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Biomedical subjects

R Clarke

Publications and source records attributed to R Clarke.

At least 145 records · Page 8Linked to original sources
PubMed1994

Perinatal factors increase breast cancer risk.

Emerging evidence suggests that breast cancer may originate during early life. In particular, offspring of mothers who during pregnancy exhibited behaviors that are associated with increased incidence of breast cancer, may be at risk. These behaviors include intake of high fat diet or alcohol, or stressful life style. We have found that neonatal exposure to handling that leads to improved ability to cope with stress, reduces 7,12-dimethylbenz(a)anthracene (DMBA)-induced mammary tumors in rats. Further, our results indicate that maternal exposure to high fat diet increases the incidence of DMBA-induced mammary tumors in female offspring. High fat diet also increases serum 17 beta-estradiol (E2) levels in pregnant animals. These results support the hypothesis that in utero concentrations of estrogens play a critical role in the vulnerability to develop breast cancer. The mechanism of estrogen action might be related to its effect on the induction of epithelial hyperplasia and altered breast differentiation. These events then increase the rate of genetic/epigenetic changes that increase the possibility of neoplastic transformation. Increased pregnancy estrogens may also lead to behavioral alterations in the offspring. This could explain the proposed association between certain behavioral patterns and increased tumorigenity. Our results in transgenic mice overexpressing transforming growth factor alpha (TGF alpha) are in accordance with this interpretation. The male TGF alpha mice exhibit elevated serum E2 levels, impaired ability to cope with stress, increased voluntary alcohol intake and high incidence of spontaneous hepatocellular tumors. These findings indicate that animal models offer a unique opportunity to investigate the role of timing of risk behaviors on breast cancer. They are also useful in the attempts to understand the mechanism of early estrogen action on mammary tumorigenesis.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1994

Sequence heterogeneity of the eae gene and detection of verotoxin-producing Escherichia coli using serotype-specific primers.

The distribution of the Escherichia coli attaching and effacing (eae) gene in strains of verotoxin-producing E. coli (VTEC) isolated from cattle and humans was studied. The majority of strains isolated from humans with bloody diarrhoea or HUS and cattle with severe diarrhoea were eae positive (82 and 83% respectively). In contrast, 59% of VTEC isolated from asymptomatic cattle were eae negative and of the remaining 41% that were eae positive, the majority were serotype O157. H7. The nucleotide sequence of the 3' end of the eae gene of enteropathogenic E. coli (EPEC) of serotype O55. H7 was found to be almost identical to that of serotype O157. H7. Specific primers are described which detect the eae sequences of VTEC serotypes O157. H7, O157. H-, and EPEC serotypes O55. H7 and O55. H-. The nucleotide sequence of the 3' end of the eae gene of serotype O111. H8 differed significantly from that of O157. H7. Primers were developed to specifically identify the eae sequences of VTEC serotypes O111. H- and O111. H8. We conclude that whereas the majority of VTEC associated with disease in cattle and humans possess the eae gene, the gene itself may not be necessary to produce haemorrhagic colitis and HUS. Sequence heterogeneity in the 3' end of eae alleles of VTEC permits specific identification of subsets of these organisms.

Adhesins, Bacterial↗
PubMed1994

Mobilisation of Ph-negative peripheral blood stem cells in CML with idarubicin and cytarabine.

Recent interest in autologous transplantation in chronic myeloid leukaemia (CML) has focused on attempts to select out putatively normal Ph-negative progenitor cells for subsequent reinfusion after high dose therapy. One way in which this may be achieved is by collecting peripheral blood stem cells (PBSCs) during the early regenerative phase following chemotherapy when Ph-negative cells seem to have a short term proliferative advantage. Data now suggest that it is possible to collect Ph-negative (and occasionally PCR negative) progenitor cells in a significant number of CML patients, a proportion of whom will go on to achieve a cytogenetic remission post-autografting. The durability of these remissions and the effect on long term survival remain to be established and at present this form of therapy should be reserved for those unsuitable for allogeneic transplantation who have failed to achieve a major cytogenetic response to interferon-alpha.

Cell Separation↗
PubMed1993

C-terminal deletion mutants of the FokI restriction endonuclease.

We have constructed two C-terminal deletion mutants of the FokI restriction endonuclease by using the polymerase-chain-reaction technique and expressed them in Escherichia coli. The two mutant proteins (MP) of 41 and 30 kDa, were purified to homogeneity and their DNA-binding properties were characterized. The 41-kDa MP specifically binds the DNA sequence, 5'-GGATG/3'-CCTAC, like the wild-type (wt) FokI, but does not cleave DNA. The 30-kDa MP does not bind DNA. The affinity of the 41-kDa MP for the DNA substrate is comparable to that of wt FokI. The 41-kDa MP interacts with its substrate like the wt FokI, as revealed by hydroxyl radical footprinting experiments. In the presence of a DNA substrate, the 41-kDa MP is cleaved by trypsin into a 30-kDa N-terminal fragment and an 11-kDa C-terminal fragment. Addition of the HPLC-purified 11-kDa C-terminal fragment to the 30-kDa MP restores its sequence-specific DNA-binding property. These results confirm that the N-terminal 41-kDa fragment of the FokI ENase constitutes the DNA recognition domain of the ENase.

Amino Acid Sequence↗
PubMed1993

MCF7/LCC2: a 4-hydroxytamoxifen resistant human breast cancer variant that retains sensitivity to the steroidal antiestrogen ICI 182,780.

The development of resistance to the antiestrogen tamoxifen occurs in a high percentage of initially responsive patients. We have developed a new model in which to investigate acquired resistance to triphenylethylenes. A stepwise in vitro selection of the hormone-independent human breast cancer variant MCF-7/LCC1 against 4-hydroxytamoxifen produced a stable resistant population designated MCF7/LCC2. MCF7/LCC2 cells retain levels of estrogen receptor expression comparable to the parental MCF7/LCC1 and MCF-7 cells. Progesterone receptor expression remains estrogen inducible in MCF7/LCC2 cells, although to levels significantly lower than observed in MCF-7 and MCF7/LCC1 cells. MCF7/LCC2 cells form tumors in ovariectomized nude mice without estrogen supplementation, and these tumors are tamoxifen resistant but can be estrogen stimulated. Significantly, MCF7/LCC2 cells have retained sensitivity to the steroidal antiestrogen ICI 182,780. These data suggest that some breast cancer patients who acquire resistance to tamoxifen may not develop cross-resistance to treatment with steroidal antiestrogens.

Animals↗
PubMed1993

Acquisition of hormone-independent growth in MCF-7 cells is accompanied by increased expression of estrogen-regulated genes but without detectable DNA amplifications.

A hormone-independent but hormone-responsive subpopulation (MCF7/MIII) of the hormone-dependent MCF-7 human breast cancer cell line (R. Clarke et al., Proc. Natl. Acad. Sci. USA 86: 3649-3653, 1989) was further passaged in ovariectomized nude mice and re-established in vitro as the continuous cell line MCF7/LCC1. The lag time to the appearance of proliferating tumors in ovariectomized animals is significantly reduced in MCF7/LCC1 when compared with MCF7/MIII cells. In gel denaturation/renaturation analysis of tumor, genomic DNA does not reveal significant differences in the pattern of detectable DNA amplifications between parent MCF-7 cells and MCF7/LCC1 cells. In the absence of estrogen, steady-state levels of phosphoinositol turnover are similar in both MCF-7 and MCF7/LCC1 cells, but turnover is increased by estrogen only in MCF-7 cells. MCF7/MIII and MCF7/LCC1, but not MCF-7 cells, express a high baseline level of the estrogen-regulated pS2 mRNA. The baseline level of expression of progesterone receptor protein, but not mRNA, is higher in MCF7/LCC1 when compared with either MCF-7 or early passage MCF7/MIII cells. However, while the estrogen receptor is also an estrogen-regulated gene, MCF7/MIII and MCF7/LCC1 cells retain estrogen receptor levels equivalent to the parental MCF-7 cells. These data indicate that progression to hormone independence can occur without major gene amplifications or a high constitutive induction of phosphoinositide metabolism. Thus, DNA amplifications may be acquired during the early initiation and/or promotional events of carcinogenesis. Significantly, acquisition of a hormone-independent but responsive phenotype in human breast cancer is associated with perturbations in the expression of specific estrogen-regulated genes.

Animals↗
PubMed1993

The invasive and metastatic properties of hormone-independent but hormone-responsive variants of MCF-7 human breast cancer cells.

We have previously isolated a series of MCF-7 human breast cancer cell variants which no longer require estrogen-supplementation for tumor growth in nude mice (Clarke et al. Proc Natl Acad Sci USA 86: 3649-3653, 1989). We now report that these hormone-independent and hormone-responsive variants (MIII, MCF7/LCC1) can invade locally from solid mammary fat pad tumors, and produce primary extensions on the surface of intraperitoneal structures including liver, pancreas, and diaphragm. Both lymphatic and hematogenous dissemination are observed, resulting in the establishing of pulmonary, bone, and renal metastases. The pattern of metastasis by MIII and MCF7/LCC1 cells closely resembles that frequently observed in breast cancer patients, and provides the first evidence of metastasis from MCF-7 cells growing in vivo without supplementary estrogen. The interexperimental incidence of metastases, and the time from cell inoculation to the appearance of metastatic disease are variable. The increased metastatic potential is not associated with an increase in either the level of laminin attachment, laminin receptor mRNA expression, or secreted type IV collagenolytic activity. We also did not detect a significant decrease in the steady-state mRNA levels of the metastasis inhibitor nm23 gene. However, when growing without estrogen in vitro, MCF7/LCC1 cells produce elevated levels of the estrogen-inducible cathepsin D enzyme.

Animals↗
PubMed1993

Hormone resistance, invasiveness, and metastatic potential in breast cancer.

Critical phenotypic changes that occur during the progression of breast cancer include the loss of hormone-dependence, acquired resistance to systemic therapies, and increased metastatic potential. We have isolated a series of MCF-7 human breast cancer variants which exhibit hormone-independent growth, antiestrogen resistance, and increased metastatic potential. Analysis of the phenotypes of these variants strongly suggests that changes in the expression of specific genes may be critical to the generation of phenotypic diversity in the process of malignant progression in breast cancer. Epigenetic changes may contribute significantly to the generation of these phenotypic changes observed during breast cancer progression. Many of the characteristics of the progressed phenotypes appear to have arisen in response to appropriate selective pressures (growth in ovariectomized nude mice; growth in the presence of antiestrogens). These observations are consistent with the concept of clonal selection and expansion in the process of malignant progression.

Breast Neoplasms↗
PubMed1993

Opposing behavioural alterations in male and female transgenic TGF alpha mice: association with tumour susceptibility.

Psychosocial factors are thought to influence risk and survival from cancer. We have previously studied specific behaviours in transgenic male CD-1 MT42 mice, which overexpress the gene encoding human transforming growth factor alpha (TGF alpha) in multiple tissues, and which develop a high incidence of spontaneous hepatocellular carcinoma. The male TGF alpha mice spent a lengthened time immobile in the swim test, were highly aggressive, had increased plasma levels of 17 beta-estradiol (E2), and reduced natural killer (NK) cell activity. The female transgenic MT42 TGF alpha mice do not develop an increased rate of tumours at any site. We hypothesised that if the alterations in male TGF alpha mice are associated with their development of hepatocellular carcinomas, female TGF alpha should not show these alterations. The data in the present study indicate that female TGF alpha mice display shortened immobility in the swim test, suggesting an improved ability to cope with stress, and appear less aggressive in the resident-intruder test than non-transgenic female CD-1 mice. The female TGF alpha mice also exhibit a 3-fold increase in the plasma levels of E2, and a 3-fold increase in NK cell activity. These findings suggest that the elevated expression of TGF alpha in the transgenic mice is associated with gender-specific behavioural alterations, and the development of spontaneous hepatocellular tumours in the males. Furthermore, TGF alpha alters hormonal and immune parameters similarly in both sexes. It remains to be determined whether the development of hepatocarcinoma in the male TGF alpha animals is associated with an impaired ability to cope with stress and elevated aggressive tendencies and/or whether manipulations leading to an impaired ability to cope with stress will promote tumourigenesis in female TGF alpha mice.

Aggression↗
PubMed1993

Utilization of non-protein nitrogen in whey-dominant formulae by low-birthweight infants.

1. The effects of increasing non-protein nitrogen intake on nitrogen balance and alpha-amino nitrogen flux rate using [15N]glycine were examined in 30 low-birthweight appropriate-for-gestational-age infants (birthweight 1.5-2.0 kg). The compositions of the three whey-dominant formulae were similar except for the ratios of non-protein nitrogen/protein nitrogen, which were 6.5:93.5, 11.4:88.6 and 17.5:82.5. 2. Infants in the three diet groups each received similar total nitrogen intakes (395 mg of N day-1 kg-1, SD 2.6; n = 3). Protein nitrogen and non-protein nitrogen intakes were different as expected. Energy absorption (449 kJ day-1 kg-1, SD 13; n = 3) did not differ significantly between groups. A similar weight gain was observed in all groups. 3. Nitrogen absorption (76%, SD 4; n = 3) was not significantly different between groups. Apparent urea balance was significantly increased and became positive in the group receiving the formula with the higher proportion of non-protein nitrogen and urea nitrogen. Nitrogen retention, however, was significantly depressed in this group, indicating decreased efficiency of nitrogen utilization at this level of non-protein nitrogen despite an enhanced urea salvage. 4. The enrichment of the 15N label in urinary urea at isotopic steady state was significantly reduced in infants receiving the highest urea-containing formula, presumably due to the dilution of 15N-labelled urea by dietary urea. No difference, however, was found in the enrichment of the 15N label in urinary ammonia. Rates of alpha-amino nitrogen flux, protein synthesis and protein breakdown calculated from the ammonia labelling did not differ significantly between groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Caseins↗
PubMed1993

Effect of chemotherapy on the energy and protein metabolism of children near the end of treatment for acute lymphoblastic leukemia.

Eight children in the final 3 mo of chemotherapy for acute lymphoblastic leukemia were studied while on oral 6-mercaptopurine (6MP) maintenance therapy and then again 4-9 mo after chemotherapy had been completed. Six of the eight were also studied a second time while on 6MP in the 24-h period after receiving intravenous methotrexate (MTX). 6MP reduced protein oxidation after a test meal and reduced fasting urinary urea excretion by enhancing the reutilization of endogenous amino acids for protein synthesis. MTX had no detectable effects on protein metabolism but reduced overnight carbohydrate utilization by enhancing fat utilization. A similar enhancement of fat utilization was evident after a test meal. The two drugs in combination resulted in effects on protein and energy metabolism that were the sum of the individual effects plus an increase in the rate of whole-body protein turnover and synthesis.

Adolescent↗
PubMed1993

Client feedback--development of an instrument for psychiatric inpatients.

A client feedback instrument and method has been developed to improve service delivery to psychiatric inpatients. Results indicate that the strongest cause of patients believing they can cope with life out of hospital is the quality of communication between staff, followed by communication with staff and satisfaction with the centre overall. A secondary finding was that good response rates can be achieved and reliable data can be gathered from psychiatric patients. Further development options and potential application to other psychiatric hospitals are discussed.

Australia↗