Eradication of Helicobacter pylori restores glutathione S-transferase activity and glutathione levels in antral mucosa.

Glutathione S-transferases (GST) and glutathione peroxidases (GPO) are important in detoxification. GST activity in the mucosa of the gastrointestinal tract is inversely correlated with the development of gastrointestinal cancer. Helicobacter pylori (H. pylori) infection has been associated with gastric cancer. We studied GST activity and the substrate glutathione (GSH) in patients with H. pylori-associated gastritis. GST activity and isoenzyme levels, GPO activity and GSH levels were studied in antral biopsies of 38 H. pylori-positive patients, before and after eradication treatment. In 31 patients in whom H. pylori was successfully eradicated, antral GST enzyme activity before therapy was 532 (465 - 598) nmol / mg protein. min (mean and 95% confidence interval) and that after therapy was 759 (682 - 836) nmol / mg protein. min (P < 0.0001). Correspondingly, levels of GST alpha and GST-P1 were higher after eradication (P < 0.001). GSH concentration significantly increased: 21.2 (16.2 - 26.2) nmol / mg protein before and 27.1 (23.6 - 30.6) nmol / mg protein after therapy (P < 0.05). In 7 patients in whom H. pylori was not eradicated, GST activity was 671 (520 - 823) nmol / mg protein. min and 599 (348 - 850) nmol / mg protein before and after treatment respectively (P = 0.32). GSH levels were 17.4 (9.0 - 25.7) nmol / mg protein and 18.2 (9.1 - 27.3) nmol / mg protein, respectively (P = 0.84). No differences in antral GPO enzyme activity, both of selenium (Se)-dependent and total GPO, before and after successful treatment were found. Eradication of H. pylori infection increases GST activity and GSH levels in antral mucosa. Low GST activity and GSH concentration due to H. pylori infection might play a role in gastric carcinogenesis.     

Glutathione S-Transferases in Small Intestinal Mucosa of Patients with Coeliac Disease

Patients with villous atrophy due to coeliac disease have an increased risk of developing small intestinal malignancies. Intestinal glutathione (GSH) and glutathione S‐transferases (GST) are involved in the protection against carcinogenesis. The aim of this study was to evaluate GSH content and GST enzyme activity in small intestinal mucosa of untreated coeliacs compared to controls. We evaluated GSH content and GST enzyme activity, including the levels of GST classes α, μ, π, θ in small intestinal biopsies of untreated coeliacs (flat mucosa, Marsh IHC, n=12) compared to normal subjects (n=23). Next, we evaluated GSH and GST's in coeliacs in remission (Marsh 0‐1, n=11), coeliacs with persisting villous atrophy while on a gluten‐free diet (partial villous atrophy, Marsh IIIA (n=5); subtotal villous atrophy, Marsh IIIB (n=6) and patients with infiltrative/crypt‐hyperplastic Marsh II lesions (n=4). Total GST enzyme activity and content of GSTa are markedly suppressed in Marsh IIIC lesions compared to controls (resp. 220±79 vs. 4641189 nmol/mg protein‐min (P<0.001) and 2.79±2.46 vs. 6.47±2.29 μg/mg protein (P<0.001). In coeliacs in remission these levels normalized. Total GST enzyme activity and GSTα levels are proportionately lowered according to the degree of mucosal pathology in Marsh II, IIIA and IIIB. (Spearman's σ correlation coefficient for total GST, ‐0.596, P<0.001; GSTα, ‐0.620, P<0.001). GSTμ, π and θ and GSH levels are not significantly different in the selected study groups of mucosal pathology compared to controls. Total GST enzyme activity and content of GSTα in small intestinal mucosa are significantly lower in untreated coeliac disease compared to controls. In Marsh II, IIIA and IIIB, GST enzyme activity and GSTα content are proportionally lower according to the degree of mucosal pathology. Normal values are seen in coeliacs in remission. This correlation between coeliac disease and a suppressed GSH/GST detoxification system may explain in part the carcinogenic risk in untreated coeliac disease.     

Low Glutathione and Glutathione S-Transferase Levels in Barrett's Esophagus as Compared to Normal Esophageal Epithelium

Patients with Barrett's esophagus, wherein squamous epithelium has been replaced by columnar epithelium, have an increased risk for developing esophageal adenocarcinoma as compared to the general population. Glutathione S-transferase (GST), a family of detoxification enzymes consisting of class α, μ, π, and θ isoforms, is involved in detoxification of carcinogens and low levels of these enzymes correlated with high cancer risk. We have now compared GST enzyme activity, GST isoenzyme composition and glutathione (GSH) content of Barrett's mucosa with that of adjacent normal squamous epithelium. Biopsy specimens of 98 patients with Barrett's esophagus were taken from both Barrett's and adjacent normal squamous epithelium. GST enzyme activity towards 1–chloro-2,4–dinitrobenzene was measured, and GST isoenzyme levels were determined by densitometrical analyses of western blots after immunodetection with monoclonal antibodies. Total GSH content was determined by high-performance liquid chromatography after conjugation with monobromobimane. Wilcoxon's signed rank test and Spearman correlation analyses were used for statistical evaluation. As compared with adjacent normal squamous epithelium, GST enzyme activity in Barrett's epithelium was reduced by 35%, and GST μ, GST π and GSH levels were reduced by 24%, 30%, and 63%, respectively. However, the minor GST α and GST θ levels were higher in Barrett's epithelium (by 625% and 33%, respectively). High levels of GSH and GSTs in general are correlated with protection against cellular or cytogenetic damage. The observed reduction in GSTs and GSH in Barrett's epithelium may therefore contribute to the increased cancer risk in this tissue.     

Glutathione S-transferases in small intestinal mucosa of patients with coeliac disease.

Patients with villous atrophy due to coeliac disease have an increased risk of developing small intestinal malignancies. Intestinal glutathione (GSH) and glutathione S-transferases (GST) are involved in the protection against carcinogenesis. The aim of this study was to evaluate GSH content and GST enzyme activity in small intestinal mucosa of untreated coeliacs compared to controls. We evaluated GSH content and GST enzyme activity, including the levels of GST classes alpha, mu, pi and theta, in small intestinal biopsies of untreated coeliacs (flat mucosa, Marsh IIIC, n = 12) compared to normal subjects (n = 23). Next, we evaluated GSH and GST's in coeliacs in remission (Marsh 0 - I, n = 11), coeliacs with persisting villous atrophy while on a gluten-free diet (partial villous atrophy, Marsh IIIA (n = 5); subtotal villous atrophy, Marsh IIIB (n = 6)) and patients with infiltrative / crypt-hyperplastic Marsh II lesions (n = 4). Total GST enzyme activity and content of GSTalpha are markedly suppressed in Marsh IIIC lesions compared to controls (resp. 220 +/- 79 vs. 464 +/- 189 nmol / mg protein*min (P < 0.001) and 2.79 +/- 2.46 vs. 6.47 +/- 2.29 mg / mg protein (P < 0.001)). In coeliacs in remission these levels normalized. Total GST enzyme activity and GSTalpha levels are proportionately lowered according to the degree of mucosal pathology in Marsh II, IIIA and IIIB. (Spearman's sigma correlation coefficient for total GST, -0.596, P < 0.001; GSTalpha, -0.620, P < 0.001). GSTmu, pi and theta and GSH levels are not significantly different in the selected study groups of mucosal pathology compared to controls. Total GST enzyme activity and content of GSTalpha in small intestinal mucosa are significantly lower in untreated coeliac disease compared to controls. In Marsh II, IIIA and IIIB, GST enzyme activity and GSTalpha content are proportionally lower according to the degree of mucosal pathology. Normal values are seen in coeliacs in remission. This correlation between coeliac disease and a suppressed GSH / GST detoxification system may explain in part the carcinogenic risk in untreated coeliac disease.     

Levels of malondialdehyde-deoxyguanosine in the gastric mucosa: relationship with lipid peroxidation, ascorbic acid, and Helicobacter pylori.

Helicobacter pylori infection is associated with elevated gastric mucosal concentrations of the lipid peroxidation product malondialdehyde and reduced gastric juice vitamin C concentrations. Malondialdehyde can react with DNA bases to form the mutagenic adduct malondialdehyde-deoxyguanosine (M(1)-dG). We aimed to determine gastric mucosal levels of M(1)-dG in relation to H. pylori infection and malondialdehyde and vitamin C concentrations. Patients (n = 124) attending for endoscopy were studied. Levels of antral mucosal M(1)-dG were determined using a sensitive immunoslot-blot technique; antral mucosal malondialdehyde was determined by thiobarbituric acid extraction, and gastric juice and antral mucosal ascorbic acid and total vitamin C were determined by high-performance liquid chromatography. Sixty-four H. pylori-positive patients received eradication therapy, and endoscopy was repeated at 6 and 12 months. Levels of M(1)-dG did not differ between subjects with H. pylori gastritis (n = 85) and those with normal mucosa without H. pylori infection (n = 39; 56.6 versus 60.1 adducts/10(8) bases) and were unaffected by age or smoking habits. Malondialdehyde levels were higher (123.7 versus 82.5 pmol/g; P < 0.001), gastric juice ascorbic acid was lower (5.7 versus 15.0 micromol/ml; P < 0.001), and antral mucosal ascorbic acid was unchanged (48.0 versus 42.7 micromol/g) in H. pylori gastritis compared with normal mucosa. Multiple regression analysis revealed that M(1)-dG increased significantly with increasing levels of malondialdehyde, antral ascorbic acid, and total antral vitamin C. M(1)-dG levels were unchanged 6 months (63.3 versus 87.0 adducts/10(8) bases; P = 0.24; n = 38) and 12 months (66.7 versus 77.5 adducts/10(8) bases; P = 0.8; n = 13) after successful eradication of H. pylori. M(1)-dG thus is detectable in gastric mucosa, but is not affected directly by H. pylori.     

Habitual consumption of fruits and vegetables: associations with human rectal glutathione S-transferase

The glutathione (GSH)/glutathione S-transferase (GST) system is an important detoxification system in the gastrointestinal tract. A high activity of this system may benefit cancer prevention. The aim of the study was to assess whether habitual consumption of fruits and vegetables, especially citrus fruits and brassica and allium vegetables, is positively associated with parameters reflecting the activity of the GSH/GST enzyme system in human rectal mucosa. GST enzyme activity, GST isoenzyme levels of GST-alpha (A1-1, A1-2 and A2-2), -mu (M1-1) and -pi (P1-1), and GSH levels were measured in rectal biopsies from 94 subjects. Diet, lifestyle, GSTM1 and GSTT1 null polymorphisms were assessed. Mean GST enzyme activity was 237 nmol/min/mg protein (SD = 79). Consumption of citrus fruits was positively associated with GST enzyme activity [difference between high and low consumption: 28.9 (95% confidence interval (CI) = 9.3-48.6) nmol/min/mg protein], but was not associated with the other parameters. A positive association with brassica vegetables was found among carriers of the GSTM1-plus genotype [difference between high and low consumption: 22.6 (95% CI = 0.2-45.0) nmol/min/mg protein], but not among GSTM1-null individuals (-25.8 nmol/min/mg protein, 95% CI = -63.3-11.8). This is in line with a positive association between consumption of brassica vegetables and GSTM isoenzyme level [difference between high and low consumption: 67.5%, 95% CI = (6.8-162.7)]. Consumption of allium vegetables was not associated with GST enzyme activity, but negatively with GSTP1-1 levels [difference between high and low consumption: -23.3%, 95% CI = (-35.5; -8.6)]. Associations were similar among those with the GSTT1-plus and GSTT1-null genotype. In conclusion, variations in habitual consumption of fruits, particularly citrus fruits, and of vegetables, in particular brassica vegetables, among those with the GSTM1-plus genotype, may contribute to variations in human rectal GST enzyme activity.     

Potential Chemoprevention Activity of Pterostilbene by Enhancing the Detoxifying Enzymes in the HT-29 Cell Line

Detoxifying enzymes are present in most epithelial cells of the human gastrointestinal tract where they protectagainst xenobiotics which may cause cancer. Induction of examples such as glutathione S-transferase (GST)and its thiol conjugate, glutathione (GSH) as well as NAD(P)H: quinoneoxidoreductase (NQO1) facilitate theexcretion of carcinogens and thus preventing colon carcinogenesis. Pterostilbene, an analogue of resveratrol, hasdemonstrated numerous pharmacological activities linked with chemoprevention. This study was conducted toinvestigate the potential of pterostilbene as a chemopreventive agent using the HT-29 colon cancer cell line tostudy the modulation of GST and NQO1 activities as well as the GSH level. Initially, our group, established theoptimum dose of 24 hours pterostilbene treatment using MTT assays. Then, effects of pterostilbene (0-50 μM)on GST and NQO1 activity and GSH levels were determined using GST, NQO1 and Ellman assays, respectively.MTT assay of pterostilbene (0-100 μM) showed no cytotoxicity toward the HT-29 cell line. Treatment increasedGST activity in the cell line significantly (p<0.05) at 12.5 and 25.0 μM. In addition, treatment at 50 μM increasedthe GSH level significantly (p<0.05). Pterostilbene also enhanced NQO1 activity significantly (p<0.05) at 12.5μM and 50 μM. Hence, pterostilbene is a potential chemopreventive agent capable of modulation of detoxifiyingenzyme levels in HT-29 cells.     

Glutathione content and glutathione-S-transferase expression in 1,3-bis(2-chloroethyl)-1-nitrosourea-resistant human malignant astrocytoma cell lines

gamma-L-glutamyl-L-cysteinylglycine (GSH) has been shown to inactivate 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) and quench DNA crosslink precursors of BCNU. Because of the central role of 2-chloroethyl-nitrosoureas in brain tumor chemotherapy, we investigated the intracellular GSH content and the expression of specific glutathione-S-transferases (GSTs) in three human malignant astrocytoma cell lines (UWR1, UWR2, and UWR3) of varying BCNU resistance to determine the interrelationship of these parameters with brain tumor BCNU resistance. GSH was assayed by ion-exchange high performance liquid chromatography after derivatization with 1-fluoro-2,4 dinitrobenzene. Both bulk and specific GST (acid, near-neutral, and basic) activities were examined using substrates that show high specificities to the different GSTs. Western blot analyses with antisera against GST-alpha, -mu, and -tau subunits were also performed on partially purified GST from the cells of each cell line. The results showed GSH content of 91, 46.5, and 28.3 nmol GSH/mg protein for UWR1, UWR2, and UWR3, respectively. Bulk GST activity (with 1-chloro-2,4-dinitrobenzene as substrate) also correlated with increasing BCNU resistance. Of the three GST classes examined by both substrate specificities and Western blotting, only the expression of the acidic form, GST-tau, correlated significantly with the rank order of BCNU resistance of the cell lines. GST-mu and -alpha were present in only trace amounts in all three cell lines.     

Glutathione S-transferase phenotypes in relation to genetic variation and fruit and vegetable consumption in an endoscopy-based population

High glutathione S-transferase (GST) activity may contribute to colorectal cancer prevention. Functional polymorphisms are known in the GSTM1, GSTT1, GSTA1 and GSTP1 genes. The influence of these GST polymorphisms and recent fruit and vegetable consumption on GST levels and activity has not been investigated simultaneously in a human population. Also, it is not clear if blood GST activity reflects rectal GST activity. Therefore, we determined GST polymorphisms in 94 patients scheduled for sigmoidoscopy. Rectal GST isoenzyme levels (GSTM1, GSTM2, GSTT1, GSTA and GSTP1) were measured by quantitative western blotting, and rectal and white blood cell total GST activities were measured spectrophotometrically using 1-chloro-2,4-dinitrobenzene (CDNB) as a substrate. Vegetable and fruit consumption was assessed by dietary record. As expected, the GSTM1 and GSTT1 deletion polymorphisms, and the GSTA1 g.-69C-->T polymorphism significantly affected the respective isoenzyme levels. Also, rectal GST isoenzyme levels differed between those with and without recent consumption of Alliaceae, Cucurbitaceae, Apiaceae and citrus fruit. Rectal GST activity, however, was not clearly influenced by fruit and vegetable consumption. It was most significantly determined by the GSTP1 c.313A-->G polymorphism; compared with the 313AA genotypes, the 313AG and 313GG genotypes showed 36 and 67 nmol/min/mg protein (P < 0.001) lower GST activity, respectively. The correlation between rectal and white blood cell GST activities was low (r = 0.40, P < 0.001), and the relevance of the various genetic and dietary factors appeared to differ between the two tissues. In conclusion, this study indicates that the GST enzyme system is influenced by both GST polymorphisms and consumption of fruits and vegetables. The latter appeared more important for individual rectal GST isoenzyme levels than for total GST activity, which could affect detoxification of isoenzyme-specific substrates. The study results do no support the use of white blood cell GST activity as a surrogate measure for rectal GST activity.     

Glutathione depletion causes cell growth inhibition and enhanced apoptosis in pancreatic cancer cells

BACKGROUND: Recent studies have demonstrated that various tumors express enhanced levels of the radical scavenger glutathione (GSH). Moreover, there are grounds for claiming that GSH plays a crucial role in cell proliferation and tumor resistance. In the current study, we investigated the relation between cell growth and GSH levels in the pancreatic adenocarcinoma cell line, AsPC-1, and the significance of GSH in tumor resistance to chemotherapy. METHODS: Cell growth in AsPC-1 was initiated through transforming growth factor-alpha (TGF-alpha) or fetal calf serum (FCS). Then, cell cycle, cell proliferation, and cellular GSH content were analyzed at different times in the presence or absence of buthionine sulfoximine (BSO). The impact of GSH on chemotherapy-induced apoptosis was studied using 5-fluorouracil or melphalan in the presence or absence of BSO. Finally, we compared the GSH content of 15 pancreatic tumor specimens with 10 normal pancreatic tissue specimens. RESULTS: Analysis of GSH in pancreatic tissues demonstrated increased GSH levels in cancerous compared with normal tissue (17.5 +/- 2.3 vs. 8. 8 +/- 1.4 nmol/mg protein; P < 0.004). Incubation of AsPC-1 with TGF-alpha or FCS resulted in cell proliferation and cell cycle activity, whereas GSH content was not altered. Incubation of GSH-depleted cells with TGF-alpha did not stimulate cell growth. In addition, GSH-depletion resulted in an increased rate of apoptosis after melphalan (6.3 +/- 0.3 % vs. 11.2 +/- 0.3 %; P < 0.001), but not after 5-fluorouracil treatment. CONCLUSIONS: Taken together, our results show enhanced GSH levels in pancreatic carcinoma and an essential role of GSH in cell proliferation and in resistance of AsPC-1 cells. Therefore, GSH-depletion may improve the efficacy of adjuvant therapy in pancreatic carcinoma.     

Determinants of Drug Response in a Cisplatin-resistant Human Lung Cancer Cell Line

To elucidate the mechanism(s) of cisplatin resistance, we have characterized a human non-small cell lung cancer cell line (PC-9/CDDP) selected from the wild type (PC-9) for acquired resistance to cisplatin. PC-9/CDDP demonstrated 28-fold resistance to cisplatin, with cross resistance to other chemotherapeutic drugs including chlorambucil (× 6.3), melphalan (× 3.7) and 3-[(4-amino-2-methyl-5-pyrimidinyl)]methyl-1-(2-chloroethyl)-1-nitrosourea (ACNU) (× 3.9). There was no expression of mdr-1 mRNA in either wild-type or resistant cells. The mRNA and protein levels of glutathione S -transferase (GST) × were similar in the two lines. A GST-μ isozyme was present in equal amounts and the activities of selenium-dependent and independent glutathione peroxidase and glutathione reductase were unchanged. The mRNA level of human metallothionein IIA and the total intracellular metallothionein levels were reduced in the resistant cells. Significantly increased intracellular glutathione (GSH) levels were found in the resistant cells (20.0 vs. 63.5 nmol/mg protein) and manipulation of these levels with buthionine sulfoximine produced a partial sensitization to either cisplatin or chlorambucil. Increased GSH probably also played a role in determining cadmium chloride resistance of the PC-9/CDDP, even though this cell line had a reduced metallothionein level. Also contributing to the cisplatin resistance phenotype was a reduced intracellular level of platinum in the PC-9/CDDP. Thus, at least two distinct mechanisms have been selected in the resistant cells which confer the phenotype and allow degrees of cross resistance to other electrophilic drugs.     

Glutathione S-transferases in normal and cancerous human colon tissue

In order to evaluate the role of the placental form of glutathione S-transferase (GST-Pi) as a tumour marker, activity andcomposition of GSTs from human colon were investi gated. GSTswere purified from normal colon mucosa and from colomc tumoursby affinity chromatography on gluta thione-agarose. After SDS-PAGEor isoelectric focusing these purified preparations revealedonly one band that comigrated with GST-Pi from human placenta.A monoclonal antibody (mAb) very specific for GST-Pi was developedand characterized. On inimunoblot this mAb stains purified GSTfrom normal and diseased colon tissue. GST activity was significantlyhigher in most cancerous (247 ? 38 nmoll mm/mg protein; n =7), compared with the corresponding normal tissues (171 ? 18nmol/min/mg protein; n = 7). In colon from patients withoutlarge bowel malignancies GST Pi is also by far the most prominentisoform detectable. In conclusion, both normal and tumorouscolon tissue predominantly express GST-Pi and therefore GST-Piis not suitable as a tumour marker for colonic carcinomas. However,the increased GST-Pi levels in colonic tumours could possiblycontribute to the relatively high resistance to anti-cancerdrugs.     

鼻咽癌患者外周血血浆可的松水平和全血还原型谷胱甘肽含量的昼夜节律

背景与目的:谷胱甘肽与细胞对抗癌药物的解毒作用以及对放射性损伤的保护机制密切相关。本文通过对鼻咽癌患者和健康志愿者外周血血浆可的松水平和全血还原型谷胱甘肽含量的昼夜节律进行研究,从而为鼻咽癌患者进行肿瘤的时间治疗提供参考资料。方法:13名鼻咽癌患者(实验组)和14名健康志愿者(正常对照组)参加本项研究。每位受试者均从中午12点开始抽取外周静脉血4.5mL,每隔4h抽血一次,在24h内共抽血6次。血浆可的松水平的检测采用放射免疫法,全血还原型谷胱甘肽含量则采用高效液相色谱法进行检测。结果:鼻咽癌患者组和正常对照组的血浆可的松水平均呈现出明显的昼夜节律的特征,且节律特性相似。两组的血浆可的松水平的峰值均出现在早晨,而谷值均出现在午夜。不同时间点时,两组的全血还原型谷胱甘肽含量不同,且均具有显著性差异(重复测量的方差分析,F=5.18,P=0.02)。余弦分析表明,鼻咽癌患者组呈现出昼夜节律变化趋势(P=0.06),峰值出现在早晨(05∶02),正常对照组也呈现出明显的昼夜节律的特征,峰值出现在早晨(07∶44±01∶56)(P<0.01)。鼻咽癌患者组外周全血中还原型谷胱甘肽的节律调整均值为(19.60±1.11)nmol/mgprotein,正常对照组为(8.95±0.46)nmol/mgprotein。结论:包括晚期患者在内的鼻咽癌患者仍然具有正常的生物节律。鼻咽癌患者全血中还原型谷胱甘肽水平呈现出昼夜节律变化的趋势,与正常对照组节律特性相似。这为临床上选择恰当的时间对患者进行化疗和放疗提供了有价值的参考资料。     

Glutathione S-transferase isoenzymes in human lung tumors

In the present studies we have compared the levels of glutathione(GSH) and GSH-related enzymes in lung tumors and correspondingnormal tissues obtained from the same individuals. We have alsoimmunologically quantitated the relative amounts of glutathioneS-transferase (or GST-P) type antigen in tumors and adjacentnormal tissues from five patients. GST activities towards 1-chloro-2,4-dinitrobenzene (CDNB) and ethacrynic acid were found to beelevated in tumors from two out of five patients (patients #1and 4), whereas the activity towards these substrates was markedlysuppressed in the tumor tissue from one of the patients (#5).Immunotitration and Western blot studies using antibodies raisedagainst -type GST isoenzymes of human lung and placenta indicatedinduction of GST -type isoenzyme in tumors from patients #1and 4 and suppression of this isoenzyme in tumor from patient#5. The tumors from patients #2 and 3 did not show any increasein GST activity or GST -type antigen. Except for the tumor frompatient #5, the GSH content was higher in the tumors from otherpatients. GSH reductase activity was found to be elevated intumors of all the patients examined in this study. These resultsindicate that GSH and GSH related enzymes are differentiallyaltered in lung tumors and GSH levels and GST - or GST-P-typeisoenzyme(s) are not uniformly elevated in all tumors.     

Helicobacter pylori antibody responses in association with eradication outcome and recurrence: a population-based intervention trial with 7.3-year follow-up in China

Objective:To identify serum biomarkers that may predict the short or long term outcomes of anti-Helicobacter pylori (H.pylori) treatment,a follow-up study was performed based on an intervention trial in Linqu County,China.Methods:A total of 529 subjects were selected randomly from 1,803 participants to evaluate total anti-H.pylori immunoglobulin G (IgG) and 10 specific antibody levels before and after treatment at 1-,2-and 7.3-year.The outcomes of anti-H.pylori treatment were also parallelly assessed by 13C-urea breath test at 45-d after treatment and 7.3-year at the end of follow-up.Results:We found the medians of anti-H.pylori IgG titers were consistently below cut-off value through 7.3 years in eradicated group,however,the medians declined in recurrence group to 1.2 at 1-year after treatment and slightly increased to 2.0 at 7.3-year.While the medians were significantly higher (＞3.0 at 2-and 7.3-year) among subjects who failed the eradication or received placebo.For specific antibody responses,baseline seropositivities of FliD and HpaA were reversely associated with eradication failure [for FliD,odds ratio (OR)=0.44,95％ confidence interval (95％ CI):0.27-0.73;for HpaA,OR=0.32,95％ CI:0.17-0.60].The subjects with multiple positive specific antibodies at baseline were more likely to be successfully eradicated in a linear fashion (Ptrend=0.006).Conclusions:Our study suggested that total anti-H.pylori IgG level may serve as a potential monitor of longterm impact on and-H.pylori treatment,and priority for H.pylori treatment may be endowed to the subjects with multiple seroposidve antibodies at baseline,especially for FliD and HapA.     

Lentinan Enhances Sensitivity of Mouse Colon 26 Tumor to cis-Diamminedichloroplatinum(II) and Decreases Glutathione Transferase Expression

We investigated the influence of a combination of lentinan, a biological response modifier, and cis-diamminedichloroplatinum(II) (CDDP) on the growth and glutathione S-transferase (GST) content of colon 26 tumor to examine whether lentinan represses GST expression and enhances the therapeutic effects of CDDP. Female CDF₁ mice inoculated subcutaneously with transplantable colon 26 adenocarcinoma cells (1X10⁶/mouse) received intraperitoneal administrations of lentinan, CDDP, or the two drugs in combination, on days 10, 14, 17 and 21 after the inoculation. On day 24, tumor weights (estimated from their length and width) were significantly lower in the CDDP+lentinan group (2.7±1.3g) than in the CDDP alone group (4.3±0.7g, p<0.05), both values being less than in the nontreated control group (7.2±1.5g). The major GST form of colon 26 tumor was identified as GST-II, the Pi class form, and a minor form as GST-III belonging to the Mu class. Both GST-II and GST-III values on day 24 were significantly decreased in the lentinan alone (0.90±0.29 and 0.26±0.11 μg/mg protein, respectively) and lentinan+CDDP groups (0.98±0.22 and 0.29±0.07 μg/mg protein), as compared with the control levels (1.39±0.20 and 0.52±0.11 μg/mg protein). However, these values were not different between the CDDP alone and lentinan+CDDP groups. Neither tissue interleukin (IL)-6, glutathione nor platinum values were different between the two groups. IL-6 values were elevated in about half of the samples treated with lentinan or CDDP and exhibited a modest inverse correlation with GST-II levels (r= - 0.46). A GST inhibitor, ethacrynic acid, enhanced the sensitivity of cultured colon 26 cells to CDDP, suggesting the possible involvement of GST in modulating the cytotoxicity of CDDP to this cell line. These results indicated that lentinan administration decreases tissue GST-II and GST-III contents and enhances the sensitivity of colon 26 tumor to CDDP.     

Glutathione S-transferase-catalyzed conjugation of bioactivated aflatoxin B(1) in human lung: differential cellular distribution and lack of significance of the GSTM1 genetic polymorphism.

Epidemiological studies suggest that aflatoxin B(1) (AFB(1)), a mycotoxin produced by certain Aspergillus species, may play a role in human respiratory cancers in occupationally-exposed individuals. AFB(1) requires bioactivation to the corresponding exo-8,9-epoxide for carcinogenicity, and glutathione S-transferase (GST)-catalyzed conjugation of the epoxide with glutathione (GSH) is a critical determinant of susceptibility to AFB(1). Of the purified human GST enzymes studied, the polymorphic hGSTM1-1 has the highest activity towards AFB(1) exo-epoxide. The influence of the GSTM1 polymorphism on AFB(1)-GSH formation, as well as the abilities of cytosols from preparations enriched in different isolated lung cell types to conjugate AFB(1)-epoxides, were examined. In whole-lung cytosols from patients undergoing clinically indicated lobectomy, GSTM1 genotype correlated with GSTM1 phenotype as determined by [(3)H]trans-stilbene oxide conjugation: GSTM1-positive = 295 +/- 31 pmol/mg/h (n = 6); GSTM1-negative = 92.8 +/- 23.3 pmol/mg/h (n = 4) (P < 0.05). In contrast, conjugation of microsome-generated [(3)H]AFB(1)-epoxides with GSH was low and variable between patients, and did not correlate with GSTM1 genotype: GSTM1-positive = 11.9 +/- 8.1, 111 +/- 66 and 510 +/- 248 fmol/mg/h (n = 6); GSTM1-negative = 15.3 +/- 16.7, 167 +/- 225 and 540 +/- 618 fmol/mg/h (n = 4) (for 1, 10 and 100 microM [(3)H]AFB(1), respectively). GSH conjugates of AFB(1) exo-epoxide and the much less mutagenic stereoisomer AFB(1) endo-epoxide were produced in a ratio of approximately 1:1 in cytosols from both whole lung and isolated cells. Total cytosolic AFB(1)-epoxide conjugation was significantly higher in fractions enriched in alveolar type II cells (3.07 +/- 1.61 pmol/mg/h) than in unseparated lung cells (0.143 +/- 0.055 pmol/mg/h) or fractions enriched in alveolar macrophages (0. 904 +/- 0.319 pmol/mg/h; n = 4) (P < 0.05). Furthermore, AFB(1)-GSH formation and percentage of alveolar type II cells in different cell fractions were correlated (r = 0.78, P < 0.05). These results demonstrate that human lung GSTs exhibit very low conjugation activity for both AFB(1)-8,9-epoxide stereoisomers, and that this activity is heterogeneously distributed among cell types, with alveolar type II cells exhibiting relatively high activity. Of the GSTs present in human peripheral lung which contribute to AFB(1) exo- and endo-epoxide detoxification, hGSTM1-1 appears to play at most only a minor role.     

甘蓝汁的抗诱变作用及其机理研究

本文通过微核试验和肝组织中谷胱甘肽－Ｓ－转移酶（ＧＳＴ）活性、还原型谷胱甘肽９ＧＳＨ）含量及细胞色素Ｐ４５０含量测定，在哺乳动物整体水平，从遗传学和生化毒理角学角度研究了甘蓝汁的抗诱变作用及其机理。结果发现，甘蓝汁对环磷酰胺（ＣＰ）诱发的小鼠骨髓多染红细胞（ＰＣＥ）微核细胞率有显著抑制作用，抑制率达４７．２％。Ｗｉｓｔａｒ大鼠饮甘蓝汁１０ｄ后，肝脏ＧＳＴ活性增加２５．０％，ＧＳＨ含量提高４７．８％     

Dexamethasone-resistant Human Pre-B Leukemia 697 Cell Line Evolving Elevation of Intracellular Glutathione Level: An Additional Resistance Mechanism

Glucocorticoids remain among the most important drugs in the treatment of acute lymphoblastic leukemia (ALL). Although the mechanisms of glucocorticoid resistance have been studied in some T-cell leukemic cell lines, less work has been done with B-cell lines. We established a dexamethasone (DEX)-resistant human pre-B lineage leukemia cell line (697/DEX) and investigated the mechanism of resistance. 697/DEX was over 430–fold more resistant to DEX compared with the parental cells (697/Neo). Overexpression of Bcl–2 protein was not observed in 697/DEX, different from the mechanism of resistance in Bcl–2–virus-infected cells (697/Bcl–2). Although the expression of p-glycoprotein (Pgp) in 697/DEX was positive, its functional activity was not detected. The numbers of glucocorticoid receptors (GR) in 697/DEX and 697/Bcl–2 were significantly lower than those in 697/Neo. In addition, 697/DEX and 697/Bcl–2 had higher levels of glutathione (GSH) than 697/Neo. In the presence of l -buthionine-( S, R )-sulfoximine (BSO), an inhibitor of GSH synthesis, both 697/DEX and 697/Bcl–2 recovered their sensitivity to DEX. Interestingly, cell death by the depletion of GSH did not involve caspase–3/7 activation in 697/Bcl–2 and 697/DEX, different from 697/Neo, suggesting a death mechanism through caspase-independent programmed cell death or necrosis. In conclusion, DEX-resistance in 697/DEX was related not only to a GR decrease, but also to an increase in intracellular GSH level in the DEX-resistant B-cell leukemia cell line. Circumvention of DEX-resistance with BSO may offer an approach to overcoming resistance to chemotherapy in B-cell lineage ALL.