Eukaryotic pathogens (Chytridiomycota and Oomycota) infecting marine microphytobenthic diatoms – a methodological comparison

Using sediment samples from the Solthörn tidal flat (southern North Sea, Germany), collected in bi‐weekly intervals from June to July 2012, a range of qualitative and quantitative screening methods for oomycete and chytrid pathogens infecting benthic diatoms were evaluated. Pre‐treatment of sediment samples using short ultrasound pulses and gradient centrifugation, in combination with CalcoFluor White, showed the best results in the visualization of both pathogen groups. The highest number of infected benthic diatoms was observed in mid July (5.8% of the total benthic diatom community). Most infections were caused by chytrids and, in a few cases, oomycetes (Lagenisma Drebes (host: Coscinodiscus radiatus Ehrenberg) and Ectrogella Zopf (hosts: Dimeregramma minor in Pritchard and Gyrosigma peisonis). Among the chytrids, sporangium morphology indicated the presence of five different morphotypes, infecting mainly epipelic taxa of the orders Naviculales (e.g., Navicula digitoradiata) and Achnanthales (e.g., Achnanthes brevipes Agardh). The presence of multiple pathogens in several epipelic diatom taxa suggests a significant role for fungal parasitism in affecting microphytobenthic diatom succession.     

Photoresponsive palladium(II) complexes with azobenzene incorporated into benzyl aryl ether dendrimers

Palladium(II) complexes, [Pd(GX-azb)₂Cl₂] (where azb = azobenzene, GX = benzyl-aryl ether dendron of generation X = 1, 2, 3), were prepared and their photophysical properties were examined. The synthesized complexes were characterized by chemical analysis, ¹H NMR and UV spectroscopy. The photochromic dendritic azobenzene ligands within the complexes [Pd(GX-azb)₂Cl₂] undergo a reversible trans/cis isomerization upon exposure to ultraviolet light.     

Functional expression of the transient receptor potential channel TRPA1, a sensor for toxic lung inhalants,in pulmonary epithelial cells

The cation channel TRPA1 functions as a chemosensory protein and is directly activated by a number of noxious inhalants. A pulmonary expression of TRPA1 has been described in sensory nerve endings and its stimulation leads to the acceleration of inflammatory responses in the lung. Whereas the function of TRPA1 in neuronal cells is well defined, only few reports exist suggesting a role in epithelial cells. The aim of the present study was therefore (1) to evaluate the expression of TRPA1 in pulmonary epithelial cell lines, (2) to characterize TRPA1-promoted signaling in these cells, and (3) to study the extra-neuronal expression of this channel in lung tissue sections. Our results revealed that the widely used alveolar type II cell line A549 expresses TRPA1 at the mRNA and protein level. Furthermore, stimulating A549 cells with known TRPA1 activators (i.e., allyl isothiocyanate) led to an increase in intracellular calcium levels, which was sensitive to the TRPA1 blocker ruthenium red. Investigating TRPA1 coupled downstream signaling cascades it was found that TRPA1 activation elicited a stimulation of ERK1/2 whereas other MAP kinases were not affected. Finally, using epithelial as well as neuronal markers in immunohistochemical approaches, a non-neuronal TRPA1 protein expression was detected in distal parts of the porcine lung epithelium, which was also found examining human lung sections. TRPA1-positive staining co-localized with both epithelial and neuronal markers underlining the observed epithelial expression pattern. Our findings of a functional expression of TRPA1 in pulmonary epithelial cells provide causal evidence for a non-neuronal TRPA1-mediated control of inflammatory responses elicited upon TRPA1-mediated registration of toxic inhalants in vivo.     

Kinase-independent phosphoramidate S1P1 receptor agonist benzyl ether derivatives

Previously published S1P receptor modulator benzyl ether derivatives have shown potential as being viable therapeutics for the treatment of neurodegenerative diseases, however, two of the most S1P₁-selective compounds are reported as being poorly phosphorylated by kinases in vivo. Phosphoramidates of BED compounds (2a, 2b) were synthesised with the aim of producing kinase-independent S1P receptor modulators. Carboxypeptidase, human serum and cell lysate processing experiments were conducted. ProTide BED analogues were found to have an acceptable level of stability in acidic and basic conditions and in vitro metabolic processing experiments showed that they are processed to the desired pharmacologically active monophosphate. The research describes the development of an entirely novel family of therapeutic agents.     

Reversible Dihydropyridine Isothiocyanate Binding to Brain Calcium Channels

Voltage-dependent calcium channels from ileal smooth muscle can be affinity-labeled with a [3H]dihydropyridine isothiocyanate radioligand. We examined the binding of this agent to brain membranes, to compare the properties of calcium channel drug binding sites in brain with those previously described in ileum. In brain, the [3H]dihydropyridine isothiocyanate labels sites that correspond in number and pharmacologic characteristics to binding sites for the classic calcium entry blocker, [3H]nitrendipine. However, in contrast to the covalent nature of dihydropyridine isothiocyanate binding in ileum, brain calcium channels are labeled reversibly. This difference in binding properties may reflect structural variations in voltage-dependent calcium channels in different tissues.     

The effects of ruthenium red,lanthanum, fluorescein isothiocyanate and trifluoperazine on vesicle transport,vesicle fusion and tip extension in pollen tubes

The effects of ruthenium red, lanthanum, fluorescein isothiocyanate and trifluoperazine, all antagonists of Ca²⁺ function in cells, have been studied in growing pollen tubes of Tradescantia virginiana. All four drugs inhibit pollen-tube growth but bring about different ultrastructural changes at the growing tips and within the cytoplasm. The results strongly support the hypothesis that Ca²⁺ plays a vital role in the mechanism of pollen-tube tip growth. The effect of ruthenium red provides evidence that sequestration of Ca²⁺ by mitochondria critically adjusts the concentration of these ions at tube tips. Fluorescein isothiocyanate appears to be a potent inhibitor of vesicle fusion at the plasma membrane, with vesicles accumulating in the tip at rates equivalent to those determined previously for their production. Both vesicle fusion and tip extension are regulated by Ca²⁺ but appear to be independently controlled processes.