Species-specific cell-matrix interactions are essential for differentiation of alveoli like structures and milk gene expression in primary mammary cells of the Cape fur seal (Arctocephalus pusillus pusillus)

Few models are in place for analysis of extreme lactation patterns such as that of the fur seals which are capable of extended down regulation of milk production in the absence of involution. During a 10–12 month lactation period, female fur seals suckle pups on shore for 2–3 days, and then undertake long foraging trips at sea for up to 28 days, resulting in the longest intersuckling bouts recorded. During this time the mammary gland down regulates milk production. We have induced Cape fur seal (Arctocephalus pusillus pusillus) mammary cells in vitro to form mammospheres up to 900 μm in diameter, larger than any of their mammalian counterparts. Mammosphere lumens were shown to form via apoptosis and cells comprising the cellular boundary stained vimentin positive. The Cape fur seal GAPDH gene was cloned and used in RT-PCR as a normalization tool to examine comparative expression of milk protein genes (αS2-casein, β-lactoglobulin and lysozyme C) which were prolactin responsive. Cape fur seal mammary cells were found to be unique; they did not require Matrigel for rapid mammosphere formation and instead deposited their own matrix within 2 days of culture. When grown on Matrigel, cells exhibited branching/stellate morphogenesis highlighting the species-specific nature of cell–matrix interactions during morphological differentiation. Matrix produced in vitro by cells did not support formation of human breast cancer cell line, PMC42 mammospheres. This novel model system will help define the molecular pathways controlling the regulation of milk protein expression and species specific requirements of the extracellular matrix in the cape fur seal.

Flower like micellar assemblies in poly(styrene)-block-poly(4-vinylpyridine)/poly(acrylic acid) complexes

The formation of rare flower like micelles in poly(styrene)-block-poly(4-vinyl pyridine)/poly(acrylic acid) (PS-b-P4VP/PAA) diblock copolymer/homopolymer complexes is reported. The self-assembly as well as the morphological changes in the complexes were induced by the addition of a high molecular weight PAA/ethanol solution into the PS-b-P4VP solution in dimethyl formamide followed by dialyses. The composition-dependent micelles were varying in size and shape with increase in PAA concentration in solution. The complex aggregates in solution were characterized by dynamic light scattering (DLS) whereas morphologies in the solid complexes were observed using transmission electron microscopy (TEM). Flower like micelles are formed in complexes at 20 wt% PAA concentration followed by 'spikey' micellar assemblies at 40 wt% PAA. The size of the micelles was found to be increased upon the addition of PAA into the block copolymer solution. Infrared studies revealed the intermolecular hydrogen bonding interactions between the complementary binding sites on PAA and the P4VP block of the block copolymer. Finally, a model was proposed to explain the self-assembly and morphological transitions in these complexes based on the experimental results obtained.

Soft-sediment deformation structures interpreted as seismite from the Middle Permian of the southern Sydney Basin, southeastern Australia

The Middle Permian Wandrawandian Siltstone of the southern Sydney Basin is well exposed along the coastline from Lagoon Head in the south to North Head in the north near Ulladulla in southern New South Wales. The unit is dominated by fossiliferous siltstone and mudstone, with abundant dropstones and minor pebbly sandstone interbeds, and contains an interval of well-preserved and extensive soft-sediment deformation structures. These deformation structures occur mainly in the middle part of the cliff sections and are bounded above and below by undeformed sedimentary units of similar lithology. A wide range of soft-sediment deformation structures have been observed, including cracks, sandstone and sandy mudstone dykes, a possible sand volcano, networks of relatively small and closely connected fissure-like structures, metre-scale complex-type slump folds, flexural stratification, concave-up depressional structures, small-scale normal faults (with displacements usually <1 m), shear planes, and breccias (pseudonodules). The slumps and associated deformations are here collectively interpreted as representing a seismite deposit attributable to penecontemporaneous deformation of soft, hydroplastic sediment layers following a liquefaction triggered by seismic shocks. The timing of the inferred earthquake events appears to correspond to the onset of a major basin-wide tectonism during the Middle Permian.

Trapping of a spiral-like intermediate of the bacterial cytokinetic protein FtsZ

The earliest stage in bacterial cell division is the formation of a ring, composed of the tubulin-like protein FtsZ, at the division site. Tight spatial and temporal regulation of Z-ring formation is required to ensure that division occurs precisely at midcell between two replicated chromosomes. However, the mechanism of Z-ring formation and its regulation in vivo remain unresolved. Here we identify the defect of an interesting temperature-sensitive ftsZ mutant (ts1) of Bacillus subtilis. At the nonpermissive temperature, the mutant protein, FtsZ(Ts1), assembles into spiral-like structures between chromosomes. When shifted back down to the permissive temperature, functional Z rings form and division resumes. Our observations support a model in which Z-ring formation at the division site arises from reorganization of a long cytoskeletal spiral form of FtsZ and suggest that the FtsZ(Ts1) protein is captured as a shorter spiral-forming intermediate that is unable to complete this reorganization step. The ts1 mutant is likely to be very valuable in revealing how FtsZ assembles into a ring and how this occurs precisely at the division site.

Taxonomy and biology of two new species of gall midge (Diptera: Cecidomyiidae) infesting Sarcocornia quinqueflora (Chenopodiaceae) in Australian salt marshes

Two new species of gall midge associated with two distinct galls on the succulent creeping shrub Sarcocornia quinqueflora are described from salt marshes in south-eastern Australia. The infestations caused by the new species hinder the growth of S. quinqueflora, the seeds of which are the major food of the critically endangered orange-bellied parrot Neophema chrysogaster. Asphondylia floriformis sp. n. Veenstra-Quah & Kolesik transforms leaf segments into flower-like galls, whereas Asphondylia sarcocorniae sp. n. Veenstra-Quah & Kolesik produces simple swellings on branches. Both galls have fungal mycelium growing in the apoplast of the gall tissue and lining the inner surface of the larval chamber where it is presumably grazed by the larva. Descriptions of the larvae, pupae, males, females and the geographical distribution of the two gall midges in south-eastern Australia are given.

In vitro bioactivity evaluation of titanium and niobium metals with different surface morphologies

Current orthopaedic biomaterials research mainly focuses on designing implants that could induce controlled, guided and rapid healing. In the present study, the surface morphologies of titanium (Ti) and niobium (Nb) metals were tailored to form nanoporous, nanoplate and nanofibre-like structures through adjustment of the temperature in the alkali-heat treatment. The in vitro bioactivity of these structures was then evaluated by soaking the treated samples in simulated body fluid (SBF). It was found that the morphology of the modified surface significantly influenced the apatite-inducing ability. The Ti surface with a nanofibre-like structure showed better apatite-inducing ability than the nanoporous or nanoplate surface structures. A thick dense apatite layer formed on the Ti surface with nanofibre-like structure after 1 week of soaking in SBF. It is expected that the nanofibre-like surface could achieve good apatite formation in vivo and subsequently enhance osteoblast cell adhesion and bone formation.

Neurotoxic, redox-competent Alzheimer's --amyloid is released from lipid membrane by methionine oxidation

The amyloid β peptide is toxic to neurons, and it is believed that this toxicity plays a central role in the progression of Alzheimer's disease. The mechanism of this toxicity is contentious. Here we report that an Aβ peptide with the sulfur atom of Met-35 oxidized to a sulfoxide (Met(O)Aβ) is toxic to neuronal cells, and this toxicity is attenuated by the metal chelator clioquinol and completely rescued by catalase implicating the same toxicity mechanism as reduced Aβ. However, unlike the unoxidized peptide, Met(O)Aβ is unable to penetrate lipid membranes to form ion channel-like structures, and β-sheet formation is inhibited, phenomena that are central to some theories for Aβ toxicity. Our results show that, like the unoxidized peptide, Met(O)Aβ will coordinate Cu₂₊ and reduce the oxidation state of the metal and still produce H2O2. We hypothesize that Met(O)Aβ production contributes to the elevation of soluble Aβ seen in the brain in Alzheimer's disease.

The raft-promoting property of virion-associated cholesterol, but not the presence of virion-associated brij 98 rafts, is a determinant of human immunodeficiency virus type 1 infectivity

Lipid rafts are enriched in cholesterol and sphingomyelin and are isolated on the basis of insolubility in detergents, such as Brij 98 and Triton X-100. Recent work by Holm et al. has shown that rafts insoluble in Brig 98 can be found in human immunodeficiency virus type 1 (HIV-1) virus-like particles, although it is not known whether raft-like structures are present in authentic HIV-1 and it is unclear whether a virion-associated raft-like structure is required for HIV replication. Independently, it was previously reported that virion-associated cholesterol is critical for HIV-1 infectivity, although the specific requirement of virion cholesterol in HIV-1 was not examined. In the present study, we have demonstrated that infectious wild-type HIV-1 contains Brij 98 rafts but only minimal amounts of Triton X-100 rafts. To directly assess the functional requirement of virion-associated rafts and various features of cholesterol on HIV-1 replication, we replaced virion cholesterol with exogenous cholesterol analogues that have demonstrated either raft-promoting or -inhibiting capacity in model membranes. We observed that variable concentrations of exogenous analogues are required to replace a defined amount of virion-associated cholesterol, showing that structurally diverse cholesterol analogues have various affinities toward HIV-1. We found that replacement of 50% of virion cholesterol with these exogenous cholesterol analogues did not eliminate the presence of Brij 98 rafts in HIV-1. However, the infectivity levels of the lipid-modified HIV-1s directly correlate with the raft-promoting capacities of these cholesterol analogues. Our data provide the first direct assessment of virion-associated Brij 98 rafts in retroviral replication and illustrate the importance of the raft-promoting property of virion-associated cholesterol in HIV-1 replication.

Inquisition of Microcystis aeruginosa and Synechocystis nanowires: characterization and modelling

Identification of extracellular conductive pilus-like structures (PLS) i.e. microbial nanowires has spurred great interest among scientists due to their potential applications in the fields of biogeochemistry, bioelectronics, bioremediation etc. Using conductive atomic force microscopy, we identified microbial nanowires in Microcystis aeruginosa PCC 7806 which is an aerobic, photosynthetic microorganism. We also confirmed the earlier finding that Synechocystis sp. PCC 6803 produces microbial nanowires. In contrast to the use of highly instrumented continuous flow reactors for Synechocystis reported earlier, we identified simple and optimum culture conditions which allow increased production of nanowires in both test cyanobacteria. Production of these nanowires in Synechocystis and Microcystis were found to be sensitive to the availability of carbon source and light intensity. These structures seem to be proteinaceous in nature and their diameter was found to be 4.5-7 and 8.5-11 nm in Synechocystis and M. aeruginosa, respectively. Characterization of Synechocystis nanowires by transmission electron microscopy and biochemical techniques confirmed that they are type IV pili (TFP) while nanowires in M. aeruginosa were found to be similar to an unnamed protein (GenBank : CAO90693.1). Modelling studies of the Synechocystis TFP subunit i.e. PilA1 indicated that strategically placed aromatic amino acids may be involved in electron transfer through these nanowires. This study identifies PLS from Microcystis which can act as nanowires and supports the earlier hypothesis that microbial nanowires are widespread in nature and play diverse roles.

Influence of fill gap on flexural strength of parts fabricated by curved layer fused deposition modeling

Rapid Prototyping Techniques (RPT) have evolved over the last decade. Novel RP techniques are being developed to improve the overall properties of parts manufactured using RPT. One such technique is the Curved layer fused deposition modeling (CLFDM) which has been developed based on the conventional Fused Deposition Modeling (FDM) technique. The CLFDM technique has gained significant amount of attention as a result of its advantages such as increased flexural strength, reduction of the stair-stepping effect and the reduction in the number of layers, especially for thin shell-like structures. This paper studies the effects of fill gap (FG) on flexural strength and bead dimension, middle-plane cross section profiles and the fracture surface and compares the results to parts made using the traditional planar layer-by-layer approach. Also, in the end some meaningful and interesting future study areas both in hardware design and software development for the CLFDM are proposed.

A comparative study of guided wave propagation in timber poles with isotropic and transversely isotropic material models

Guided wave (GW) has been used for many years in non-destructive testing (NDT). There are various ways to generate the guided wave, including impact or impulse either manually or using devices. Although the method of impact or impulse is considered to be simple and practical in guided wave generation, it produces waves with broadband frequencies, which often make analysis much more difficult. The frequency bandwidth produced by manual impacts is usually at the low end, and is therefore justified when dealing with one dimensional wave propagation assumption in low strain integrity testing of cylindrical structures. Under such assumption if the velocity is known accurately, NDTs can produce reasonably good results for the condition assessment of the structure. However, for guided wave propagation in timber pole-like structures, it is rather complicated as timber is an orthotropic material and wave propagation in an orthotropic medium exhibits different characteristics from that in isotropic medium. It is possible to obtain solutions for guided wave propagation in orthotropic media for cylindrical structures, even though the orthotropic material greatly complicates GW propagation. In this paper, timber has been considered as a transversely isotropic (i.e. simplified orthotropic) material and a comparative study of GW propagation in a timber pole is conducted considering isotropic and transversely isotropic modelling. Phase velocity, group velocity and attenuation are the main parameters for this comparative study. Moreover, tractionfree situation and embedded geotechnical condition are also taken into consideration to evaluate the effect of boundary. Displacement profile, wave propagation pattern and power flow at particular frequency are utilized to determine different displacement components of longitudinal and flexural waves along and across the timber pole. Effect of temperature and moisture content (in terms of modulus of elasticity) in timber pole is also compared to show the variation in phase velocity.

Controllable drug uptake and nongenomic response through estrogen-anchored cyclodextrin drug complex

Breast cancer is a leading killer of women worldwide. Cyclodextrin-based estrogen receptor-targeting drug-delivery systems represent a promising direction in cancer therapy but have rarely been investigated. To seek new targeting therapies for membrane estrogen receptor-positive breast cancer, an estrogen-anchored cyclodextrin encapsulating a doxorubicin derivative Ada-DOX (CDE1-Ada-DOX) has been synthesized and evaluated in human breast cancer MCF-7 cells. First, we synthesized estrone-conjugated cyclodextrin (CDE1), which formed the complex CDE1-Ada-DOX via molecular recognition with the derivative adamantane-doxorubicin (Ada-DOX) (Kd =1,617 M(-1)). The structure of the targeting vector CDE1 was fully characterized using (1)H- and (13)C-nuclear magnetic resonance, mass spectrometry, and electron microscopy. CDE1-Ada-DOX showed two-phase drug-release kinetics with much slower release than Ada-DOX. The fluorescence polarization analysis reveals that CDE1-Ada-DOX binds to recombinant human estrogen receptor α fragments with a Kd of 0.027 µM. Competition assay of the drug complex with estrogen ligands demonstrated that estrone and tamoxifen competed with CDE1-Ada-DOX for membrane estrogen receptor binding in MCF-7 cells. Intermolecular self-assembly of CDE1 molecules were observed, showing tail-in-bucket and wire-like structures confirmed by transmission electronic microscopy. CDE1-Ada-DOX had an unexpected lower drug uptake (when the host-guest ratio was >1) than non-targeting drugs in MCF-7 cells due to ensconced ligands in cyclodextrins cavities resulting from the intermolecular self-assembly. The uptake of CDE1-Ada-DOX was significantly increased when the host-guest ratio was adjusted to be less than half at the concentration of CDE1 over 5 µM due to the release of the estrone residues. CDE1 elicited rapid activation of mitogen-activated protein kinases (p44/42 MAPK, Erk1/2) in minutes through phosphorylation of Thr202/Tyr204 in MCF-7 cells. These results demonstrate a targeted therapeutics delivery of CDE1-Ada-DOX to breast cancer cells in a controlled manner and that the drug vector CDE1 can potentially be employed as a molecular tool to differentiate nongenomic from genomic mechanism.

Upper lower triassic stromatolite from Anhui, South China: geobiologic features and paleoenvironmental implications

Widespread stromatolites and other microbialite deposits characterize Lower Triassic marine successions worldwide. This study documents a stromatolite deposit, 1.1. m thick, from the upper Spathian (Lower Triassic) of the Susong area, South China. The stromatolite comprises distinct laminated domes in the basal part and columns at the upper part. Dark laminae are loosely spaced and interlayered with thicker light colored laminae. Diffusive dark colored laminae are composed of peloidal micrite that grade locally into microclotted structures, and yield copies of bacteria clump-like and coccoid-like spheroid aggregates. The former are characterized by cloudy, micrite nuclei rimmed by coarse-grained, euhedral sparry calcite crystals, while the latter are comprised of solid calcite crystal nuclei coated with rather thin micrite envelopes. The cloudy, micrite nuclei resemble bacteria clump-like structures observed in present-day travertine. Both the coccoid-like spheroids and bacteria clump-like structures are surrounded by coarse-grained euhedral calcite crystals, suggesting a similar accretion mechanism. Both spherical structures therefore could be crucial in the accretion of the Susong stromatolite. The laminated/microclotted structures are interpreted as the result of variation in timing of lithification relative to the timing of the decay of microbes. Micro-analysis also unravels the common occurrence of authigenic micro-quartz crystals in association with Fe-bearing illite clay minerals in the stromatolite columns. Their coalescing nature with each other, together with the associated pyrite grains, strongly support the formation of micro-quartz crystals from microbial reduction of an Fe-bearing smectite precursor by sulfate reducing bacteria. A comparison of the Susong stromatolite with its counterparts from the upper Lower Triassic strata in Dajiang, South China reveals many similarities in stromatolite microstructures, suggesting that a harsh, euxinic-anoxic environment resulting in the bloom of sulfate reducing bacteria most likely extended into the latest Spathian in South China.