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B Schutte

Publications and source records attributed to B Schutte.

At least 55 records · Page 3Linked to original sources
PubMed1992

Effects of butylated hydroxyanisole on arachidonic acid and linoleic acid metabolism in relation to gastrointestinal cell proliferation in the rat.

In order to determine the effect of oral administration of 2(3)-tert-butyl-4-hydroxyanisole (BHA; dose-level: 1.5% BHA of the diet) on arachidonic acid (AA) and linoleic acid (LA) metabolism in correlation with changes in gastrointestinal cell kinetics, we coadministered two inhibitors of prostaglandin H synthase, acetylsalicylic acid (ASA) and indomethacin (IM), to rats. Coadministration of ASA (0.2%) and IM (0.002%) in the drinking water, resulted in a significant reduction of the BHA-induced enhancement of cell proliferation in forestomach and glandular stomach. ASA completely counteracted the effect of BHA on labeling indices in colon/rectum whereas IM exhibited no effect in this organ. Both inhibitors had no direct effect on cell kinetics in the control groups. ASA, and to a lesser degree IM, inhibited prostaglandin E2 release in all tissues examined. Whereas ASA did inhibit lipoxygenase-mediated metabolism of AA in forestomach tissue, ASA did not affect the release of AA- and LA-derived hydroxy fatty acids in glandular stomach and colon/rectum. IM did not affect lipoxygenase production. BHA, however, appeared to be a strong inhibitor of both routes of AA metabolism. While ASA nor IM affected LA metabolism, BHA inhibited both prostaglandin H synthase-mediated and lipoxygenase-mediated metabolism of AA and LA. A causal role of AA or LA metabolites in the process of cell proliferation enhancement induced by BHA, can therefore be excluded. Prostaglandin H synthase may, however, be involved in BHA activation by converting the hydroquinone metabolite of BHA to the corresponding quinone by redox cycling, which is probably accompanied by reactive intermediate production.

Animals↗
PubMed1992

The effect of avidin-biotin interactions in detection systems for in situ hybridization.

The effect of avidin-biotin interactions in several detection systems for the non-radioactive in situ hybridization (ISH) technique was studied in a model system using a transitional cell carcinoma line and a biotinylated DNA probe. We performed fluorescence ISH to unravel the individual steps in a sensitive and frequently used amplification method which makes use of the alternating cytochemical detection layers of fluorescein isothiocyanate-conjugated avidin (AvFITC) and biotinylated goat anti-avidin (BioGAA) antibodies to detect the hybridized and biotinylated probe. Our experiments revealed that BioGAA antibodies bind with their antigen binding sites and not with their biotin moieties to avidin molecules that have already interacted with the DNA probe. The probable working mechanism of this amplification method is presented in a model. Furthermore, we used a peroxidase staining technique to compare with each other the sensitivity of several other detection systems in which avidin-biotin interactions play an important role, e.g., the avidin-biotinylated peroxidase complex (ABC) system. The experiments show that avidin molecules can not be efficiently used to interconnect two biotinylated molecular layers, since their introduction leads to firmly closed cytochemical networks. Such a closed network is already formed between the hybridized and biotinylated DNA probe and a first detection layer of avidin molecules, as appears from the finding that biotinylated molecules could hardly be coupled to these avidin molecules in a following detection layer. Therefore, the results presented here provide us with new insight into the molecular basis of cytochemical network formation. This will enable us to choose the proper procedures for increasing the sensitivity of ISH detection systems.

Avidin↗
PubMed1992

Chromosomal localization of transfected genes by a combination of hot banding and fluorescence in situ hybridization.

We describe the combination of hot banding with fluorescence in situ hybridization as a rapid and efficient method to identify integration sites of transfected DNA sequences in chromosomes. As a test system we used SW480 EJ2, a clonal cell line obtained after transfection of SW480 with pSV2neoEJ, a plasmid containing a point-mutated, c-Ha-RAS oncogene. Nick-translated probes were compared with random primed-labeled probes to evaluate their relative efficiency in fluorescence in situ hybridization. The fluorescence signals were quantified in interphase nuclei by confocal scanning laser microscopy. Nick-translated probes were found to yield better results. Hot banding followed by fluorescence in situ hybridization localized the integration site of pSV2neoEJ in SW480 EJ2 at the site of a translocation on a marker chromosome Xp+. The combination of fluorescence in situ hybridization and hot banding can be used to (a) rapidly and efficiently analyze integration sites in large numbers of transfectants, (b) assess the clonality of transfected cell lines, and (c) localize the site of integration of transfected genes in the recipient genome.

Animals↗
PubMed1992

A novel fluorescence detection method for in situ hybridization, based on the alkaline phosphatase-fast red reaction.

We have used naphthol-ASMX-phosphate and Fast Red TR in combination with alkaline phosphatase (APase) to produce fluorescent precipitated reaction products in a non-radioactive in situ hybridization (ISH) method. To obtain optimal and discrete localization of the strongly red fluorescent ISH signals, the enzyme precipitation procedure was optimized. The optimal reaction time and the concentrations of substrate and capture agent were determined. Furthermore, polyvinyl alcohol (PVA) was used to increase the viscosity of the reaction mixture and thus to reduce diffusion of the reaction product. Our results show that the APase-Fast Red detection method has at least the same sensitivity as currently observed in other immunofluorescent detection systems. A single copy DNA sequence of 15.8 KB could be localized with high efficiency in metaphase spreads and in interphase nuclei. Double labeling procedures, in which the FITC- and azo-dye fluorescence are combined, are also feasible. The red fluorescent ISH signals showed hardly any fading as compared with FITC fluorescence on exposure to either light from the mercury-arc lamp or laser light. Therefore, these red fluorescent signals with a virtually permanent character allow a better analysis and three-dimensional localization of such cytochemically detected genomic fractions by means of confocal scanning laser microscopy as compared with the use of FITC, TRITC, or Texas Red as label.

Alkaline Phosphatase↗
PubMed1991

Biological effects of short-term feeding to rats of repeatedly used deep-frying fats in relation to fat mutagen content.

The effects of short-term feeding of mutagen containing, heated deep-frying oils on urinary and faecal mutagenicity, plasma clinical biochemical parameters, peroxidative effects and cell proliferative indices in the gastro-intestinal tract were determined in rats. Repeatedly used frying oils [a saturated fatty acid-rich coconut oil (CO) and a polyunsaturated fatty acid (PUFA)-rich (greater than 60% PUFA) vegetable frying oil (PO)] were administered to groups of seven rats at a level of 10% (by weight) in the diet for 4 wk; control groups were fed equal amounts of the unheated oils. Both heated oils showed direct-acting mutagenicity to Salmonella tester strain TA97; heated PO was also mutagenic to strain TA100. Both heated CO and heated PO contained detectable amounts of thiobarbituric acid-reactive substances (TBA-RS). In heated PO, hydroperoxides of linoleic acid were also present. In groups fed heated oils the mutagenicity of urine and faeces to strain TA97 was not found to be increased in comparison with the control groups. Faecal mutagenicity to strain TA100 was also unaffected by consumption of heated oils. Urinary excretion of TA100 mutagens was significantly increased in rats fed heated PO. Plasma alkaline phosphatase activity was clearly raised in rats fed heated PO, in comparison with rats fed unheated oils or heated CO. In addition, other clinical biochemical plasma parameters showed a tendency to be increased in rats fed heated PO, indicating hepatic and renal cellular toxicity. Urinary and faecal excretion of thiobarbituric acid-reactive substances (TBA-RS) were also slightly, but not significantly, increased in rats fed heated PO. Feeding heated CO to rats did not result in increased plasma enzyme activities and excretion of TBA-RS, nor in increased cell proliferation in gastro-intestinal tissues. Cell proliferation of the oesophageal tissues were slightly, but significantly, increased in rats fed heated PO, in comparison with the group fed unheated PO. Tissues of the glandular stomach and colon/rectum did not show significantly enhanced cell proliferation in the group fed heated PO. The results obtained in this study indicated that consumption of heated oils containing TA100 mutagens and oxidation products of linoleic acid produced indications of cellular damage to liver and kidneys, and increased urine mutagenicity, as well as enhanced cell proliferation in the oesophagus.

Administration, Oral↗
PubMed1991

Modulation by dietary factors of BHA-induced alterations in cell kinetics of gastro-intestinal tract tissues in rats.

To determine the effects of dietary ethanol or fibre on 2(3)-tert-butyl-4-hydroxyanisole (BHA)-induced alterations in cell kinetics in gastro-intestinal tract tissues, groups of six male Wistar rats were fed diets containing 0% (control) or 1.5% BHA for 2 wk. One group fed 1.5% BHA and one pair-fed control group received 10% ethanol in the drinking-water; two similarly fed groups received drinking-water only. Another group fed 1.5% BHA and a pair-fed control group received a diet supplemented with 20% cellulose; two similar groups received no fibre supplementation. Cell kinetics in the forestomach, glandular stomach and oesophageal tissue were determined, after 14 days, by bivariate 5-bromo-deoxyuridine/DNA analysis using immunocytochemistry and flow cytometry. In the fibre experiment, colorectal tissue was also examined. In both experiments the labelling indices in all the gastro-intestinal tract tissues were significantly altered in the BHA-fed groups compared with the corresponding control groups. In the ethanol experiment no statistically significant difference in the labelling indices was observed in the forestomach or glandular stomach between the two control groups or between the two BHA-fed groups. However, intake of ethanol-supplemented drinking-water induced increases in oesophageal labelling indices in rats fed a BHA-free diet. Thus 14 days of simultaneous ethanol administration has no effect on BHA-induced alterations in cell kinetics in the oesophagus, glandular stomach or forestomach of rats. In the forestomach and colorectal tissue, a high-cellulose diet resulted in a significant decrease in the BHA-induced elevation of labelling indices. Thus dietary cellulose provides a partial protection against the proliferation-enhancing effects of BHA in the rat gastro-intestinal tract.

Animals↗
PubMed1990

Colonic mucosal changes in nude mice associated with orthotopic xenografts of human colon cancer cells.

We used the nude mouse tumour xenograft model to study the pathogenesis of mucosa alterations in the large bowel surrounding a carcinoma. In mouse colonic mucosa overlying HT-29 colonic carcinoma xenografts in the caecum, the crypts were elongated in comparison with those in distant mucosa and also frequently showed a shift towards sialomucin production. These features, which are comparable with socalled transitional mucosa (TM) in man, were absent in control animals inoculated with Indian Ink instead of HT-29 cells. Although the localization of the proliferative cell compartment in mouse colonic mucosa overlying HT-29 xenografts appeared to be confined to the lower half of the crypt as in normal mucosa, the relative length of the DNA synthesizing cell compartment along the crypts was slightly elongated. These data strongly suggest that TM should be regarded as a secondary phenomenon rather than a premalignant change in large intestinal epithelium and that higher proliferative activity of epithelial cells contributes little to the elongation of crypts in TM.

Animals↗
PubMed1990

Dose-dependent effects of short-term dietary administration of the food additive butylated hydroxyanisole on cell kinetic parameters in rat gastro-intestinal tract.

Groups of ten 5-week old male Wistar rats were fed a diet containing 0, 0.25, 0.50, 0.75, 1.0 or 2.0% butylated hydroxyanisole (BHA) ad libitum for 2 weeks; another group of rats served as a pair-fed control (PFC) group for the 2% BHA-fed animals. Subsequently, rats were injected i.p. with the thymidine-analogue bromodeoxyuridine (BrdU) which was incorporated into the DNA of cells during DNA synthesis. Cell kinetic parameters in gastro-intestinal tract tissues were determined by means of bivariate BrdU/DNA analysis applying flow-cytometry to randomized tissue samples or by applying immunohistology to randomized tissue sections. In the forestomach, glandular stomach, small intestine and colon/rectum, mean tissue labelling index (LI) was significantly increased in rats on a diet supplemented with 2% BHA, in comparison with rats fed the basal diet (0% BHA) ad libitum or restricted to the mean daily intake of 2% BHA-fed rats (PFC). In the oesophagus of rats fed 2% BHA, the LI was significantly higher in comparison with their PFC group, but not with the group of rats fed 0% BHA ad libitum. In rat forestomach, an apparent no observed effect level for ad libitum fed rats was found at 0.5% BHA (LI) and at 0.75% BHA (potential doubling time). Thus, the oesophagus, glandular stomach, small intestine and large bowel, in addition to the forestomach, are possible target tissues in rats for the proliferation enhancing effects of BHA. At the time of termination of the experiment, plasma BHA concentrations were dose dependently increased and were in the range that is easily attained in man after ingestion of a dose equal to the acceptable daily intake for BHA (0.5 mg/kg).

Animals↗
PubMed1990

Cellular proliferation at the site of organ allografts and the influence of immunosuppressive therapy.

In this study we investigated cellular proliferation of T cells and macrophages at the site of rat cardiac allografts and determined the influence of immunosuppressive therapy on the proliferative characteristics of these cell types. A bromodeoxyuridine-labeling technique was used that allowed both the accurate detection of proliferative activity and the phenotypic characterization of cellular infiltrates within grafted tissues. In untreated recipients (BN----Lewis), T cytotoxic/suppressor cells as well as T helper cells showed proliferative activity at the site of the graft. The percentage of OX8-positive cells within the graft that showed proliferation ranged from 15% to 37%. The percentage of W3/25-positive cells within the graft that showed proliferation ranged from 25% to 30%. In contrast, macrophages hardly showed proliferative activity within the graft; only 1-4% of the macrophages stained positive for bromodeoxyuridine. From these observations it is concluded that the graft serves as a nonlymphoid tissue site, wherein lymphocytes can freely proliferate and expand. To study the influence of immunosuppressive therapy on cellular proliferation, the steroid budesonide, 120 micrograms/kg/day, was administered for 13 days (MST, 20 days). During effective immunosuppressive therapy, still a remarkable amount of infiltrating cells was present within the grafts. Moreover, immunosuppressive treatment did not primarily appear to affect the proliferative capacity of the individual cell types. OX8-positive cells as well as W3/25-positive cells clearly showed proliferative activity within the treated grafts. However, despite the presence of these proliferative cells, signs of graft destruction were absent during immunosuppressive therapy. This finding may shed new light on the effect of steroids at the site of the graft and their role in the prevention of tissue destruction.

Animals↗
PubMed1989

Cytokinetic analysis of lung cancer by bromodeoxyuridine labeling of cytology specimens.

Recent flow cytometric (FCM) studies have indicated the prognostic value of S-phase cells (SPF) in lung cancer. More refined cytokinetic analysis can be obtained by dual-parameter FCM, labeling S-phase cells with 5-bromodeoxyuridine (BrdUrd), which can be detected using a monoclonal antibody (MoAb) to BrdUrd. Tumor cells obtained through bronchoscopic brush were incubated for 1 hr in RPMI 1640 medium with 10% fetal calf serum and 10 microM BrdUrd. After fixation in ethanol, pepsin treatment, and DNA denaturation, the nuclei were stained with anti-BrdUrd MoAb and propidium iodide. From 14 of 20 patients, sufficient material was obtained (three adenocarcinoma and seven squamous cell, one giant cell, and three small cell carcinoma). The measured SPF ranged from 5.2% to 26%. The labeling index (LI), calculated as the ratio of the number of BrdUrd-labeled cells to the total number of aneuploid cells, or diploid cells in the case of a diploid tumor, ranged from 1.2% to 16.7%; LI and SPF correlated significantly (r = 0.69). In this study, we have demonstrated the feasibility of determining the actively DNA-synthesizing cells on brush material from lung cancer cells. In addition, some extra information can be obtained about the SPF population, including the fraction of unlabeled SPF, which could be of prognostic significance.

Bromodeoxyuridine↗
PubMed1989

Butylated hydroxyanisole-induced alterations in cell kinetic parameters in rat forestomach in relation to its oxidative cytochrome P-450-mediated metabolism.

Four groups of six male Wistar rats (85 +/- 7 g) were fed a diet containing 0% (control) or 2% of the carcinogenic food antioxidant butylated hydroxyanisole (BHA) for 2 weeks. In this experiment, feeding 2% BHA is equivalent to a dose of 2.1 +/- 0.3 g BHA/kg/day. One 0% BHA and one 2% BHA-fed group of rats were daily injected i.p. with the cytochrome P-450 inducer phenobarbital (PB; 60 mg/kg) in saline. These two groups were encoded 0PB and 2PB respectively. Simultaneously, two control groups of rats were injected i.p. with saline only (0 and 2 respectively). PB administration increased relative weight, cytochrome P-450 content and ethoxycoumarin-0-deethylase activity of livers as compared to control rats. In addition, cytochrome P-450-mediated oxidative demethylation of BHA into tert-butyl-hydroquinone (TBHQ), monitored as urinary TBHQ excretion, was significantly increased in PB-induced rats as compared to non-induced rats (0.59 +/- 0.19 versus 0.37 +/- 0.09%; P less than 0.05). The mean labelling index (LI) and potential doubling time (Tpot) in rat forestomach were significantly (P less than 0.01) altered in groups of rats fed 2% BHA as compared to their appropriate control groups. No differences in cell kinetic parameters between either the two control groups (0, 0PB) or between the 2% BHA-fed groups (2, 2PB) was observed. Thus, although an increase in oxidative demethylation of BHA as a response to PB administration is evident, biotransformation of BHA into TBHQ is not correlated to changes in cell kinetic parameters in rat forestomach. Moreover, in rats oxidative cytochrome P-450-mediated demethylation of BHA into TBHQ appears not to be related to the oral dose of BHA. This indicates that oxidative cytochrome P-450-mediated biotransformation of BHA does not contribute to the tumorigenicity of BHA in rat forestomach.

Animals↗
PubMed1988

Cytomegalovirus alters the von Willebrand factor content in human endothelial cells.

Cultured human endothelial cells were infected with human cytomegalovirus AD 169 and Kerr. The infection resulted in the appearance of viral antigens in the nuclei of about 10% of the endothelial cells and in the concomitant disappearance of vWF from the infected cells. No differences were observed between endothelial cells from different sources (umbilical cord veins or arteries, adult veins).

Antigens, Viral↗
PubMed1988

A multivariate analysis of pathologic prognostic indicators in large bowel cancer.

A multivariate analysis of the pathologic data of 350 patients with primary colorectal cancer was performed. In addition to conventional parameters such as shape and size of the primary tumor, central node involvement, angioinvasive growth, grade, and stage, new variables such as the immunoreactivity patterns of carcinoembryonic antigen (CEA), CA 19-9, mucin, serotonin, secretory component (SC), and the DNA index were tested for their potential prognostic value. Every variable except CA 19-9, serotonin, and DNA showed significant prognostic information in univariate analysis. However, in the multivariate analysis stage was the predictive factor with the highest hazard ratio, but absence of central node involvement, tumors with diameters between 3.5 cm and 6 cm, exophytic tumor growth, well-differentiated tumors, tumors with CEA immunoreactivity, absence for staining with serotonin, and diploid tumors also were included in the relative risk model. Thus, the aforementioned variables appear to play a role in the establishment of a prognostic index.

Aged↗
PubMed1988

Xenografting of normal colonic mucosa in athymic mice.

Colonic mucosa was implanted subcutaneously into nude mice in order to investigate the potential use of xenografts as an in vivo model system for the study of colonic epithelial proliferation and differentiation. The xenografts were followed for 5 weeks. Proliferation was studied by bromodeoxyuridine (BrdU) incorporation followed by visualization of the DNA-synthesizing cells with an anti BrdU monoclonal antibody. Differentiated cells were visualized by histochemical staining of goblet- and enterochromaffin cells and of columnar cells by immunoperoxidase staining for carcinoembryonic antigen (CEA), secretory component, and serotonin. In different strains of mice different observations were made. Less immunocompetent strains (Balb c nu/nu, NMRI nu/nu) developed abscesses at the site of the xenograft as well as wasting disease. These phenomena almost never occurred in CD-1 nu/nu mice. The success rate was highest in CD-1 nu/nu mice. After about 1 week, only crypt base cells remained vital and started to repopulate crypts with epithelial cells showing normal colonic differentiation features such as CEA, secretory component, and serotonin immunoreactivity and mainly sulphomucin production. After longer periods of time, crypts started to form cyst-like structures due to accumulation of secretion products and dead cells. DNA-synthesizing cells were seen in the basal areas of recognizable crypts and in the cyst-like structures in a random distribution. These results indicate that normal colon mucosa xenografted into nude mice maintains its proliferative and differentiating capacity for at least 5 weeks.

Animals↗
PubMed1988

Flow cytometric analysis of DNA ploidy level in paraffin-embedded tissue of non-small-cell lung cancer.

Investigations regarding the prognostic value of DNA content (ploidy) and proliferative characteristics [percentage of cells in S-phase or S-phase fraction (SPF)] have been greatly facilitated by the application of flow cytometry (FCM) using nuclei isolated from paraffin-embedded tissue. We have applied this technique to tumor sections from patients presenting with non-small-cell lung cancer (NSCLC) in 1980 and 1981. From 67 out of 115 patients material of sufficient quantity and quality was obtained to perform DNA-FCM. A multivariate analysis including stage of disease (UICC), age, tumor histology and treatment modality was performed to examine the prognostic significance of DNA-FCM in NSCLC. Aneuploidy was found in 65% of cases. In our study, the DNA content was not related to histology, stage of disease or treatment modality, nor to the length of survival (log rank test P = 0.62). Calculation of SPF was possible in 49/67 cases. The SPF was not related to histology, stage of disease or treatment modality, but a significant prognostic value was found for survival; patients with a high SPF died earlier (P = 0.04) and this was especially true for squamous cell carcinoma (P = 0.02). This study demonstrates the prognostic importance of DNA-FCM-derived information in NSCLC using a multivariate analysis; however further prospective studies in larger patient populations are needed.

Adenocarcinoma↗