Agonist-stimulated oscillations and cycling of intracellular free calcium in individual cultured muscle cells

Receptor regulation of [Ca2+]i was monitored in individual BC3H-1 muscle cells with intracellularly trapped fura-2 using digital imaging analysis techniques. Activation of alpha 1-adrenergic or H1-histaminergic receptors resulted in multiple bursts, or oscillations, of elevated [Ca2+]i with an average interval frequency of approximately 1.8 min-1. The duration of oscillatory behavior was generally more prolonged in response to phenylephrine than in response to histamine. Additionally, a larger fraction of the cells responded with [Ca2+]i oscillations to phenylephrine (approximately 90%) than to histamine (approximately 60%), although the majority of cells produced oscillations in response to both agonists. In most cells, the receptor-mediated [Ca2+]i oscillations continued for several minutes in the absence of extracellular Ca2+, although the amplitude of the individual peaks gradually decreased. The activation of [Ca2+]i oscillations by H1-receptors was more dependent upon extracellular Ca2+ than those elicited by alpha 1-receptors, reflecting the greater dependency of the histaminergic response on Ca2+ influx. Readdition of Ca2+ to the incubation buffer resulted in the resumption of the [Ca2+]i oscillations. These results indicate that considerable cycling of Ca2+ between the cytoplasm and the endoplasmic reticulum must occur. Receptor-mediated [Ca2+]i oscillations were much more prevalent in subconfluent cells than in confluent cells, possibly due to increased coupling of the cells at higher densities. The cells were capable of responding independently of one another, since sister cells displayed unique temporal responses immediately following cell division. Thus, the linkage of receptor occupancy to [Ca2+]i elevation is a functionally unique property for each individual cell and can be influenced by epigenetic factors.     

Alignment of clonedamiE gene ofPseudomonas aeruginosa with theN-terminal sequence of amidase

A restriction enzyme map was constructed for 5.1-kb fragment of Pseudomonas aeruginosa DNA inserted into plasmid pBR322. Restriction enzyme sites were matched to the N-terminal amino acid sequence of amidase to obtain alignment of the amiE gene within the cloned fragment.     

Cytochrome c4 from Pseudomonas aeruginosa

The dihaem cytochrome c₄ from Pseudomonas aeruginosa has been crystallized in space group P6₅22 with cell dimensions $\text{a = b = 62.4}\text{A}\text{?}\text{, c = 174.2}\text{A}\text{?}$, and one molecule per asymmetric unit. Two heavy-atom derivatives, UO₂(NO₃)₂ and K₂Pt(NO₂)₄, which substitute at one and three sites, respectively, have allowed a low-resolution electron density map to be obtained. This shows clearly the two domains of the molecule.     

Characterization of Boron Deficiency in Soybeans

Boron-deficient soybean roots have a rosette appearance at the terminals caused by the death of the root primordium and the initiation of new root primordia. The root tips show some darkening, swelling, and then a collapse of the tissue. The petiole s of B-deficient plants are very brittle. Physiologically, B-deficiency in HA-soybeans does not appear to involve an interference in the transport of photosynthate (C¹⁴ distribution) or precursors required for citrate synthesis in the roots. More Ca 45 was found in the tops of B-sufficient than B-deficient plants, but the opposite trend existed in the root sap. Boron deficiency symptoms were accentuated by maintaining a relatively low pH of the prenutrient solution by the addition of K salts.     

Development of the Mouse Dermal Adipose Layer Occurs Independently of Subcutaneous Adipose Tissue and Is Marked by Restricted Early Expression of FABP4

The laboratory mouse is a key animal model for studies of adipose biology, metabolism and disease, yet the developmental changes that occur in tissues and cells that become the adipose layer in mouse skin have received little attention. Moreover, the terminology around this adipose body is often confusing, as frequently no distinction is made between adipose tissue within the skin, and so called subcutaneous fat. Here adipocyte development in mouse dorsal skin was investigated from before birth to the end of the first hair follicle growth cycle. Using Oil Red O staining, immunohistochemistry, quantitative RT-PCR and TUNEL staining we confirmed previous observations of a close spatio-temporal link between hair follicle development and the process of adipogenesis. However, unlike previous studies, we observed that the skin adipose layer was created from cells within the lower dermis. By day 16 of embryonic development (e16) the lower dermis was demarcated from the upper dermal layer, and commitment to adipogenesis in the lower dermis was signalled by expression of FABP4, a marker of adipocyte differentiation. In mature mice the skin adipose layer is separated from underlying subcutaneous adipose tissue by the panniculus carnosus. We observed that the skin adipose tissue did not combine or intermix with subcutaneous adipose tissue at any developmental time point. By transplanting skin isolated from e14.5 mice (prior to the start of adipogenesis), under the kidney capsule of adult mice, we showed that skin adipose tissue develops independently and without influence from subcutaneous depots. This study has reinforced the developmental link between hair follicles and skin adipocyte biology. We argue that because skin adipocytes develop from cells within the dermis and independently from subcutaneous adipose tissue, that it is accurately termed dermal adipose tissue and that, in laboratory mice at least, it represents a separate adipose depot.     

The Discovery of orally available thrombin inhibitors : Studies towards the optimisation of CGH1668

The chemical optimisation of CGH1668 1 is described employing an in vivo model of absorption to determine the influence on bioavailability of single point modifications to five key molecular templates. The discovery of an orally bioavailable and selective thrombin inhibitor, 24, highlights the utility of this approach.     

BGX: a fully Bayesian integrated approach to the analysis of Affymetrix GeneChip data

We present Bayesian hierarchical models for the analysis of Affymetrix GeneChip data. The approach we take differs from other available approaches in two fundamental aspects. Firstly, we aim to integrate all processing steps of the raw data in a common statistically coherent framework, allowing all components and thus associated errors to be considered simultaneously. Secondly, inference is based on the full posterior distribution of gene expression indices and derived quantities, such as fold changes or ranks, rather than on single point estimates. Measures of uncertainty on these quantities are thus available. The models presented represent the first building block for integrated Bayesian Analysis of Affymetrix GeneChip data: the models take into account additive as well as multiplicative error, gene expression levels are estimated using perfect match and a fraction of mismatch probes and are modeled on the log scale. Background correction is incorporated by modeling true signal and cross-hybridization explicitly, and a need for further normalization is considerably reduced by allowing for array-specific distributions of nonspecific hybridization. When replicate arrays are available for a condition, posterior distributions of condition-specific gene expression indices are estimated directly, by a simultaneous consideration of replicate probe sets, avoiding averaging over estimates obtained from individual replicate arrays. The performance of the Bayesian model is compared to that of standard available point estimate methods on subsets of the well known GeneLogic and Affymetrix spike-in data. The Bayesian model is found to perform well and the integrated procedure presented appears to hold considerable promise for further development.     

Epidermal Notch signalling: differentiation, cancer and adhesion

The Notch pathway plays an important role in regulating epidermal differentiation. Notch ligands, receptors and effectors are expressed in a complex and dynamic pattern in embryonic and adult skin. Genetic ablation or activation of the pathway reveals that Notch signalling promotes differentiation of the hair follicle, sebaceous gland and interfollicular epidermal lineages and that Notch acts as an epidermal tumour suppressor. Notch signalling interacts with a range of other pathways to fulfil these functions and acts via RBP-Jkappa dependent and independent mechanisms. The effects on differentiation can be cell autonomous and non-autonomous, and Notch contributes to stem cell clustering via modulation of cell adhesion.