Validation of DNA methylation profiling in formalin-fixed paraffin-embedded samples using the Infinium HumanMethylation450 Microarray

A formalin-fixed paraffin-embedded (FFPE) sample usually yields highly degraded DNA, which limits the use of techniques requiring high-quality DNA, such as Infinium Methylation microarrays. To overcome this restriction, we have applied an FFPE restoration procedure consisting of DNA repair and ligation processes in a set of paired fresh-frozen (FF) and FFPE samples. We validated the FFPE results in comparison with matched FF samples, enabling us to use FFPE samples on the Infinium HumanMethylation450 Methylation array.     

Cytotoxic and antiprotozoal activity of flavonoids from Lonchocarpus spp.

A number of flavonoids isolated from Lonchocarpus spp. were evaluated for their antiprotozoal and cytotoxic activity. Flavone 6 and chalcone 7 were found to be the most active against Leishmania parasites and against cell cultures of Leukemia P388DI and adenocarcinoma prostate PC-3.     

A Toxoplasma type 2C serine-threonine phosphatase is involved in parasite growth in the mammalian host cell

Toxoplasma gondii is a human protozoan parasite that belongs to the phylum of Apicomplexa and causes toxoplasmosis. As the other members of this phylum, T. gondii obligatory multiplies within a host cell by a peculiar type of mitosis that leads to daughter cell assembly within a mother cell. Although parasite growth and virulence have been linked for years, few molecules controlling mitosis have been yet identified and they include a couple of kinases but not the counteracting phosphatases. Here, we report that in contrast to other animal cells, type 2C is by far the major type of serine threonine phosphatase activity both in extracellular and in intracellular dividing parasites. Using wild type and transgenic parasites, we characterized the 37 kDa TgPP2C molecule as an abundant cytoplasmic and nuclear enzyme with activity being under tight regulation. In addition, we showed that the increase in TgPP2C activity significantly affected parasite growth by impairing cytokinesis while nuclear division still occurred. This study supports for the first time that type 2C protein phosphatase is an important regulator of cell growth in T. gondii.     

Variation in the diet of the viperine snake Natvix mama in relation to prey availability

The diet of the viperine snake was compared with food availability in the Ebro Delta, a wetland largely occupied by rice fields, in 1990 and 1991. Snake selection of prey type and size was studied seasonally and by snake group: males, females and immature snakes. Overall, feeding activity (percentage of individuals with prey and number of prey per stomach) increased with food availability. Diet analysis showed that viperine snakes mainly foraged on the green frog Rana perezi (adults and tadpoles) and the carp Cyprinus earpio. Conversely, viperine snakes rejected the mosquito fish Gambusia holbroki which is the most abundant species in autumn, when Natrix maura has a low feeding activity. Statistical comparisons between viperine snake diet and prey availability showed that males selected small carp, immature snakes selected tadpoles and, in spring, females selected frogs. The selection of small carp by males may reflect a sexual divergence of trophic niche related to sexual size dimorphism, as females are larger than males. As tadpoles are presumably easier to catch than fish, tadpole selection by immature individuals may reflect variance in capture abilities. In spring, the selection of frogs by females overlapped with vitellogenesis, suggesting that females compensate for the cost of reproduction by selecting green frogs, which have a greater biomass and higher energy content than fish. Carps eaten in spring were smaller than in summer. Moreover, in summer viperine snakes selected smaller carp than the available mean size. This divergent tendency between carp size selection and carp size availability reveals how seasonal diet shifts in prey size selection may be a response to an increase in prey size.     

Isolation and characterization of microsatellites in Pacific white shrimp Penaeus (Litopenaeus) vannamei

Five polymorphic microsatellite loci were characterized for Penaeus (Litopenaeus) vannamei. Loci were isolated using a partial Sau3A1 genomic library by the sequencing of randomly selected clones and by a biotinylated (CT)₁₀ and (GT)₁₀ probes screening procedure. The last strategy resulted in the most useful data. About 40% of the clones showed a previously reported satellite/microsatellite (PVS1), reducing the chance of finding new microsatellite regions. Whereas two of the microsatellite loci with more than 10 alleles will be useful for mating analysis in a breeding program, the others might prove useful for population genetic studies.     

Ultrastructural studies of endocrine-like cells in the fundic gastric mucosa of the bullfrog,Rana catesbeiana

Summary This study is concerned with electron-microscopic observations on endocrine or paracrine cells in the fundic gastric mucosa of the bullfrog. Also, an attempt was made to identify the histamine-releasing cells involved in the secretagogue response. At least three distinct endocrine-like cell types were found. The classification is based on the appearance of secretory granules and other organelles, and the relationship of endocrine-like cells with other cells in the tissue. The amphibian endocrine-like cells resemble the ECL, D and EC cells of mammals. Type-I (ECL) cells showed degranulation after repeated stimulation with tetragastrin (TG), acetylcholine (ACh) and K⁺ depolarizing solution, all of which release histamine.     

Thromboxane receptor blockade improves cyclosporine nephrotoxicity in rats

Cyclosporine A (CyA) nephorotoxicity is associated with impaired renal hemodynamic funtion and increased production of the vasoconstrictor eicosanoid thromboxane A₂ (TxA₂). In CyA toxic rats, renal dysfunction cna be partially reversed by inhibitors of thromoboxane sysnthase. However, interpretation of these results is complicated since inhibitance of thromboxane synthase may cause accumulation of prostaglandin endoperoxides that can act as partial agonists at the TxA₂ receptor and may blunt the efficacy of treatment. Furthermore, these endoperoxides may be used as substrate for production of vasodilator prostaglandins causing beneficial effects on hemodynamics which are independent of thromboxane inhibition. To more specially examine the role of TxA₂ in CyA toxicity, we investigated the effects of the thromboxane receptor antagonist GR32191 on renal hemodynamics in a rat model of CyA nephrotoxicity. In this model, administration of CyA resulted in a significant decrease in glomerular filtration rate (GFR) 2.85±0.26 [CyA] vs 6.82±0.96 ml/min/kg [vehicle]; p<0.0005) and renal blood flow (RBF) (21.6±2.31 [CyA] vs 31.8±3.60 ml/min/kg [vehicle]; p<0.025). Renal vascular resistance (RVR) was significantly higher in rats given CyA compared to animals treated with CyA vehicle (5.32±0.55 [cyCyA] vs 3.54±0.24 mm Hg/min/ml/kg [vehicle]; p<0.05). These hemodynamic alterations were associated with a significant increase in urinary excretion of unmetabolized, “native” thromboxane B₂ (TxB₂ (103±18 [CyA] vs 60±16 pg/hour [vehicle]; p<0.05). Only minimal histomorphologic changes were apparent by light microscopic examination of kidneys from both CyA and vehicle treated animals. However, with immunoperoxidase staining, a significantly greater number of cells experssing the rat common leukocyte antigen was found in the renal interstitium of rats given CyA^*. There was no detectable increase in monocytes/macrophages in the kidneys of CyA toxic animals. In rats treated with CyA, intraarterial infusion of GR32191 at maximally tolerated doses significanlty increased GFR and RBD, and decreased RVR. Although both RBF and RVR were restored to levels not different from controls, GFR remained significantly reduced following administration of GR32191. These data suggest that the potent vasoconstrictor TxA₂ plays an important role in mediating renal dysfunction in CyA nephrotoxicity. However, other factors may be important in producing nephrotoxicity associated with CyA.     

A physiological analogy of the niche for projecting the potential distribution of plants

Aim  To develop a physiologically based model of the plant niche for use in species distribution modelling. Location  Europe. Methods  We link the Thornley transport resistance (TTR) model with functions which describe how the TTR’s model parameters are influenced by abiotic environmental factors. The TTR model considers how carbon and nutrient uptake, and the allocation of these assimilates, influence growth. We use indirect statistical methods to estimate the model parameters from a high resolution data set on tree distribution for 22 European tree species. Results  We infer, from distribution data and abiotic forcing data, the physiological niche dimensions of 22 European tree species. We found that the model fits were reasonable (AUC: 0.79–0.964). The projected distributions were characterized by a false positive rate of 0.19 and a false negative rate 0.12. The fitted models are used to generate projections of the environmental factors that limit the range boundaries of the study species. Main conclusions  We show that physiological models can be used to derive physiological niche dimensions from species distribution data. Future work should focus on including prior information on physiological rates into the parameter estimation process. Application of the TTR model to species distribution modelling suggests new avenues for establishing explicit links between distribution and physiology, and for generating hypotheses about how ecophysiological processes influence the distribution of plants.     

Homodimerization as a molecular switch between low and high efficiency PrPC cell surface delivery and neuroprotective activity

PrP^C is associated with a variety of functions, and its ability to interact with a multitude of partners, including itself, may largely explain PrP multifunctionality and the lack of consensus on the genuine physiological function of the protein in vivo. In contrast, there is a consensus in the literature that alterations in PrP^C trafficking and intracellular retention result in neuronal degeneration. In addition, a proteolytic modification in the late secretory pathway termed the α-cleavage induces the secretion of PrPN1, a PrP^C-derived metabolite with fascinating neuroprotective activity against toxic oligomeric Aβ molecules implicated in Alzheimer disease. Thus, studies focusing on understanding the regulation of PrP^C trafficking to the cell surface and the modulation of α-cleavage are essential. The objective of this commentary is to highlight recent evidences that PrP^C homodimerization stimulates trafficking of the protein to the cell surface and results in high levels of PrPN1 secretion. We also discuss a hypothetical model for these results and comment on future challenges and opportunities.