α−α Cross-Links Increase Fibrin Fiber Elasticity and Stiffness

Fibrin fibers, which are ∼100 nm in diameter, are the major structural component of a blood clot. The mechanical properties of single fibrin fibers determine the behavior of a blood clot and, thus, have a critical influence on heart attacks, strokes, and embolisms. Cross-linking is thought to fortify blood clots; though, the role of α–α cross-links in fibrin fiber assembly and their effect on the mechanical properties of single fibrin fibers are poorly understood. To address this knowledge gap, we used a combined fluorescence and atomic force microscope technique to determine the stiffness (modulus), extensibility, and elasticity of individual, uncross-linked, exclusively α–α cross-linked (γQ398N/Q399N/K406R fibrinogen variant), and completely cross-linked fibrin fibers. Exclusive α–α cross-linking results in 2.5× stiffer and 1.5× more elastic fibers, whereas full cross-linking results in 3.75× stiffer, 1.2× more elastic, but 1.2× less extensible fibers, as compared to uncross-linked fibers. On the basis of these results and data from the literature, we propose a model in which the α-C region plays a significant role in inter- and intralinking of fibrin molecules and protofibrils, endowing fibrin fibers with increased stiffness and elasticity

Accuracy and Precision of Age Estimates for Pallid Sturgeon from Pectoral Fin Rays

Accurate age information is critical to the biological understanding and management of most fish species, but particularly for species of concern, such as the pallid sturgeon Scaphirhynchus albus. The accuracy and precision of pallid sturgeon age estimates from pectoral fin ray sections has never been established, yet all accumulated age information for the species was collected using this technique. To examine the accuracy and precision of age estimates, 16 pectoral fin ray samples from age-6 pallid sturgeon were obtained from Gavins Point National Fish Hatchery, South Dakota. The fin rays were sectioned, mounted, and independently examined twice by each of two readers. Only 28.1% of the age estimates accurately reflected the known age of the fish. Multiple readings of the same sample by the same reader (within-reader precision) only agreed 25% of the time, differences being as great as 5 years between the two estimates. Between-reader agreement was 46.9%, the two readers\u27 estimates of the same fish differing by as much as 2 years. Because of low accuracy and precision, estimated ages from pallid sturgeon pectoral fin rays should be viewed with caution

Dynamic properties of noise and Her6 levels are optimized by miR-9, allowing the decoding of the Her6 oscillator

This work was supported by a Wellcome Trust Senior Research Fellowship to NP (106185/Z/14/Z).Noise is prevalent in biology and has been widely quantified using snapshot measurements. This static view obscures our understanding of dynamic noise properties and how these affect gene expression and cell state transitions. Using a CRISPR/Cas9 Zebrafish her6::Venus reporter combined with mathematical and in vivo experimentation, we explore how noise affects the protein dynamics of Her6, a basic helix‐loop‐helix transcriptional repressor. During neurogenesis, Her6 expression transitions from fluctuating to oscillatory at single‐cell level. We identify that absence of miR‐9 input generates high‐frequency noise in Her6 traces, inhibits the transition to oscillatory protein expression and prevents the downregulation of Her6. Together, these impair the upregulation of downstream targets and cells accumulate in a normally transitory state where progenitor and early differentiation markers are co‐expressed. Computational modelling and double smFISH of her6 and the early neurogenesis marker, elavl3, suggest that the change in Her6 dynamics precedes the downregulation in Her6 levels. This sheds light onto the order of events at the moment of cell state transition and how this is influenced by the dynamic properties of noise. Our results suggest that Her/Hes oscillations, facilitated by dynamic noise optimization by miR‐9, endow progenitor cells with the ability to make a cell state transition.Publisher PDFPeer reviewe

The secretion inhibitor Exo2 perturbs trafficking of Shiga toxin between endosomes and the trans-Golgi network

The small-molecule inhibitor Exo2 {4-hydroxy-3-methoxy-(5,6,7,8-tetrahydrol[1]benzothieno[2,3-d]pyrimidin-4-yl)hydraz-one benzaldehyde} has been reported to disrupt the Golgi apparatus completely and to stimulate Golgi–ER (endoplasmic reticulum) fusion in mammalian cells, akin to the well-characterized fungal toxin BFA (brefeldin A). It has also been reported that Exo2 does not affect the integrity of the TGN (trans-Golgi network), or the direct retrograde trafficking of the glycolipid-binding cholera toxin from the TGN to the ER lumen. We have examined the effects of BFA and Exo2, and found that both compounds are indistinguishable in their inhibition of anterograde transport and that both reagents significantly disrupt the morphology of the TGN in HeLa and in BS-C-1 cells. However, Exo2, unlike BFA, does not induce tubulation and merging of the TGN and endosomal compartments. Furthermore, and in contrast with its effects on cholera toxin, Exo2 significantly perturbs the delivery of Shiga toxin to the ER. Together, these results suggest that the likely target(s) of Exo2 operate at the level of the TGN, the Golgi and a subset of early endosomes, and thus Exo2 provides a more selective tool than BFA for examining membrane trafficking in mammalian cells

Long-Term Adult Feline Liver Organoid Cultures for Disease Modeling of Hepatic Steatosis

Hepatic steatosis is a highly prevalent liver disease, yet research is hampered by the lack of tractable cellular and animal models. Steatosis also occurs in cats, where it can cause severe hepatic failure. Previous studies demonstrate the potential of liver organoids for modeling genetic diseases. To examine the possibility of using organoids to model steatosis, we established a long-term feline liver organoid culture with adult liver stem cell characteristics and differentiation potential toward hepatocyte-like cells. Next, organoids from mouse, human, dog

α−α Cross-Links Increase Fibrin Fiber Elasticity and Stiffness

Fibrin fibers, which are ∼100 nm in diameter, are the major structural component of a blood clot. The mechanical properties of single fibrin fibers determine the behavior of a blood clot and, thus, have a critical influence on heart attacks, strokes, and embolisms. Cross-linking is thought to fortify blood clots; though, the role of α–α cross-links in fibrin fiber assembly and their effect on the mechanical properties of single fibrin fibers are poorly understood. To address this knowledge gap, we used a combined fluorescence and atomic force microscope technique to determine the stiffness (modulus), extensibility, and elasticity of individual, uncross-linked, exclusively α–α cross-linked (γQ398N/Q399N/K406R fibrinogen variant), and completely cross-linked fibrin fibers. Exclusive α–α cross-linking results in 2.5× stiffer and 1.5× more elastic fibers, whereas full cross-linking results in 3.75× stiffer, 1.2× more elastic, but 1.2× less extensible fibers, as compared to uncross-linked fibers. On the basis of these results and data from the literature, we propose a model in which the α-C region plays a significant role in inter- and intralinking of fibrin molecules and protofibrils, endowing fibrin fibers with increased stiffness and elasticity

Long-Term Adult Feline Liver Organoid Cultures for Disease Modeling of Hepatic Steatosis.

Hepatic steatosis is a highly prevalent liver disease, yet research is hampered by the lack of tractable cellular and animal models. Steatosis also occurs in cats, where it can cause severe hepatic failure. Previous studies demonstrate the potential of liver organoids for modeling genetic diseases. To examine the possibility of using organoids to model steatosis, we established a long-term feline liver organoid culture with adult liver stem cell characteristics and differentiation potential toward hepatocyte-like cells. Next, organoids from mouse, human, dog, and cat liver were provided with fatty acids. Lipid accumulation was observed in all organoids and interestingly, feline liver organoids accumulated more lipid droplets than human organoids. Finally, we demonstrate effects of interference with β-oxidation on lipid accumulation in feline liver organoids. In conclusion, feline liver organoids can be successfully cultured and display a predisposition for lipid accumulation, making them an interesting model in hepatic steatosis research

A systematic review, meta-analysis and metaregression of the effect of protein supplementation on resistance training-induced gains in muscle mass and strength in healthy adults

Objective We performed a systematic review, metaanalysis and meta-regression to determine if dietary protein supplementation augments resistance exercise training (RET)-induced gains in muscle mass and strength. Data sources A systematic search of Medline, Embase, CINAHL and SportDiscus. Eligibility criteria Only randomised controlled trials with RET ≥6 weeks in duration and dietary protein supplementation. Design Random-effects meta-analyses and metaregressions with four a priori determined covariates. Twophase break point analysis was used to determine the relationship between total protein intake and changes in fat-free mass (FFM). ResultsData from 49 studies with 1863 participants showed that dietary protein supplementation significantly (all p Summary/conclusion Dietary protein supplementation significantly enhanced changes in muscle strength and size during prolonged RET in healthy adults. Increasing age reduces and training experience increases the efficacy of protein supplementation during RET. With protein supplementation, protein intakes at amounts greater than ~1.6 g/kg/day do not further contribute RET-induced gains in FFM