Array comparative genomic hybridization analysis of genomic alterations in breast cancer subtypes

In this study, we performed high-resolution array comparative genomic hybridization with an array of 4153 bacterial artificial chromosome clones to assess copy number changes in 44 archival breast cancers. The tumors were flow sorted to exclude non-tumor DNA and increase our ability to detect gene copy number changes. In these tumors, losses were more frequent than gains, and gains in 1q and loss in 16q were the most frequent alterations. We compared gene copy number changes in the tumors based on histologic subtype and estrogen receptor (ER) status, i.e., ER-negative infiltrating ductal carcinoma, ER-positive infiltrating ductal carcinoma, and ER-positive infiltrating lobular carcinoma. We observed a consistent association between loss in regions of 5q and ER-negative infiltrating ductal carcinoma, as well as more frequent loss in 4p16, 8p23, 8p21, 10q25, and 17p11.2 in ER-negative infiltrating ductal carcinoma compared with ER-positive infiltrating ductal carcinoma (adjusted P values < or = 0.05). We also observed high-level amplifications in ER-negative infiltrating ductal carcinoma in regions of 8q24 and 17q12 encompassing the c-myc and c-erbB-2 genes and apparent homozygous deletions in 3p21, 5q33, 8p23, 8p21, 9q34, 16q24, and 19q13. ER-positive infiltrating ductal carcinoma showed a higher frequency of gain in 16p13 and loss in 16q21 than ER-negative infiltrating ductal carcinoma. Correlation analysis highlighted regions of change commonly seen together in ER-negative infiltrating ductal carcinoma. ER-positive infiltrating lobular carcinoma differed from ER-positive infiltrating ductal carcinoma in the frequency of gain in 1q and loss in 11q and showed high-level amplifications in 1q32, 8p23, 11q13, and 11q14. These results indicate that array comparative genomic hybridization can identify significant differences in the genomic alterations between subtypes of breast cancer.     

Ischemic stroke after using over the counter products containing ephedra

Dietary supplements containing Ephedra used for weight loss and physical performance enhancement such as "herbal ecstasy" are widely available, and it is estimated that at least 1% of the adult population have taken these products. Ephedra products including Ephedra alkaloids such as phenylpropanolamine or other ephedrine compounds are sold under different names such as Metabolife 356, Ripped Fuel, Thermadrene, and Shape-Fast Plus. Over 2 years, five patients with ischemic infarctions associated with use of Ephedra products were evaluated at Indiana University Hospital. Ephedrine, like other sympathomimetic agents, predisposes patients to both ischemic and hemorrhagic strokes. People who take over the counter Ephedra products that claim to boost weight loss, increase energy, or bolster physical performance are at risk of adverse events including ischemic and hemorrhagic strokes.     

Early and reversible neuropathology induced by tetracycline-regulated lentiviral overexpression of mutant huntingtin in rat striatum

The ability to overexpress full-length huntingtin or large fragments represents an important challenge to mimic Huntington's pathology and reproduce all stages of the disease in a time frame compatible with rodent life span. In the present study, tetracycline-regulated lentiviral vectors leading to high expression levels were used to accelerate the pathological process. Rats were simultaneously injected with vectors coding for the transactivator and wild type (WT) or mutated huntingtin (TRE-853-19Q/82Q) in the left and right striatum, respectively, and analyzed in the 'on' and 'off' conditions. Overexpression of TRE-853-19Q protein or residual expression of TRE-853-82Q in 'off' condition did not cause any significant neuronal pathology. Overexpressed TRE-853-82Q protein led to proteolytic release of N-terminal htt fragments, nuclear aggregation, and a striatal dysfunction as revealed by decrease of DARPP-32 staining but absence of NeuN down-regulation. The differential effect on the DARPP-32/NeuN neuronal staining was observed as early as 1 month after injection and maintained at 3 months. In contrast, expression of a shorter htt form (htt171-82Q) did not require processing prior formation of nuclear aggregates and caused decrease of both DARPP-32 and NeuN neuronal markers at one month post-injection suggesting that polyQ pathology may be dependent on protein context. Finally, the reversibility of the pathology was assessed. Huntingtin expression was turn 'on' for 1 month and then shut 'off' for 2 months. Recovery of DARPP-32 immunoreactivity and clearance of huntingtin aggregates were observed in animals treated with doxycycline. These results suggest that a tetracycline-regulated system may be particularly attractive to model Huntington's disease and induce early and reversible striatal neuropathology in vivo.     

Development of displacement binding and GTPgammaS scintillation proximity assays for the identification of antagonists of the micro-opioid receptor

This article describes the development of micro-opioid receptor (MOR) binding and GTPgammaS functional SPAs as improved screening tools for the identification of MOR antagonists. Opioid receptors are members of the seven-transmembrane G protein-coupled receptor (GPCR) family and are involved in the control of various aspects of human physiology, including pain, stress, reward, addiction, respiration, gastric motility, and pituitary hormone secretion. Activation of the MOR initiates intracellular signaling pathways leading to a reduction in intracellular cyclic AMP levels, inhibition of calcium channels, and activation of potassium channels resulting in a reduction of the excitability of neurons. Characterization of opioid receptor ligand binding has traditionally been accomplished through the use of low throughput filtration-based binding assays, whereas functional activity has been based upon cyclic AMP measurements or filtration-based GTPgammaS functional assays. This report describes the development of a MOR displacement binding SPA using the radiolabeled antagonist [(3)H]diprenorphine ((3)H-DPN). The assay was optimized using statistical experimental design and demonstrates the stability and robustness necessary for HTS. The assay was biased toward the identification of MOR antagonists through the addition of Na(+). Our assay conditions also minimized the phenomenon of ligand depletion, a problem commonly observed in low-volume assays using high receptor-expressing cell lines. The optimized procedure revealed (3)H-DPN affinity constants at the MOR that were consistent with results obtained using filtration methods (K(D) (SPA) = 1.89 +/- 0.24 nM, K(D) (filtration) = 1.88 +/- 0.35 nM). The binding SPA identified known opioid receptor modulators contained within the Library of Pharmacological Active Compounds (LOPAC) cassette, and the GTPgammaS scintillation proximity assay (SPA) was used to confirm the functional activity of the LOPAC antagonists acting at the MOR. Conversion of the ligand binding and GTPgammaS functional assays to a homogeneous SPA generated a simple assay with dramatically increased throughput. Data from the development and implementation of the displacement binding and GTPgammaS functional SPAs are presented.     

Extended access to amphetamine self-administration increases impulsive choice in a delay discounting task in rats

Background  

d-Amphetamine (AMPH) is a widely prescribed attention deficit hyperactivity disorder medication, but little is known about its effects on impulsive choice with escalated use.     

A narrative review on women and cardiac rehabilitation: Program adherence and preferences for alternative models of care

Despite the preponderance of evidence on the numerous benefits of CR, it remains largely under-utilized in women. The objective of this narrative review was to summarize and synthesize the literature on women and CR with regard to outcomes, adherence, and preferences for alternative models of CR. Studies of the effectiveness of CR have generally revealed no major differences between men and women. However, female-specific data are lacking on the effect of CR on mortality and morbidity. Research suggests that women and men may be equally likely to prefer home-based to hospital-based CR services. Women's preferences for and outcomes in, women-only CR are beginning to be uncovered. Discussing program model options with female cardiac patients and referring to preferred types may be the appropriate approach until further evidence is available.     

In vitro investigation of pig cells for resistance to human antibody‐mediated rejection

Although human complement‐dependent cytotoxicity (CDC) of α1,3‐galactosyltransferase gene‐knockout (GTKO) pig cells is significantly weaker than that of wild‐type (WT) cells, successful xenotransplantation will require pigs with multiple genetic modifications. Sera from healthy humans were tested by (i) flow cytometry for binding of IgM/IgG, and (ii) CDC assay against peripheral blood mononuclear cells and porcine aortic endothelial cells from five types of pig – WT, GTKO, GTKO transgenic for H‐transferase (GTKO/HT), WT transgenic for human complement regulatory protein CD46 (CD46) and GTKO/CD46. There was significantly higher mean IgM/IgG binding to WT and CD46 cells than to GTKO, GTKO/HT, and GTKO/CD46, but no difference between GTKO, GTKO/HT, and GTKO/CD46 cells. There was significantly higher mean CDC to WT than to GTKO, GTKO/HT, CD46, and GTKO/CD46 cells, but no difference between GTKO and GTKO/HT. Lysis of GTKO/CD46 cells was significantly lower than that of GTKO or CD46 cells. CD46 expression provided partial protection against serum from a baboon sensitized to a GTKO pig heart. GTKO/CD46 cells were significantly resistant to lysis by human serum and sensitized baboon serum. In conclusion, the greatest protection from CDC was obtained by the combination of an absence of Gal expression and the presence of CD46 expression, but the expression of HT appeared to offer no advantage over GTKO. Organs from GTKO/CD46 pigs are likely to be significantly less susceptible to CDC.