Theoretical and practical considerations for the measurement of P-glycoprotein function in acute myeloid leukemia

This paper summarizes experimental data and theoretical considerations, that are important for the measurement of P-glycoprotein (Pgp) function in acute myeloid leukemia (AML). The data are presented in subdivisions based on the techniques used, which will facilitate finding specific information. Based on our extensive experience with Pgp analysis, which includes radioactive assays, flow cytometry and fluorescence microscopy, we recommend a flow cytometry-based assay, that measures the effect of 2 microM PSC 833 on rhodamine 123 (R123) accumulation as the most practical and sensitive functional Pgp test. In combination with the flow cytometric measurement of Pgp using an antibody against an extracellular epitope (eg MRK16), this offers a sensitive and reproducible method for Pgp detection in AML, which is also rapid and practical. Furthermore, an R123 accumulation assay is specific for Pgp, because R123 is transported much less efficiently by the multidrug resistance protein (MRP) than by Pgp. Another probe of similar sensitivity and specificity is 3,3'-diethyloxacarbocyanine iodide. Alternatively, especially for the analysis of small numbers of cells (for example sorted subpopulations of leukemic cells), convenient and sensitive procedures are being developed by using DNA-binding Pgp substrates which remain fixed in the nuclei of the cells upon formaldehyde exposure for quantitative fluorescence laser scanning microscopy with image analysis. Less experimental data have been published to establish the optimal conditions for dual parameter flow cytometry (Pgp function, in eg Pgp+ or CD34+ cells). However, laboratories with flow cytometry experience will be able to implement this useful option to analyze subpopulations of cells.     

Dietary fats and lung cancer risk among women: the Missouri Women's Health Study (United States)

In a population-based case-control study of women in Missouri (United States), most of whom were smokers, we obtained information on adult diet to evaluate the effects of dietary fats in relation to lung cancer risk. All newly diagnosed, primary lung cancer cases among women 35 to 84 years of age reported to the Missouri Cancer Registry from 1 January 1993 to 31 January 1994 were invited to participate, as were population-based controls. The analysis focused on interviews obtained from 624 controls and 587 cases. In-person interviews were obtained from 99.0 percent of controls and 60.6 percent of cases. Age and energy-adjusted relative risks suggested a direct relation between risk of lung cancer and intake of dietary fats (e.g., total fat, saturated fat) and frequency of meat consumption. After adjusting for confounders, dietary fats were no longer associated with risk, but the adverse effect of frequent consumption of meat persisted. Risk was elevated about 90 percent (95 percent confidence interval = 1.2-3.0) among women in the highest quintile of red meat intake compared with those in the lowest quintile. Risk estimates associated with red meat consumption, however, were dependent on interview status; the effect was restricted to cases whose dietary information was provided by proxy. In summary, after adjusting for potential confounders and removing data obtained from proxy respondents, dietary fats and consumption of red meat were not associated with lung cancer risk among women in Missouri.     

Energy availability and mammary carcinogenesis: effects of calorie restriction and exercise

The purpose of this experiment was to compare the carcinogenic response in the mammary gland among groups of rats whose energy metabolism had been modulated by restricting dietary calories and/or by increasing energy expenditure via exercise. Female F344 rats (n = 132) were injected i.p. with 1-methyl-1-nitrosomethylurea (50 mg/kg at 50 and 57 days of age) and were randomized into one of four treatment groups: (i) unrestricted, sedentary; (ii) calorie-restricted, sedentary; (iii) unrestricted, exercised; (iv) calorie-restricted, exercised. The targeted level of calorie-restricted was 20% and exercise was achieved by treadmill-running (20 m/min at a 15% grade for 30 min, 5 days/week). During the 20.5 week study, rats were palpated twice a week for detection of mammary tumors and urine was collected for determination of 24-h cortical steroid excretion. At the end of the study, all mammary lesions were histologically classified. Carcass composition and carcass energy were determined. Mammary carcinogenesis was inhibited among calorie-restricted, sedentary rats compared with unrestricted, sedentary rats (79% inhibition, P < 0.001). No inhibition of carcinogenesis was observed among exercised rats (unrestricted or calorie-restricted) relative to the unrestricted, sedentary rats. Within the present experimental design, exercise had no effect on carcinogenesis despite significant reductions of carcass fat and carcass energy among both groups of rats that exercised. Cortical steroid level was significantly higher only in calorie-restricted, sedentary rats (P < 0.05). These results do not support the hypothesis that reductions of body weight gain, carcass fat or carcass energy are sufficient conditions to inhibit mammary carcinogenesis. The results do suggest that changes in urinary cortical steroid excretion may predict whether an energy-related intervention is likely to alter mammary carcinogenesis.     

Kennedy's disease: clinical and molecular study of two Italian families

Kennedy's disease, or spinal and bulbar muscular atrophy (SBMA), is a rare X-linked motoneuron disorder with variable signs of androgen insensitivity. It is associated with the expansion of a trinucleotide CAG repeat within the androgen receptor (AR) gene. We here report our clinical and molecular findings in two Italian families with Kennedy's disease. The increased size of the CAG repeat was demonstrated in four affected males and seven carrier females.     

Capillary instabilities in thin films

Very thin films, less than 100 nm-thick, are used in a variety of applications, including as catalysts and for thin film reactions to form patterned silicides in electronic devices. Because of their high surface to volume ratio, these very thin films are subject to cap-illary instability and can agglomerate well below their melting temperatures. In order to develop a general understanding of agglomeration in very thin films, we have studied initially continuous and patterned films of gold on fused silica substrates. Two in situ techniques were used to monitor agglomeration: 1) heating and video recording in a transmission electron microscope, and 2) measurement of the intensity of laser light transmitted through a sample heated in a furnace. Electron microscopy allowed inves-tigation of the role of the microstructure of the Au film and analysis of light transmis-sion during heating allowed determination of temperature-dependent and film-thick-ness-dependent agglomeration rates. These results will be described along with models for the agglomeration process.     

Prediction of stress and displacement fields in stress concentration regions by least squares asymptotic analysis of displacement data

This paper demonstrates that highly accurate predictions of the stress fields, including the peak stress of a stress concentration region, can be obtained easily by a least squares asymptotic analysis (LSAA) of even a relatively sparse set of displacement data from points in this zone lying sufficiently far from the boundary to avoid 'edge effects'.     

Angiogenesis promoted by vascular endothelial growth factor: regulation through alpha1beta1 and alpha2beta1 integrins

Vascular endothelial growth factor (VEGF), also known as vascular permeability factor, is a cytokine of central importance for the angiogenesis associated with cancers and other pathologies. Because angiogenesis often involves endothelial cell (EC) migration and proliferation within a collagen-rich extracellular matrix, we investigated the possibility that VEGF promotes neovascularization through regulation of collagen receptor expression. VEGF induced a 5- to 7-fold increase in dermal microvascular EC surface protein expression of two collagen receptors-the alpha1beta1 and alpha2beta1 integrins-through induction of mRNAs encoding the alpha1 and alpha2 subunits. In contrast, VEGF did not induce increased expression of the alpha3beta1 integrin, which also has been implicated in collagen binding. Integrin alpha1-blocking and alpha2-blocking antibodies (Ab) each partially inhibited attachment of microvascular EC to collagen I, and alpha1-blocking Ab also inhibited attachment to collagen IV and laminin-1. Induction of alpha1beta1 and alpha2beta1 expression by VEGF promoted cell spreading on collagen I gels which was abolished by a combination of alpha1-blocking and alpha2-blocking Abs. In vivo, a combination of alpha1-blocking and alpha2-blocking Abs markedly inhibited VEGF-driven angiogenesis; average cross-sectional area of individual new blood vessels was reduced 90% and average total new vascular area was reduced 82% without detectable effects on the pre-existing vasculature. These data indicate that induction of alpha1beta1 and alpha2beta1 expression by EC is an important mechanism by which VEGF promotes angiogenesis and that alpha1beta1 and alpha2beta1 antagonists may prove effective in inhibiting VEGF-driven angiogenesis in cancers and other important pathologies.