Cellular, biochemical, and molecular changes during encystment of free-living amoebae

Free-living amoebae are protozoa found in soil and water. Among them, some are pathogenic and many have been described as potential reservoirs of pathogenic bacteria. Their cell cycle is divided into at least two forms, the trophozoite and the cyst, and the differentiation process is named encystment. As cysts are more resistant to disinfection treatments than trophozoites, many studies focused on encystment, but until recently, little was known about cellular, biochemical, and molecular modifications operating during this process. Important signals and signaling pathways at play during encystment, as well as cell responses at the molecular level, have been described. This review summarizes our knowledge and focuses on new findings.     

Islet Endothelial Activation and Oxidative Stress Gene Expression Is Reduced by IL-1Ra Treatment in the Type 2 Diabetic GK Rat

Background

Inflammation followed by fibrosis is a component of islet dysfunction in both rodent and human type 2 diabetes. Because islet inflammation may originate from endothelial cells, we assessed the expression of selected genes involved in endothelial cell activation in islets from a spontaneous model of type 2 diabetes, the Goto-Kakizaki (GK) rat. We also examined islet endotheliuml/oxidative stress (OS)/inflammation-related gene expression, islet vascularization and fibrosis after treatment with the interleukin-1 (IL-1) receptor antagonist (IL-1Ra).

Methodology/Principal Findings

Gene expression was analyzed by quantitative RT-PCR on islets isolated from 10-week-old diabetic GK and control Wistar rats. Furthermore, GK rats were treated s.c twice daily with IL-1Ra (Kineret, Amgen, 100 mg/kg/day) or saline, from 4 weeks of age onwards (onset of diabetes). Four weeks later, islet gene analysis and pancreas immunochemistry were performed. Thirty-two genes were selected encoding molecules involved in endothelial cell activation, particularly fibrinolysis, vascular tone, OS, angiogenesis and also inflammation. All genes except those encoding angiotensinogen and epoxide hydrolase (that were decreased), and 12-lipoxygenase and vascular endothelial growth factor (that showed no change), were significantly up-regulated in GK islets. After IL-1Ra treatment of GK rats in vivo, most selected genes implied in endothelium/OS/immune cells/fibrosis were significantly down-regulated. IL-1Ra also improved islet vascularization, reduced fibrosis and ameliorated glycemia.

Conclusions/Significance

GK rat islets have increased mRNA expression of markers of early islet endothelial cell activation, possibly triggered by several metabolic factors, and also some defense mechanisms. The beneficial effect of IL-1Ra on most islet endothelial/OS/immune cells/fibrosis parameters analyzed highlights a major endothelial-related role for IL-1 in GK islet alterations. Thus, metabolically-altered islet endothelium might affect the β-cell microenvironment and contribute to progressive type 2 diabetic β-cell dysfunction in GK rats. Counteracting islet endothelial cell inflammation might be one way to ameliorate/prevent β-cell dysfunction in type 2 diabetes.     

Cyclic strain promotes shuttling of PYK2/Hic-5 complex from focal contacts in osteoblast-like cells

We showed that cyclic strain (CS) of osteoblastic cells induced tyrosine phosphorylation of two homologous tyrosine kinases FAK and PYK2, and of two homologous adaptor proteins paxillin and Hic5, with similar kinetics. Immunostaining showed that all four proteins were localized to focal contacts in controls. In contrast, the dynamics of their subcellular localization observed after CS differed. While FAK and paxillin remained at the focal contact, Hic-5 and PYK2 translocated outside ventral focal contacts as early as 30 min after CS and were sequestered by the cytoskeleton. Co-immunoprecipitation showed that the association of PYK2/Hic-5 and PYK2/FAK increased with time after strain while that of paxillin and Hic-5 decreased. Altogether these results suggested that CS regulates focal contact activity in osteoblasts by modulating PYK2-containing complexes in particular by shuttling out of the focal contact the adaptor Hic-5 and favoring the anchorage of FAK within contacts.     

A higher-order entity formed by the flexible assembly of RAP1 with TRF2

Telomere integrity is essential to maintain genome stability, and telomeric dysfunctions are associated with cancer and aging pathologies. In human, the shelterin complex binds TTAGGG DNA repeats and provides capping to chromosome ends. Within shelterin, RAP1 is recruited through its interaction with TRF2, and TRF2 is required for telomere protection through a network of nucleic acid and protein interactions. RAP1 is one of the most conserved shelterin proteins although one unresolved question is how its interaction may influence TRF2 properties and regulate its capacity to bind multiple proteins. Through a combination of biochemical, biophysical and structural approaches, we unveiled a unique mode of assembly between RAP1 and TRF2. The complete interaction scheme between the full-length proteins involves a complex biphasic interaction of RAP1 that directly affects the binding properties of the assembly. These results reveal how a non-DNA binding protein can influence the properties of a DNA-binding partner by mutual conformational adjustments.     

Early Medieval Muslim Graves in France: First Archaeological,Anthropological and Palaeogenomic Evidence

The rapid Arab-Islamic conquest during the early Middle Ages led to major political and cultural changes in the Mediterranean world. Although the early medieval Muslim presence in the Iberian Peninsula is now well documented, based in the evaluation of archeological and historical sources, the Muslim expansion in the area north of the Pyrenees has only been documented so far through textual sources or rare archaeological data. Our study provides the first archaeo-anthropological testimony of the Muslim establishment in South of France through the multidisciplinary analysis of three graves excavated at Nimes. First, we argue in favor of burials that followed Islamic rites and then note the presence of a community practicing Muslim traditions in Nimes. Second, the radiometric dates obtained from all three human skeletons (between the 7th and the 9th centuries AD) echo historical sources documenting an early Muslim presence in southern Gaul (i.e., the first half of 8th century AD). Finally, palaeogenomic analyses conducted on the human remains provide arguments in favor of a North African ancestry of the three individuals, at least considering the paternal lineages. Given all of these data, we propose that the skeletons from the Nimes burials belonged to Berbers integrated into the Umayyad army during the Arab expansion in North Africa. Our discovery not only discusses the first anthropological and genetic data concerning the Muslim occupation of the Visigothic territory of Septimania but also highlights the complexity of the relationship between the two communities during this period.     

Effects of the humic substances of de-inking paper sludge on the antagonism between two compost bacteria and Pythium ultimum

We investigated the in vitro influence of humic substances (HS) extracted from de-inking paper sludge compost on the inhibition of Pythium ultimum by two compost bacteria, Rhizobium radiobacter (Agrobacterium radiobacter) and Pseudomonas aeruginosa. When low concentrations (5 or 50 mg l(-1)) of HS were added to the culture medium, fungal inhibition by R. radiobacter significantly increased (P<0.01) by 2-3%. In contrast, these low levels of HS had no effect on P. ultimum inhibition by P. aeruginosa. The Fe, chelated by HS, was in part responsible for the decrease of P. ultimum inhibition by the bacteria when increasing amounts of HS were added in the culture medium. The addition of 500 mg l(-1) of humic acids isolated from de-inking paper sludge compost or from fossil origin completely eliminated the inhibition of P. ultimum by R. radiobacter. This Fe effect also stimulated growth of R. radiobacter and reduced its siderophore production in a minimal medium supplemented with HS as sole source of Fe. The results showed that HS influence microbial antagonism when added to a culture medium. However, this effect varies with different factors such as the type of bacteria, concentration of HS, molecular weight and Fe content.     

Carbonyl scavenger and antiatherogenic effects of hydrazine derivatives

Reactive carbonyl compounds (RCC) generated by polyunsaturated fatty acid oxidation alter progressively cellular and tissular proteins by forming adducts on free amino groups and thiol residues (carbonyl stress). Carbonyl scavengers may neutralize RCC, but their protective effect in atherosclerosis has not been extensively studied. We report the carbonyl scavenger and antiatherogenic properties of hydrazine derivatives, namely hydralazine, an antihypertensive drug, isoniazid, an antituberculosis agent, and two antidepressants, phenelzine and iproniazid. These drugs were poorly efficient in preventing the oxidation of LDL mediated by smooth muscle cells (SMCs), but inhibited the toxicity of UV-oxidized LDL (oxLDL) and of 4-hydroxynonenal (4-HNE). Hydrazine derivatives prevented the formation of foam cells resulting from LDL oxidation in human macrophagic U937 cells, and blocked the carbonyl stress in SMCs, by inhibiting the decrease in free amino group content, the increase in carbonylated proteins, and the formation of 4-HNE adducts on PDGFR. Experimental studies carried out on apoE-/- mice supplemented with drugs (30 mg/L in drinking water) showed a significant carbonyl stress inhibition correlated with a net reduction of atherosclerotic lesion development. In conclusion, these data indicate that hydrazine derivatives exhibit carbonyl scavenger and antiatherogenic properties, which opens novel therapeutical approaches for atherosclerosis and its cardiovascular complications.     

A New Combination with D-Cateslytin to Eradicate Root Canal Pathogens

International Journal of Peptide Research and Therapeutics - The success of endodontic treatments depends on the elimination of intracanal pathogens. Since irrigation and instrumentation can only...     

Motions and structural variability within toxins: implication for their use as scaffolds for protein engineering

Animal toxins are small proteins built on the basis of a few disulfide bonded frameworks. Because of their high variability in sequence and biologic function, these proteins are now used as templates for protein engineering. Here we report the extensive characterization of the structure and dynamics of two toxin folds, the "three-finger" fold and the short alpha/beta scorpion fold found in snake and scorpion venoms, respectively. These two folds have a very different architecture; the short alpha/beta scorpion fold is highly compact, whereas the "three-finger" fold is a beta structure presenting large flexible loops. First, the crystal structure of the snake toxin alpha was solved at 1.8-A resolution. Then, long molecular dynamics simulations (10 ns) in water boxes of the snake toxin alpha and the scorpion charybdotoxin were performed, starting either from the crystal or the solution structure. For both proteins, the crystal structure is stabilized by more hydrogen bonds than the solution structure, and the trajectory starting from the X-ray structure is more stable than the trajectory started from the NMR structure. The trajectories started from the X-ray structure are in agreement with the experimental NMR and X-ray data about the protein dynamics. Both proteins exhibit fast motions with an amplitude correlated to their secondary structure. In contrast, slower motions are essentially only observed in toxin alpha. The regions submitted to rare motions during the simulations are those that exhibit millisecond time-scale motions. Lastly, the structural variations within each fold family are described. The localization and the amplitude of these variations suggest that the regions presenting large-scale motions should be those tolerant to large insertions or deletions.