Functional domains of the human epididymal protease inhibitor, eppin

Eppin has two potential protease inhibitory domains: a whey acid protein or four disulfide core domain and a Kunitz domain. The protein is also reported to have antibacterial activity against Gram-negative bacteria. Eppin and its whey acid protein and Kunitz domains were expressed in Escherichia coli and their ability to inhibit proteases and kill bacteria compared. The Kunitz domain inhibits elastase (EC 3.4.21.37) to a similar extent as intact eppin, whereas the whey acid protein domain has no such activity. None of these fragments inhibits trypsin (EC 3.4.21.4) or chymotrypsin (EC 3.4.21.1) at the concentrations tested. In a colony forming unit assay, both domains have some antibacterial activity against E. coli, but this was not to the same degree as intact eppin or the two domains together. When bacterial respiratory electron transport was measured using a 2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide assay, eppin and its domains caused an increase in the rate of respiration. This suggests that the mechanism of cell killing may be partly through the permeablization of the bacterial inner membrane, resulting in uncoupling of respiratory electron transport and consequent collapse of the proton motive force. Thus, we conclude that although both of eppin's domains are involved in the protein's antibacterial activity, only the Kunitz domain is required for selective protease inhibition.     

Investigation of the effect of Interleukin-1 receptor antagonist (IL-1ra) on markers of inflammation in non-ST elevation acute coronary syndromes (The MRC-ILA-HEART Study)

Background

Point of care testing (PoCT) may be a useful adjunct in the management of chronic conditions in general practice (GP). The provision of pathology test results at the time of the consultation could lead to enhanced clinical management, better health outcomes, greater convenience and satisfaction for patients and general practitioners (GPs), and savings in costs and time. It could also result in inappropriate testing, increased consultations and poor health outcomes resulting from inaccurate results. Currently there are very few randomised controlled trials (RCTs) in GP that have investigated these aspects of PoCT.

Design/Methods

The Point of Care Testing in General Practice Trial (PoCT Trial) was an Australian Government funded multi-centre, cluster randomised controlled trial to determine the safety, clinical effectiveness, cost effectiveness and satisfaction of PoCT in a GP setting. The PoCT Trial covered an 18 month period with the intervention consisting of the use of PoCT for seven tests used in the management of patients with diabetes, hyperlipidaemia and patients on anticoagulant therapy. The primary outcome measure was the proportion of patients within target range, a measure of therapeutic control. In addition, the PoCT Trial investigated the safety of PoCT, impact of PoCT on patient compliance to medication, stakeholder satisfaction, cost effectiveness of PoCT versus laboratory testing, and influence of geographic location.

Discussion

The paper provides an overview of the Trial Design, the rationale for the research methodology chosen and how the Trial was implemented in a GP environment. The evaluation protocol and data collection processes took into account the large number of patients, the broad range of practice types distributed over a large geographic area, and the inclusion of pathology test results from multiple pathology laboratories. The evaluation protocol developed reflects the complexity of the Trial setting, the Trial Design and the approach taken within the funding provided. The PoCT Trial is regarded as a pragmatic RCT, evaluating the effectiveness of implementing PoCT in GP and every effort was made to ensure that, in these circumstances, internal and external validity was maintained.

Trial Registration

12612605000272695     

Localization of Protein Aggregation in Escherichia coli Is Governed by Diffusion and Nucleoid Macromolecular Crowding Effect

Aggregates of misfolded proteins are a hallmark of many age-related diseases. Recently, they have been linked to aging of Escherichia coli (E. coli) where protein aggregates accumulate at the old pole region of the aging bacterium. Because of the potential of E. coli as a model organism, elucidating aging and protein aggregation in this bacterium may pave the way to significant advances in our global understanding of aging. A first obstacle along this path is to decipher the mechanisms by which protein aggregates are targeted to specific intercellular locations. Here, using an integrated approach based on individual-based modeling, time-lapse fluorescence microscopy and automated image analysis, we show that the movement of aging-related protein aggregates in E. coli is purely diffusive (Brownian). Using single-particle tracking of protein aggregates in live E. coli cells, we estimated the average size and diffusion constant of the aggregates. Our results provide evidence that the aggregates passively diffuse within the cell, with diffusion constants that depend on their size in agreement with the Stokes-Einstein law. However, the aggregate displacements along the cell long axis are confined to a region that roughly corresponds to the nucleoid-free space in the cell pole, thus confirming the importance of increased macromolecular crowding in the nucleoids. We thus used 3D individual-based modeling to show that these three ingredients (diffusion, aggregation and diffusion hindrance in the nucleoids) are sufficient and necessary to reproduce the available experimental data on aggregate localization in the cells. Taken together, our results strongly support the hypothesis that the localization of aging-related protein aggregates in the poles of E. coli results from the coupling of passive diffusion-aggregation with spatially non-homogeneous macromolecular crowding. They further support the importance of “soft” intracellular structuring (based on macromolecular crowding) in diffusion-based protein localization in E. coli.     

Hydrogel-Forming Microneedles Prepared from “Super Swelling” Polymers Combined with Lyophilised Wafers for Transdermal Drug Delivery

We describe, for the first time, hydrogel-forming microneedle arrays prepared from “super swelling” polymeric compositions. We produced a microneedle formulation with enhanced swelling capabilities from aqueous blends containing 20% w/w Gantrez S-97, 7.5% w/w PEG 10,000 and 3% w/w Na₂CO₃ and utilised a drug reservoir of a lyophilised wafer-like design. These microneedle-lyophilised wafer compositions were robust and effectively penetrated skin, swelling extensively, but being removed intact. In in vitro delivery experiments across excised neonatal porcine skin, approximately 44 mg of the model high dose small molecule drug ibuprofen sodium was delivered in 24 h, equating to 37% of the loading in the lyophilised reservoir. The super swelling microneedles delivered approximately 1.24 mg of the model protein ovalbumin over 24 h, equivalent to a delivery efficiency of approximately 49%. The integrated microneedle-lyophilised wafer delivery system produced a progressive increase in plasma concentrations of ibuprofen sodium in rats over 6 h, with a maximal concentration of approximately 179 µg/ml achieved in this time. The plasma concentration had fallen to 71±6.7 µg/ml by 24 h. Ovalbumin levels peaked in rat plasma after only 1 hour at 42.36±17.01 ng/ml. Ovalbumin plasma levels then remained almost constant up to 6 h, dropping somewhat at 24 h, when 23.61±4.84 ng/ml was detected. This work represents a significant advancement on conventional microneedle systems, which are presently only suitable for bolus delivery of very potent drugs and vaccines. Once fully developed, such technology may greatly expand the range of drugs that can be delivered transdermally, to the benefit of patients and industry. Accordingly, we are currently progressing towards clinical evaluations with a range of candidate molecules.     

菲律宾Boracay岛和Mambukal岛三种共栖狐蝠对人类活动的行为和激素反应

栖息地丧失和捕猎导致许多大型狐蝠濒危。尽管在东半球热带地区已经禁止捕猎和采伐,狐蝠的数量仍然在下降。既能维护当地居民利益又能保护狐蝠的折衷对策是生态旅游。然而,人类活动对狐蝠的影响还是未知的。菲律宾Boracay和Mambukal都是旅游区,前者游人少而稳定,后者游人密集且变异较大。我们用非损伤取样法研究了这两个旅游区三种狐蝠的生理紧张情况。在Mambukal,当狐蝠外出采食时,采集狐蝠的粪样,实验室分析粪样肾上腺皮质激素代谢物(GCM)浓度。我们观察记录了紧张反应等一些行为,同时记录了环境因素和人类活动情况。结果表明狐蝠已经习惯了游人的活动,表现在:(1)我们发现两个旅游区游人数量和活动强度差异显著,但是两地狐蝠的行为没有明显的差异;并且在有干扰和无干扰的日期之间,狐蝠的行为也无显著差异;两个旅游区的狐蝠行为表达一致;(2)各种行为与测定的粪样GCM浓度都不相关;(3)尽管在不同研究地点人类活动变异很大,我们发现人类活动并未影响粪样GCM浓度;不过,栖息在狐蝠群中心位置个体的粪样GCM浓度低于在群外围个体的GCM浓度;(4)一些环境因素(如干扰以及与栖息群的距离)影响狐蝠一些行为(如不安、身体护理和哈欠)的表达。如果干扰没有造成狐蝠紧张反应,那么生态旅游将不失为一种保护濒危狐蝠栖息地的理想方案。不过,我们还应认识到,人类干扰对狐蝠行为的影响可能比本研究观察到的更加复杂。因此,对于保护濒危狐蝠,时刻保持干扰最小是最好的选择。     

Laboratory evaluation of certain natural compounds against the melon ladybird beetle,Epilachna chrysomelina F. attacking cucurbit plants

Neem Azal-T/S, Neudosan and Spruzit flüssig at different concentrations were evaluated against the second and fourth instar larvae of E. chrysomelina under laboratory conditions. In case of acute effects, the highest mortality was recorded when larvae were offered squash leaves treated with the highest concentration (2%) of Neem Azal-T/S, and the lowest mortality was, however recorded after treating them with the lowest concentration (0.25%). On the other hand, the second instar larvae were more susceptible to any compound at any concentration than the fourth instar one. One per cent Neudosan formulation showed the highest toxicity to both the 2nd and 4th instar larvae. As the concentration of this compound decreased the mortality percentage of treated larvae decreased to reach the minimum at the lowest concentration (0.25%). The same trend could be applied for Spruzit flüssig compound. When Neem Azal-T/S was compared with Neudosan, the later at any concentration was more toxic than the former. Generally, Neudosan and Spruzit flüssig seemed to be active against E. chrysomelina during the first three days after treatment, then, deterioration occurred among this compound. In case of latent effects, the duration of the treated fourth instar larvae of E. chrysomelina with any tested compound, was prolonged. An increase in the concentration caused prolongation of this period to reach the longest at the highest concentration. This prolongation, was more pronounced after Neem Azal-T/S treatment followed by Neudosan and Spruzit flüssig. The same trend could be applied for the duration of the pupal stage as being affected by the different concentrations of the tested compounds which were applied to the fourth instar larvae. However, adults of E. Chrysomelina treated as fourth instar larvae with high concentrations of Neem Azal-T/S showed some adult malformations. The sex ratios of adults produced from fourth instar larvae treated with different concentrations of the tested natural compounds were about 1?:?1, except for two cases; 1% Neudosan where the number of females were double the number of males and 0.5% Spruzit flüssig where the number of males were double the number of females. An increase in the concentration of any tested compound caused an obvious decrease in the percentage of emerged adults. The least emergence percentage was recorded in the case of Spruzit flüssig, followed by Neudosan and Neem Azal-T/S. Treating the fourth instar larvae with the different concentrations of the tested natural compounds significantly shortened the longevity of produced adults and decreased the fecundity.     

Can the presence of curved forms of the diatom Aulacoseira ambigua in the Nile (Egypt) and Vaal (South Africa) Rivers be ascribed to similar water quality conditions?

Spiral colonies of the diatom Aulacoseira ambigua were assigned the rank of forma (Aulacoseira ambigua f. japonica). This spiral-shaped, colonial, centric diatom has limited geographical distribution and is currently reported to occur in only a few countries in the world. In Africa this species was described for the first time from the Vaal River, South Africa, but recent research on the Nile River also revealed its presence in Egypt. Physical and chemical data on water quality in these two river systems were compared to determine whether the presence of this uncommon ‘phenoecodeme’ could be ascribed to similar environmental conditions. Results indicate that the rivers are dissimilar with regard to many variables, but both rivers provide turbid, warm and eutrophic waters with medium to high mineral content and it was concluded that these factors favour and sustain the growth of this curved form.     

Weight loss is not mandatory for exercise-induced effects on health indices in females with metabolic syndrome

The aim of this study was to investigate the impact of moderate aerobic training on functional, anthropometric, biochemical, and health-related quality of life (HRQOL) parameters on women with metabolic syndrome (MS). Fifteen untrained women with MS performed moderate aerobic training for 15 weeks, without modifications of dietary behaviours. Functional, anthropometric, biochemical, control diet record and HRQOL parameters were assessed before and after the training. Despite body weight maintenance, the patients presented decreases in waist circumference (P = 0.001), number of MS components (P = 0.014), total cholesterol (P = 0.049), HDL cholesterol (P = 0.004), LDL cholesterol (P = 0.027), myeloperoxidase activity (P = 0.002) and thiobarbituric acid-reactive substances levels (P = 0.006). There were no differences in total energy, carbohydrate, protein and lipid intake pre- and post-training. Furthermore, improvements in the HRQOL subscales of physical functioning (P = 0.03), role-physical (P = 0.039), bodily pain (P = 0.048), general health (P = 0.046) and social functioning scoring (P = 0.011) were reported. Despite the absence of weight loss, aerobic training induced beneficial effects on functional, anthropometric, biochemical and HRQOL parameters in women with MS.