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Background AZD8055 is a potent orally available mTOR kinase inhibitor with in vitro and in vivo antitumour activity against a range of tumour types. Preclinical studies showed that AZD8055 induced a dose-dependent pharmacodynamic effect in xenograft models in vivo but a lack of understanding of the relative contributions of the maximum inhibition of the biomarkers and the duration of inhibition to the antitumour effect limited the rational design of experiments to optimize the dose and schedules. Methods In this study, a mathematical modeling approach was developed to relate pharmacodynamics and antitumour activity using preclinical data generated in mice bearing U87-MG xenografts. Results Refinement and validation of the model was carried out in a panel of additional human tumour xenograft models with different growth rates and different sensitivity to AZD8055 (from partial growth inhibition to regression). Finally, the model was applied to accurately predict the efficacy of high, intermittent dosing schedules of AZD8055. Conclusion Overall, this new model linking pharmacokinetics, pharmacodynamic biomarkers and efficacy across multiple tumour xenografts with different sensitivity to AZD8055 was able to identify the optimal dose and route of administration to maximize the antitumour efficacy in pre-clinical models and its potential for translation into man.

Minimising losses to pests and diseases is essential for producing sufficient food to feed our rapidly growing population. The necrotrophic fungus Botrytis cinerea triggers devastating pre- and post-harvest yield losses in tomato (Solanum lycopersicum). Current control methods are based on the pre-harvest use of fungicides, which are limited by strict legislation. Here, we have tested whether induction of resistance by β-aminobutyric acid (BABA) at different developmental stages, provides an alternative strategy to protect tomato fruit post-harvest against B. cinerea. Soil-drenching plants with BABA once fruit had already formed, had no impact on tomatoes susceptibility to B. cinerea. Whereas BABA application to seedlings was found to significantly reduce the post-harvest infection of fruit. This resistance response was not associated with a yield reduction, however there was a delay in fruit ripening. Untargeted metabolomics unravelled differences between fruit from water and BABA-treated plants, demonstrating that BABA triggered a defence-associated metabolomics profile that was long-lasting. Targeted analysis of defence hormones suggested a role of abscisic acid (ABA) in the resistance phenotype. Post-harvest application of ABA to the fruit of water-treated plants induced susceptibility to B. cinerea. This phenotype was absent from the ABA exposed fruit of BABA-treated plants, suggesting a complex role of ABA in the BABA-induced resistance phenotype. A final targeted metabolomic analysis detected trace residues of BABA accumulated in the red fruit. Overall, we have demonstrated that β-aminobutyric acid induces post-harvest resistance in tomato fruit against B. cinerea with no penalties in yield. This article is protected by copyright. All rights reserved.

Although topical TLR7 therapies such as imiquimod have proved successful in the treatment of dermatological malignancy, systemic delivery may be required for optimal immunotherapy of nondermatological tumors. We report that intravenous delivery of the novel small molecule TLR7 agonist, DSR-6434, leads to the induction of type 1 interferon and activation of T and B lymphocytes, NK and NKT cells. Our data demonstrate that systemic administration of DSR-6434 enhances the efficacy of ionizing radiation (IR) and leads to improved survival in mice bearing either CT26 or KHT tumors. Of the CT26 tumor-bearing mice that received combined therapy, 55% experienced complete tumor resolution. Our data reveal that these long-term surviving mice have a significantly greater frequency of tumor antigen specific CD8+ T cells when compared to age-matched tumor-naive cells. To evaluate therapeutic effects on spontaneous metastases, we showed that combination of DSR-6434 with local IR of the primary tumor significantly reduced metastatic burden in the lung, when compared to time-matched cohorts treated with IR alone. The data demonstrate that systemic administration of the novel TLR7 agonist DSR-6434 in combination with IR primes an antitumor CD8+ T-cell response leading to improved survival in syngeneic models of colorectal carcinoma and fibrosarcoma. Importantly, efficacy extends to sites outside of the field of irradiation, reducing metastatic load. Clinical evaluation of systemic TLR7 therapy in combination with IR for the treatment of solid malignancy is warranted. What's new? Recent evidence suggests that damage from ionizing radiation (IR) can render tumor cells immunogenic. Unfortunately, established tumors often suppress this anti-tumor immune response. Combination therapy with IR and immune-modulators such as Toll-like-receptor (TLR) family agonists may overcome this problem. In this proof-of-concept study, the authors examined one such small-molecule drug, called DSR-6434. They found that systemic administration of DSR-6434 can enhance the effectiveness of radiotherapy in mice, and that this occurs via the generation of tumor-specific immune responses. Easily delivered drugs that activate TLR-family molecules may thus offer a promising therapeutic approach.

RATIONALE: Metabonomic studies use complex biological samples (blood plasma/serum, tissues, etc.) that when analysed with high-performance liquid chromatography/mass spectrometry (HPLC/MS) or nuclear magnetic resonance (NMR) generate profiles that may contain many thousands of features. These profiles can be difficult to interpret with the majority of the features contributing little to the study. As such there is an argument for the development of techniques that can simplify the problem by targeting particular classes of compounds. METHODS: In this study ultra-performance liquid chromatography/inductively coupled plasma mass spectrometry (UPLC/ICP-MS) was used to profile tumour tissue and plasma samples for phosphorus- and sulfur-containing metabolites. These samples were xenograft tumours (derived from breast, lung and colon cell lines) and plasma obtained from nude mice. Plasma was also obtained from non-tumour-bearing mice as a control. Due to isobaric interferences this method took advantage of the dynamic reaction cell within the ICP-MS system to react the phosphorus and sulfur ions with oxygen. The PO+ and SO+ ions were then monitored free of interferences. The total phosphorus and sulfur within each sample was also recorded using flow injection ICP-MS. A robust quality control system based on pooled sample replicate analysis was used throughout the study. RESULTS: Determination of the total phosphorus and sulfur content of each sample was sufficient in itself for statistical differentiation between the majority of the cell lines analysed. Subsequent reversed-phase chromatographic profiling of the organic tumour and plasma extracts revealed the presence of a number of well-retained phosphorus-containing compounds that showed tumour-specific profiles. Reversed-phase profiling was not suitable for the sulfur-containing compounds which eluted with the solvent front. CONCLUSIONS: This study has shown the potential use of UPLC/ICP-MS to differentiate between tumour cell lines, using both plasma and tumour tissue samples, based solely on metabolites that contain phosphorus or sulfur. Whilst further work is required to identify these compounds this methodology shows the ability of the described methods to provide targets for future biomarker discovery studies.

Global amphibian declines are a major element of the current biodiversity crisis. Monitoring changes in the distribution and abundance of target species is a basic component in conservation decision making and requires robust and repeatable sampling. For EU member states, surveillance of designated species, including the common frog Rana temporaria, is a formal requirement of the ‘EC Habitats & Species Directive’. We deployed established methods for estimating frog population density at local water bodies and extrapolated these to the national and ecoregion scale. Spawn occurred at 49.4% of water bodies and 70.1% of independent 500-m survey squares. Using spawn mat area, we estimated the number of adult breeding females and subsequently the total population assuming a sex ratio of 1:1. A negative binomial model suggested that mean frog density was 23.5 frogs ha−1 [95% confidence interval (CI) 14.9–44.0] equating to 196 M frogs (95%CI 124 M–367 M) throughout Ireland. A total of 86% of frogs bred in drainage ditches, which were a notably common feature of the landscape. The recorded distribution of the species did not change significantly between the last Article 17 reporting period (1993–2006) and the current period (2007–2011) throughout the Republic of Ireland. Recording effort was markedly lower in Northern Ireland, which led to an apparent decline in the recorded distribution. We highlight the need to coordinate biological surveys between adjacent political jurisdictions that share a common ecoregion to avoid apparent disparities in the quality of distributional information. Power analysis suggested that a reduced sample of 40–50 survey squares is sufficient to detect a 30% decline (consistent with the International Union for Conservation of Nature Category of ‘Vulnerable’) at 80% power providing guidance for minimizing future survey effort. Our results provide a test case for other EU member states to follow when conducting future conservation assessments for R. temporaria and other clump-spawning amphibians.

Polymeric medical devices widely used in orthopedic surgery play key roles in fracture fixation and orthopedic implant design. Topographical modification and surface micro-roughness of these devices regulate cellular adhesion, a process fundamental in the initiation of osteoinduction and osteogenesis. Advances in fabrication techniques have evolved the field of surface modification; in particular, nanotechnology has allowed the development of nanoscale substrates for the investigation into cell-nanofeature interactions. In this study human osteoblasts (HOBs) were cultured on ordered nanoscale pits and random nano "craters" and "islands". Adhesion subtypes were quantified by immunofluorescent microscopy and cell-substrate interactions investigated via immuno-scanning electron microscopy. To investigate the effects of these substrates on cellular function 1.7 k microarray analysis was used to establish gene profiles of enriched STRO-1+ progenitor cell populations cultured on these nanotopographies. Nanotopographies affected the formation of adhesions on experimental substrates. Adhesion formation was prominent on planar control substrates and reduced on nanocrater and nanoisland topographies; nanopits, however, were shown to inhibit directly the formation of large adhesions. STRO-1+ progenitor cells cultured on experimental substrates revealed significant changes in genetic expression. This study implicates nanotopographical modification as a significant modulator of osteoblast adhesion and cellular function in mesenchymal populations.

The results of 88 cadaver-donor renal first transplants from the Newcastle Transplantation Centre have been reviewed after a minimum period of 6 months post transplant. They show the significant benefit of two antigen identity at the second HL-A locus between kidney donor and recipient. Pretransplant blood transfusion showed a beneficial effect on kidney survival. Cases with preformed cytotoxic antibodies showed a favourable outcome compared to cases without antibody, provided the crossmatch was negative. Rh(D) positive recipients showed a significant association with satisfactory outcome when compared to the Rh(D) negative recipients.

We apply a recently developed method for controlling the spreading of cultured cells using electron beam lithography (EBL) to create polymethylmethacrylate (PMMA) substrata with repeating nanostructures. There are indications that the reduced cell spreading on these substrata, compared with planar PMMA, results from a reduced adhesivity since there are fewer adhesive structures and fewer of their associated stress fibres. The reduced cell spreading also results in a reduced nuclear area and a closer spacing of centrosomes within the nucleus, suggesting that the tension applied to the nucleus is reduced as would be expected from the reduction in stress fibres. In order to obtain further evidence for this, we have used specific inhibitors of components of the cytoskeleton and have found effects comparable with those induced by the new substrata. We have also obtained evidence that these subtrata result in downregulation of gene expression which suggests that this may be due to the changed tension on the nucleus: an intriguing possibility that merits further investigation.

The ability of cells to alter their genomic regulation in response to mechanical conditioning or through changes in morphology and the organization of the interphase nuclei are key questions in cell biology. Here, two nanotopographies have been used as a model surfaces to change cell morphology in order to investigate spatial genomic changes within the nuclei of fibroblasts. Initially, centromeres for chromosome pairs were labeled and the average distance on different substrates calculated. Further to this, Affymetrix whole genome GeneChips were used to rank genomic changes in response to topography and plot the whereabouts on the chromosomes these changes were occurring. It was seen that as cell spreading was changed, so were the positions along the chromosomes that gene regulations were being observed. We hypothesize that as changes in cell and thus nuclear morphology occur, that this may alter the probability of transcription through opening or closing areas of the chromosomes to transcription factors.

Clinical studies often produce fresh tissue samples, which ideally should be immediately snap frozen for storage and subsequent analysis. However, this is often not practically possible, and there is inevitably a time period during which the sample is stored on ice. The delay in freezing may allow endogenous degradation of proteins to occur, affecting 2-D gel protein profiles. This study aims to investigate the type and extent of this degradation by examining how the time-to-freezing delay alters prostatic tissue protein profile. The prostate carcinoma-3 cell line (PC-3), prostate cancer xenografts and canine prostate were used with fluorescence 2-D DIGE to assess protein degradation. It was found that 30-min processing time had minimal effects on the protein profile. Longer delays had little visible effect, but subtle alterations in protein profile began to accumulate as time increased. These data support the practice of completing tissue processing as rapidly as possible, and indicate that short processing times do not notably perturb the 2-D gel spot pattern from prostatic tissue.

Glucocorticoids regulate plasticity and survival of hippocampal neurons. Aberrant exposure to this steroid hormone can result in neurodegeneration, perhaps secondary to disruption of calcium homeostasis. Calbindin, a calcium-binding protein that buffers excess calcium, may protect against neurodegeneration resulting from overabundance of intracellular calcium. In this study, we examined whether chronic treatment (1 year) with cortisol enhances hippocampal calbindin expression in primates. Calbindin is a marker for inhibitory neurons and the dentate gyrus is known to adopt an inhibitory phenotype in response to extreme conditions such as seizures. Thus, we hypothesized that chronic cortisol exposure may also promote a GABAergic phenotype. Therefore, we examined the expression of the GABA-synthesizing enzyme glutamic acid decarboxylase. The expression of brain-derived neurotrophic factor, which is responsive to glucocorticoids, was also examined. Our results demonstrate significant increases in calbindin, glutamic acid decarboxylase and brain-derived neurotrophic factor in several regions of the primate hippocampus, including the dentate gyrus and CA3, in response to chronic cortisol exposure. These results suggest that chronic cortisol exposure may shift the balance towards a GABAergic phenotype, perhaps as part of a compensatory feedback mechanism to dampen the initial excitatory effects of glucocorticoids in the hippocampus.

Background The proportion of births by cesarean section in Australia has recently increased by 35 percent, rising from 17 percent in 1990 to 23 percent in 2000. Unlike previous research, which has focused on women's role in increasing rates of cesarean section, this study aimed to explore the existence of wider cultural norms of acceptance of cesarean section in the Australian community, and the implications these might have for rising rates. Methods A postal self-completion questionnaire was sent to a consecutive sample of 148 women who delivered 7 weeks earlier at the Women's and Children's Hospital, a tertiary-referral public maternity hospital in metropolitan Adelaide, South Australia. These women were recruited at 18 weeks' gestation, and had been involved in a wider study. The questionnaire included three sections, one section on agreement with statements pertaining to cultural acceptance of cesarean section, a second on personal consideration of cesarean section in early pregnancy, and a third including sociodemographic questions. Results Of the total sample, 92 women responded to the questionnaire (response rate 62%). Of 6 items relating to community acceptance of cesarean section, responses ranged from 71.4 percent agreement ("common for people to think that cesarean section offers an easier way of giving birth") to 23.1 percent agreement ("the media seems to portray cesarean section as a better option than vaginal delivery"). The option of having a cesarean section was considered by almost 15 percent (10/68) of women early in their pregnancy. For the vast majority (8/10) this consideration was clinically based. Conclusions This investigation demonstrated that these Australian women, independent of sociodemographic variables such as age and education level, agreed that cesarean section was perceived as an easy, convenient way of giving birth.

In the 1970s, four trials failed to demonstrate any mortality reduction using a combination of chest X-ray (CXR) and/or sputum cytology. The recent early lung cancer action project (ELCAP) demonstrated that modern screening is capable of detecting Stage I lung cancers. Bronchial epithelial changes leading up to cancers are now being understood to include histologic changes and genetic alterations. Emerging molecular markers detected in sputum and serum show promise in the future of lung cancer screening.

Background In chronic renal failure, metabolic acidosis is associated with increased whole body protein degradation. In rats this effect of acidosis occurs in skeletal muscle and is associated with increased ubiquitin mRNA expression. This has not been demonstrated in humans. Materials and methods Six patients with chronic renal failure and acidosis underwent muscle biopsy before and after 1 month's treatment with sodium bicarbonate. RNA was extracted from the biopsy, and the expression of the genes for ubiquitin and the proteasome component, C2, were measured by Northern blotting. Results and conclusions There was no significant difference in the expression of ubiquitin or C2 after bicarbonate treatment. This is contrast with results from animal models of acidosis and some other catabolic conditions in humans. This may reflect the complexity of the ubiquitin-dependent pathway, and it may be that changes in ubiquitin expression are only seen with more severe and/or acute changes in pH.

Background: Although forebrain pro-opiomelanocortin (POMC)–producing neurons seem to mediate or modulate many responses to ethanol consumption, changes in activity of this opiomelanocortinergic system in response to chronic ethanol consumption, withdrawal, and subsequent abstinence remain unresolved. Methods: We investigated the effects of chronic daily ethanol consumption, withdrawal, and subsequent abstinence on adult male Sprague-Dawley rat forebrain opiomelanocortinergic activity as reflected by changes in hypothalamic POMC messenger RNA (mRNA) content by using a well characterized liquid diet model that we have previously demonstrated to accurately simulate not only daily oral ethanol consumption quantity and pattern, but also both neuroendocrine and behavioral changes characteristic of actively drinking and subsequently abstinent alcoholics. Results: After 7 weeks of daily ethanol consumption at night and withdrawal during the day, evening mediobasal hypothalamus POMC mRNA concentrations were suppressed versus both ad libitum–fed and pair-fed controls. Morning POMC mRNA concentrations were also suppressed versus ad libitum–fed controls and tended to be decreased versus pair-fed controls. Three weeks after gradual removal of ethanol from the diet, mediobasal hypothalamus POMC mRNA concentrations were increased relative to ad libitum–fed and pair-fed controls. Plasma concentrations of corticosterone, testosterone, and leptin were also altered by the daily ethanol/withdrawal treatment and by subsequent abstinence. Conclusions: Because each of these hormones has been demonstrated to modify forebrain POMC gene expression under some conditions, the overall changes in forebrain opiomelanocortinergic regulation in response to chronic daily ethanol/withdrawal and subsequent abstinence probably reflect, at least in part, regulation by multiple endocrine mechanisms, together with responses to stress, development of tolerance during chronic daily ethanol consumption, and rebound of function after termination of this consumption. Overall, the demonstrated changes in forebrain POMC gene expression are consistent with significant roles for forebrain opiomelanocortinergic regulation in mediating alcohol dependence, propensity to relapse, and the alcohol deprivation effect.

COGNITIVE FUNCTION IN COGNITIVELY NORMAL ADULTS Elaine R. Peskind, Ge (Gail) Li, Charles W. Wilkinson, Jane Shofer, Lucy Y. Wang, Carl Sikkema, Joseph F. Quinn, Douglas Galasko, Murray A. Raskind, Thomas J. Montine, VA Puget Sound Health Care System and University of Washington, Seattle, Washington, United States; University of Washington, School of Medicine, Seattle, Washington, United States; University of Washington, Seattle, Washington, United States; VA Puget Sound Health Care System, Seattle, Washington, United States; 5 Oregon Health and Sciences University and Portland VA Medical Center, Portland, Oregon, United States; University of California San Diego, La Jolla, California, United States. Contact e-mail: gli@u.washington.edu

Background: Hypothalamo-pituitary-adrenal (HPA) function has been demonstrated to be compromised for weeks and even months after alcoholics cease ethanol consumption. Because nonalcoholic subjects with family history-associated increased risk for alcoholism also exhibit compromised HPA function, it is not clear whether defects in the HPA axis of abstinent alcoholics reflect a preexisting condition that may be responsible for increased risk for alcohol abuse versus a persisting adaptational change in response to prolonged alcohol abuse. Consequently, we investigated whether chronic daily ethanol consumption and withdrawal by male Sprague Dawley® rats would induce persistent HPA changes consistent with those demonstrated in abstinent alcoholics. Methods and Results: In an initial experiment in which ethanol (5%, w/v) was incrementally introduced to liquid diet over a 1 week period followed by 4 weeks of chronic ethanol consumption, not only ethanol-treated rats but also pair-fed control rats exhibited decreased (p < 0.05 vs. ad-libitum-fed controls) anterior pituitary pro-opiomelanocortin (POMC) mRNA concentrations and associated decreases in plasma corticosterone and adrenocorticotropin (ACTH) levels for at least 3 weeks after gradual withdrawal of ethanol from the diet. Pair-feeding-induced decreases (p < 0.05) in thymus and spleen weights suggested that the pair-fed controls were likely stressed in this model, probably in response to the marked and irregular suppression of liquid diet consumption immediately after introduction of ethanol. Consequently, a second model was developed in which ethanol was introduced to the liquid diet much more gradually (i.e., over 3 weeks). In contrast with the rapid ethanol-introduction model, this more prolonged ethanol introduction followed by 4 weeks of chronic daily ethanol consumption increased plasma corticosterone levels (p < 0.05), increased adrenal gland weight (p < 0.05), and decreased thymus and spleen weights (both p < 0.01) without altering any of these parameters in the pair-fed controls. Three weeks after gradual withdrawal of ethanol from the diet, anterior pituitary POMC mRNA concentrations were suppressed (p < 0.05) and thymus and spleen weights were increased (p < 0.05) versus both pair-fed and ad-libitum-fed controls, accompanied by trends for decreased basal plasma corticosterone and adrenal weights. Conclusions: Chronic daily ethanol treatment induced changes in the HPA axis that persisted for at least 3 weeks after complete cessation of ethanol consumption. These persistent alterations in the HPA axis are similar to the aberrant HPA regulation of abstinent alcoholics, sons of alcoholics, Lewis rats, and individuals who suffer from posttraumatic stress disorder and some types of depression, that is, categories of individuals who all exhibit increased risk for high ethanol consumption. Thus, these chronic daily ethanol-induced persistent changes in the HPA axis may have significant roles in alcohol abstinence syndrome and may increase vulnerability to relapse.

To assess acute effects of alcohol on forebrain and pituitary opiomelanocortinergic regulation, a model was developed in which experienced (previously introduced to ethanol administration, so the subjective response was not a novel stimulus) male Sprague-Dawley rats received pulsatile intragastric ethanol infusions during the dark (active) photophase to produce and sustain (for 3 hr) behaviorally relevant (0, 40 to 70, 80 to 110, or 120 to 150 mg/dl) plasma ethanol levels. The effects of alcohol on hypothalamo-pituitary-adrenal (H-P-A) axis function were biphasic with respect to dosage (inhibition with low dosage and stimulation with higher dosages) and time (initial stimulation with higher dosages was followed by rapid return to control levels even though elevated plasma ethanol levels were maintained). The effects of alcohol on H-P-A activation were also inconsistent; some of the animals did not appear to respond even though elevated (i.e., >100 mg/dl) plasma ethanol levels were produced. Induction of moderate (80 to 110 mg/dl) plasma ethanol levels acutely (within 30 min) increased immunoreactive (i) β-endorphin concentrations in the ventral tegmental area of the brain; higher (120 to 150 mg/dl) plasma ethanol levels increased iβ-endorphin concentrations in both the ventral tegmental area and the nucleus accumbens, whereas iss-endorphin concentrations were not significantly altered in other brain areas. High (120 to 150 mg/dl) plasma ethanol levels also increased mediobasohypothalamic pro-opiomelanocortin (biosynthetic precursor of forebrain β-endorphin) mRNA concentrations at 3 and 6 hr after initiation of ethanol infusions. Results demonstrate that atraumatic induction of physiologically meaningful plasma alcohol levels by gastric ethanol infusion activates the forebrain opiomelanocortinergic opioid system and exerts complex effects on the interrelated H-P-A system, consistent with evidence that these systems may interact to mediate or modulate some responses to alcohol ingestion.

We describe fabrication methods and the characterisation and use of extracellular microelectrode arrays for the detection of action potentials from neurons in culture. The 100 microns2 platinised gold microelectrodes in the 64 electrode array detect the external current which flows during an action potential with S:N ratios of up to 500:1, giving a maximum recorded signal of several millivolts. The performance of these electrodes is enhanced if good sealing of the cells over the electrodes is obtained and further enhanced if the electrodes and the cells lie in a deep groove in the substratum. The electrodes can be used for both recording and stimulation of activity in cultured neurons and for recording from multiple sites on a single cell. The use of such electrodes to obtain recordings from invertebrate neurons is described. The particular advantages of these electrodes, their long term stability, non-invasive nature, high packing density, and utility in stimulation, are demonstrated.

The role of the cytoskeleton and cell attachments in the alignment of baby hamster kidney fibroblasts to ridge and groove substratum topography was investigated using confocal scanning microscopy. This was carried out with normal cells and cells treated with the cytoskeleton modifiers cytochalasin D, colcemid, and taxol. Actin was localised with fluorescent phalloidin. Tubulin, vinculin, and intracellular adhesion molecule-1 were visualised by indirect immunofluorescence. The spreading, elongation, and orientation of the cells after 24 h of culture in these conditions were measured on grooves of 5, 10, and 25 microns width and 0.5, 1, 2, and 5 microns depth. We have also observed events over the first 30 min of cell attachment. Five minutes after cell attachment, F-actin condensations were seen close to the intersection of groove wall and ridge top, that is, at a topographic discontinuity. The condensations were often at right angles to the groove edge and showed a periodicity of 0.6 microns. Vinculin arrangement at the early stages of cell spreading was similar to that of actin. Organisation of the microtubule system followed later, becoming obvious at about 30 min after cell plating. The Curtis and Clark theory (that cells react to topography primarily at lines of discontinuity in the substratum by actin nucleation) is supported by these results. The use of cytoskeletal poisons did not entirely abolish cell reaction to grooves. Colcemid increased cell spreading and reduced cell orientation and elongation. Cytochalasin D reduced cell spreading, orientation, and elongation. Taxol reduced cell elongation but did not affect cell spreading and orientation. We conclude that the aggregation of actin along groove/ridge boundaries is a primary driving event in determining fibroblast orientation on microgrooved substrata.

Background. Adenocarcinoma of the uterine cervix typically is an aggressive neoplasm with a propensity for early invasion and dissemination. Little data are available correlating histologic, histochemical, or immunocytochemical parameters with the biologic behavior of this neoplasm. Specifically, the implication of expression of estrogen and progesterone receptors in cervical adenocarcinoma is essentially undefined. Methods. An immunocytochemical hormone receptor assay using specific monoclonal antibodies against estrogen receptors (ER) and progesterone receptors (PgR) was used to study paraffin-embedded specimens from 54 patients with primary cervical adenocarcinomas. The specimens were evaluated for heterogeneity and intensity of staining. An attempt also was made to study the relationship between the pattern of hormone receptor expression and other established prognostic indicators. Results. In all patients, diffuse positive staining of benign endocervical epithelial or stromal cells was observed. Positive immunostaining was seen in the adenocarcinoma specimens of 16 of 54 (30%) patients for ER and 19 of 54 (35%) patients for PgR. Expression of ER and PgR statistically correlated with each other (P = 0.0001). Endocervical-type adenocarcinoma had the highest degree of staining for both ER and PgR. Clear cell carcinomas and intestinal-type carcinomas were negative for both receptors. Positivity for ER and PgR inversely correlated with histologic grade as defined by the International Federation of Gynecology and Obstetrics (FIGO). The relationship between hormone receptor expression and FIGO stage was not statistically significant. Survival was associated with clinical stage (P = 0.004) and with immunocytochemical status of ER expression (P = 0.032) and PgR expression (P = 0.009). Conclusions. This study of 54 specimens from patients with cervical adenocarcinoma suggests that positive expression of ER and PgR is associated with prolonged survival.

Erythrocyte sodium-lithium countertransport (SLC) activity, membrane fluidity, plasma trigly-ceride and cholesterol were measured in hyperlipidaemic patients and normal subjects. Fluidity was assessed by the fluorescence anisotropy (inversely related to fluidity) of the probes 1,6-diphenyl-1,3,5-hexatriene (DPH) and 1,4-trimethylammonium-3,5-hexatriene (TMA-DPH). In a second group of patients the maximum velocity (Vmax) and external sodium affinity constant (km) of SLC was also measured. In the first group of patients, SLC activity was increased compared with the controls (0.279 ± 0.019 vs. 0.213 ± 0.013, P= 0.006) as was membrane fluidity in the deep hydrophobic regions (DPH anisotropy 0.211 ± 0.0007 vs. 0.215 ± 0.0011, P= 0.007). There was a strong correlation between SLC and DPH anisotropy (Rs= -0.72, P= < 0.001) which was due to the correlation between Vmax and DPH anisotropy (Rs=-0.90, P= < 0.001). Increases in Vmax of SLC in hyperlipidaemic patients may be due to differences in lipid organisation in the deep hydrophobic regions of the membrane which may affect the turnover rate of the transporter.

Many amphibian species have experienced substantial population declines or have disappeared al- together during the last several decades at a number of amphibian survey sites in Central and South Amer- ica. Our study addresses the use of trends in solar UV-B radiation exposure (280-320 nm) at these sites over the last two decades, derived from the Total Ozone Mapping Spectrometer satellite data. It is intended to dem- onstrate a role for satellite observations in determining whether UV-B radiation is a contributing factor in amphibian declines. We used these data to calculate the daily erythemal (sunburning) UV-B, or UV-B ery , expo- sure at the latitude, longitude, and elevation of each of 20 survey sites. The annually averaged UV-B ery dose, as well as the maximum values, have been increasing in both Central and South America, with higher levels reached at the Central American sites. The annually averaged UV-B ery exposure increased significantly from 1979-1998 at all 11 Central American sites we examined ( r 2 � 0.60-0.79; p � 0.015), with smaller but sig- nificant increases at five of the nine South American sites ( r 2 � 0.24-0.42; p � 0.05). The number of days having the highest UV-B exposure ( � 6.75 kJ/m 2 /day) increased in both regions from � 40 days per year to approximately 58 days per year in 1998 ( r 2 � 0.24-0.42; p � 0.001). In Central America, the contribution of these very high UV-B ery exposure levels to the annual UV-B ery total increased from approximately 5 to approx- imately 15% over the 19-year period, but actual daily exposures for each species are unknown. A UV-B ratio, the highest monthly UV-B exposure relative to the annual average for the highest UV-B category ( � 6.75 kJ/m 2 ), increased in both regions over this time period ( r 2 � 0.73; p � 0.001). This UV index was consistently higher for Central America, where species declines have been the most severe. These results should justify further re- search into whether UV-B ery radiation plays a role in amphibian population declines and extinctions. We dis- cuss synergy among UV-B radiation and other factors, especially those associated with alterations of water chemistry (e.g., acidification) in aqueous habitats.

Objective To establish the changes in erythrocyte sodium lithium countertransport (SLC) with advancing normal pregnancy and to determine if these changes were different in pregnancy induced hypertension (PIH). The changes in both groups were assessed in relation to haemodynamic changes. Design SLC, mean arterial pressure (MAP), cardiac output (CO) and total peripheral vascular resistance (TPVR) were determined serially during normal pregnancy and cross-sectionally in PIH. Women were studied again 20 weeks after delivery where possible. Setting Routine antenatal clinic and antenatal ward of a regional reference centre. Subjects Fifty-one normal primigravid women were studied serially and 41 primigravid women with PIH were studied at time of diagnosis. Results During normal pregnancy SLC (mmol Li/h/l cells) increased from a nonpregnant value of 0.24 ± 0.02 (mean ± SEM) to 0.32 ± 0.02 at 14 weeks, and 0.37 ± 0.02 at 20 weeks gestation. This was maintained until 38 weeks (0.40 ± 0.02). The increase until 20 weeks occurred at the time of greatest change in CO (5.10 ± 0.18 to 6.79 ± 0.20 l/min) and TPVR (1327 ± 58 to 969 ± 33 dyn/s/cm−5). The decrease in TPVR with a rise in SLC is opposite to the relation reported in essential hypertension so that a functional relation is unlikely. However, the changes within pregnancy were positively correlated (r = 0.43, P

The operating theory of the gas centrifuge is developed under both steady and unsteady state behavior. It is shown that countercurrent flow patterns in a gas centrifuge enhance the possible separation. The effect of self-diffusion of the gaseous mixtures is shown to be significant with low molecular weight gases but of less significance with heavy gases such as uranium isotopes. The time required to develop steady state pressure and concentration profiles is in the order of minutes, even with no net flow to and from the machine.

For situations in which capillary condensation of water occurs (e.g., within porous media), the thermodynamics can usually be studied by means of the Laplace and Young equations and a compressibility equation. We show here that the compressibility equation for the liquid does not reflect the criteria of equal gas and liquid phase potential changes at equilibrium, for departures from a specified reference state. Carman (1953) concluded that the capillary condensate can exist in a state of tension for large vapor-liquid pressure differentials and may exhibit physical properties that differ substantially from normal values. The familiar Kelvin equation is used to calculate the capillary curvature and is derived from the general Laplace and Young relationship. Melrose (1966) has obtained this relationship by matching the coefficients of the internal free energy and hydrostatic balances of the system shown in Figure 1.

The emulsion polymerization of styrene initiated by gamma rays was studied using competitive particle growth and conventional techniques. With gamma rays, the rate of freeradical generation is independent of temperature, and the absorption of energy is uniform throughout the system. From the competitive growth experiments, the particle growth rates were similar to those of persulfate ioninitiated systems and different from those of benzoyl peroxide- initiated systems. From the conventional experiments, the number of particles increased approxidately as the square of the emulsifier concentration for both gamma snd persulfate ion-initiated polymerizations. With gamma rays, the number of particles decreased with increasing temperature; the opposite was observed for persulfate. This may be explained by the relative rates of free- radical generation of each system; for gamma rays this rate is independent of temperature, while for persulfate, the activation energy for initiator decomposition is greater than that for polymerization propagation. For the gamma ray system, the calculated values of k/sub p/ increased with increasing particle size; the activation energy was 7.2 ( sigma = 0.59) kcal/mole. The calculated values of k/sub t/ at 50 deg C were 10/sup 4/ to 10/sup 5/ l/mole/ sec. (auth)