Multispectral imaging for determination of astaxanthin concentration in salmonids

Multispectral imaging has been evaluated for characterization of the concentration of a specific cartenoid pigment; astaxanthin. 59 fillets of rainbow trout, Oncorhynchus mykiss, were filleted and imaged using a rapid multispectral imaging device for quantitative analysis. The multispectral imaging device captures reflection properties in 19 distinct wavelength bands, prior to determination of the true concentration of astaxanthin. The samples ranged from 0.20 to 4.34 g per g fish. A PLSR model was calibrated to predict astaxanthin concentration from novel images, and showed good results with a RMSEP of 0.27. For comparison a similar model were built for normal color images, which yielded a RMSEP of 0.45. The acquisition speed of the multispectral imaging system and the accuracy of the PLSR model obtained suggest this method as a promising technique for rapid in-line estimation of astaxanthin concentration in rainbow trout fillets.     

Inhibition of polyamine synthesis reduces the growth rate and delays the expression of differentiated phenotypes in primary cultures of embryonic mesoderm from chick

Summary Inhibition of polyamine synthesis in early chick embryos blocks their development at gastrulation. Analyses of arrested embryos show that mesodermal outgrowth and differentiation are drastically impaired. To study these effects in greater detail, we have used primary cultures of embryonic mesoderm from chick. The cultures were treated with -difluoromethylornithine (DFMO), an enzyme-activated irreversible inhibitor of ornithine decarboxylase, the first and rate-limiting enzyme in polyamine synthesis. In control culture medium, mesodermal cells retained their in ovo outgrowth behavior and differentiation pattern. Addition of 10 mM DFMO to the culture medium, however, retarded attachment and outgrowth, and reduced the rate of proliferation of the mesodermal cells. Furthermore, the expression of differentiated phenotypes, such as beating heart tissue, erythroid cells, and adipocyte-like cells, was delayed. Simultaneous addition of 100 M putrescine prevented or reduced the effects of DFMO, showing that these were indeed caused by polyamine deficiency. In the DFMO-treated mesoderm, DNA synthesis was markedly suppressed by the first day. Similar effects on RNA and protein synthesis developed at a later time. Our data suggest that a reduction in the concentrations of the polyamines decreases the rate of mesodermal cell proliferation, and as a conseqence delays the expression of differentiated phenotypes.     

Displacement of sterols from sterol/sphingomyelin domains in fluid bilayer membranes by competing molecules

The formation of sterol and palmitoyl sphingomyelin enriched ordered domains in a fluid bilayer was examined using domain selective fluorescent reporter molecules (cholestatrienol and trans-parinaric acid containing lipids) together with a quencher molecule in the fluid phase. The aim of the study was to explore how stable the ordered domains were and how different, biologically interesting, membrane intercalators could affect domain stability and sterol distribution between domains. We show that sterols easily can be displaced from ordered domains by a variety of saturated, single- and double-chain membrane intercalators with a small polar group as a common denominator. Of the two-chain intercalators examined, both palmitoyl ceramide and palmitoyl dihydroceramide were effective in displacing sterols from ordered domains. Of the single-chain intercalators, hexadecanol and hexadecyl amide displaced the sterol from sterol/sphingomyelin domains, whereas palmitic acid, sphingosine and sphinganine failed to do so. All molecules examined stabilized the sphingomyelin-rich domains, as reported by trans-parinaric-sphingomyelin and by scanning calorimetry. Parallels between the displacement of sterol from ordered domains in our model membrane system and the ability of the above mentioned molecules to alter the chemical activity and distribution of sterols in biological membranes are discussed.     

TGF-beta induced G(1) cell cycle arrest requires the activity of the proteasome pathway. Transforming growth factor

Transforming growth factor-beta (TGF-beta) induces a potent G(1)/S-phase cell cycle arrest of epithelial cells by inhibiting the activities of cyclin D- and cyclin E-associated kinase complexes. Downregulation of the kinase activities is mediated by induction of cyclin dependent kinase (CDK) inhibitor p15(Ink4b) which blocks CDK4 and CDK6 kinases and leads to binding of p27(Kip1) to CDK2-cyclin E complex. Levels of several of these factors are controlled by the ubiquitin-proteasome pathway. We demonstrate here that proteasomal inhibitors release the cells from TGF-beta imposed G(1)-phase arrest and instigate the entry of the cells into S-phase. Proteasomal inhibitors are shown to specifically increase the activity of the cyclin D-kinase complex by increasing the levels of p27(Kip1) and cyclin D and by maintaining CDK4/6 protein levels leading to phosphorylation of the retinoblastoma protein without increasing cyclin E-associated kinase activity. The results indicate caution in the potential therapeutic use of the proteasome inhibitors due to unscheduled initiation of DNA replication in the presence of a physiological growth inhibitor.     

A recruited protease is involved in catabolism of pyrimidines

In nature, the same biochemical reaction can be catalyzed by enzymes having fundamentally different folds, reaction mechanisms and origins. For example, the third step of the reductive catabolism of pyrimidines, the conversion of N-carbamyl-β-alanine to β-alanine, is catalyzed by two β-alanine synthase (βASase, EC 3.5.1.6) subfamilies. We show that the “prototype” eukaryote βASases, such as those from Drosophila melanogaster and Arabidopsis thaliana, are relatively efficient in the conversion of N-carbamyl-βA compared with a representative of fungal βASases, the yeast Saccharomyces kluyveri βASase, which has a high K_m value (71 mM). S. kluyveri βASase is specifically inhibited by dipeptides and tripeptides, and the apparent K_i value of glycyl-glycine is in the same range as the substrate K_m. We show that this inhibitor binds to the enzyme active center in a similar way as the substrate. The observed structural similarities and inhibition behavior, as well as the phylogenetic relationship, suggest that the ancestor of the fungal βASase was a protease that had modified its profession and become involved in the metabolism of nucleic acid precursors.