BRCA1 Attenuates Progesterone Effects on Proliferation and NFκB Activation in Normal Human Mammary Epithelial Cells.

Germline mutations in the breast cancer susceptibility gene BRCA1, encoding a tumor suppressor protein, greatly enhance the risk of breast and ovarian cancer. This tissue-specificity implicates the role of ovarian hormones. Indeed, BRCA1 has been demonstrated to regulate the signalling axis of the hormone, progesterone, and its receptor, the progesterone receptor (PR), and progesterone action has been implicated in BRCA1-related tumorigenesis. BRCA1 also plays important roles in oxidative stress and activating nuclear factor kappaB (NFκB) signalling pathways. Like wildtype BRCA1 function, PR signalling has also been shown to inhibit NFκB activation. Although PR and BRCA1 networks are known to interact, their interaction at the level of NFκB activation in the human breast is not understood. This study investigates the effect of reduced BRCA1 expression on proliferation and NFκB activation in human breast cells, and the impact of progesterone on these effects. The major findings are that: 1) Reduced BRCA1 levels inhibit cell growth in normal human mammary cells and breast cancer cells; 2) Reduced BRCA1 levels stimulated inflammatory targets and NFκB activity in normal human mammary cells; 3) Wildtype BRCA1 inhibited the pro-proliferative effects of progesterone in normal mammary epithelial cells, and; 4) Progesterone attenuated BRCA1-mediated NFκB activation in normal human mammary cells. These data have important implications for our understanding of progesterone action in BRCA1 mutation carriers, and how inhibition of this action may potentially delay tumorigenesis or impart a more favourable prognosis.

The Use of Recombinant Tissue Plasminogen Activator (rTPA) in The Treatment of Fibrinous Pleuropneumonia in Horses: 25 Cases (2007-2012).

BACKGROUND: Information about treatment protocols, adverse effects and outcomes with intrapleural recombinant tissue plasminogen activator (rTPA) use in horses with fibrinous pleuropneumonia is limited. HYPOTHESIS/OBJECTIVES: Describe factors that contribute to clinical response and survival of horses treated with rTPA intrapleurally. ANIMALS: Horses with bacterial pneumonia and fibrinous pleural effusion diagnosed by ultrasonography, that were treated with rTPA intrapleurally. METHODS: Retrospective multicenter case series from 2007-2012. Signalment, history, clinical and laboratory evaluation, treatment, and outcome obtained from medical records. Regression analysis used to identify associations between treatments and outcomes. RESULTS: Thirty three hemithoraces were treated in 25 horses, with 55 separate treatments. Recombinant tissue plasminogen activator (375-20,000 µg/hemithorax) was administered 1-4 times. Sonographically visible reduction in fibrin mat thickness, loculations, fluid depth, or some combination of these was seen in 32/49 (65%) treatments. Response to at least 1 treatment was seen in 17/20 (85%) horses with sonographic follow-up evaluation after every treatment. Earlier onset of rTPA treatment associated with increased survival odds. No association was found between cumulative rTPA dose or number of rTPA doses and survival, development of complications, duration of hospitalization or total charges. Clinical evidence of hypocoagulability or bleeding was not observed. Eighteen horses (72%) survived to discharge. CONCLUSIONS AND CLINICAL IMPORTANCE: Treatment with rTPA appeared safe and resulted in variable changes in fibrin quantity and organization within the pleural space. Recombinant tissue plasminogen activator could be a useful adjunct to standard treatment of fibrinous pleuropneumonia, but optimal case selection and dosing regimen remain to be elucidated.

Direct binding to antigen-coated beads refines the specificity and cross-reactivity of four monoclonal antibodies that recognize polymorphic epitopes of HLA class I molecules.

Monoclonal antibodies with specificity for human leukocyte antigen (HLA) class I determinants of HLA were originally characterized using serological assays in which the targets were cells expressing three to six HLA class I variants. Because of this complexity, the specificities of the antibodies were defined indirectly by correlation. Here we use a direct binding assay, in which the targets are synthetic beads coated with 1 of 111 HLA class I variants, representing the full range of HLA-A, -B and -C variation. We studied one monoclonal antibody with monomorphic specificity (W6/32) and four with polymorphic specificity (MA2.1, PA2.1, BB7.2 and BB7.1) and compared the results with those obtained previously. W6/32 reacted with all HLA class I variants. MA2.1 not only exhibits high specificity for HLA-A*02, -B*57 and -B*58, but also exhibited cross-reactivity with HLA-A*11 and -B*15:16. At low concentration (1 µg/ml), PA2.1 and BB7.2 were both specific for HLA-A*02 and -A*69, and at high concentration (50 µg/ml) exhibited significant cross-reactions with HLA-A*68, -A*23 and -A*24. BB7.1 exhibits specificity for HLA-B*07 and -B*42, as previously described, but reacts equally well with HLA-B*81, a rare allotype defined some 16 years after the description of BB7.1. The results obtained with cell-based and bead-based assays are consistent and, in combination with amino acid sequence comparison, increase understanding of the polymorphic epitopes recognized by the MA2.1, PA2.1, BB7.2 and BB7.1 antibodies. Comparison of two overlapping but distinctive bead sets from two sources gave similar results, but the overall levels of binding were significantly different. Several weaker reactions were observed with only one of the bead sets.

Progesterone and estrogen receptors segregate into different cell subpopulations in the normal human breast.

Progesterone is critical in normal breast development and its synthetic derivatives are emerging as major drivers of breast cancer risk. The recent demonstration that progesterone regulates the stem cell compartment in the murine mammary gland, despite the absence of progesterone receptor (PR) in mammary stem cells, highlights the fact that PR distribution in progenitor cell subsets in the human breast remains to be conclusively shown. By utilising two independent cell sorting strategies to fractionate cells into distinct subpopulations enriched for different cell lineage characteristics, we have demonstrated a consistent enrichment of PR transcripts, relative to estrogen receptor transcripts, in the bipotent progenitor subfraction in the normal human breast. We have also shown co-expression of both steroid hormone receptors with basal markers in a subset of human breast cells, and finally we have demonstrated that PR+ bipotent progenitor cells are estrogen-insensitive, and that estrogen regulates PR in mature luminal cells only.

Review: Immunogenetics of human placentation.

Natural killer (NK) cells are a population of lymphocytes that function in both immune defense and reproduction. Diversifying NK cell phenotype and function are interactions between NK cell receptors and major histocompatibility complex (MHC) class I ligands. As a consequence of strong and variable selection these ligand-receptor systems are polymorphic, rapidly evolving, and considerably species-specific. Counterparts to the human system of HLA class I ligands and killer cell immunoglobulin-like receptors (KIR) are present only in apes and Old World monkeys. HLA-C, the dominant ligand for human KIR and the only polymorphic HLA class I expressed by trophoblast, is further restricted to humans and great apes. Even then, the human system appears qualitatively different from that of chimpanzees, in that it has evolved a genetic balance between particular groups of receptors and ligands that favor reproductive success and other groups of receptors and ligands that have been correlated with disordered placentation. Human populations that have survived successive episodes of epidemic disease and population bottlenecks maintain a breadth of diversity for KIR and HLA class I, implying that loss of such diversity disfavors long-term survival of a human population.

Distribution of flunixin meglumine and firocoxib into aqueous humor of horses.

BACKGROUND: Nonsteroidal anti-inflammatory drugs (NSAIDs) are commonly used systemically for the treatment of inflammatory ocular disease in horses. However, little information exists regarding the ocular penetration of this class of drugs in the horse. OBJECTIVE: To determine the distribution of orally administered flunixin meglumine and firocoxib into the aqueous humor of horses. ANIMALS: Fifteen healthy adult horses with no evidence of ophthalmic disease. METHODS: Horses were randomly assigned to a control group and 2 treatment groups of equal sizes (n = 5). Horses assigned to the treatment groups received an NSAID (flunixin meglumine, 1.1 mg/kg PO q24h or firocoxib, 0.1 mg/kg PO q24h for 7 days). Horses in the control group received no medications. Concentrations of flunixin meglumine and firocoxib in serum and aqueous humor and prostaglandin (PG) E(2) in aqueous humor were determined on days 1, 3, and 5 and aqueous : serum ratios were calculated. RESULTS: Firocoxib penetrated the aqueous humor to a significantly greater extent than did flunixin meglumine at days 3 and 5. Aqueous : serum ratios were 3.59 ± 3.32 and 11.99 ± 4.62% for flunixin meglumine and firocoxib, respectively. Ocular PGE(2) concentrations showed no differences at any time point among study groups. CONCLUSIONS AND CLINICAL IMPORTANCE: Both flunixin meglumine and firocoxib penetrated into the aqueous humor of horses. This study suggests that orally administered firocoxib penetrates the aqueous humor better than orally administered flunixin meglumine at label dosages in the absence of ocular inflammation. Firocoxib should be considered for the treatment of inflammatory ophthalmic lesions in horses at risk for the development of adverse effects associated with nonselective NSAID administration.

Repetitive stimulation of the common peroneal nerve as a diagnostic aid for botulism in foals.

BACKGROUND: Botulism is a potentially fatal paralytic disorder for which definitive diagnosis is difficult. OBJECTIVES: To determine if repetitive stimulation of the common peroneal nerve will aid in the diagnosis of botulism in foals. ANIMALS: Four control and 3 affected foals. METHODS: Validation of the test in healthy foals for its comparison in foals with suspected botulism. Controls were anesthetized and affected foals were sedated to avoid risks of anesthesia. The common peroneal nerve was chosen for its superficial location and easy access. Stimulating electrodes were placed along the common peroneal nerve. For recording, the active and reference electrodes were positioned over the midpoint and distal end of the extensor digitorum longus muscle, respectively. Repeated supramaximal stimulation of the nerve was performed utilizing a range of frequencies (1-50 Hz). Data analysis consisted of measuring the amplitude and area under the curve for each M wave and converting these values into percentages of decrement or increment based on the comparison of subsequent potentials to the initial one (baseline) within each set. RESULTS: A decremental response was seen at all frequencies in control foals. Decremental responses also were observed in affected foals at low frequencies. An incremental response was seen in all affected foals at 50 Hz. CONCLUSIONS AND CLINICAL IMPORTANCE: Decreased baseline M wave amplitudes with incremental responses at high rates are supportive of botulism. Repetitive nerve stimulation is a safe, simple, fast, and noninvasive technique that can aid in the diagnosis of suspected botulism in foals.

Serum cardiac troponin I concentrations transiently increase in rats given rosiglitazone.

Rosiglitazone, a peroxisome proliferator-activated receptor γ (PPARγ) agonist of the thiazolidinedione class, is a major insulin-sensitizing drug widely used to treat type-2 diabetes. Rosiglitazone causes myocardial hypertrophy in rodents and increases the risk of cardiac events in man. To better characterize its cardiac effects, male Wistar rats were orally administered 0, 10 or 80 mg/kg/day rosiglitazone. Myocardial gene expression profiling, hematology, histopathology and clinical chemistry, including measurement of serum cardiac troponin (cTn) I concentration with the ultrasensitive assay, were evaluated after 6 and 24h and 7 and 14 days of dosing. Heart weight was increased 10% after 7 days and 16% after 14 days of dosing at 80 mg/kg/day in the absence of microscopic changes. At the transcriptomic level, the number of differentially expressed probes was small: it was most at 24h in rats given 80 mg/kg rosiglitazone with 356 differentially regulated probes (fold change >1.3 fold, p<0.05). Also, gene categories typically associated with myocardial damage were not over-represented. Most importantly, serum cTnI concentrations in 5/9 rats after 7 days of dosing at 80 mg/kg/day were above the upper limit of serum cTnI concentration. cTnI concentrations after 14 days of dosing were similar between rats given the vehicle and rosiglitazone at 80 mg/kg. This is the first study to detect increases of serum cTnI concentrations in rats administered rosiglitazone. In light of reported cardiac events in patients chronically dosed with PPARγ agonists, our results support serum cTnI concentrations as an early biomarker of cardiac liability.

The antiproliferative effects of progestins in T47D breast cancer cells are tempered by progestin induction of the ETS transcription factor Elf5.

Prolactin and progesterone act together to regulate mammary alveolar development, and both hormones have been implicated in breast cancer initiation and progression. Here we show that Elf5, a prolactin-induced ETS transcription factor that specifies the mammary secretory cell lineage, is also induced by progestins in breast cancer cells via a direct mechanism. To define the transcriptional response to progestin elicited via Elf5, we made an inducible Elf5 short hairpin-RNA knock-down model in T47D breast cancer cells and used it to prevent the progestin-induction of Elf5. Functional analysis of Affymetrix gene expression data using Gene Ontologies and Gene Set Enrichment Analysis showed enhancement of the progestin effects on cell cycle gene expression. Cell proliferation assays showed a more efficacious progestin-induced growth arrest when Elf5 was kept at baseline levels. These results showed that progestin induction of Elf5 expression tempered the antiproliferative effects of progestins in T47D cells, providing a further mechanistic link between prolactin and progestin in the regulation of mammary cell phenotype.

Priming of familiar and unfamiliar visual objects over delays in young and older adults.

Although priming of familiar stimuli is usually age invariant, little is known about how aging affects priming of preexperimentally unfamiliar stimuli. Therefore, this study investigated the effects of aging and encoding-to-test delays (0 min, 20 min, 90 min, and 1 week) on priming of unfamiliar objects in block-based priming paradigms. During the encoding phase, participants viewed pictures of novel objects (Experiments 1 and 2) or novel and familiar objects (Experiment 3) and judged their left-right orientation. In the test block, priming was measured using the possible-impossible object-decision test (Experiment 1), symmetric-asymmetric object-decision test (Experiment 2), and real-nonreal object-decision test (Experiment 3). In Experiments 1 and 2, young adults showed priming for unfamiliar objects at all delays, whereas older adults whose baseline task performance was similar to that of young adults did not show any priming. Experiment 3 found no effects of age or delay on priming of familiar objects; however, priming of unfamiliar objects was only observed in the young participants. This suggests that when older adults cannot rely on preexisting memory representations, age-related deficits in priming can emerge.

Bias effects in the possible/impossible object decision test with matching objects.

In the possible/impossible object decision test, priming has consistently been found for structurally possible, but not impossible, objects, leading Schacter, Cooper, and Delaney (1990) to suggest that priming relies on a system that represents the global 3-D structure of objects. Using a modified design with matching objects to control for the influence of episodic memory, Ratcliff and McKoon (1995) and Williams and Tarr (1997) found negative priming for impossible objects (i.e., lower performance for old than for new items). Both teams argued that priming derives from (1) episodic memory for object features and (2) bias to respond "possible" to encoded objects or their possible parts. The present study applied the matched-objects design to the original Schacter and Cooper stimuli-same possible objects and matching impossible figures-with minimal procedural variation. The data from Experiment 1 only partially supported the bias models and suggested that priming was mediated by both local and global structural descriptions. Experiment 2 showed that negative priming for impossible objects derived from the structural properties of these objects, not from the influence of episodic memory on task performance. Supplemental materials for this study may be downloaded from mc.psychonomic-journals.org/content/supplemental.

Aging does not affect brain patterns of repetition effects associated with perceptual priming of novel objects.

This study examined how aging affects the spatial patterns of repetition effects associated with perceptual priming of unfamiliar visual objects. Healthy young (n = 14) and elderly adults (n = 13) viewed four repetitions of structurally possible and impossible figures while being scanned with blood oxygenation level-dependent functional magnetic resonance imaging. Although explicit recognition memory for the figures was reduced in the elder subjects, repetition priming did not differ across the two age groups. Using multivariate linear modeling, we found that the spatial networks of regions that demonstrated repetition-related increases and decreases in activity were identical in both age groups, although there was a trend for smaller magnitude repetition effects in these networks in the elder adults for objects that had been repeated thrice. Furthermore, repetition-related reductions in activity in the left inferior frontal cortex for possible objects correlated with repetition-related facilitation in reaction time across both young and elder subjects. Repetition-related increases of an initially negative response were observed for both object types in both age groups in parts of the default network, suggesting that less attention was required for processing repeated stimuli. These findings extend prior studies using verbal and semantic picture priming tasks and support the view that perceptual repetition priming remains intact in later adulthood because the same spatial networks of regions continue to show repetition-related neural plasticity across the adult life span.

Global familiarity of visual stimuli affects repetition-related neural plasticity but not repetition priming.

In this study, we tested the prediction of the component process model of priming [Henson, R.N. (2003). Neuroimaging studies of priming. Prog Neurobiol, 70 (1), 53-81] that repetition priming of familiar and unfamiliar objects produces qualitatively different neural repetition effects. In an fMRI study, subjects viewed four repetitions of familiar objects and globally unfamiliar objects with familiar components. Reliable behavioral priming occurred for both item types across the four presentations and was of a similar magnitude for both stimulus types. The imaging data were analyzed using multivariate linear modeling, which permits explicit testing of the hypothesis that the repetition effects for familiar and unfamiliar objects are qualitatively different (i.e., non-scaled versions of one another). The results showed the presence of two qualitatively different latent spatial patterns of repetition effects from presentation 1 to presentation 4 for familiar and unfamiliar objects, indicating that familiarity with an object's global structural, semantic, or lexical features is an important factor in priming-related neural plasticity. The first latent spatial pattern strongly weighted regions with a similar repetition effect for both item types. The second pattern strongly weighted regions contributing a repetition suppression effect for the familiar objects and repetition enhancement for the unfamiliar objects, particularly the posterior insula, superior temporal gyrus, precentral gyrus, and cingulate cortex. This differential repetition effect might reflect the formation of novel memory representations for the unfamiliar items, which already exist for the familiar objects, consistent with the component process model of priming.

Microscopic morphology and the origins of the membrane maturation model of Golgi apparatus function.

The membrane maturation (flow differentiation) model of Golgi apparatus function embodies concepts of saccule formation at one face of the Golgi apparatus from membranes derived from endoplasmic reticulum and utilization of saccules in vesicle formation at the opposite face for delivery to the plasma membrane as existing saccules are displaced from one position within the stack to another. Derivation of the model came almost entirely from light and electron microscopy. Especially important were observations that passage through the Golgi apparatus was accompanied by differentiation of membranes from endoplasmic reticulum-like to plasma membrane-like across the polarity axis of the stacked saccules. The concept of coparticipation of endoplasmic reticulum and/or nuclear envelope, transition, and secretory vesicles and other pre- and post-Golgi apparatus structures through the operation of an integrated endomembrane system was essential to the model. Dynamic aspects confirmed initially by autoradiographing and cell fractionation studies have been corroborated in newer approaches of fluorescent labeling and with living cells.

An event-related fMRI study of the neural networks underlying repetition suppression and reaction time priming in implicit visual memory.

Unfamiliar line drawings were presented to subjects three times during BOLD fMRI scanning. A set of brain areas was detected in which the effect of stimulus repetition on the evoked fMRI response depended on whether or not the drawing could be conceived as a coherent three-dimensional structure. Differential repetition effects were found in the neural response to drawings of both structurally possible and impossible objects. This differential effect of repetition was related to the amount of reaction time priming on the concurrent task involving decisions about three-dimensional structure in the possible but not in the impossible objects. These results point to different neurophysiological processing mechanisms for structurally possible and impossible images and demonstrate neural plasticity that predicts behavioral priming for structurally possible images.

Sources of errors on visuoperceptual tasks: role of education, literacy, and search strategy.

The current study explored possible sources of demographic effects through analyses of errors from modified formats of the Benton Visual Retention Test (BVRT) completed by African American elders. Results indicate that: (1) reading level was a stronger predictor of BVRT performance than years of education; (2) on the single-item matching format of the task, individuals with lower reading levels disproportionately produced errors on items that differed in geometric, rather than spatial features; and (3) on a multiple-choice matching format, individuals with lower reading levels committed more errors on items where the target was located in the lower half of a 2 x 2 matrix.