, EC 1.2.1.12) by its specific enzyme activity, N-terminal amino acid sequence, and Western blotting. Incubation of purified GAPDH with ALCK resulted in a decrease of GAPDH activity, but not in a decrease in the amount of GAPDH. The ALCK-induced GAPDH decrease in the cell extract was abrogated by co-incubation with a serine protease inhibitor, diisopropyl fluorophosphate, suggesting that GAPDH was first inactivated by ALCK, and subsequently degraded by a serine protease(s). GAPDH degradation was also observed in U937 cell cultures in the presence of ALCK. The significance of GAPDH inhibition in the apoptotic process is discussed.     

alpha-Chlorohydrin inhibits glyceraldehyde-3-phosphate dehydrogenase in multiple organs as well as in sperm.

Numerous studies have documented inhibitory effects of alpha-chlorohydrin (ACH) on glyceraldehyde-3-phosphate dehydrogenase (G3PDH) activity in spermatozoa. A sperm-specific G3PDH isoform has been described. The possibility that ACH may inhibit G3PDH in cell types other than sperm was investigated in this work. In addition, the onset of ACH-induced epididymal toxicity was described. Changes to epididymal histology occurred 6 h following a single dose of ACH (50 mg/kg po) and were confined to the proximal initial segment. By 24 h, no epithelial cells lined the basement membrane of that region. Three h after ACH administration (50 mg/kg po), G3PDH activity was significantly decreased in sperm (85%) as well as in kidney (31%), liver (49%), and epididymis (35%). Enzyme activity remained inhibited at 6 and 24 h. G3PDH was immunolocalized in the epididymis and staining was highest in the efferent ducts and initial segment as well as in smooth muscle. Since G3PDH is a microtubule-associated protein and microtubule-dependent endocytosis occurs in the epididymis, beta-tubulin was also immunolocalized. beta-tubulin densely stained the apical region of initial segment and caput epithelial cells. Disruption of beta-tubulin immunostaining correlated with the localization and onset of the lesion. Co-localization of G3PDH and beta-tubulin immunostaining was not observed although both antibodies most densely stained the initial segment. Our data indicate that histologic changes to the proximal initial segment of the epididymis occur rapidly, but subsequent to G3PDH inhibition. Moreover, ACH inhibition of G3PDH is not confined to sperm, although the sperm enzyme is most sensitive to inhibition.     

Effect of an inactivator of glyceraldehyde-3-phosphate dehydrogenase, a fortuitous arsenate reductase, on disposition of arsenate in rats.

The environmentally prevalent arsenate (AsV) is reduced in the body to the much more toxic arsenite (AsIII). Recently, we have demonstrated that the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH) catalyzes the reduction of AsV in the presence of glutathione, yet the role of GAPDH in AsV reduction in vivo is unknown. Therefore, we examined the effect of (S)-alpha-cholorhydrin (ACH), which forms a GAPDH-inhibitory metabolite, on the reduction of AsV in rats. These studies confirmed the in vitro role of GAPDH as an AsV reductase, inasmuch as 3 h after administration of ACH (100 or 200 mg/kg, ip) to rats both the cytosolic GAPDH activity and the AsV-reducing activity dramatically fell in the liver, moderately decreased in the kidneys, and remained unchanged in the muscle. Moreover, the AsV-reducing activity closely correlated with the GAPDH activity in the hepatic cytosols of control and ACH-treated rats. Two confounding effects of ACH (i.e., a slight fall in hepatic glutathione levels and a rise in urinary AsV excretion) prompted us to examine its influence on the disposition of injected AsV (50 micromol/kg, iv) in rats with ligated bile duct as well as in rats with ligated bile duct and renal pedicles. These experiments demonstrated that the hepatic retention of AsV significantly increased, and the combined levels of AsV metabolites (i.e., AsIII plus methylated arsenicals) in the liver decreased in response to ACH; however, ACH failed to delay the disappearance of AsV from the blood of rats with blocked excretory routes. Thus, the GAPDH inactivator ACH inhibits AsV reduction by the liver, but not by the whole body, probably because the impaired hepatic reduction is compensated for by hepatic and extrahepatic AsV-reducing mechanisms spared by ACH. It is most likely that ACH inhibits hepatic AsV reduction predominantly by inactivating GAPDH in the liver; however, a slight ACH-induced glutathione depletion may also contribute. While this study seems to support the conclusion that GAPDH in the liver is involved in AsV reduction in rats, confirmation of the in vivo role of GAPDH as an AsV reductase is desirable.