The connecting cilium inner scaffold provides a structural foundation that protects against retinal degeneration

Inherited retinal degeneration due to loss of photoreceptor cells is a leading cause of human blindness. These cells possess a photosensitive outer segment linked to the cell body through the connecting cilium (CC). While structural defects of the CC have been associated with retinal degeneration, its nanoscale molecular composition, assembly, and function are barely known. Here, using expansion microscopy and electron microscopy, we reveal the molecular architecture of the CC and demonstrate that microtubules are linked together by a CC inner scaffold containing POC5, CENTRIN, and FAM161A. Dissecting CC inner scaffold assembly during photoreceptor development in mouse revealed that it acts as a structural zipper, progressively bridging microtubule doublets and straightening the CC. Furthermore, we show that Fam161a disruption in mouse leads to specific CC inner scaffold loss and triggers microtubule doublet spreading, prior to outer segment collapse and photoreceptor degeneration, suggesting a molecular mechanism for a subtype of retinitis pigmentosa.

Inherited retinal degeneration due to loss of photoreceptor cells is a leading cause of human blindness. Ultrastructure expansion microscopy on mouse retina reveals the presence of a novel structure inside the photoreceptor connecting cilium, the inner scaffold, that protects the outer segment against degeneration.     

The chemotaxonomic and medicinal significance of phenolic acids in Arctopus and Alepidea (Apiaceae subfamily Saniculoideae)

The occurrence of (R)-3′-O-β-d-glucopyranosylrosmarinic acid, rosmarinic acid and caffeic acid in two important South African medicinal plants is reported for the first time. (R)-3′-O-β-d-Glucopyranosylrosmarinic acid and rosmarinic acid were isolated and identified in several samples from three species of the genus Arctopus L. (sieketroos) and three species of the genus Alepidea F. Delaroche (ikhathazo), both recently shown to be members of the subfamily Saniculoideae of the family Apiaceae. The compounds occur in high concentrations (up to 15.3 mg of (R)-3′-O-β-d-glucopyranosylrosmarinic acid per g dry wt) in roots of Arctopus. Our results provide a rationale for the traditional uses of these plants, as the identified compounds are all known for their antioxidant activity, with rosmarinic acid further contributing to a wide range of biological activities. Furthermore, we confirm the idea that (R)-3′-O-β-d-glucopyranosylrosmarinic acid is a useful chemotaxonomic marker for the subfamily Saniculoideae.     

Waterborne pathogen detection by use of oligonucleotide-based microarrays

A small-oligonucleotide microarray prototype was designed with probes specific for the universal 16S rRNA and cpn60 genes of several pathogens that are usually encountered in wastewaters. In addition to these two targets, wecE-specific oligonucleotide probes were included in the microarray to enhance its discriminating power within the Enterobacteriaceae family. Universal PCR primers were used to amplify variable regions of 16S rRNA, cpn60, and wecE genes directly in Escherichia coli and Salmonella enterica serovar Typhimurium genomic DNA mixtures (binary); E. coli, S. enterica serovar Typhimurium, and Yersinia enterocolitica genomic DNA mixtures (ternary); or wastewater total DNA. Amplified products were fluorescently labeled and hybridized on the prototype chip. The detection sensitivity for S. enterica serovar Typhimurium was estimated to be on the order of 0.1% (10(4) S. enterica genomes) of the total DNA for the combination of PCR followed by microarray hybridization. The sensitivity of the prototype could be increased by hybridizing amplicons generated by PCR targeting genes specific for a bacterial subgroup, such as wecE genes, instead of universal taxonomic amplicons. However, there was evidence of PCR bias affecting the detection limits of a given pathogen as increasing amounts of a different pathogen were spiked into the test samples. These results demonstrate the feasibility of using DNA microarrays in the detection of waterborne pathogens within mixed populations but also raise the problem of PCR bias in such experiments.     

Sphingosine in apoptosis signaling

The sphingolipid metabolites ceramide, sphingosine, and sphingosine 1-phosphate contribute to controlling cell proliferation and apoptosis. Ceramide and its catabolite sphingosine act as negative regulators of cell proliferation and promote apoptosis. Conversely, sphingosine 1-phosphate, formed by phosphorylation of sphingosine by a sphingosine kinase, has been involved in stimulating cell growth and inhibiting apoptosis. As the phosphorylation of sphingosine diminishes apoptosis, while dephosphorylation of sphingosine 1-phosphate potentiates it, the role of sphingosine as a messenger of apoptosis is of importance. Herein, the effects of sphingosine on diverse signaling pathways implicated in the apoptotic process are reviewed.     

Advanced glycation end-products disrupt human endothelial cells redox homeostasis: new insights into reactive oxygen species production

Advanced glycation end-products (AGEs) trigger multiple metabolic disorders in the vessel wall that may in turn lead to endothelial dysfunction. The molecular mechanisms by which AGEs generate these effects are not completely understood. Oxidative stress plays a key role in the development of deleterious effects that occur in endothelium during diabetes. Our main objectives were to further understand how AGEs contribute to reactive oxygen species (ROS) overproduction in endothelial cells and to evaluate the protective effect of an antioxidant plant extract. The human endothelial cell line EA.hy926 was treated with native or modified bovine serum albumin (respectively BSA and BSA-AGEs). To monitor free radicals formation, we used H₂DCF-DA, dihydroethidium (DHE), DAF-FM-DA and MitoSOX Red dyes. To investigate potential sources of ROS, nicotinamide adenine dinucleotide phosphate (NADPH) oxidase and mitochondrial inhibitors were used. The regulation of different types of ROS by the polyphenol-rich extract from the medicinal plant Doratoxylon apetalum was also studied for a therapeutic perspective. BSA-AGEs exhibited not only less antioxidant properties than BSA, but also pro-oxidant effects. The degree of albumin glycoxidation directly influenced oxidative stress through a possible communication between NADPH oxidase and mitochondria. D. apetalum significantly decreased intracellular hydrogen peroxide and superoxide anions mainly detected by H₂DCF-DA and DHE respectively. Our results suggest that BSA-AGEs promote a marked oxidative stress mediated at least by NADPH oxidase and mitochondria. D. apetalum plant extract appeared to be an effective antioxidant compound to protect endothelial cells.