From a peptide lead to an orally active peptidomimetic fibrinogen receptor antagonist

Antagonists of the platelet fibrinogen receptor (GP IIb/IIIa receptor) are expected to be a new promising class of antithrombotic agents. The binding of fibrinogen to the fibrinogen receptor depends on an Arg-Gly-Asp-Ser (RGDS) tetrapeptide recognition motif. Structural modifications of the RGDS lead have led to the discovery of a non-peptide RGD mimetic GP IIb/IIIa antagonist 20 (S 1197). Compound 20 inhibits dose-dependently and reversibly human platelet aggregation. Modeling studies based on structure–activity data revealed the following structural features of the drug as important for receptor binding: the amidino group, the carboxylate group, hydrophobic substitutions at the carboxyl-terminus and at the side chain carrying the positive charge, the carboxyl-terminal NH group of the -amino acid as a hydrogen bond donor and one oxygen atom of the hydantoin as a hydrogen bond acceptor. The ethyl ester prodrug of 20 (S 5740) is an orally active antithrombotic agent which has the potential to be used to treat and prevent thrombotic diseases in humans.     

2′-phosphodiesterase and 2′,5′-oligoadenylate synthetase activities in the lowest metazoans,sponge [porifera]

Sponges [porifera], the most ancient metazoans, contain modules related to the vertebrate immune system, including the 2′,5′-oligoadenylate synthetase (OAS). The components of the antiviral 2′,5′-oligoadenylate (2–5A) system (OAS, 2′-Phosphodiesterase (2′-PDE) and RNAse L) of vertebrates have not all been identified in sponges. Here, we demonstrate for the first time that in addition to the OAS activity, sponges possess a 2′-PDE activity, which highlights the probable existence of a premature 2–5A system. Indeed, Suberites domuncula and Crella elegans exhibited this 2–5A degrading activity. Upon this finding, two out of three elements forming the 2–5A system have been found in sponges, only a endoribonuclease, RNAse L or similar, has to be found. We suspect the existence of a complex immune system in sponges, besides the self/non-self recognition system and the use of phagocytosis and secondary metabolites against pathogens.     

Cryotherapy with liquid nitrogen versus topical salicylic acid application for cutaneous warts in primary care: randomized controlled trial

Background

Cryotherapy is widely used for the treatment of cutaneous warts in primary care. However, evidence favours salicylic acid application. We compared the effectiveness of these treatments as well as a wait-and-see approach.

Methods

Consecutive patients with new cutaneous warts were recruited in 30 primary care practices in the Netherlands between May 1, 2006, and Jan. 26, 2007. We randomly allocated eligible patients to one of three groups: cryotherapy with liquid nitrogen every two weeks, self-application of salicylic acid daily or a wait-and-see approach. The primary outcome was the proportion of participants whose warts were all cured at 13 weeks. Analysis was on an intention-to-treat basis. Secondary outcomes included treatment adherence, side effects and treatment satisfaction. Research nurses assessed outcomes during home visits at 4, 13 and 26 weeks.

Results

Of the 250 participants (age 4 to 79 years), 240 were included in the analysis at 13 weeks (loss to follow-up 4%). Cure rates were 39% (95% confidence interval [CI] 29%–51%) in the cryotherapy group, 24% (95% CI 16%–35%) in the salicylic acid group and 16% (95% CI 9.5%–25%) in the wait-and-see group. Differences in effectiveness were most pronounced among participants with common warts (n = 116): cure rates were 49% (95% CI 34%–64%) in the cryotherapy group, 15% (95% CI 7%–30%) in the salicylic acid group and 8% (95% CI 3%–21%) in the wait-and-see group. Cure rates among the participants with plantar warts (n = 124) did not differ significantly between treatment groups.

Interpretation

For common warts, cryotherapy was the most effective therapy in primary care. For plantar warts, we found no clinically relevant difference in effectiveness between cryotherapy, topical application of salicylic acid or a wait-and-see approach after 13 weeks. (ClinicalTrial.gov registration no. ISRCTN42730629)Cutaneous warts are common.^1–3 Up to one-third of primary school children have warts, of which two-thirds resolve within two years.^4,5 Because warts frequently result in discomfort,⁶ 2% of the general population and 6% of school-aged children each year present with warts to their family physician.^7,8 The usual treatment is cryotherapy with liquid nitrogen or, less frequently, topical application of salicylic acid.^9–12 Some physicians choose a wait-and-see approach because of the benign natural course of warts and the risk of side effects of treatment.^10,11A recent Cochrane review on treatments of cutaneous warts concluded that available studies were small, poorly designed or limited to dermatology outpatients.^10,11 Evidence on cryotherapy was contradictory,^13–18 whereas the evidence on salicylic acid was more convincing.^19–23 However, studies that compared cryotherapy and salicylic acid directly showed no differences in effectiveness.^24,25 The Cochrane review called for high-quality trials in primary care to compare the effects of cryotherapy, salicylic acid and placebo.We conducted a three-arm randomized controlled trial to compare the effectiveness of cryotherapy with liquid nitrogen, topical application of salicylic acid and a wait-and-see approach for the treatment of common and plantar warts in primary care.     

Inhibition of amylase secretion from differentiated AR4-2J pancreatic acinar cells by an actin cytoskeleton controlled protein tyrosine phosphatase activity.

Disruption of the actin cytoskeleton in AR4-2J pancreatic acinar cells led to an increase in cytosolic protein tyrosine phosphatase activity, abolished bombesin-induced tyrosine phosphorylation and reduced bombesin-induced amylase secretion by about 45%. Furthermore, both tyrosine phosphorylation and amylase secretion induced by phorbol ester-induced activation of protein kinase C were abolished. An increase in the cytosolic free Ca2+ concentration by the Ca2+ ionophore A23187 had no effect on tyrosine phosphorylation but induced amylase release. Only when added together with phorbol ester, the same level of amylase secretion as with bombesin was reached. This amylase secretion was inhibited by about 40%, by actin cytoskeleton disruption similar to that induced by bombesin. We conclude that actin cytoskeleton-controlled protein tyrosine phosphatase activity downstream of protein kinase C activity regulates tyrosine phosphorylation which in part is involved in bombesin-stimulated amylase secretion.     

A compact unc45b‐promoter drives muscle‐specific expression in zebrafish and mouse

Summary: Gene therapeutic approaches to cure genetic diseases require tools to express the rescuing gene exclusively within the affected tissues. Viruses are often chosen as gene transfer vehicles but they have limited capacity for genetic information to be carried and transduced. In addition, to avoid off‐target effects the therapeutic gene should be driven by a tissue‐specific promoter in order to ensure expression in the target organs, tissues, or cell populations. The larger the promoter, the less space will be left for the respective gene. Thus, there is a need for small but tissue‐specific promoters. Here, we describe a compact unc45b promoter fragment of 195 bp that retains the ability to drive gene expression exclusively in skeletal and cardiac muscle in zebrafish and mouse. Remarkably, the described unc45b promoter fragment not only drives muscle‐specific expression but presents heat‐shock inducibility, allowing a temporal and spatial quantity control of (trans)gene expression. Here, we demonstrate that the transgenic expression of the smyd1b gene driven by the unc45b promoter fragment is able to rescue the embryonically lethal heart and skeletal muscle defects in smyd1b‐deficient flatline mutant zebrafish. Our findings demonstrate that the described muscle‐specific unc45b promoter fragment might be a valuable tool for the development of genetic therapies in patients suffering from myopathies. genesis 54:431–438, 2016. © 2016 The Authors. Genesis Published by Wiley Periodicals, Inc.     

RhoA regulates peroxisome association to microtubules and the actin cytoskeleton

The current view of peroxisome inheritance provides for the formation of new peroxisomes by both budding from the endoplasmic reticulum and autonomous division. Here we investigate peroxisome-cytoskeleton interactions and show by proteomics, biochemical and immunofluorescence analyses that actin, non-muscle myosin IIA (NMM IIA), RhoA, Rho kinase II (ROCKII) and Rab8 associate with peroxisomes. Our data provide evidence that (i) RhoA in its inactive state, maintained for example by C. botulinum toxin exoenzyme C3, dissociates from peroxisomes enabling microtubule-based peroxisomal movements and (ii) dominant-active RhoA targets to peroxisomes, uncouples the organelles from microtubules and favors Rho kinase recruitment to peroxisomes. We suggest that ROCKII activates NMM IIA mediating local peroxisomal constrictions. Although our understanding of peroxisome-cytoskeleton interactions is still incomplete, a picture is emerging demonstrating alternate RhoA-dependent association of peroxisomes to the microtubular and actin cytoskeleton. Whereas association of peroxisomes to microtubules clearly serves bidirectional, long-range saltatory movements, peroxisome-acto-myosin interactions may support biogenetic functions balancing peroxisome size, shape, number, and clustering.     

Increased tolerance of Litopenaeus vannamei to white spot syndrome virus (WSSV) infection after oral application of the viral envelope protein VP28

It has been generally accepted that invertebrates such as shrimp do not have an adaptive immune response system comparable to that of vertebrates. However, in the last few years, several studies have suggested the existence of such a response in invertebrates. In one of these studies, the shrimp Penaeus monodon showed increased protection against white spot syndrome virus (WSSV) using a recombinant VP28 envelope protein of WSSV. In an effort to further investigate whether this increased protection is limited to P. monodon or can be extended to other penaeid shrimp, experiments were performed using the Pacific white shrimp Litopenaeus vannamei. As found with P. monodon, a significantly lower cumulative mortality for VP28-fed shrimp was found compared to the controls. These experiments demonstrate that there is potential to use oral application of specific proteins to protect the 2 most important cultured shrimp species, P. monodon and L. vannamei, against WSSV. Most likely, this increased protection is based on a shared and, therefore, general defence mechanism present in all shrimp species. This makes the design of intervention strategies against pathogens based on defined proteins a viable option for shrimp culture.     

Local and relational judgements of surface colour: constancy indices and discrimination performance

Colour constancy is generally assumed to arise from a combination of perceptual constancy mechanisms operating to partially discount illumination changes and relational mechanisms involved in judging the colour relationships between object surfaces. Here we examined the characteristics of these mechanisms using a 'yes/no' task. Subjects judged whether a target colour patch embedded in an array of coloured patches (a) stayed the same across a simulated temporal illuminant change (local colour judgement), or (b) changed in a manner consistent with the illuminant change (relational colour judgement). The colour of the target patch remained constant in one-third of the trials, changed in accord with the illuminant shift in another third, and shifted partially with the illuminant change in the remaining third. We found that perceptual constancy was relatively weak and relational constancy strong, as assessed using a modified colour constancy index. Randomising the spatial positions of coloured patches across the illuminant change did not affect subjects' constancy indices. Application of signal detection analysis revealed some otherwise hidden effects. In the case of relational judgements, subjects adopted more conservative criteria (fewer true and false positives) with randomisation, maintaining a constant level of discrimination performance (d'). For local judgements, randomisation led to small increases in performance but no changes in criteria. We conclude that signal detection theory provides a useful tool to supplement conventional approaches to understanding colour constancy.