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Biomedical subjects

J M Gee

Publications and source records attributed to J M Gee.

At least 55 records · Page 3Linked to original sources
PubMed1997

Comparative studies of the mitogenic effects of epidermal growth factor and transforming growth factor-alpha and the expression of various growth factors in neoplastic and non-neoplastic prostatic cell lines.

BACKGROUND: The role of growth factors in prostate cell growth has been investigated as these peptides may be involved in the autonomous growth of hormone-independent prostate cancer. METHODS: Responses of neoplastic (PC-3 and CPA) and non-neoplastic (CAPE) prostatic cell lines to epidermal growth factor (EGF) and transforming growth factor-alpha (TGF-alpha) were determined using clonogenic and growth curve analysis. The constitutive expression of EGF, TGF-alpha, and TGF-beta 1-3 mRNA was examined using Northern blotting and EGF and TGF-alpha protein levels were determined immunohistochemically. RESULTS: Growth curve and clonogenic analysis indicated that EGF and TGF-alpha were mitogenic in each cell line. The magnitude of the clonogenic response varied between the cell lines, with CPA cells showing the greatest growth increases. CPA cells also displayed the highest levels of EGF and TGF-alpha mRNA and protein. TGF-beta 1 mRNA was detected in the order of magnitude, PC-3 > CPA > CAPE. Furthermore, PC-3 and CPA cells expressed TGF-beta 3 and TGF-beta 2 transcripts respectively. In each cell line, the expression of any growth factor mRNA was not affected by exogenous EGF. CONCLUSIONS: The growth responses of the cell lines to EGF and TGF-alpha did not correlate with their constitutive levels of EGF and TGF-alpha mRNA and protein, thus whilst growth factors may be important in malignant cell growth, other pathways may also be involved in the autocrine regulation of cell proliferation.

Animals↗
PubMed1997

Effect of saponin on the transmucosal passage of beta-lactoglobulin across the proximal small intestine of normal and beta-lactoglobulin-sensitised rats.

The ability of saponins and glycoalkaloids to permeabilise the mammalian intestinal barrier has been previously demonstrated in vitro, leading to the hypothesis that membranolytic saponins may facilitate transfer to the tissues of otherwise excluded macromolecules. An enhanced uptake of, for instance, potentially allergenic species from the lumen is one of the factors that may affect the induction of food allergy, and its presentation in already sensitised individuals. In the experiments described here, an increase in the transmucosal uptake of the milk allergen beta-lactoglobulin (beta LG) was assessed in non-sensitised and sensitised Brown Norway rats in the presence of Gypsophila saponin. Isolated jejunal loops were exposed in vivo to either beta LG followed by saponin, saponin followed by beta LG or the two compounds simultaneously. Portal vein blood samples were collected and assayed for beta LG and rat mucosal mast cell protease (RCMP II) activity. Mucosal tissue was also examined histologically and assayed for histamine content. Sham-operated animals, exposed to physiological buffer alone, were included as controls and beta LG measurements corrected for this component which was negligible. No transfer of beta LG occurred in the absence of saponin in non-sensitised rats, whereas a significant enhancement was observed in the presence of saponin. beta LG was detected in the portal circulation of sensitised rats exposed to beta LG alone; however addition of saponin to the intestinal lumen further enhanced this uptake, possibly by an independent mechanism. Histological examination of the mucosal epithelium exposed to saponin revealed damage, especially at the villus tips. Mucosal histamine and serum RCMP II concentrations were consistent with the differences observed between sensitised and non-sensitised animals. It is concluded that exposure to food constituents capable of permeabilising the mucosal epithelium may increase the risk of sensitisation to dietary antigens.

Animals↗
PubMed1997

Soluble non-starch polysaccharides derived from complex food matrices do not increase average lipid droplet size during gastric lipid emulsification in rats.

The creation of a finely dispersed lipid emulsion is essential for efficient hydrolysis of dietary triglycerides. The effectiveness of emulsification within the stomach depends upon the shear force generated by gastric motility and the concentration of emulsifiers present in the gastric contents. Other dietary constituents can modify these factors, and previous studies have suggested that the presence of soluble non-starch polysaccharides (NSP) during digestion might increase the average size of intraluminal emulsion droplets. In the present study, we developed a new technique for the isolation and analysis of intraluminal lipid emulsions by optical diffraction analysis. The method was applied to rats fed powdered semipurified diets that were free of all NSP or supplemented with insoluble cellulose, guar gum, or NSP derived from apple, carrot or rolled oats. Cellulose had no significant effect on emulsion size, and there was no evidence that the average sizes of lipid droplets in the gastric fundus or antrum were higher than control values in rats fed diets supplemented with any source of soluble NSP. In the groups fed oats and cooked carrot NSP, the mean droplet diameters approached half the values for diets free of NSP or containing insoluble cellulose. The difference between rats fed NSP from cooked carrot and those fed cellulose was significant in the proximal stomach (P < 0.05), and that between rats fed raw oats and rats fed cellulose was significant in the distal stomach (P < 0.05). Soluble dietary fiber does not inhibit lipid or cholesterol absorption via any inhibition of lipid emulsification.

Absorption↗
PubMed1997

Use of reverse transcription-polymerase chain reaction methodology to detect estrogen-regulated gene expression in small breast cancer specimens.

We describe the development and use of a sensitive reverse transcription-PCR (RT-PCR) procedure to detect novel estrogen-regulated gene expression in small clinical breast cancer samples, in which such study would be extremely difficult by any other molecular or immunocytochemical means. Assay optimization for pLIV1, estrogen receptors (ERs), progesterone receptors, and pS2 gene products was carried out on 50 primary breast cancers for which comparative Northern analysis and immunocytochemical data were available. Using 27 amplification cycles and a 0.5 microM primer concentration, varying expressions of the gene products were recorded simultaneously with a constant densitometric signal for a coamplified endogenous control gene (alpha-actin). Good concordances were subsequently observed between pLIV1 status generated by RT-PCR and both Northern analysis (P = 0.002) and ER status by immunocytochemistry (P = 0.0244). Agreement was also noted between ER (P = 0.002), progesterone receptor (P = 0.0005), and pS2 (P = 0. 0023) RT-PCR and immunocytochemical methodologies. The RT-PCR assays were then applied to 10 needle core trucut biopsies in which similar relationships were obtained. Our results justify the future use of this RT-PCR methodology to examine new estrogen-regulated genes in small breast cancer samples, and it is envisaged that this technology will prove invaluable in many future breast cancer studies.

Biomarkers, Tumor↗
PubMed1996

Effects of short-term antiestrogen treatment of primary breast cancer on estrogen receptor mRNA and protein expression and on estrogen-regulated genes.

Effects of the pure antiestrogen ICI182780 and tamoxifen on ER-protein, ER-mRNA, and estrogen-regulated mRNA expression were analysed using matched pretreatment core-cut biopsies and post-treatment mastectomy samples from 43 ER positive human breast cancers. Sixteen controls received either no preoperative treatment (n = 9) (7 days) or placebo (n = 7) (median 21 days) prior to primary surgery. Nineteen patients received ICI182780 6 mg/day (n = 10) or 18 mg/day (n = 9) for 7 days. Eight patients were given preoperative tamoxifen (4 x 40 mg-day 1, 20 mg/day thereafter, median 21 days). ER-protein expression was assessed on pre and post treatment samples by immunocytochemistry. ER, pS2, pLIV1, and actin-mRNA expression was determined by northern analysis on post-treatment samples only. ER-mRNA levels were similar to controls following ICI182780 or tamoxifen treatment. However ER-protein levels were significantly suppressed by ICI182780, particularly at the higher dosage (p = 0.0013). Tamoxifen had no significant effect on ER-protein levels. The ER-mRNA and ER-protein contents of control tumors were linearly related (Spearman r = 0.719, p = 0.006). A similar relationship between pretreatment protein and post ICI182780 treatment mRNA levels was observed (r = 0.652, p = 0.005). However, comparison of post ICI182780 treatment protein and mRNA results shows a loss of linearity through a reduction in protein without concurrent loss of mRNA (r = 0.28, p = 0.257). pS2 mRNA hybridization was lower in ICI182780 treated samples than controls (Mann-Whitney p = 0.035) but was unaffected by tamoxifen. pLIV1 mRNA hybridization was uninfluenced by either treatment. Short term exposure of breast tumors to ICI182780 appears to produce a greater inhibition of estrogen-induced transcriptional events than tamoxifen. These effects appear to occur without a concurrent reduction in ER mRNA levels.

Antineoplastic Agents, Hormonal↗
PubMed1996

Brown Norway rat model of food allergy: effect of plant components on the development of oral sensitization.

The Brown Norway (BN) rat was examined as a model for investigating factors that influence the development of food allergy. An antigen dose-response curve for the production of antigen-specific reaginic antibody (IgE) induced through the oral route was determined. Animals were dosed orally with 1.0, 2.5, 5.0, 7.5, 10.0 and 12.0 mg ovalbumin/ml (0.5 ml/100 g twice a week for 6 wk). To promote IgE production the adjuvant carrageenan was administered once a week by the i.p. route. The effect on oral sensitization of 1.5 mg Gypsophila sp. saponin/ml administered together with the antigen on oral sensitization was examined in animals treated with 2.5, 6.0 or 10.0 mg ovalbumin/ml. The number of animals producing antigen specific reaginic antibody in response to 2.5 mg ovalbumin/ml was significantly increased (P < 0.01) in the group that received saponin with 2.5 mg ovalbumin/ml. These studies indicate that the BN rat is a sensitive model for the investigation of allergic reactions to food and has the potential to determine the impact of other dietary factors on the development of oral sensitization.

Adjuvants, Immunologic↗
PubMed1996

Short-term effects of pure anti-oestrogen ICI 182780 treatment on oestrogen receptor, epidermal growth factor receptor and transforming growth factor-alpha protein expression in human breast cancer.

Expression of oestrogen receptor (ER), epidermal growth factor receptor (EGFR) and transforming growth factor-alpha (TGF alpha) proteins was assessed by immunocytochemistry on primary breast cancer specimens obtained before and following short-term (7-day) presurgical exposure to pure anti-oestrogen (7 alpha- [9- (4,4,5,5,5-pentafluoropentylsulphinyl) nonyl] estra-1,3,5, (10)-triene-3,17 beta-diol, ICI 182780) treatment and compared with no-treatment controls. Paired needle-core and mastectomy samples were obtained from 21 patients. Effects of ICI 182780 (10(-7)M) on MCF7 breast cancer cell ER, EGFR and TGF alpha expression were also examined over 14 days. ER protein was significantly suppressed by ICI 182780 in vivo (P = 0.009) and comparative analysis of short term ICI 182780 effects in vitro, using ER-positive MCF7 cells, gave largely equivalent results. EGFR and TGF alpha protein levels were unaltered by treatment. ICI 182780 suppresses ER without a concomitant rise in either EGFR or TGF alpha.

Breast Neoplasms↗
PubMed1996

Fermentable carbohydrate modulates postprandial enteroglucagon and gastrin release in rats.

We studied the effects of a fermentable sugar-alcohol (lactitol) on the concentrations of enteroglucagon and gastrin in the blood of rats for 7.5 h after feeding. The control and treatment groups were fed on semi-purified diets containing either non-fermentable cellulose or lactitol respectively, at 100 g/kg. Compared with the cellulose-fed group, the animals fed with lactitol had higher levels of enteroglucagon (5-10 times higher than control; P < 0.05) and lower serum gastrin (70-80% of control; P < 0.05) for several hours after the withdrawal of feed. In contrast, varying the level of dietary lipid (maize oil) over a range of 8-120 g/kg had no effect on the release of either peptide. These results suggest that poorly absorbed fermentable dietary carbohydrate stimulates postprandial plasma enteroglucagon and inhibits serum gastrin release in the rat. The mechanism is uncertain but an endocrine response by the colon to fermentation products seems probable.

Animals↗
PubMed1996

Fermentable carbohydrates elevate plasma enteroglucagon but high viscosity is also necessary to stimulate small bowel mucosal cell proliferation in rats.

Enteroglucagon is a collective term for a small family of peptides derived from proglucagon by post-translational processing in the L-cells of the distal small intestine and colon. There is evidence that it inhibits gastric secretion, and high levels of enteroglucagon occur in plasma during intestinal adaptation, which suggests that it may also function as a trophic factor for the intestine. Certain types of soluble non-starch polysaccharide (dietary fiber) stimulate the release of enteroglucagon in rats but the mechanism is unknown. In this study we explored the importance of the viscosity and fermentability of nonabsorbed carbohydrates as determinants of plasma enteroglucagon and mucosal cell proliferation in the distal ileum of rats. Replacement of cellulose (10 g/kg) with guar gum in a semisynthetic diet led to a prompt and sustained rise in plasma enteroglucagon concentrations. Our initial hypothesis that this was a consequence of delayed nutrient absorption was disproven by the fact that hydroxypropylmethylcellulose (HPMC), a viscous but nonfermentable polysaccharide, had no effect on plasma enteroglucagon under the same conditions. In contrast, the nondigestible disaccharide lactitol led to a prolonged rise in plasma enteroglucagon, similar to that observed with guar gum. Lactitol is nonviscous, but highly fermentable, and we conclude that fermentable carbohydrate is an important stimulus for the release of enteroglucagon under our experimental conditions. There was no evidence that enteroglucagon released by this mechanism exerted trophic effects on the distal small intestinal mucosa.

Animals↗
PubMed1995

Immunocytochemical localization of Fos protein in human breast cancers and its relationship to a series of prognostic markers and response to endocrine therapy.

The nuclear transcription factor Fos is inducible by both steroid hormones and peptide growth factors. It thus forms a potential point of interaction between steroid hormone- and growth factor-directed pathways and may be critical in the subversion of steroid hormone control in breast cancer. In this light, the present study has used immunocytochemistry to demonstrate in clinical primary breast cancer that Fos expression is indeed significantly associated with a failure to respond to endocrine therapy, with preliminary analysis revealing a survival advantage for those patients whose tumours lacked Fos. Sustained elevated levels of Fos expression were significantly associated with further factors, notably peptide growth factors and their receptors (e.g., EGFR, TGF alpha), as well as with the proliferation marker Ki-67, which have been linked previously to endocrine insensitivity in breast cancer. In contrast, there appeared to be a trend for Fos to be absent in those tumours expressing markers of endocrine responsiveness (e.g., oestrogen receptor [ER], and also ER-mediated markers i.e., PR, pS2 or bcl-2). Interestingly, many of these trends were maintained in ER+ patients, suggesting that Fos may be of importance in directing loss of endocrine sensitivity in ER+ disease.

Breast Neoplasms↗
PubMed1995

A cautionary note regarding the application of Ki-67 antibodies to paraffin-embedded breast cancers.

Immunocytochemical studies examining the Ki-67 proliferation marker in paraffin-embedded material have recently been made possible by the availability of several antibodies, notably MIB-1, which are readily applicable to microwaved sections. Using breast cancer material, the present study examines correlations shown by these new paraffin assays and also by PCNA (proliferating cell nuclear antigen), an existing marker of proliferation, with the established Ki-67 cryosection assay. Paraffin sections were microwaved prior to incubation with Ki-67 or MIB-1 antibodies. Signal detection was carried out with a biotinylated secondary antibody, peroxidase-conjugated streptavidin, and DAB/H2O2 chromogen. The results suggest that caution is required when studying proliferation in paraffin-embedded breast cancers by immunostaining using Ki-67 antibodies. Nuclear staining in wax sections (Ki-Par, MIB-1, PCNA) greatly exceeded that in cryosections (Ki-Froz) and thus correlations were notably absent between Ki-Par or PCNA immunostaining and the routine Ki-Froz assay. Immunostaining with MIB-1 or PCNA may, however, be useful to assess proliferation if cut-offs are applied to eliminate weak immunostaining associated with wax sections. Thus, an approximately linear relationship was seen between MIB-1/Ki-Froz, which was improved if only moderately or moderately/strongly MIB-1-positive cells were scored. Similarly, a significant correlation was also revealed between PCNA/Ki-Froz if such a cut-off was applied.

Antibodies, Monoclonal↗
PubMed1995

Differential expression of oestrogen regulated genes in breast cancer.

Pathological endpoints such as tumour size, lymph node status and vascular invasion remain the most useful guides in selecting treatment strategies for breast cancer. There is a need, however, to further investigate the molecular mechanisms that determine the properties of an individual tumour e.g., hormone responsiveness and probability of metastasis. While numerous prognostic factors have now been identified few have contributed to defining clinical response to therapy. Oestrogen-regulated genes are likely to be important since they not only define a functional oestrogen receptor, but alterations in their expression might provide insights into the mechanisms involved in tumour progression and loss of endocrine sensitivity. Recently an oestrogen responsive gene, pLIV1, has been isolated and shown to be expressed in ER+ disease where it appears to predict nodal involvement. The present paper describes aspects of its regulation and discusses the potential role of this and other genes in the development of endocrine resistance.

Breast Neoplasms↗
PubMed1995

Cell proliferation in prostatic carcinoma: comparative analysis of Ki-67, MIB-1 and PCNA.

Antibodies to assess the proliferative index of tumours are being increasingly employed together with established markers for prognostic evaluation. This study set out to compare three cell proliferation markers, Ki-67, MIB-1 and PCNA, utilizing a semi-quantitative method of assessment, in 20 human prostatic carcinomas. The streptavidin-biotin immunostaining system was used for the monoclonal antibodies MIB-1 and PCNA and an indirect immunoperoxidase assay for the monoclonal antibody Ki-67. Significant correlations were found between the expression of Ki-67 in frozen tissues and MIB-1 in formal saline-fixed wax-embedded tissues (p = 0.0003); between Ki-67 and PCNA expression in Bouin's-fixed tissues (p < or = 0.0001); and MIB-1 (formalin-saline-fixed tissues) and PCNA (Bouin's-fixed tissues) (p < or = 0.0001). A more intense nuclear staining pattern with less heterogeneity was observed for MIB-1 compared with PCNA, suggesting the antibody of choice, on formal saline-fixed tissues, is MIB-1, which closely correlated with Ki-67, a marker we have previously shown to be of prognostic value in prostatic carcinoma.

Antibodies, Monoclonal↗