In situ conformation of spider silk proteins in the intact major ampullate gland and in solution

To understand the spinning process of dragline silk by spiders, the protein conformation before spinning has to be determined. Raman confocal spectromicroscopy has been used to study the conformation of the proteins in situ in the intact abdominal major ampullate gland of Nephila clavipes and Araneus diadematus spiders. The spectra obtained are typical of natively unfolded proteins and are very similar to that of a mixture of recombinant silk proteins. Vibrational circular dichroism reveals that the conformation is composed of random and polyproline II (PPII) segments with some alpha-helices. The alpha-helices seem to be located in the C-terminal part whereas the repetitive sequence is unfolded. The PPII structure can significantly contribute to the efficiency of the spinning process in nature.     

Effect of coculturing canine notochordal,nucleus pulposus and mesenchymal stromal cells for intervertebral disc regeneration

IntroductionEarly degenerative changes in the nucleus pulposus (NP) are observed after the disappearance of notochordal cells (NCs). Thus, it has been suggested that NCs play an important role in maintaining the NP and may have a regenerative potential on other cells of the NP. As the number of resident NP cells (NPCs) decreases in a degenerating disc, mesenchymal stromal (stem) cells (MSCs) may be used for cell supplementation. In this study, using cells of one species, the regenerative potential of canine NCs was assessed in long-term three-dimensional coculture with canine NPCs or MSCs.MethodsCanine NCs and canine NPCs or MSCs were cocultured in alginate beads for 28 days under hypoxic and high-osmolarity conditions. Cell viability, cell morphology and DNA content, extracellular matrix production and expression of genes related to NC markers (Brachyury, KRT18) and NP matrix production (ACAN, COL2A1, COL1A1) were assessed after 1, 15 and 28 days of culture.ResultsNCs did not completely maintain their phenotype (morphology, matrix production, gene expression) during 28 days of culture. In cocultures of NPCs and NCs, both extracellular matrix content and anabolic gene expression remained unchanged compared with monoculture groups, whereas cocultures of MSCs and NCs showed increased glycosaminoglycan/DNA. However, the deposition of these proteoglycans was observed near the NCs and not the MSCs. Brachyury expression in the MSC and NC coculture group increased in time. The latter two findings indicate a trophic effect of MSCs on NCs rather than vice versa.ConclusionsNo regenerative potential of canine NCs on canine NPCs or MSCs was observed in this study. However, significant changes in NC phenotype in long-term culture may have resulted in a suboptimal regenerative potential of these NCs. In this respect, NC-conditioned medium may be better than coculture for future studies of the regenerative potential of NCs.

Electronic supplementary material

The online version of this article (doi:10.1186/s13075-015-0569-6) contains supplementary material, which is available to authorized users.     

Naphthalene/quinoline amides and sulfonylureas as potent and selective antagonists of the EP4 receptor

Two new series of EP₄ antagonists based on naphthalene/quinoline scaffolds have been identified as part of our on-going efforts to develop treatments for inflammatory pain. One series contains an acidic sulfonylurea pharmacophore, whereas the other is a neutral amide. Both series show subnanomolar intrinsic binding potency towards the EP₄ receptor, and excellent selectivity towards other prostanoid receptors. While the amide series generally displays poor pharmacokinetic parameters, the sulfonylureas exhibit greatly improved profile. MF-592, the optimal compound from the sulfonylurea series, has a desirable overall preclinical profile that suggests it is suitable for further development.     

The identification of substituted benzothiophene derivatives as PGE(2) subtype 4 receptor antagonists: From acid to non-acid

We disclose herein our preliminary SAR study on the identification of substituted benzothiophene derivatives as PGE₂ subtype 4 receptor antagonists. A potent EP₄ antagonist 6a (K_i = 1.4 nM with 10% HSA) was identified. Furthermore, we found that an acidic group was not essential for the EP₄ antagonizing activity in the series and neutral replacements were identified. This opens a new direction for future EP₄ antagonist design.     

Sensory experience differentially modulates the mRNA expression of the polysialyltransferases ST8SiaII and ST8SiaIV in postnatal mouse visual cortex

Polysialic acid (PSA) is a unique carbohydrate composed of a linear homopolymer of α-2,8 linked sialic acid, and is mainly attached to the fifth immunoglobulin-like domain of the neural cell adhesion molecule (NCAM) in vertebrate neural system. In the brain, PSA is exclusively synthesized by the two polysialyltransferases ST8SiaII (also known as STX) and ST8SiaIV (also known as PST). By modulating adhesive property of NCAM, PSA plays a critical role in several neural development processes such as cell migration, neurite outgrowth, axon pathfinding, synaptogenesis and activity-dependent plasticity. The expression of PSA is temporally and spatially regulated during neural development and a tight regulation of PSA expression is essential to its biological function. In mouse visual cortex, PSA is downregulated following eye opening and its decrease allows the maturation of GABAergic synapses and the opening of the critical period for ocular dominance plasticity. Relatively little is known about how PSA levels are regulated by sensory experience and neuronal activity. Here, we demonstrate that while both ST8SiaII and ST8SiaIV mRNA levels decrease around the time of eye opening in mouse visual cortex, only ST8SiaII mRNA level reduction is regulated by sensory experience. Using an organotypic culture system from mouse visual cortex, we further show that ST8SiaII gene expression is regulated by spiking activity and NMDA-mediated excitation. Further, we show that both ST8SiaII and ST8SiaIV mRNA levels are positively regulated by PKC-mediated signaling. Therefore, sensory experience-dependent ST8SiaII gene expression regulates PSA levels in postnatal visual cortex, thus acting as molecular link between visual activity and PSA expression.     

Étude chimiotaxonomique de deux espéces nouvelles de Hazunta (Apocynaceae)

Twenty-two indole alkaloids have been isolated from two new Hazunta species (Apocynaceae). Apparently, five of them are bis-indoles of a previously unknown structure.     

Systematic position of some nearctic Heligmosomoidea (Nematoda: Trichostrongylina) from the U.S. National Parasite collection and their description

The systematic position of some heligmosomoid nematodes from rodents, deposited in the U.S. National Parasite Collection (USNPC), is revised, mainly through the study of their synlophe, which in all cases was unknown or insufficiently described. The material was registered as different species of Longistriata Schulz, 1926, a genus whose representatives are only parasitic in Holarctic insectivores. Longistriata norvegica Dikmans, 1935, parasitic in Rattus sp. becomes a synonym of Hassalstrongylus aduncus (Chandler, 1932). Specimens registered as Longistriata dalrymplei Dikmans, 1935, from Ondatra zibethicus, are confirmed to belong to Carolinensis (Travassos, 1937). Specimens registered as Longistriata noviberiae Dikmans, 1935, parasitic in Sylvilagus floridanus alacer, were found to belong to Vexillata, and Vexillata noviberiae n. comb. is here proposed. This is the first record of a species of Vexillata in a lagomorph. Other specimens registered as Longistriata norvegica, parasitic in Geomys floridanus austrinus, were also found to be an undescribed species of Vexillata (Hall, 1916), which is named Vexillata chitwoodi n. sp. This is similar to Vexillata chabaudi Yoyotte-Vado, 1972, Vexillata petteri Durette-Desset, 1970, Vexillata scorzai Guerrero, 1984 and Vexillata tejerai Guerrero, 1984, all having the same number of cuticular ridges (4 dorsal, 5 ventral) and the division of the dorsal ray at its apex. The most related species is V. chabaudi, which is differentiated from the new species by rays 4 not curved distally, by thick rays 8, and by a dorsal ray enlarged at the level of the arising of rays 8.