Stable transfection of the oestrogen receptor gene into a human osteosarcoma cell line

The oestrogen receptor (ER) gene was introduced into an ER-negative osteoblast-like osteosarcoma cell line HTB 96 by transfection. A number of clones were isolated which expressed ER at levels of up to 70 fmol/mg cytosol protein as determined by immunoassay. Scatchard analysis of the binding of [3H]17 beta-oestradiol in cytosols demonstrated the presence of high affinity binding sites, with a dissociation constant of 0.08-0.13 nM at 4 degrees C. High levels of a 3 kb ER mRNA are produced by the clones, which have gene copy numbers ranging from 2 to greater than 10. Functional receptor activity has been demonstrated by co-transfection of a plasmid containing the chloramphenicol acetyl transferase (CAT) gene linked to an oestrogen response element. Induction of CAT activity is observed in the presence of added oestradiol and is concentration-dependent. The transfected ER is also able to affect endogenous cellular function as several ER-positive clones, but not HTB 96 cells, are growth inhibited by oestradiol in the concentration range 10(-9)-10(-7) M. These effects on growth are not induced by other classes of steroids and are reversible by antioestrogens. No endogenous genes have yet been identified which are oestrogen-regulated in ER-transfected clones.     

Growth of food-borne pathogenic bacteria in oil-in-water emulsions: I—Methods for investigating the form of growth

Methods are presented for investigating the site and form of growth of bacteria in model oil-in-water emulsions and in dairy cream. Following growth of the bacteria, the continuous aqueous phase is gelled using agarose and the oil phase removed using a mixture of chloroform and methanol. Using this method, the authors have found that Listeria monocytogenes, Salmonella typhimurium and Yersinia enterocolitica grow in the form of colonies in concentrated oil-in-water emulsions. Colonies of L. monocytogenes and Y. enterocolitica also form in artificially-inoculated fresh and tinned dairy cream. If information about the precise site of growth is not required, the authors have discovered that intact colonies can be liberated from the model emulsions by dissolving away the oil phase with chloroform: methanol.     

Male and female flowers of the dioecious plant sorrel show different patterns of MADS box gene expression.

Male and female flowers of the dioecious plant sorrel (Rumex acetosa) each produce three whorls of developed floral organs: two similar whorls of three perianth segments and either six stamens (in the male) or a gynoecium consisting of a fertile carpel and two sterile carpels (in the female). In the developing male flower, there is no significant proliferation of cells in the center of the flower, in the position normally occupied by the carpels of a hermaphrodite plant. In the female flower, small stamen primordia are formed. To determine whether the organ differences are associated with differences in the expression of organ identity genes, cDNA clones representing the putative homologs of B and C function MADS box genes were isolated and used in an in situ hybridization analysis. The expression of RAD1 and RAD2 (two different DEFICIENS homologs) in males and females was confined to the stamen whorl; the lack of expression in the second, inner perianth whorl correlated with the sepaloid nature of the inner whorl of perianth segments. Expression of RAP1 (a PLENA homolog) occurred in the carpel and stamen whorls in very young flower primordia from both males and females. However, as soon as the inappropriate set of organs ceased to develop, RAP1 expression became undetectable in those organs. The absence of expression of RAP1 may be the cause of the arrest in organ development or may be a consequence.     

Microcalorimetric investigations of the interactions of small ions with arterial elastin

The enthalpies of interactions of porcine arterial elastin with alkali metal and alkali earth halides and sulphates were investigated by means of flow microcalorimetry and the stoichiometry measured using radiotracer techniques. In aqueous solutions, all alkali earth halides interacted exothermically at concentrations ranging from 0.01 to 2.5M. All the alkali metal halides, particularly NaCl, exhibited complex concentration-dependent interactions, exothermic at low concentrations and endothermic at high concentrations. Both the anion and cation contributed to the response, although the anion seemed to dominate. SO interacted most strongly of the anions tested. All interactions were reversible in the sense that repeat experiments gave identical results, but the enthalpy of “adsorption” was generally different from that of “desorption.” The enthalpy of interaction depended on the conformation of the elastin in a salt-specific manner. For example, CaCl₂ and MgCl₂ interacted similarly in water but very differently in 1 : 1 water : methanol. © 1994 John Wiley & Sons, Inc.     

Using elasticity/temperature relationships to characterise gelling carrageenans

Carrageenan characterisation by chemical means is long and complex. ¹³C NMR is undoubtedly the technique providing the most information, but the apparatus is expensive and measurement times are long. For several important problems in carrageenan characterisation, rheological methods provide quantitative information, comparable, for practical purposes, with that obtained by NMR. In particular, the apparent elastic modulus (G_app) is followed as a function of temperature, during cooling.Small amounts of kappa carrageenan in supposedly pure samples of the iota form have a great impact on the rheology and apparent ion selectivity of the latter. Using the elasticity/temperature relationship, as little as 2% of the kappa form can be detected in samples of iota carrageenan.The fraction of precursor sugars is another important parameter in the gelling of carrageenans. The form of the elasticity/temperature function is strongly dependent on the fraction of precursor sugars.Finally, it is shown that the elasticity/temperature relationship can provide a useful fingerprint of heterogeneous carrageenans.     

Aerolysin — the ins and outs of a model channel-forming toxin

Aerolysin is one of a large group of bacterial proteins that can kill target cells by forming discrete channels in their plasma membranes. The toxin has many properties in common with the porins of the Gram-negative bacterial outer membrane, including an extensive amount of β-structure, a high proportion of hydrophilic amino acid side-chains and no hydrophobic stretches in the primary structure. It also oligomerizes to produce an insertion-competent state. Aerolysin is secreted as a dimer by members of the Aeromonas family. It binds to a high-affinity receptor on the target cell that has recently been shown to be a glycosylphosphatidylinositol-anchored glycoprotein. Binding is followed by heptamerization to form a structure that we propose contains a β-barrel which can insert into the membrane and produce a channel.     

Decompression comparison of helium and hydrogen in rats

Lillo, R. S., E. C. Parker, and W. R. Porter.Decompression comparison of helium and hydrogen in rats.J. Appl. Physiol. 82(3): 892-901, 1997.The hypothesis that there are differences in decompression riskbetween He and H₂ wasexamined in 1,607 unanesthetized male albino rats subjected to dives on2% O₂-balance He or 2%O₂-balanceH₂ (depths  50 ATA, bottom times  60 min). The animals were decompressed to 10.8 ATA with profilesvarying from rapid to slow, with up to four decompression stops of up to 60 min each. Maximum likelihood analysis was used to estimate therelative decompression risk on a per unit pressure basis (termed "potency") and the rate of gas uptake and elimination, bothfactors affecting the decompression sickness risk, from a specific dive profile. H₂ potency for causingdecompression sickness was found to be up to 35% greater than that forHe. Uptake rates were unresolvable between the two gases with the timeconstant (TC) estimated at ~2-3 min, leading to saturation inboth cases in <15 min. Washout of both gases was significantly slowerthan uptake, with He washout (TC ~1.5-3 h) substantially slowerthan H₂ washout (TC ~0.5 h). Itis unknown whether the decompression advantage of the faster washout ofH₂ or the disadvantage of itsincreased potency, observed in the rat, would be important for humandiving.

     

Predisposing factors in systemic and central nervous system candidiasis: Histopathological and cultural observations in the rat

Four atypical isolates of Microsporum canis, three from humans and one from a cat, were obtained from North-West London. These and a further human isolate were compared with each other and with a typical isolate of the fungus. Immediately after isolation the atypical isolates were very labile, but were stabilised after a few subcultures from selected sectors. The stable forms differed from each other, but all had a tendency to brown rather than yellow pigmentation, to feathery submerged mycelium and to abnormal macroconidia. The macroscopic appearance and texture of the colonies depended on the density, orientation and branching pattern of the submerged mycelium.In recent years similar brown, feathery forms of M. canis have been reported from monkeys but not from cats. It is suggested that all such isolates may be culturally stable forms of a very unstable strain, probably feline in origin, which has yet to be described.