Activity and expression of progesterone metabolizing 5α-reductase, 20α-hydroxysteroid oxidoreductase and 3α(β)-hydroxysteroid oxidoreductases in tumorigenic (MCF-7, MDA-MB-231, T-47D) and nontumorigenic (MCF-10A) human breast cancer cells

BACKGROUND: Recent observations indicate that human tumorous breast tissue metabolizes progesterone differently than nontumorous breast tissue. Specifically, 5α-reduced metabolites (5α-pregnanes, shown to stimulate cell proliferation and detachment) are produced at a significantly higher rate in tumorous tissue, indicating increased 5α-reductase (5αR) activity. Conversely, the activities of 3α-hydroxysteroid oxidoreductase (3α-HSO) and 20α-HSO enzymes appeared to be higher in normal tissues. The elevated conversion to 5α-pregnanes occurred regardless of estrogen (ER) or progesterone (PR) receptor levels. To gain insight into these differences, the activities and expression of these progesterone converting enzymes were investigated in a nontumorigenic cell line, MCF-10A (ER- and PR-negative), and the three tumorigenic cell lines, MDA-MB-231 (ER- and PR-negative), MCF-7 and T-47D (ER- and PR-positive). METHODS: For the enzyme activity studies, either whole cells were incubated with [(14)C]progesterone for 2, 4, 8, and 24 hours, or the microsomal/cytosolic fraction was incubated for 15–60 minutes with [(3)H]progesterone, and the metabolites were identified and quantified. Semi-quantitative RT-PCR was employed to determine the relative levels of expression of 5αR type1 (SRD5A1), 5αR type 2 (SRD5A2), 20α-HSO (AKR1C1), 3α-HSO type 2 (AKR1C3), 3α-HSO type 3 (AKR1C2) and 3β-HSO (HSD3B1/HSD3B2) in the four cell lines using 18S rRNA as an internal control. RESULTS: The relative 5α-reductase activity, when considered as a ratio of 5α-pregnanes/4-pregnenes, was 4.21 (± 0.49) for MCF-7 cells, 6.24 (± 1.14) for MDA-MB-231 cells, 4.62 (± 0.43) for T-47D cells and 0.65 (± 0.07) for MCF-10A cells, constituting approximately 6.5-fold, 9.6-fold and 7.1 fold higher conversion to 5α-pregnanes in the tumorigenic cells, respectively, than in the nontumorigenic MCF-10A cells. Conversely, the 20α-HSO and 3α-HSO activities were significantly higher (p < 0.001) in MCF-10A cells than in the other three cell types. In the MCF-10A cells, 20α-HSO activity was 8-14-fold higher and the 3α-HSO activity was 2.5-5.4-fold higher than in the other three cell types. The values of 5αR:20α-HSO ratios were 16.9 – 32.6-fold greater and the 5αR:3α-HSO ratios were 5.2 – 10.5-fold greater in MCF-7, MDA-MB-231 and T-47D cells than in MCF-10A cells. RT-PCR showed significantly higher expression of 5αR1 (p < 0.001), and lower expression of 20α-HSO (p < 0.001), 3α-HSO2 (p < 0.001), 3α-HSO3 (p < 0.001) in MCF-7, MDA-MB-231 and T-47D cells than in MCF-10A cells. CONCLUSION: The findings provide the first evidence that the 5αR activity (leading to the conversion of progesterone to the cancer promoting 5α-pregnanes) is significantly higher in the tumorigenic MCF-7, MDA-MB-231 and T-47D breast cell lines than in the nontumorigenic MCF-10A cell line. The higher 5αR activity coincides with significantly greater expression of 5αR1. On the other hand, the activities of 20α-HSO and 3α-HSO are higher in the MCF-10A cells than in MCF-7, MDA-MB-231 and T-47D cells; these differences in activity correlate with significantly higher expression of 20α-HSO, 3α-HSO2 and 3α-HSO3 in MCF-10A cells. Changes in progesterone metabolizing enzyme expression (resulting in enzyme activity changes) may be responsible for stimulating breast cancer by increased production of tumor-promoting 5α-pregnanes and decreased production of anti-cancer 20α – and 3α-4-pregnenes

Robust isothermal electric switching of interface magnetization: A route to voltage-controlled spintronics

Roughness-insensitive and electrically controllable magnetization at the (0001) surface of antiferromagnetic chromia is observed using magnetometry and spin-resolved photoemission measurements and explained by the interplay of surface termination and magnetic ordering. Further, this surface in placed in proximity with a ferromagnetic Co/Pd multilayer film. Exchange coupling across the interface between chromia and Co/Pd induces an electrically controllable exchange bias in the Co/Pd film, which enables a reversible isothermal (at room temperature) shift of the global magnetic hysteresis loop of the Co/Pd film along the magnetic field axis between negative and positive values. These results reveal the potential of magnetoelectric chromia for spintronic applications requiring non-volatile electric control of magnetization.Comment: Single PDF file: 27 pages, 6 figures; version of 12/30/09; submitted to Nature Material

Reproducibility of the airway response to an exercise protocol standardized for intensity, duration, and inspired air conditions, in subjects with symptoms suggestive of asthma

<p>Abstract</p> <p>Background</p> <p>Exercise testing to aid diagnosis of exercise-induced bronchoconstriction (EIB) is commonly performed. Reproducibility of the airway response to a standardized exercise protocol has not been reported in subjects being evaluated with mild symptoms suggestive of asthma but without a definite diagnosis. This study examined reproducibility of % fall in FEV₁and area under the FEV₁time curve for 30 minutes in response to two exercise tests performed with the same intensity and duration of exercise, and inspired air conditions.</p> <p>Methods</p> <p>Subjects with mild symptoms of asthma exercised twice within approximately 4 days by running for 8 minutes on a motorized treadmill breathing dry air at an intensity to induce a heart rate between 80-90% predicted maximum; reproducibility of the airway response was expressed as the 95% probability interval.</p> <p>Results</p> <p>Of 373 subjects challenged twice 161 were positive (≥10% fall FEV₁on at least one challenge). The EIB was mild and 77% of subjects had <15% fall on both challenges. Agreement between results was 76.1% with 56.8% (212) negative (< 10% fall FEV₁) and 19.3% (72) positive on both challenges. The remaining 23.9% of subjects had only one positive test. The 95% probability interval for reproducibility of the % fall in FEV₁and AUC_0-30min was ± 9.7% and ± 251% for all 278 adults and ± 13.4% and ± 279% for all 95 children. The 95% probability interval for reproducibility of % fall in FEV₁and AUC_{0-30 min}for the 72 subjects with two tests ≥10% fall FEV₁was ± 14.6% and ± 373% and for the 34 subjects with two tests ≥15% fall FEV₁it was ± 12.2% and ± 411%. Heart rate and estimated ventilation achieved were not significantly different either on the two test days or when one test result was positive and one was negative.</p> <p>Conclusions</p> <p>Under standardized, well controlled conditions for exercise challenge, the majority of subjects with mild symptoms of asthma demonstrated agreement in test results. Performing two tests may need to be considered when using exercise to exclude or diagnose EIB, when prescribing prophylactic treatment to prevent EIB and when designing protocols for clinical trials.</p

Expression of progesterone metabolizing enzyme genes (AKR1C1, AKR1C2, AKR1C3, SRD5A1, SRD5A2) is altered in human breast carcinoma

BACKGROUND: Recent evidence suggests that progesterone metabolites play important roles in regulating breast cancer. Previous studies have shown that tumorous tissues have higher 5α-reductase (5αR) and lower 3α-hydroxysteroid oxidoreductase (3α-HSO) and 20α-HSO activities. The resulting higher levels of 5α-reduced progesterone metabolites such as 5α-pregnane-3,20-dione (5αP) in tumorous tissue promote cell proliferation and detachment, whereas the 4-pregnene metabolites, 4-pregnen-3α-ol-20-one (3αHP) and 4-pregnen-20α-ol-3-one (20αDHP), more prominent in normal tissue, have the opposite (anti-cancer-like) effects. The aim of this study was to determine if the differences in enzyme activities between tumorous and nontumorous breast tissues are associated with differences in progesterone metabolizing enzyme gene expression. METHODS: Semi-quantitative RT-PCR was used to compare relative expression (as a ratio of 18S rRNA) of 5αR type 1 (SRD5A1), 5αR type 2 (SRD5A2), 3α-HSO type 2 (AKR1C3), 3α-HSO type 3 (AKR1C2) and 20α-HSO (AKR1C1) mRNAs in paired (tumorous and nontumorous) breast tissues from 11 patients, and unpaired tumor tissues from 17 patients and normal tissues from 10 reduction mammoplasty samples. RESULTS: Expression of 5αR1 and 5αR2 in 11/11 patients was higher (mean of 4.9- and 3.5-fold, respectively; p < 0.001) in the tumor as compared to the paired normal tissues. Conversely, expression of 3α-HSO2, 3α-HSO3 and 20α-HSO was higher (2.8-, 3.9- and 4.4-fold, respectively; p < 0.001) in normal than in tumor sample. The mean tumor:normal expression ratios for 5αR1 and 5αR2 were about 35–85-fold higher than the tumor:normal expression ratios for the HSOs. Similarly, in the unmatched samples, the tumor:normal ratios for 5αR were significantly higher than the ratios for the HSOs. CONCLUSIONS: The study shows changes in progesterone metabolizing enzyme gene expression in human breast carcinoma. Expression of SRD5A1 (5αR1) and SRD5A2 (5αR2) is elevated, and expression of AKR1C1 (20α-HSO), AKR1C2 (3α-HSO3) and AKR1C3 (3α-HSO2) is reduced in tumorous as compared to normal breast tissue. The changes in progesterone metabolizing enzyme expression levels help to explain the increases in mitogen/metastasis inducing 5αP and decreases in mitogen/metastasis inhibiting 3αHP progesterone metabolites found in breast tumor tissues. Understanding what causes these changes in expression could help in designing protocols to prevent or reverse the changes in progesterone metabolism associated with breast cancer

Fixation strength of biocomposite wedge interference screw in ACL reconstruction: effect of screw length and tunnel/screw ratio. A controlled laboratory study

<p>Abstract</p> <p>Background</p> <p>Primary stability of the graft is essential in anterior cruciate ligament surgery. An optimal method of fixation should be easy to insert and provide great resistance against pull-out forces.</p> <p>A controlled laboratory study was designed to test the primary stability of ACL tendinous grafts in the tibial tunnel. The correlation between resistance to traction forces and the cross-section and length of the screw was studied.</p> <p>Methods</p> <p>The tibial phase of ACL reconstruction was performed in forty porcine tibias using digital flexor tendons of the same animal. An 8 mm tunnel was drilled in each specimen and two looped tendons placed as graft. Specimens were divided in five groups according to the diameter and length of the screw used for fixation. Wedge interference screws were used. Longitudinal traction was applied to the graft with a Servohydraulic Fatigue System. Load and displacement were controlled and analyzed.</p> <p>Results</p> <p>The mean loads to failure for each group were 295,44 N (Group 1; 9 × 23 screw), 564,05 N (Group 2; 9 × 28), 614,95 N (Group 3; 9 × 35), 651,14 N (Group 4; 10 × 28) and 664,99 (Group 5; 10 × 35). No slippage of the graft was observed in groups 3, 4 and 5. There were significant differences in the load to failure among groups (ANOVA/P < 0.001).</p> <p>Conclusions</p> <p>Longer and wider interference screws provide better fixation in tibial ACL graft fixation. Short screws (23 mm) do not achieve optimal fixation and should be implanted only with special requirements.</p

Autism as a disorder of neural information processing: directions for research and targets for therapy

The broad variation in phenotypes and severities within autism spectrum disorders suggests the involvement of multiple predisposing factors, interacting in complex ways with normal developmental courses and gradients. Identification of these factors, and the common developmental path into which theyfeed, is hampered bythe large degrees of convergence from causal factors to altered brain development, and divergence from abnormal brain development into altered cognition and behaviour. Genetic, neurochemical, neuroimaging and behavioural findings on autism, as well as studies of normal development and of genetic syndromes that share symptoms with autism, offer hypotheses as to the nature of causal factors and their possible effects on the structure and dynamics of neural systems. Such alterations in neural properties may in turn perturb activity-dependent development, giving rise to a complex behavioural syndrome many steps removed from the root causes. Animal models based on genetic, neurochemical, neurophysiological, and behavioural manipulations offer the possibility of exploring these developmental processes in detail, as do human studies addressing endophenotypes beyond the diagnosis itself

Plasma protein C levels in immunocompromised septic patients are significantly lower than immunocompetent septic patients: a prospective cohort study

Introduction: Activated Protein C [APC] improves outcome in immunocompetent patients with severe sepsis particularly in those who are perceived to have high mortality risk. Before embarking on a trial of APC administration in immunocompromised septic patients, a preliminary study on plasma levels of protein C in this cohort is essential