Lethal effect of protamine and histone on competent Bacillus subtilis cells. Inhibition of genetic transformation by protamine in sublethal concentration.

Summary Under experimental conditions of genetic transformation, protamine and total histone were bactericidal for Bacillus subtilis cells. The abilities to cause lethality were very similar for both, either protamine or histone, with no antagonistic effects amongst these natural polycations. With both basic proteins acting simultaneously the enhancement was higher than a summation of the separate lethal effects. Sublethal concentration of protamine added at the beginning of transformation time, produced a strong inhibition of transforming efficiency. The same concentration added later than 10 min from the start of transformation had no inhibitory effect. These facts together with the absence of inhibition by simple pretreatment of DNA alone as well as the cell protection by protamine against lytic activity of lysozyme, suggest a protamine-cell surface interaction which impedes DNA uptake events.     

Endogenous acid ceramidase protects epithelial cells from Porphyromonas gingivalis-induced inflammation in vitro

Ceramidases are a group of enzymes that degrade pro-inflammatory ceramide by cleaving a fatty acid to form anti-inflammatory sphingosine lipid. Thus far, acid, neutral and alkaline ceramidase isozymes have been described. However, the expression patterns of ceramidase isoforms as well as their role in periodontal disease pathogenesis remain unknown. In this study, expression patterns of ceramidase isoforms were quantified by real-time PCR and immunohistochemistry in gingival samples of patients with periodontitis and healthy subjects, as well as in EpiGingivalTM-3D culture and OBA-9 gingival epithelial cells both of which were stimulated with or without the presence of live Porphyromonas gingivalis (ATCC 33277 strain). A significantly lower level of acid ceramidase expression was detected in gingival tissues from periodontal patients compared to those from healthy subjects. In addition, acid-ceramidase expression in EpiGingival? 3D culture and OBA-9?cells was suppressed by stimulation with P. gingivalis in vitro. No significant fluctuation was detected for neutral or alkaline ceramidases in either gingival samples or cell cultures. Next, to elucidate the role of acid ceramidase in P. gingivalis-induced inflammation in vitro, OBA-9?cells were transduced with adenoviral vector expressing the human acid ceramidase (Ad-ASAH1) gene or control adenoviral vector (Ad-control). In response to stimulation with P. gingivalis, ASAH1-over-expressing OBA-9?cells showed significantly lower mRNA expressions of caspase-3 as well as the percentage of Annexin V-positive cells, when compared with OBA-9?cells transduced with Ad-control vector. Furthermore, in response to stimulation with P. gingivalis, ASAH1-over-expressing OBA-9?cells produced less TNF-α, IL-6, and IL1β pro-inflammatory cytokines than observed in OBA-9?cells transduced with Ad-control vector. Collectively, our data show the novel discovery of anti-inflammatory and anti-apoptotic effects of acid ceramidase in host cells exposed to periodontal bacteria, and the attenuation of the expression of host-protective acid ceramidase in periodontal lesions.     

How old are you—Evaluation of age reading methods for the invasive round goby (Neogobius melanostomus,Pallas 1814)

In the Baltic, the first observation of the round goby (Neogobius melanostomus, Pallas 1814) was made in 1990. Within the past decade the species became invasive and spread rapidly throughout the Baltic Sea. Studies about the fishes potential impacts on resident species promote the need for an increasing knowledge of their basic stock structures such as growth rates, longevity and mortality, which all rely on accurate estimates of age. Former studies on the round goby have used several different age reading techniques. In this study, we compared three standard otolith preparation methods for ageing and present the best procedure for the invasive round goby. The results showed significant differences in age estimates of the same fish between the different preparation methods and between readers. The estimation of the first annulus, the first year, was the most problematic. The overall agreement was lowest when reading the whole otoliths while the best performance was achieved with sectioned and stained preparation method. Depending on method used the growth estimates also differed. The results question comparability between previous studies and highlight the importance of harmonised aging procedures for the round goby for obtaining correct estimates of population parameters such as growth rate, age at maturity, and longevity.     

Human amylin proteotoxicity impairs protein biosynthesis,and alters major cellular signaling pathways in the heart,brain and liver of humanized diabetic rat model in vivo

Introduction

Chronic hypersecretion of the 37 amino acid amylin is common in type 2 diabetics (T2D). Recent studies implicate human amylin aggregates cause proteotoxicity (cell death induced by misfolded proteins) in both the brain and the heart.

Objectives

Identify systemic mechanisms/markers by which human amylin associated with cardiac and brain defects might be identified.

Methods

We investigated the metabolic consequences of amyloidogenic and cytotoxic amylin oligomers in heart, brain, liver, and plasma using non-targeted metabolomics analysis in a rat model expressing pancreatic human amylin (HIP model).

Results

Four metabolites were significantly different in three or more of the four compartments (heart, brain, liver, and plasma) in HIP rats. When compared to a T2D rat model, HIP hearts uniquely had significant DECREASES in five amino acids (lysine, alanine, tyrosine, phenylalanine, serine), with phenylalanine decreased across all four tissues investigated, including plasma. In contrast, significantly INCREASED circulating phenylalanine is reported in diabetics in multiple recent studies.

Conclusion

DECREASED phenylalanine may serve as a unique marker of cardiac and brain dysfunction due to hyperamylinemia that can be differentiated from alterations in T2D in the plasma. While the deficiency in phenylalanine was seen across tissues including plasma and could be monitored, reduced tyrosine was seen only in the brain. The 50 % reduction in phenylalanine and tyrosine in HIP brains is significant given their role in supporting brain chemistry as a precursor for catecholamines (dopamine, norepinephrine, epinephrine), which may contribute to the increased morbidity and mortality in diabetics at a multi-system level beyond the effects on glucose metabolism.

     

β cell membrane remodelling and procoagulant events occur in inflammation‐driven insulin impairment: a GLP‐1 receptor dependent and independent control

Inflammation and hyperglycaemia are associated with a prothrombotic state. Cell‐derived microparticles (MPs) are the conveyors of active procoagulant tissue factor (TF) and circulate at high concentration in diabetic patients. Liraglutide, a glucagon‐like peptide (GLP)‐1 analogue, is known to promote insulin secretion and β‐cell preservation. In this in vitro study, we examined the link between insulin impairment, procoagulant activity and plasma membrane remodelling, under inflammatory conditions. Rin‐m5f β‐cell function, TF activity mediated by MPs and their modulation by 1 μM liraglutide were examined in a cell cross‐talk model. Methyl‐β‐cyclodextrine (MCD), a cholesterol depletor, was used to evaluate the involvement of raft on TF activity, MP shedding and insulin secretion as well as Soluble N‐éthylmaleimide‐sensitive‐factor Attachment protein Receptor (SNARE)‐dependent exocytosis. Cytokines induced a two‐fold increase in TF activity at MP surface that was counteracted by liraglutide. Microparticles prompted TF activity on the target cells and a two‐fold decrease in insulin secretion via protein kinase A (PKA) and p38 signalling, that was also abolished by liraglutide. Large lipid raft clusters were formed in response to cytokines and liraglutide or MCD‐treated cells showed similar patterns. Cells pre‐treated by saturating concentration of the GLP‐1r antagonist exendin (9‐39), showed a partial abolishment of the liraglutide‐driven insulin secretion and liraglutide‐decreased TF activity. Measurement of caspase 3 cleavage and MP shedding confirmed the contribution of GLP‐1r‐dependent and ‐independent pathways. Our results confirm an integrative β‐cell response to GLP‐1 that targets receptor‐mediated signalling and membrane remodelling pointing at the coupling of insulin secretion and inflammation‐driven procoagulant events.     

Integration and scaling of UV‐B radiation effects on plants: from molecular interactions to whole plant responses

A process based model integrating the effects of UV‐B radiation to molecular level processes and their consequences to whole plant growth and development was developed from key parameters in the published literature. Model simulations showed that UV‐B radiation induced changes in plant metabolic and/or photosynthesis rates can result in plant growth inhibitions. The costs of effective epidermal UV‐B radiation absorptive compounds did not result in any significant changes in plant growth, but any associated metabolic costs effectively reduced the potential plant biomass. The model showed significant interactions between UV‐B radiation effects and temperature and any factor leading to inhibition of photosynthetic production or plant growth during the midday, but the effects were not cumulative for all factors. Vegetative growth were significantly delayed in species that do not exhibit reproductive cycles during a growing season, but vegetative growth and reproductive yield in species completing their life cycle in one growing season did not appear to be delayed more than 2–5 days, probably within the natural variability of the life cycles for many species. This is the first model to integrate the effects of increased UV‐B radiation through molecular level processes and their consequences to whole plant growth and development.     

Comparison of two methods for one-step in-straw thawing and direct transfer of cattle blastocysts

The aim of this experiment was to evaluate the feasibility of dilution of the cryoprotectant by a sucrose solution in the straw followed by direct transfer without any selection of the embryos. A comparison was made between the method described by Renard et al. (7) and a method slightly modified from that published by Leibo (8). The feasibility is proved by a mean pregnancy rate of 41.4%. Although the difference was not statistically significant, the method described by Renard et al. (7) led to a higher pregnancy rate (44.7%) than that of Leibo (8) (38.5%).     

Representatives of the genus Triploporella (Dasycladales, calcareous algae) in the Lower Cretaceous limestones of Romania

Recent studies on the Lower Cretaceous deposits located in various areas of the Romanian Carpathians resulted in the identification of several specimens of dasycladalean algae assigned to the genus Triploporella including Triploporella carpatica Bucur, Triploporella cf. praturlonii Barattolo, Triploporella cf. steinmannii Barattolo, Triploporella sp. 1, Triploporella sp. 2, and Triploporella n. sp. This paper provides arguments on their taxonomic assignment, together with discussions on the Triploporella species described in the literature and their paleobiogeographic significance.