The sugar-chain heterogeneity of human gamma-glutamyl transferases from the reproductive system and kidney

The sugar-chain heterogeneity of gamma-glutamyl transferase (gamma-GTP, EC 2.3.2.2) from the human reproductive system (seminal plasma, prostate and testis) and kidney was investigated using the serial lectin affinity technique and their properties were compared. According to the results of serial lectin affinity chromatography, a possible sugar chains of enzymes from reproductive system were mainly of the hybrid type without fucose linkages to the innermost GlcNAc and/or the biantennary complex type sugar chains and a few were of the multiantennary complex-type with branched GlcNAc (beta 1-4) Man and bisecting complex type sugar chains. On the contrary, the major sugar chains of kidney gamma-GTP were of the multiantennary complex type and/or bisecting complex type sugar chains. Results of isoelectric focusing showed the gamma-GTP bound multiantennary complex type sugar chains to be the most acidic glycoprotein. Moreover, the biantennary type sugar chains were slightly more acidic than the high mannose and/or hybrid type sugar chains, varying with the degree of sialylation.     

Comparative studies on the properties of purified gamma-glutamyl transferase from human reproductive system and the kidney

Gamma-glutamyl transferase (GGT) of the human seminal plasma and reproductive tissues was purified and its properties were compared to those of the enzyme from kidney. A single band of GGT was obtained by polyacrylamide gel electrophoresis. Purification was 1080-fold for seminal plasma, 206-fold for prostate, 608-fold for testis and 382-fold for kidney. Similar Km value (0.87-1.06 mM) and optimum pH (8.2-8.5) were obtained for the enzymes of the four different sources. Their thermal stabilities were identical. However, inhibitions by Zn2+ and Cu2+ were different between kidney and reproductive system GGT. Molecular mass of the native enzyme was 78 kDa for seminal plasma, prostate and testis and 79 kDa and 105 kDa for kidney. The subunit molecular masses of the enzymes from seminal plasma, prostate and kidney consisted of three proteins, suggesting the precursor form, and the heavy and light subunits of the mature form.     

Correlation between estrogen receptor protein and carcinoembryonic antigen in normal and carcinomatous human breast tissue.

We determined estrogen receptor protein and carcinoembryonic antigen in cytosols prepared from 189 human breast carcinoma tissues, 85 benign or normal breast biopsies, and 101 tissue specimens metastatic from breast carcinoma. Carcinoembryonic antigen was observed in 70% of the primary carcinomatous tissues, 15% of the benign or normal specimens, and 51% of the metastases. Ninety-six of the 189 primary carcinomatous specimens with increased concentrations of carcinoembryonic antigen were also positive for estrogen receptor protein, whereas 67 of the 72 benign or normal biopsies with low concentrations of carcinoembryonic antigen were also negative for estrogen receptor protein. All five fenign specimens with positive estrogen receptor protein and normal carcinoembryonic antigen concentrations were from fibroadenomas. The concordance between estrogen receptor protein and carcinoembryonic antigen in the primary carcinomatous tissue was 66%, in metastatic carcinoma 51%, and in benign and normal tissue 85%.     

A novel gamma-glutamyl transpeptidase in renal carcinoma in comparison with normal kidney enzyme.

A novel gamma-glutamyl transpeptidase was found in a renal carcinoma tissue. This enzyme electrophoresed more quickly than that of normal kidney, but more slowly than normal liver enzyme. Their Rf values are 0.46, 0.33, and 0.52 for the renal carcinoma, normal kidney and normal liver enzyme, respectively. After treatment of the renal carcinoma and normal kidney enzyme with neuraminidase, the renal carcinoma gamma-glutamyl transpeptidase still electrophoresed slightly faster than that of normal kidney. The catalytic properties of the renal carcinoma gamma-glutamyl transpeptidase were almost the same as those of normal kidney except for molecular weight; the molecular weight for the renal carcinoma was estimated to be about 130 000, while that of normal kidney was about 90 000. These results may mean that the enzyme of the renal carcinoma is different from that of normal kidney in chemical constituents other than sialic acid. Of 10 patients with renal carcinoma examined electrophoretically, 5 posessed this novel enzyme in their renal carcinoma tissues.     

Differences between purified ectopic and normal alpha subnits of human glycoprotein hormones.

"Ectopic" proteins, not distinguished immunologically from the common alpha subunit of the human glycoprotein hormones, were purified approximately 10,000-fold from a gastric carcinoid tumor (A.L.-alpha) and from tissue culture medium of bronchogenic carcinoma cell lines (ChaGo-alpha). The purified A.L.-alpha was homogeneous by sodium dodecyl sulfate (SDS) gel electrophoresis while the purified ChaGo-alpha showed multiple components, some of which represented aggregated species. In SDS gel electrophoresis, the apparent molecular weights of A.L.-alpha (15,000) and dithioerythritol-reduced ChaGo-alpha (13,000) were significantly lower than those of the alpha subunits of human chorionic gonadotropin (hCG-alpha), luteinizing hormone, follicle-stimulating hormone, or thyroid-stimulating hormone (22,000-23,000). Binding experiments with [35S]-SDS suggested that these apparent differences in molecular weight resulted, at least in part, from diminished binding of the SDS by the normal compared to the ectopic alpha subunits. In gel chromatography, the apparent molecular weights of A.L.-alpha (27,000) and ChaGo-alpha (30,000) were slightly higher than those of normal alpha subunits (23,000-24,000). Both A.L.-alpha and ChaGo-alpha were not distinguished from hCG-alpha in ion-exchange chromatography. The composition of A.L.-alpha was similar to that of hCG-alpha in 13 amino acids but showed decreased phenylalanine and increased valine; glucosamine was identified in both A.L.-alpha and hCG-alpha. Under conditions in which hCG-alpha combined with the hCG beta subunit (hCG-beta) to produce 95% of the expected gonadotropin-binding activity in a rat testis radioreceptor-assay, A.L.-alpha incubation with hCG-beta resulted in only 2% of the expected activity, and ChaGo-alpha incubation with hCG-beta produced no detectable activity. These characteristics of ectopic alpha subunits may reflect abnormalities of neoplastic protein synthesis or carbohydrate attachment which result in polypeptides with chemical and immunologic similarity to normal subunits but with differences in physical and combining properties; alternatively, the ectopic subunits may represent as yet unrecognized alpha precursor forms.     

Chemical nature of intestinal-type alkaline phosphatase in human kidney.

Approximately 10% of the alkaline phosphatase activity in human kidney is derived from the intestinal-type alkaline phosphatase isoform, which can be differentiated from adult intestinal alkaline phosphatase by selective reactivity with monoclonal antibodies. The NH2-terminal sequence of the renal intestinal-type alkaline phosphatase was shown to be identical to sequences of the adult and meconial alkaline phosphatases except for the NH2-terminal valine residue, which is missing in the renal intestinal-type enzyme. Incubation of purified meconial alkaline phosphatase with kidney homogenate resulted in removal of the NH2-terminal valine residue, indicating the presence of aminopeptidases in kidney that catalyze this hydrolysis. Furthermore, the oligosaccharide chains of the renal intestinal-type alkaline phosphatase were shown to differ from those of meconial and adult intestinal alkaline phosphatases, as revealed by lectin affinity chromatography. The heterogeneity of the intestinal-type alkaline phosphatase can therefore be generated both by partial peptide bond hydrolysis and differences in glycosylation.     

Lack of value of serum gamma-glutamyl transferase in the diagnosis of hepatobiliary disease.

Serum gamma-glutamyl transferase (GGT, EC. 2.3.2.2. was measured in 173 patients with diseases of the hepatobiliary system (including metastatic cancer) and in 90 patients who were subsequently shown to have primary diseases of other etiology. All patients had been selected because they had abnormal alkaline phosphatase, aspartate aminotransferase or bilirubin on SMA 12/60 screening. Serum GGT was elevated in 97% of patients with primary hepatobiliary disease. The magnitude of the increase in GGT was variable in all groups and was unhelpful in differential diagnosis, even between medical and surgical cases. Moreover, GGT was abnormal in 69 patients who did not have primary hepatobiliary disease (77%), an incidence higher than that for other enzyme tests performed. We conclude that because GGT was more susceptible than other tests to spurious elevation in the absence of hepatobiliary disease and was unhelpful in differential diagnosis, it has little value apart from monitoring alcohol abuse and enzyme induction.     

The activity of gamma-glutamyl transferase in the serum of multiple sclerosis and other neurological diseases.

1. In the sera of 142 neurological patients, including 35 cases with multiple sclerosis (MS), and 20 normal controls the gamma-glutamyltransferase (gamma-GT) activity was determined. The result of this investigation was compared with the "combined hippuric acid test" formely often used in MS patients. 2. The MS patients are the only group which significantly differs from the normal controls. There are also small differences between them and some of the other neurological patient groups. About one-third of the MS patients show moderately pathological serum activities. On the other hand, patients with a definite liver affection, especially chronic alcoholics, regularly show an even more important elevation of the values, which makes this test a sensitive tool for the screening of alcohol-induced liver intoxications. 3. Although the gamma-GT test in not quite as sensitive in indicating organic defects in MS patients, it seems to reflected a disturbed liver function in the same way as the more complicated "combined hippuric acid test". It may therefore be used as a simple tool of finding and following up organic defects in MS patients.     

The association between an increased level of gamma-glutamyl transferase and systolic blood pressure in diabetic subjects

Gamma-glutamyl transferase (GGT) is an enzyme present in serum and on most cell surfaces and serves as an oxidative stress marker. Although serum GGT is associated with hypertension development, little data are available on the associations between GGT and hypertension among populations with diabetes mellitus (DM). Our aim was to investigate the potential association between the changes in systolic or diastolic blood pressure (SBP/DBP) and the GGT level in type 2 DM subjects, in comparison with non-DM subjects. In 179 non-DM and 177 DM subjects, SBP/DBP, body mass index (BMI), fasting plasma glucose, serum asparate aminotransferase, alanine aminotransferase and GGT were measured at the baseline and after a 1-year period. Between these 2-measurement points, in non-DM subjects, SBP and DBP levels were significantly increased, while GGT tended to increase. In contrast, in DM subjects, the mean levels of SBP, DBP and GGT remained unchanged. Multivariate analysis revealed that in non-DM subjects the degree of increase in SBP was significantly and positively correlated to that of GGT (beta = 0.165), along with age and BMI. Likewise, the increase in DBP was correlated to that of GGT in non-DM subjects (beta = 0.170). In contrast, in DM subjects, the degree of increase in SBP was significantly correlated to that of only GGT (beta = 0.166). These results suggest that the presence of DM may attenuate the effects of GGT on DBP.     

Effect of anticonvulsant drugs on urinary excretion of gamma-glutamyl transferase in women.

Excretion of gamma-glutamyl transferase was found to be significantly higher in a group of epileptic patients on long-term anticonvulsant therapy than in normal subjects. The losses, which may reflect renal enzyme induction, were unrelated to drug dose or duration of therapy and were significantly lower than those found in normal healthy pregnant women at term.     

The enzymatic mechanisms for deoxythymidine synthesis in human leukocytes: IV. Comparisons between normal and leukemic leukocytes

(1) Synthesis of deoxythymidine by either direct transfer of deoxyribosyl to thymine (pyrimidine deoxyribosyltransferase) or by a coupled deoxynucleoside phosphorylase mechanism is approximately twofold greater with normal leukocyte extracts (55 to 88% granulocytes) than with extracts prepared from leukocytes obtained from patients with chronic myelogenous leukemia. Activities in lymphocytes (normal or leukemic) are one-fifth the activity of normal granulocytes.(2) The lower activity in chronic myelogenous leukemia remains at 50% of normal even when patients are in hematologic remission with a normal per cent mature granulocytes in the peripheral blood.(3) The leukemic enzyme could not be distinguished from the normal by pH optima, thermal stability, or kinetic properties. The Km's for the deoxyribosyl acceptor and deoxyribosyl donors were identical for both enzymes. Both are subject to substrate inhibition by thymine and to inhibition by purine bases with similar Ki's. In addition, the transferase component of both the leukemic and the normal cell enzyme is activated by phosphate and arsenate. It appears, therefore, that there is no qualitative difference between the enzyme obtained from leukocytes of patients with chronic myelogenous leukemia and the enzyme obtained from normal leukocytes, suggesting that the difference in total cell activity is due to an actual decrease in amount of enzyme in chronic myelogenous leukemia or to a mixed cell population, one with a normal quantity of enzyme and the other with little or no active enzyme.(4) In both the normal cell and the leukemic cell extracts, transferase and phosphorylase activities could not be separated. The ratio of the two activities remained constant over a 140- and a 230-fold purification in normal and leukemic cell extracts, respectively. These and other observations indicate that transferase and phosphorylase activities are associated with the same protein.(5) The metabolism of pyrimidine and purine deoxynucleosides is similar for normal and leukemic cells. Catabolism of all deoxynucleosides tested was by direct phosphorolysis, except for deoxyadenosine which required initial deamination to deoxyinosine before phosphorolysis. In contrast to the greater rates of pyrimidine deoxynucleoside synthesis and cleavage with normal leukocyte extracts, the rates of purine deoxynucleoside synthesis and cleavage were approximately twofold greater with extracts prepared from cells of patients with chronic myelogenous leukemia. There was no significant difference in the rate of phosphorolytic cleavage of pyrimidine nucleosides (uridine) between the CML and normal leukocyte extracts.     

A short review of techniques for the localisation of gamma-glutamyl transferase isoenzymes after electrophoresis.

The development of a sensitive method for the localisation of gamma-glutamyl transferase isoenzymes after electrophoresis on various media is described. Recommendations on final conditions are given for optimal concentrations of substrate, acceptor and buffer, and for pH of the reaction mixture. The reaction conditions are compared with other isoenzyme localisation techniques previously reported.     

Clinical usefulness of the simultaneous localisation of the isoenzymes of alkaline phosphate and gamma-glutamyl transferase

Simultaneous localisation of the isoenzymes of alkaline phosphatase and gamma-glutamyl transferase was carried out on 173 serum samples from patients with hepatobiliary and bone disease to determine (i) if a better discrimination of the underlying pathology could be obtained by such an approach, and (ii) if the pathology suggested by the alkaline phosphatase isoenzyme separation could help in determining the cellular location of the gamma-glutamyl transferase isoenzymes. It was found that the localisation of the gamma-glutamyl transferase isoenzymes had, with a few exceptions, little advantage over the measurement of the total activity but that some evidence concerning the cellular location of the gamma-glutamyl transferase isoenzymes could be gained by such an approach. There was support for the view that the alpha 2-migrating isoenzyme is derived from the liver parenchymal cytoplasm, but little support for the view that the alpha 1-migrating isoenzyme is predominantly the induced microsomal enzyme or that the beta-migrating isoenzyme is the result of impaired biliary secretion.     

脑梗死患者与健康体检者颅外段脑动脉血管结构和血流动力学变化的差异

背景颈动脉粥样硬化患者中脑梗死发生率较高,但缺少大样本的研究,尤其缺少血流动力学变化的研究.目的研究脑梗死患者颅外段脑动脉血管结构和血流动力学变化特点,并与健康人进行差异比较.设计以诊断为依据,病例对照研究.地点和对象1999-01/2002-05中国铁道建筑总公司总医院住院患者300例(脑梗死组),其中男242例,女58例.2000-02/2001-10在本院体检健康者105例(健康组),男50例,女55例.主要观察指标血管内径及颈总动脉内壁(内膜与中膜之和);血流参数测定收缩期最大血流速度(Vmax)、舒张期最小血流速度(Vmin)、血管阻力指数(RI)、收缩期颈内动脉血流速度与颈总动脉血流速度的比值(VICA/VCCA).结果脑梗死患者梗死侧血管粥样硬化斑发生率(55.8%)明显高于非梗死侧(46.3%),差异有显著性意义(x2=4.487,P＜0.05),侧向符合率高;其血流参数的改变以Vmin,VICA/VCCA及RI最为明显,差异均有高度显著性意义(P＜0.01～0.001).结论脑梗死患者梗死侧血管粥样硬化斑的发生率明显高于非梗死侧,侧向符合率高;血流参数的改变以MIN明显减慢、RI明显增高为主要特点.     

Serum gamma-glutamyl transferase activity in chronic renal failure during regular hemodialysis and after successful renal transplantation.

Serum gamma-glutamyl transferase activity (gamma-GT) was measured in 108 uraemic patients, 110 patients on regular haemodialysis and 71 sucessfully transplanted patients. The incidence of abnormal results was 11, 14 and 28%, respectively. The results in the non-dialyzed and dialyzed patients indicate that in uraemia serum gamma-GT activity remains normal and as such, significant hepatic microsomal enzyme induction does not appear to occur. Raised serum gamma-GT in uraemic patients and after renal transplantation should suggest concurrent, added, pathology such as hepatobiliary disease or the administration of hepatic microsomal enzyme inducing drugs.