Expression of glutathione S-transferases and cytochrome P450 in normal and tumor breast tissue

The level of expression of glutathione S-transferases (GSTs) and cytochrome P450s in breast tissue are potentially important determinants in both the susceptibility of this tissue to the mutagenic effects of chemical carcinogens and in the response of breast tumors to chemotherapy. In this study we have investigated the expression of these proteins in 41 tumor and surrounding normal breast tissue samples by measurement of substrate metabolism. Western blot analysis and immunohistochemistry. In addition, we have quantitated the concentration of alpha, mu and pi class GST subunits using radioimmunoassay. All three classes of GST were expressed in breast tissue. The pi and mu class enzymes preponderate. Both the polymorphic mu class GST as well as a further form, present in all individuals, were found in high concentration. The polymorphic mu class GST was expressed in approximately 50% of the samples, which is consistent with the frequency of this polymorphism in the population and therefore does not appear to be a factor in susceptibility to this disease. Interestingly, although levels of the alpha class GST were very low, in two tumor samples extremely high levels of the B1B1 subunit were detected. Immunohistochemical studies showed significant variability in the localization of the pi class of GST between normal epithelial cells, infiltrating plasma cells and tumor cells, and in some samples GST pi appeared to be almost absent from the tumor tissue. No direct, or inverse correlation was found between GST pi concentration determined by radioimmunoassay and estrogen receptor levels. However, when studied by immunohistochemistry estrogen receptor negative tumors did tend to have higher GST pi content. The only cytochrome P450 detectable by Western blot analysis was a member of the P450IIC gene family. This was apparently distinct from the P450IIC proteins expressed in the liver and was detected in normal and tumor tissues to a similar extent.     

Expression of transforming growth factor-alpha in gastric carcinoma and normal gastric mucosa cells.

The presence of transforming growth factor-alpha (TGF-alpha) was studied in a total of 120 consecutive patients with gastric carcinomas. This immunohistochemical study found TGF-alpha expression in 60% (n = 72) of carcinomas. It also was detected in normal gastric epithelial mucosa in 36% (n = 43) of specimens. There was no significant statistical correlation between TGF-alpha expression and clinical features (e.g., patient age and sex) or pathologic features (e.g., tumor stage, grade, and localization or growth pattern according to an earlier histologic classification). Expression of TGF-alpha by these tumor cells had no influence on prognosis. It was neither a histopathologic marker of high malignancy nor a useful prognostic tool. It had no influence on the invasive growth pattern of gastric carcinoma.     

Enhanced expression of glutathione S-transferases in colorectal carcinoma compared to non-neoplastic mucosa

Ten paired samples of primary human colorectal carcinoma and adjacent non-neoplastic mucosa were analysed for total glutathione S-transferase (GST) activities as determined by 1-chloro-2,4-dinitrobenzene assays. These tissues were also investigated for the expression of acidic (pi), basic (alpha) and neutral (mu) GSTs using Western blotting procedures and immunohistochemical staining. For each of the paired samples examined the total GST activity was higher in tumour than in adjacent non-neoplastic mucosa. Western blotting, using an antibody against acidic GST also showed strong immunoreactivity in all the samples with more intense reactions in tumour compared to mucosa in nine out of the ten paired samples. Low levels of basic GST were also expressed in all samples of tumour and mucosa. Neutral GST was not detectable in two samples of tumour and corresponding mucosa, but low levels of expression were demonstrated in the remaining eight. Immunohistochemical staining for acidic GST showed a dark brown reaction in all tumour cells; in non-neoplastic mucosa there was positive immunoreactivity for epithelial cells situated deep within the crypts and a negative reaction for surface epithelial cells. Immunohistochemical staining for basic GST was negative except for one sample of tumour and two of mucosa. Neutral GST was expressed only in two samples of tumour and two samples of mucosa. We therefore conclude that there is enhanced expression of GSTs, acidic GST being the predominant form, in tumour compared to normal mucosa, in keeping with a role for GSTs in colonic carcinogenesis and acquired or innate drug resistance.     

Expression of differential nitric oxide synthase isoforms in human normal gastric mucosa and gastric cancer tissue

The present study investigated the expression and distribution of three isoforms of nitric oxide synthase (NOS) in different anatomical regions of the human stomach and in gastric neoplastic tissues by immunohistochemistry using specific antibodies. Intracellular localization of individual isoenzymes of NOS was detected in normal gastric mucosa. Gastric cancer tissues had a marked reduction of all three NOS isoforms expression. The expression of the endothelial NOS, neuronal NOS and inducible NOS in the tumor tissue was significantly lower than in normal gastric mucosa (P = 0.01, P = 0.02, P < 0.01, respectively). In the tumor tissue the expression of inducible NOS was significantly lower than the expression of both constitutive forms of NOS (P < 0.01). There was a tendency to higher expression of both constitutive forms of NOS in earlier stages T2 of the tumor compared to advanced T4 tumor. In contrast, the expression of inducible NOS was higher than in the advanced T4 tumor than in the earlier stages T2 of the tumor. The mapping of the expression of endothelial NOS, neuronal NOS and inducible NOS in human stomach showed higher expression of NOS isoforms in the distal third than in the proximal third of the stomach (P = 0.03, P = 0.04, P = 0.01, respectively). We conclude that there is greater expression of NOS in the stomach corpus and in antrum than in the proximal third of the normal human stomach mirroring the anatomical predilection of common pathological changes in this part of the human stomach. Furthermore, there was loss of the expression of individual isoenzymes in gastric neoplasms.      

Correlation of glutathione S-transferases with overall survival in patients with gastric carcinoma

Glutathione S-transferases (GST) are enzymes involved in the detoxification of xenobiotics and are divided into four subclasses, Alpha, Mu, Pi, and Theta. Most human gastrointestinal tumors contain increased amounts of GST Pi and GST enzyme activity. The relationship between GST parameters and tumor and patient characteristics, including overall survival, were studied retrospectively in normal and malignant gastric tissue from 49 patients with primary gastric carcinoma. Twelve patients (24%) were alive at the end of the study with a mean follow-up time of 4.1+/-0.4 years. Levels of GST Alpha, Mu, Pi and GST enzyme activity were not related to tumor stage, localization and diameter of the tumor, number of eosinophils in the tumor, presence of intestinal metaplasia in normal gastric mucosa, or gender and age of the patient. Optimal dichotomization and uni- and multivariate analyses were done with the Cox proportional hazard model. None of the clinicopathological parameters were associated with survival, except the number of eosinophils in the tumor. In contrast, high levels of GST Pi in both normal mucosa (Hazard ratio 3.0, p=0.02) and in gastric carcinoma (HR 2.2, p=0.05) and the presence of GST Mu in normal (HR 0.4, p=0.05) and malignant (HR 0.3, p=0.009) gastric tissue were found to have a significant prognostic value, independent from the clinicopathological parameters, when added separately to a Cox model. In conclusion, the levels of GST Mu and Pi in both normal or carcinomatous gastric tissue have an independent prognostic impact on overall survival.     

胃癌组织、癌前病变及正常胃黏膜组织中EphB2的表达及病理学意义

目的探讨EphB2受体在胃癌组织、癌前病变中的表达及对血管生成和肿瘤生物学行为的影响。方法应用链霉素亲生物素-过氧化酶免疫组织化学法(S-P)检测75例胃癌组织、45例癌前病变中EphB2的表达情况并与30例正常胃黏膜组织对照,采用CD34标记血管内皮细胞,计算微血管密度(MVD)。结果EphB2在胃癌组织及癌前病变表达高于正常胃黏膜组织,分别为64.0%、33.3%和10.0%,三者差异存在显著性(H=29.113,P<0.001)。EphB2在胃癌组织不同临床病理分期表达不同,分别为0、33.3%、52.2%、80.0%和100.0%,随临床病理分期增加表达率呈上升趋势(H=23.518,P<0.001)。CD34(以MVD表示)在胃癌组织、癌前病变及正常胃黏膜表达分别为67.61±13.13、33.92±3.60和21.29±3.28,呈下降趋势(F=318.831,P<0.001)。EphB2与CD34(以MVD表示)在胃癌组织中皆呈高表达,两者呈正相关(rs=0.915,P<0.001)。结论EphB2在胃癌组织中表达并随临床病理分期增加表达率呈上升趋势,EphB2在部分癌前病变中表达,EphB2参与肿瘤血管形成。     

Quantification of induction of rat oesophageal, gastric and pancreatic glutathione and glutathione S-transferases by dietary anticarcinogens

Four dietary, naturally occurring anticarcinogens (flavone,coumarin,     

Expression and different polarity of aminopeptidase N in normal human colonic mucosa and colonic tumors.

Expression and cellular localization of brush-border enzymes (aminopeptidase N, dipeptidylpeptidase IV, lactase, maltase) in normal human colon, colonic polyps and malignant intestinal tumors were investigated with a panel of monoclonal antibodies reacting with either native or denatured proteins. The enzymes were detected on cryostat sections by indirect immunofluorescence staining, or affinity-purified and analyzed by gel electrophoresis and immunoblotting. Dipeptidylpeptidase IV, lactase and maltase were absent from all samples examined, while aminopeptidase N (APN) was detected at the basal membrane of the epithelial cells in most specimens of colon obtained from individuals free of intestinal tumors. In contrast, APN was frequently localized at the luminal membrane of the surface epithelium in large-intestinal mucosa distal to tumors, adenomas and hyperplastic polyps, and from members of hereditary colon cancer syndrome families. APN was also expressed in colonic tumors, where it was present in an apical cell membrane location in 3/23 adenomas and 14/35 adenocarcinomas examined. No correlation was found between tumor-cell invasiveness (classified by "Dukes" stage) and expression or cellular location of aminopeptidase N. Histologically, all positive tumors were moderately or well differentiated. These results suggest that aminopeptidase N is normally expressed in adult human colon, but epithelial cells in the large and small intestine differ in their ways of sorting this enzyme intracellularly and eventually inserting it into different aspects of their surface membrane, a process which may be altered at an early stage of carcinogenesis.     

Glutathione and glutathione S-transferases in Barrett's epithelium.

Glutathione content, enzyme activity and isoenzyme composition of glutathione S-transferases were assayed in normal and Barrett''s esophageal epithelium of ten patients with Barrett''s esophagus. In addition, gastric and duodenal specimens from the same patients were also investigated. Glutathione content, glutathione S-transferase enzyme activity as well as glutathione S-transferase pi content were all significantly lower in Barrett''s epithelium as compared to normal esophageal mucosa. In contrast, glutathione S-transferase class alpha enzymes are markedly expressed in Barrett''s epithelium, whereas only low amounts are present in normal esophageal epithelium. Glutathione and glutathione S-transferase composition in Barrett''s epithelium show striking similarities with gastric epithelium, whereas duodenal epithelium is provided with considerable higher amounts of glutathione and glutathione S-transferases, except for levels of glutathione S-transferase class pi, which are lower. A significant negative correlation exists between glutathione S-transferase enzyme activity in the mucosa along the gastrointestinal tract, and the tumour incidence. Since glutathione and glutathione S-transferase are correlated with protection against cellular or cytogenetic damage, the low content of glutathione and glutathione S-transferases in the Barrett''s esophagus may be a factor of relevance for the increased tumour risk in this tissue.     

Genetic polymorphisms of glutathione S-transferases and the risk of adult brain tumors: a meta-analysis.

BACKGROUND: Studies investigating the association between genetic polymorphisms of glutathione S-transferases (GST) and risk of adult brain tumors have reported conflicting results. The rationale of this meta-analysis was to determine whether GST variants increase the susceptibility of adult brain tumors by pooling data. METHODS: Two investigators independently searched the HuGENet database, MEDLINE, EMBASE, conference articles, and manually reviewed bibliographies of retrieved articles. Papers were included if they were observational studies investigating the influence of GSTM1, GSTT1, GSTP1 I105V, or GSTP1 A114V on the development of adult brain cancers. Potential sources of heterogeneity between studies were explored in a meta-regression. RESULTS: We identified eight eligible studies, which included 1,630 cases of glioma, 245 cases of meningioma, and 7,151 controls. Using the random effects model, there was no association between any of the GST variants and the risk of glioma [overall odds ratio (OR), 1.08; 95% confidence interval (95% CI), 0.95-1.22]. Subgroup analyses also showed no relationship between GST variants and histopathologic groups; the overall ORs were 1.13 (95% CI, 0.88-1.43) for high-grade glioma and 1.08 (95% CI, 0.76-1.55) for low-grade glioma. A random effects meta-regression suggested that the use of in-hospital controls produced larger effect estimates in glioma than the use of population controls (overall OR, 1.30; 95% CI, 1.03-1.65). The T1 null genotype was significantly associated with a risk of meningioma (OR, 1.95; 95% CI, 1.02-3.76), but the M1 variant was not. CONCLUSION: This study did not suggest any relationship between GST variants and risks of glioma; the T1 null genotype may influence the susceptibility of meningioma, but larger studies are needed to substantiate this relationship.     

Levels and expressions of orotate phosphoribosyltransferase in gastric carcinoma and normal gastric mucosa tissues

Background Orotate phosphoribosyltransferase (OPRT; EC 2.4.2.10), a key enzyme that catalyzes one of the primary steps in the phosphorylation of fluoropyrimidine, was recently recognized as an important enzyme that determines the anticancer effects of the dihydropyrimidine dehydrogenase-inhibitory fluoropyrimidine, S-1. Methods Levels of OPRT were examined in 97 gastric carcinoma tissues and 65 normal gastric mucosa tissues obtained from patients with gastric carcinoma. The relation between OPRT levels and clinicopathological variables was evaluated, and correlations of OPRT with thymidylate synthase and dihydropyrimidine dehydrogenase levels in gastric carcinoma tissues were evaluated. Results Although OPRT levels were high in well-differentiated and localized carcinomas, they were not correlated with other clinicopathological variables or with the pathological stage of gastric carcinoma. Levels of OPRT were significantly higher in gastric carcinoma tissue than in normal gastric mucosa. OPRT levels were not correlated with levels of either thymidylate synthase or dihydropyrimidine dehydrogenase. In samples of gastric carcinoma tissues and normal gastric mucosa tissues obtained simultaneously from 24 patients, no correlation was found between OPRT levels in gastric carcinoma and levels in normal gastric mucosa. Conclusion These results suggest that the OPRT level is significantly higher in gastric carcinoma tissue than in normal gastric mucosa and that the OPRT level in gastric carcinoma is a novel variable that is independent of the levels of other previously known enzymes related to 5-fluorouracil (FU) metabolism.     

谷胱甘肽S-转移酶-π在胃癌中的表达及其意义

目的 了解GST-π在胃癌中的表达及其与胃癌生物学行为的关系。方法 应用免疫组化法检测85例行根治性切除的胃癌组织中GST-π的表达情况。结果 GST-π在胃癌组织中的阳性表达率为71.8％。其阳性表达与肿瘤的浸润深度及淋巴结转移个数有关,与大体类型及生长方式无关,与病理组织分型无关。在临床分期中,Ⅰ、Ⅱ、Ⅲ期与Ⅳ期比较差异有显著性。结论 通过监测GST-π在胃癌组织中的表达可以起到预测预后的作用,同时对了解耐药程度、选择合理的化疗药物、实行个体化治疗及进行耐药逆转治疗具有十分重要的意义。     

Placental form of glutathione S-transferase in normal and diseased human uterine cervical mucosa

Expression of the human placental form of glutathione S-transferase (GST-pi), an enzyme proposed as a marker for human and experimental neoplasia, was immunohistochemically evaluated in 51 samples of 'normal' and diseased adult human uterine cervix. Five fetal uteri were also studied. GST-pi positivity was detected in 54, 92, 95 and 83% of the 'normal', non-neoplastic, cervical intra-epithelial neoplasia (CIN) and cancer cases respectively. All five fetal uteri and the positive 'normal' adult cases presented cells immunostained for GST-pi throughout the thickness of the mucosa, including the basal layer. Some non-neoplastic conditions like inflammation, repair and metaplasia and some dysplastic and neoplastic lesions showed areas of positively stained cells within an otherwise negative tissue, indicating a phenotypic heterogeneity regarding the enzyme expression. Our results confirm that GST-pi has a fetal character and indicate that it may appear in the adult cervical squamous epithelia under 'normal' or pathological conditions not necessarily linked to the process of carcinogenesis. Therefore it cannot be used as a marker for cervical epithelial neoplasia.     

O6-alkyltransferase activity in normal human gastric mucosa

The spectrum of activity of the DNA repair enzyme O6-alkyltransferase has been studied in a large series of normal stomachs in order to establish the baseline range of values for this enzyme. Sixty-eight patients with histologically normal stomachs were biopsied during the course of upper gastrointestinal endoscopy and the biopsies assayed for O6-alkyl-transferase activity. A wide spectrum of activity was found with values ranging from 38 fmol O6-guanine extracted/mg protein to over 400 fmol/mg. This suggests that there may be wide inter-individual differences in susceptibility to alkylating actions in the human gastric mucosa.     

CARCINOGENESIS: Expression of cytochrome P450s and glutathione S-transferases in human esophagus with squamous-cell carcinomas

In order to clarify the expression of cytochrome P450 and glutathioneS-transferase in human esophagus, 41 samples of human esophaguswith squamous-cell carcinoma were investigated by immunoblotanalysis and enzyme assays. Cytochrome P450 1A2/1 was clearlyexpressed in micro-somes, and the amount in samples with tumoroustissue was significantly greater than that in samples withouttumorous tissues or in liver; cytochrome P450 2B6 and 3A4/3were expressed polymorphically. Aryl hydrocarbon hydroxylaseactivity was detected in microsomes and was greater in samplesfrom smokers than non-smokers. Patients who both smoked anddrank alcohol, however, had activity similar to that of patientswithout these habits. Glutathione S-transferase Ml and A1/2protein existed polymorphically in cytosol, and glutathioneS-transferase Pl-1 was detected in all samples.The frequencyof expression of the glutathione S-transferase A1/2 proteinwas greater in patients with Ml protein than in those without;no difference in the expression was seen for glutathione S-transferasePl-1. Neither smoking nor drinking influenced the expressionor activity of glutathione S-transferase. Our data support theidea that some carcinogens can be directly activated or inactivatedin human esophageal epithelium.     

Glutathione S-transferases and glutathione in human head and neck cancer

Glutathione S-transferase (GST) enzyme activity, GST isoenzymecomposition and glutathione (GSH) concentration were assessedin normal and squamous cell carcinoma specimens of 14 patientswith oral or oropharyngeal cancer and 11 patients with laryngealcancer. Comparing malignant with normal oral/oropharyngeal tissues,no significant differences in GSH content, GST enzyme activityor isoenzyme composition were found. However, some tumours hadup to 3-fold increased GST enzyme activities and 11 malignantsamples over-expressed GST-     

Gene Expression Profile Differences in Gastric Cancer and Normal Gastric Mucosa by Oligonucleotide Microarrays

OBJECTIVE To study the difference of gene expression in gastric cancer (T) and normal tissue of gastric mucosa (C), and to screen for associated novel genes in gastric cancers by oligonucleotide microarrays. METHODS U133A (Affymetrix, Santa Clara, CA) gene chip was used to detect the gene expression profile difference in T and C. Bioinformatics was used to analyze the detected results. RESULTS When gastric cancers were compared with normal gastric mucosa, a total of 270 genes were found with a difference of more than 9 times in expression levels. Of the 270 genes, 157 were up-regulated (Signal Log Ratio [SLR] ≥3), and 113 were down-regulated (SLR ≤-3). Using a classification of function, the highest number of gene expression differences related to enzymes and their regulatory genes (67, 24.8%), followed by signal-transduction genes (43,15.9%). The third were nucleic acid binding genes (17, 6.3%), fourth were transporter genes (15, 5.5%) and fifth were protein binding genes (12, 4.4%). In addition there were 50 genes of unknown function, accounting for 18.5%. The five above mentioned groups made up 56.9% of the total gene number. CONCLUSION The 5 gene groups (enzymes and their regulatory proteins, signal transduction proteins, nucleic acid binding proteins, transporter and protein binding) were abnormally expressed and are important genes for further study in gastric cancers.     

Distribution of beta-human chorionic gonadotropin-positive cells in noncancerous gastric mucosa and in malignant gastric tumors.

The authors examined the localization and behavior of beta-human chorionic gonadotropin (HCG)-positive cells in human gastric noncancerous mucosa and in gastric malignant tumors, using immunohistochemistry and the anti-beta-HCG antibody. The beta-HCG-positive cells were located mainly in the antral mucosa and were generally restricted to the neck portion of the pyloric glands, although a few were present in fundic glands of the gastric body. The beta-HCG-immunoreactive cells were found in gastric carcinomas in 53% of the 92 cases examined. These cells were observed more often in advanced carcinomas that were histologically poorly differentiated than in early carcinomas or in well-differentiated tumors, but this prevalence had no statistical significance. The presence of the beta-HCG-positive cells in the gastric carcinomas suggested no appreciable prognostic significance, even quantitatively. In the syncytiotrophoblast-like tumor cells seen in four gastric tumor samples with histologic features of a choriocarcinoma, immunoreactivity to the beta-HCG was striking. There was, however, no recognizable dominance in the number of beta-HCG-reactive cells in the noncancerous mucosa around the tumor.