A facile method for urinary phenylalanine measurement on paper-based lab-on-chip for PKU therapy monitoring.

A miniaturized paper-based lab-on-chip (LoC) was developed for the facile measurement of urinary Phe (phenylalanine) level on PKU (Phenylketonuria) treated patient. This system permits the monitoring of Phe in a dynamic range concentration of 20-3000 µM.

An innovative chemical strategy for PCR-free genetic detection of pathogens by an integrated electrochemical biosensor.

An innovative chemical strategy integrated in a miniaturized electrochemical device was developed for sensitive detection of a pathogen genome (HBV virus) without any amplification step. The results show a limit of detection comparable to the standard qRT-PCR method (20 copies per reaction), paving the way to future development of genetic PoC devices addressing automatized and low-cost molecular diagnostics.

Constitutively expressed c-myb abrogates the requirement for insulinlike growth factor 1 in 3T3 fibroblasts.

The proto-oncogene c-myb, whose expression is usually limited to cells of the hematopoietic lineages, can be expressed in fibroblasts if placed under the control of a constitutive promoter, such as the simian virus SV40 early promoter. 3T3 cells carrying a constitutively expressed human c-myb were found to grow in 1% serum or in a serum-free, platelet-derived growth factor-supplemented medium, whereas the parent cell line, BALB/c 3T3, needed insulinlike growth factor 1 (IGF-1) in addition to platelet-derived growth factor for growth. myb-carrying cells, however, could not grow in platelet-poor plasma. In fibroblasts, therefore, a constitutively expressed c-myb can abrogate the requirement for platelet-poor plasma or IGF-1. When 3T3 cells constitutively expressed both c-myc and c-myb, they could grow in serum-free medium without added growth factors. The ability of c-myb to abrogate in fibroblasts the IGF-1 requirement seems to be due to its ability to induce overexpression of IGF-1, as indicated by an increase in steady-state levels of IGF-1 mRNA. These results have some important implications; for instance, they suggest a commonality of pathways for entry into S phase in different cell types and the possibility of a myb-like or myb-equivalent gene product of critical importance for entry of fibroblasts into S phase.

Ionic strength-controlled hybridization and stability of hybrids of KRAS DNA single-nucleotides: A surface plasmon resonance study.

The discrimination of a fully matched, unlabeled KRAS wild-type (WT) (C-G) target sample with respect to three of the most frequent KRAS codon mutations (G12 S (C-A), G12 R (C-C), G12C (C-T)) was investigated using an optimized detection strategy involving surface plasmon resonance (SPR), based on optimized probe-surface density and ionic strength control. The changes observed in the SPR signal were always larger for WT compared with the single-mismatch target DNA oligonucleotides, and were aligned with the theoretical energy differences between the base pair C-G, C-T, C-A, C-C. Hybridization rates of ∼106M-1s-1 were detected without the introduction of high temperature and labels, usually needed in conventional hybridization methods. One hundred percent mutation discrimination of the matched KRAS wild-type (C-G) sequence with respect to three mismatched G12C (C-T), G12 S (C-A), G12 R (C-C) target sequences was achieved.

3alpha-hydroxy-5alpha-pregnan-20-one in the midbrain ventral tegmental area mediates social, sexual, and affective behaviors.

Progestins mediate the onset and duration of lordosis, the mating posture of female rodents, through actions in the hypothalamus and ventral tegmental area. In the hypothalamus, progesterone has traditional, "genomic" actions via intracellular progestin receptors. In the ventral tegmental area, 3alpha-hydroxy-5alpha-pregnan-20-one has "non-genomic" actions independent of progestin receptors to facilitate lordosis that involve GABA(A)/benzodiazepine receptors, NMDA type glutamate receptors, and/or dopamine receptors. 3alpha-Hydroxy-5alpha-pregnan-20-one levels also change with behavioral and/or environmental stimuli and may have a role in other reproductively-relevant behaviors, such as affiliation, exploration, and anxiety (socio-sexual behaviors). Data are reviewed that support the notion that: 1) effects of 3alpha-hydroxy-5alpha-pregnan-20-one in the midbrain ventral tegmental area facilitate lordosis and other reproductively-relevant behaviors. 2) 3alpha-Hydroxy-5alpha-pregnan-20-one, formed in the ventral tegmental area from metabolism of progestins, produced peripherally by endocrine glands, or centrally from biosynthesis in glial cells mediates socio-sexual behaviors. 3) 3alpha-Hydroxy-5alpha-pregnan-20-one's actions at GABA(A)/benzodiazepine receptors, NMDA type glutamate receptors, and dopamine receptors in the ventral tegmental area are important for lordosis; however, effects at these substrates on socio-sexual behaviors have not been elucidated. Given 3alpha-hydroxy-5alpha-pregnan-20-one's involvement in stress responses, its putative role as a homeostatic regulator and in the pathophysiology and treatment of neuropsychiatric disorders is discussed.

In the ventral tegmental area, cyclic AMP mediates the actions of progesterone at dopamine type 1 receptors for lordosis of rats and hamsters.

Progesterone-facilitated lordosis is enhanced by activation of, and inhibited by antagonism of, dopamine type 1 receptors (D1) in the ventral tegmental area (VTA). Given that D1 activation leads to increases in cyclic AMP (cAMP), we hypothesised that, in the VTA, progesterone's actions on lordosis that involve D1 are mediated, in part, by cAMP. In Experiment 1, naturally receptive rats and hamsters were pretested for lordosis, infused with the cAMP analogue 8-bromo-cAMP (200 ng) or vehicle to the VTA, and tested again 30 min later. In Experiments 2 and 3, ovariectomised, oestradiol (10 microg) + progesterone (0 or 100 microg)-primed rats and oestradiol (10 microg) + progesterone (0 or 200 microg)-primed hamsters were pretested for lordosis and infused with 8-bromo-cAMP (200 ng), the adenylyl cyclase inhibitor 2',5'-dideoxyadenosine (12 microM) or vehicle to the VTA. Subjects were tested again 30 min later. In Experiment 4, oestradiol + progesterone-primed rats and hamsters were pretested and infused with the D1 agonist SKF38393 (0 or 100 ng) to the VTA. Thirty minutes later, subjects were tested again and infused with 2',5'-dideoxyadenosine (12 microM) or vehicle. Subjects were tested again 30 min later. VTA infusions of 8-bromo-cAMP enhanced lordosis of naturally receptive or hormone-primed rats and hamsters and 2',5'-dideoxyadenosine decreased lordosis of oestradiol + progesterone-primed rats and hamsters. D1-mediated increases in progesterone-facilitated lordosis were reduced by 2',5'-dideoxyadenosine. These data suggest that progesterone-facilitated lordosis of rats and hamsters may be modulated by D1 and cAMP activity in the VTA.

Increase in cellular RNA in cells infected with DNA oncogenic viruses.

The amount of RNA per cell has been measured by flow microfluorometry in cultured cells stimulated by serum or infected with DNA oncogenic viruses (SV40, polyoma virus, and adenovirus). While all four agents stimulate, although to a different extent, cellular DNA synthesis in quiescent cells, the accumulation of cellular RNA varies. Serum, SV40, and polyoma virus cause a marked increase in total cellular RNA that is already apparent in G1 cells before entry into S. On the other hand, adenovirus 2, while capable of stimulating cellular DNA synthesis, fails to detectably increase the amount of RNA per cell. These results suggest that adenovirus 2 may act on quiescent cells through mechanisms different from those of serum, SV40, or polyoma virus.

Inhibiting biosynthesis and/or metabolism of progestins in the ventral tegmental area attenuates lordosis of rats in behavioural oestrus.

In the ventral tegmental area (VTA), lordosis of rats is facilitated by 5alpha-pregnan-3alpha-ol-20-one (3alpha,5alpha-THP). Central 3alpha,5alpha-THP results from metabolism of peripheral progesterone, from the ovaries and/or adrenals, by sequential enzymatic activity of 5alpha-reductase and 3alpha-hydroxysteroid oxidoreductase (3alpha-HSOR). In addition, in glial cells, cholesterol is converted into pregnenolone by the P450 side-chain cleavage enzyme (P450scc), which is then metabolized to progesterone by 3beta-hydroxysteroid dehydrogenase, and subsequently reduced to 3alpha,5alpha-THP. We hypothesize that, in the VTA, formation of 3alpha,5alpha-THP by both metabolism and biosynthesis is necessary for facilitation of lordosis of female rats. In Experiment 1, naturally-receptive rats received bilateral VTA infusions of a P450scc inhibitor, digitoxin (1 microg/side); a 5alpha-reductase inhibitor, finasteride (10 microg/side); digitoxin (1 microg/side)+finasteride (10 microg/side); or vehicle and were tested 3 h later for lordosis. In Experiment 2, the effects of VTA infusions of digitoxin, finasteride, digitoxin+finasteride, or vehicle on lordosis and midbrain and plasma 3alpha,5alpha-THP levels were examined. In Experiment 3, we investigated whether infusions of 3alpha,5alpha-THP to the VTA reinstated lordosis and midbrain 3alpha,5alpha-THP levels following administration of inhibitors. VTA infusions of digitoxin, finasteride, or digitoxin+finasteride, significantly and similarly reduced lordosis and midbrain, but not plasma 3alpha,5alpha-THP levels, compared to vehicle. Following receipt of inhibitor infusions, 3alpha,5alpha-THP to the VTA restored lordosis and midbrain 3alpha,5alpha-THP levels. These data suggest that, in the VTA, both central biosynthesis of progesterone and metabolism of progesterone (from central and/or peripheral sources) to 3alpha,5alpha-THP are important for mediating lordosis of rats.

Mitochondrial benzodiazepine receptors in the ventral tegmental area modulate sexual behaviour of cycling or hormone-primed hamsters.

Hamsters are highly dependent upon the central actions of progesterone (P4) for facilitation of sexual behaviour. In the ventral tegmental area (VTA), P4 has actions through its neurosteroid metabolite 5alpha-pregnan-3alpha-ol-20-one (3alpha,5alpha-THP). The effects of enhancing or inhibiting neurosteroidogenesis (and thereby 3alpha,5alpha-THP concentrations), through manipulations of mitochondrial benzodiazepine receptors, in the VTA on socio-sexual behaviour of female hamsters were examined. Intact, naturally receptive hamsters and ovariectomized (OVX), hormone-primed hamsters were unilaterally infused via chronic guide cannula to the VTA with the mitochondrial benzodiazepine receptor antagonist 1-(2-chlorophenyl)-N-methyl-N-(1-methylpropyl)-3-isoquinolinecarboximide (PK-11195) or the mitochondrial benzodiazepine receptor agonist, N,N-dihexyl-2-(4-fluorophenyl)indole-30-acetamide (FGIN 1-27) and tested for sexual responsiveness and lordosis. PK-11195 (5.6, 11.2 or 22.4 nm) to the VTA attenuated sexual responsiveness of naturally receptive or oestradiol benzoate (EB) + P4-primed hamsters compared to vehicle. In addition, FGIN 1-27 (11.4 nm) infusions to the VTA increased sexual responsiveness and lordosis of cycling or OVX, EB + P4-primed hamsters, compared to vehicle infusions. In OVX, EB + P4-primed hamsters, decrements in sexual responsiveness produced by VTA infusions of PK-11195 (5.6 nm) were attenuated by VTA infusions of 3alpha,5alpha-THP. VTA infusions of PK-11195 (5.6 nm) or FGIN 1-27 (11.4 nm), respectively, decreased and increased midbrain levels of 3alpha,5alpha-THP compared to each other. Together, these findings indicate that manipulating actions of mitochondrial benzodiazepine receptors in the VTA can augment and inhibit neurosteroidogenesis and sexual responsiveness of hormone-primed and naturally receptive hamsters.

The photochemistry of flutamide and its inclusion complex with beta-cyclodextrin. Dramatic effect of the microenvironment on the nature and on the efficiency of the photodegradation pathways.

The photochemistry of the anticancer drug flutamide (FM), 2-methyl-N-[4-nitro-3-(trifluoromethyl)phenyl]propanamide, in homogeneous media and in the beta-cyclodextrin (beta-CD) cavity has been investigated. The photoreactivity of the free molecule has been rationalized on the basis of an intramolecular nitro to nitrite rearrangement followed by cleavage of the nitrite intermediate. The twisted geometry of the nitro group with respect to the aromatic plane plays a key role in triggering such a photoprocess. Incorporation of FM in the beta-CD cavity leads to dramatic effects on both the efficiency and the nature of the photochemical deactivation pathways of the guest molecule. A 20-fold increase in the FM photodecomposition quantum yield and the formation of photoproducts originated by both reduction of the nitro group and cleavage of the amide bond were observed in the presence of the macrocycle. Such a behavior cannot be attributed exclusively to the micropolarity of beta-CD and/or to its role as a reactant. The induced circular dichroism spectra and the nature of the photoproducts formed in these experimental conditions provide indications that the photoreactivity in the beta-CD microenvironment could likely be mediated by structural changes of FM upon complexation.

Estrous cycle and sex differences in performance on anxiety tasks coincide with increases in hippocampal progesterone and 3alpha,5alpha-THP.

Sex differences and estrous cycle variations in anxiolytic-like behaviors and progestin concentrations were examined. Proestrous (n=22), estrous (n=19), diestrous (n=20), and male (n=18) Long-Evans rats were tested in horizontal crossing, open field, elevated plus-maze, emergence, holeboard, social interaction, tailflick, pawlick, and defensive burying tasks. Concentrations of plasma and hippocampal progesterone and 5alpha-pregnan-3alpha-ol-20-one (3alpha,5alpha-THP) were measured by radioimmunoassay in behaviorally tested (proestrus n=11, estrus n=8, diestrus n=9, male n=7) and yoked non-tested rats (proestrus n=11, estrus n=8, diestrus n=10, male n=8). Proestrous females exhibited more anxiolytic-like behavior than all other groups on the elevated plus-maze, social interaction, and defensive burying tasks. Proestrous females had significantly shorter latencies to emerge from a cylinder than did estrous and diestrous females, but not males. Proestrous and estrous females entered significantly more peripheral and total squares in a brightly-lit open field than did males. While proestrous females had a tendency to make more beam breaks than did males in the horizontal crossing task, there were no differences between groups on the holeboard task. There was a tendency for proestrous females to have longer tailflick latencies than diestrous and male rats; however, on the pawlick task there were no differences among the groups. Plasma and central progesterone and 3alpha,5alpha-THP of tested and non-tested rats were not different. Proestrous females had significantly higher plasma and hippocampal progesterone and 3alpha,5alpha-THP levels than all other groups. These data demonstrate that proestrous increases in anxiolytic-like behavior coincide with elevated circulating and hippocampal progestin concentrations.

Women in the public sector: cancer mortality.

Very few mortality occupational studies of female workers have been published, even though the number of women in the workforce has increased dramatically to reach more than 45%. Public sector employees comprise 12% of the female workforce. This article reports the findings of a retrospective cohort mortality study of 1371 full-time female employees of the city of Buffalo, New York, who were employed between January 1, 1950, and October 1, 1979, and have worked a minimum of 5 years. The majority of women were hired in the 1940s and early 1950s and began employment after age 30. This predominantly white-collar, service-oriented female cohort demonstrated significantly lower all-cause mortality than that expected based on US mortality rates for white females. The highest observed numbers of cancer deaths were for breast, digestive, and respiratory cancers. The majority of person-years were accrued in clerical and service occupations. Higher-than-expected mortality was shown for reproductive and respiratory cancers among professional employees; digestive cancers, diseases of the nervous system, and pneumonia among clerical workers; and cancers of the lung and brain and diseases of the digestive system, especially ulcers, among service employees. Because these results are based on few observed cases, they must be interpreted cautiously.

Environmental risk factors of breast cancer.

Breast cancer is women's most ubiquitous cancer. The role of dietary factors is controversial, but there is limited evidence for such occupational risk factors as employment in the pharmaceutical industry and as a beautician. Ionizing radiation probably increases the risk. Exposure to chlorinated hydrocarbon pesticides, chlorinated solvents, and polychlorinated biphenyls may be risk factors, although the evidence is insufficient. Data on low-frequency electromagnetic fields are inconclusive. Tobacco smoking may be a risk factor, but the effect may depend on N-acetyltransferase 2 genetic polymorphisms. There are yet unidentified determinants, probably environmental, that may act via estrogenic activity or through other mechanisms. The etiology may vary according to the joint estrogen and progesterone receptor status of the tumor. P53 mutation frequency varies considerably in breast cancer populations, which may reflect variation in exogenous exposures. Epidemiology research on breast cancer needs to consider subtypes of the disease, lifetime exposure assessment, host susceptibility, and adjustment for reproductive and menstrual history.

Risk of premenopausal breast cancer in association with occupational exposure to polycyclic aromatic hydrocarbons and benzene.

OBJECTIVES: This study examined the relationship between risk of premenopausal breast cancer and occupational exposure to benzene and polycyclic aromatic hydrocarbons (PAH) and whether the proposed relationship between PAH and breast cancer differed by tumor estrogen receptor (ER) status. METHODS: In a case-referent study of premenopausal breast cancer, occupational histories and other information were obtained through interviews, and job-exposure matrices were used to assess exposure to PAH and benzene. RESULTS: A dose-response relationship for the probability of exposure to benzene [low: odds ratio (OR) 1.64, 95% confidence interval (95% CI) 0.64-4.21; high: OR 1.95, 95% CI 1.14-3.33) and to PAH (low: OR 1.56, 95% CI 0.78-3.12; high: OR 2.40, 95% CI 0.96-6.01). Risk increased with duration of exposure to benzene, but not to PAH. A dose-response relationship was not evident for the intensity of exposure to benzene or to PAH. When analyses were stratified by tumor ER status, PAH exposure was related to a greater increase in the risk of ER-positive (OR 2.27, 95% CI 1.14-4.54) than ER-negative (OR 1.12, 95% CI 0.47-2.64) breast cancer. Risk of ER-positive, but not ER-negative, tumors increased with the probability of exposure to PAH. CONCLUSIONS: The findings suggest an association between risk and occupational exposure to benzene. Although it was difficult to study PAH independently of benzene, there was some suggestion of an association between PAH exposure and ER-positive tumors. These data should be interpreted with caution because of the limitations of this study, including low-response rates and small numbers of exposed persons.

A comparative evaluation of police suicide rate validity.

The authors assessed sensitivity, specificity, and predictive value of official police suicide rates and compared them to municipal workers. Deaths officially classified as suicide, accidental, and undetermined were submitted to a panel of medical examiners for validation. Six cases originally in the accident and undetermined rubric were reclassified as suicide. Official police suicide rates had less sensitivity (83.3% compared to 92.3%) of actual suicides than municipal worker rates. Police suicide rates also showed a lower negative proportion than municipal worker rates (86.2% compared to 98.7%). A generalizable sensitivity proportion equation for assessing suicide rates in other police groups is presented.

Expression of cell cycle-dependent genes and proliferative state of lymphocytes in aging.

The authors analyzed the expression of some genes involved in the control of T lymphocyte proliferation in a group of healthy elderly subjects. They focused their attention on genes involved in the G(0)/G(1) transition (TK, PCNA, H3, IL2-R) and showed decreased expression in the TK, H3 and IL2-R genes. Using flow cytofluorimetry, delayed transition from the G(0)/G(1) to the S stage was observed.

Electrochemical Switching of Chromogenic Monolayers Self-Assembled on Transparent Platinum Electrodes.

Transparent, ultrathin Pt electrodes permit the simultaneous electrochemical and spectroscopic investigation of self-assembled monolayers of electrochromic compounds. Voltage stimulations applied to the Pt substrate reversibly alter the redox state of the chemisorbed molecules and, hence, modulate the intensity of the light transmitted through the Pt/monolayer assembly.