Increased corticosterone levels in mice subjected to the rat exposure test

In recent years, there has been a notable interest in studying prey-predator relationships to develop rodent-based models for the neurobehavioral aspects of stress and emotion. However, despite the growing use of transgenic mice and results showing important differences in the behavioral responses of rats and mice, little research has been conducted regarding the responses of mice to predators. The rat exposure test (RET), a recently developed and behaviorally validated prey-predator (mouse-rat)-based model, has proven to be a useful tool in evaluating the defensive responses of mice facing rats. To further validate the RET, we investigated the endocrine and behavioral responses of mice exposed to this apparatus. We first constructed a plasma corticosterone secretion curve in mice exposed to a rat or to an empty cage (control). Rat-exposed mice showed a pronounced rise in corticosterone levels that peaked 15 min from the beginning of the predator exposure. The corticosterone levels and behavioral responses of mice exposed to a rat or to a toy in the RET apparatus were then measured. We observed high plasma corticosterone levels along with clear avoidance behaviors represented by decreases in tunnel and surface area exploration and increases in risk assessment behaviors and freezing. This strongly suggests that the test elicits a repertoire of behavioral responses compatible with an aversion state and indicates that it is a promising model for the evaluation of prey-predator interactions. However, more physiological, neurochemical, and pharmacological studies are needed to further validate the test.     

Manganese(II) complexes of di-2-pyridinylmethylene-1,2-diimine di-Schiff base ligands: Structures and reactivity

Manganese(II) complexes [Mn(L)X₂] were prepared and characterized, where L is a neutral di-Schiff base ligand incorporating pyridylimine donor arms, including (1R,2R)-N,N′-bis(2-pyridylmethylidene)-1,2-diphenylethylenediimine (L¹), (1R,2R)-N,N′-bis(6-methyl-2-pyridylmethylidene)-1,2-cyclohexyldiimine (L²), or (1R,2R)-, (1S,2S)- or racemic N,N′-bis(2-pyridylmethylidene)-1,2-cyclohexyldiimine (L³), and X⁻ =  or Cl⁻. Product complexes were structurally characterized, specifically including [Mn(R,R-L¹)(NCCH₃)₃](ClO₄)₂, [Mn(R,R-L²)(OH₂)₂](ClO₄)₂ and racemic [Mn(L³)Cl₂]. The first of these complexes features a heptacoordinate ligand field in a distorted pentagonal bipyramid, and the latter two are hexacoordinate, but retain equatorially monovacant pentagonal bipyramidal structures. Complexes [Mn(L³)X₂] (X⁻ = Cl⁻, ) were reacted with the primary phosphine FcCH₂PH₂ (Fc = -C₅H₄FeC₅H₅), H₂O and ethyldiazoacetate (EDA). The first two substrates prompted reactivity at a single ligand imine bond, resulting in hydrophosphination and hydrolysis, respectively. Complexes of the derivative ligands were also structurally characterized. Evidence for EDA activation was obtained by electrospray ionization mass spectrometry, but catalytic carbene transfer was not obtained.     

Generation of stable cell clones expressing mixtures of human antibodies

Therapeutic monoclonal antibodies, a highly successful class of biological drugs, are conventionally manufactured in mammalian cell lines. A recent approach to increase the therapeutic effectiveness of monoclonal antibodies has been to combine two or more of them; however this increases the complexity of development and manufacture. To address this issue a method to efficiently express multiple monoclonal antibodies from a single cell has been developed and we describe here the generation of stable cell clones that express high levels of a human monoclonal antibody mixture. PER.C6® cells were transfected with a combination of plasmids containing genes encoding three different antibodies. Clones that express the three corresponding antibody specificities were identified, subcloned, and passaged in the absence of antibiotic selection pressure. At several time points, batch production runs were analyzed for stable growth and IgG production characteristics. The majority (11/12) of subclones analyzed expressed all three antibody specificities in constant ratios with total IgG productivity ranging between 15 and 20 pg/cell/day under suboptimal culture conditions after up to 67 population doublings. The growth and IgG production characteristics of the stable clones reported here resemble those of single monoclonal antibody cell lines from conventional clone generation programs. We conclude that the methodology described here is applicable to the generation of stable PER.C6® clones for industrial scale production of mixtures of antibodies. Biotechnol. Bioeng. 2010;106: 741–750. © 2010 Wiley Periodicals, Inc.     

Structure–function relationships of the antigenicity of mycolic acids in tuberculosis patients

Cell wall mycolic acids (MA) from Mycobacterium tuberculosis (M.tb) are CD1b presented antigens that can be used to detect antibodies as surrogate markers of active TB, even in HIV coinfected patients. The use of the complex mixtures of natural MA is complicated by an apparent antibody cross-reactivity with cholesterol. Here firstly we report three recombinant monoclonal scFv antibody fragments in the chicken germ-line antibody repertoire, which demonstrate the possibilities for cross-reactivity: the first recognized both cholesterol and mycolic acids, the second mycolic acids but not cholesterol, and the third cholesterol but not mycolic acids. Secondly, MA structure is experimentally interrogated to try to understand the cross-reactivity. Unique synthetic mycolic acids representative of the three main functional classes show varying antigenicity against human TB patient sera, depending on the functional groups present and on their stereochemistry. Oxygenated (methoxy- and keto-) mycolic acid was found to be more antigenic than alpha-mycolic acids. Synthetic methoxy-mycolic acids were the most antigenic, one containing a trans-cyclopropane apparently being somewhat more antigenic than the natural mixture. Trans-cyclopropane-containing keto- and hydroxy-mycolic acids were also found to be the most antigenic among each of these classes. However, none of the individual synthetic mycolic acids significantly and reproducibly distinguished the pooled serum of TB positive patients from that of TB negative patients better than the natural mixture of MA. This argues against the potential to improve the specificity of serodiagnosis of TB with a defined single synthetic mycolic acid antigen from this set, although sensitivity may be facilitated by using a synthetic methoxy-mycolic acid.     

High temperature pulses decrease indirect chilling injury and elevate ATP levels in the flesh fly, Sarcophaga crassipalpis

Indirect chilling injury commonly occurs during long-term exposure to low temperature in many organisms including insects. A previous study revealed increased rates of survival and reduced cold injury in flesh flies, Sarcophaga crassipalpis, that experienced an intermittent pulse of high temperature during a low-temperature regiment. We extended these studies by determining survival rates and ATP levels for flies that had undergone continuous long-term exposure at 0 °C versus those experiencing a 24-h warming pulse of either 15 or 20 °C. Survival among flies that had undergone a warming pulse was significantly greater than for flies that were maintained continuously at 0 °C. Furthermore, ATP levels of flies that had experienced a warming pulse were significantly higher than those of flies maintained at 0 °C. These data suggest that brief warming pulses during long-term cold storage allow regeneration of energy reserves that promote survival and reduce indirect chilling injury.     

Chibby interacts with NBPF1 and clusterin, two candidate tumor suppressors linked to neuroblastoma

The NBPF genes are members of a gene family that underwent a remarkable increase in their copy number during recent primate evolution. The NBPF proteins contain 5 to 40 copies of a domain known as the NBPF repeat or DUF1220. Very little is known about the function of these domains or about the NBPF proteins. We performed a yeast two-hybrid screening with the aminoterminal domain of NBPF11 and found that Chibby, a documented repressor of Wnt signaling, interacts with multiple NBPF proteins. More specifically, a coiled-coil region in the NBPF proteins interacts with the coiled-coil domain in the carboxyterminal region of Chibby. Nonetheless, this interaction did not influence the repressor function of Chibby in a TOPFLASH reporter assay. Using Chibby as bait in a new yeast two-hybrid screening, we identified clusterin as a binding protein. Chibby and clusterin were co-immunoprecipitated with NBPF1, suggesting the formation of a tri-molecular complex. Although we have not pinpointed the role of these mutual interactions, the possible formation of a macromolecular complex of three candidate tumor suppressor proteins, including the enigmatic NBPF1, points at important functional implications.     

Fish–environment Associations in the Coastal Waters of Andros Island, The Bahamas

Synopsis We surveyed fish communities and corresponding environmental conditions at three broadly similar coastal sites of eastern Andros Island, The Bahamas over a summer–winter–summer sequence to assess the relationship between detailed environmental features and fish species patterns. Environmental variables included covers of various benthic flora components, benthic flora diversity, floral canopy height, micro-crustacean diversity and density, water temperature, extent of destructive land-use and extent of invasion by human-introduced exotic terrestrial plants. Correspondence analysis (CA) indicated that spatial (site) differences in environmental characteristics were greater than temporal (seasonal) differences. Detrended canonical correspondence analysis (DCCA) was used to assess the strength of relationships between the environmental characteristics and the distribution patterns of 25 fish species. Environmental features deemed to be most important in influencing fish species patterns included benthic flora canopy height, extent of invasion by exotic terrestrial plants, cover of Batophora oerstedii, cover of Thalassia testudinum, turf cover, water temperature, micro-crustacean diversity, and micro-crustacean density. Based upon similarities in distribution patterns, fish species formed four clusters which, ultimately, reflected similarities in species’ feeding habits and preferences for habitats that likely maximize foraging success. We conclude that fish distribution patterns are related to environmental characteristics, and that anthropogenic coastal activity, by influencing coastal benthic characteristics, may influence the distribution and abundances of fish species in coastal habitats.     

A dynamic spatial gradient of Hebbian learning in dendrites

Backpropagating action potentials (bAPs) are an important signal for associative synaptic plasticity in many neurons, but they often fail to fully invade distal dendrites. In this issue of Neuron, Sj?str?m and H?usser show that distal propagation failure leads to a spatial gradient of Hebbian plasticity in neocortical pyramidal cells. This gradient can be overcome by cooperative distal synaptic input, leading to fundamentally distinct Hebbian learning rules for distal versus proximal synapses.