Depletion of the gut microbiota differentially affects the impact of whey protein on high-fat diet-induced obesity and intestinal permeability

Acknowledgement: The authors thank Fiona Crispie and Amanda Brechon from Teagasc Moorepark Food Research Centre for their assistance in 16S library preparation and sequencing. The authors thank Thomaz Bastiaanssen for generating the pictures for caecal metabolomics analysis. The authors thank Joana Pereira, Eoin Sherwin and Marina Shverer for helping with the Ussing chambers experiment. S. B. was funded under the Teagasc Walsh Fellowship scheme (grant number 2016007). K. N. N. was supported by Teagasc, Ireland and in part by a research grant from Science Foundation Ireland (SFI) under grant numbers SFI/16/BBSRC/3389 and BBSRC under the grant number BB/P009875/1 (to K.N.N. and J.R.S). Funding information Teagasc Walsh Fellowship Programme, Grant/Award Number: 2016007; Science Foundation Ireland, Grant/Award Number: SFI/16/BBSRC/3389; Biotechnology and Biological Sciences Research Council, Grant/Award Number: BB/P009875/1Peer reviewedPublisher PD

Vangl2 disruption alters the biomechanics of late spinal neurulation leading to spina bifida in mouse embryos

peer-reviewedHuman mutations in the planar cell polarity component VANGL2 are associated with the neural tube defect spina bifida. Homozygous Vangl2 mutation in mice prevents initiation of neural tube closure, precluding analysis of its subsequent roles in neurulation. Spinal neurulation involves rostral-to-caudal ‘zippering’ until completion of closure is imminent, when a caudal-to-rostral closure point, ‘Closure 5’, arises at the caudal-most extremity of the posterior neuropore (PNP). Here, we used Grhl3Cre to delete Vangl2 in the surface ectoderm (SE) throughout neurulation and in an increasing proportion of PNP neuroepithelial cells at late neurulation stages. This deletion impaired PNP closure after the ∼25-somite stage and resulted in caudal spina bifida in 67% of Grhl3Cre/+Vangl2Fl/Fl embryos. In the dorsal SE, Vangl2 deletion diminished rostrocaudal cell body orientation, but not directional polarisation of cell divisions. In the PNP, Vangl2 disruption diminished mediolateral polarisation of apical neuroepithelial F-actin profiles and resulted in eversion of the caudal PNP. This eversion prevented elevation of the caudal PNP neural folds, which in control embryos is associated with formation of Closure 5 around the 25-somite stage. Closure 5 formation in control embryos is associated with a reduction in mechanical stress withstood at the main zippering point, as inferred from the magnitude of neural fold separation following zippering point laser ablation. This stress accommodation did not happen in Vangl2-disrupted embryos. Thus, disruption of Vangl2-dependent planar-polarised processes in the PNP neuroepithelium and SE preclude zippering point biomechanical accommodation associated with Closure 5 formation at the completion of PNP closure

Integrin-Mediated Focal Anchorage Drives Epithelial Zippering during Mouse Neural Tube Closure.

Epithelial fusion is a key process of morphogenesis by which tissue connectivity is established between adjacent epithelial sheets. A striking and poorly understood feature of this process is "zippering," whereby a fusion point moves directionally along an organ rudiment. Here, we uncover the molecular mechanism underlying zippering during mouse spinal neural tube closure. Fusion is initiated via local activation of integrin β1 and focal anchorage of surface ectoderm cells to a shared point of fibronectin-rich basement membrane, where the neural folds first contact each other. Surface ectoderm cells undergo proximal junction shortening, establishing a transitory semi-rosette-like structure at the zippering point that promotes juxtaposition of cells across the midline enabling fusion propagation. Tissue-specific ablation of integrin β1 abolishes the semi-rosette formation, preventing zippering and causing spina bifida. We propose integrin-mediated anchorage as an evolutionarily conserved mechanism of general relevance for zippering closure of epithelial gaps whose disturbance can produce clinically important birth defects

Protein quality and quantity influence the effect of dietary fat on weight gain and tissue partitioning via host-microbiota changes

ACKNOWLEDGMENTS This work was supported by a research grant from Science Foundation Ireland (SFI) under grant SFI/16/BBSRC/3389, BBSRC under grant number BB/ P009875/1 (to K.N.N. and J.R.S.), and in part by SFI and the Department of Agriculture, Food and Marine under grant 16/RC/3835 (to VistaMilk). We thank Conall Strain, David Mannion, and John Leech for contributing to the metabolomics analysis. We thank Alina Kondrashina for help with the Milliplex system.Peer reviewedPublisher PD

The role of proteoglycans in the initiation of neural tube closure

Neurulation is the embryonic process that gives rise to the neural tube (NT), the precursor of the brain and spinal cord. Recent work has emphasised the importance of proteoglycans in convergent extension movements and NT closure in lower vertebrates. The current study is focused on the role of proteoglycans in the initiation of NT closure in mammals, termed closure 1. In this project, the initial aim was to characterise the ‘matrisome’, or in vivo extracellular matrix (ECM) composition, during mammalian neurulation. Tissue site of mRNA expression and protein localisation of ECM components, including proteoglycans, were then investigated showing their distinct expression patterns prior to and after the onset of neural tube closure. The expression analysis raised various hypothesis that were subsequently tested, demonstrating that impaired sulfation of ECM proteoglycan chains worsens the phenotype of planar cell polarity (PCP) mutant loop tail (Vangl2Lp) predisposed to neural tube defects. Exposure of Vangl2Lp/+ embryos to chlorate, an inhibitor of glycosaminoglycan sulfation, during ex vivo whole embryo culture prevented NT closure, converting Vangl2Lp/+ to the mutant Vangl2Lp/Lp pathophenotype. The same result was obtained by exposure of Vangl2Lp/+ + embryos to chondroitinase or heparitinase. Taken together, it indicated that the PCP pathway functionally interacts with chondroitin and heparan sulfate proteoglycans during initiation of NT closure. In order to investigate the possible role of proteoglycans in mammalian convergent extension, the node of Vangl2Lp/+ embryos was labelled with DiO. The study revealed that the PCP-proteoglycan interaction is mediated independently of convergent extension. The failure of neural fold apposition and reduced Fgfr1 signalling was proposed as potential causative mechanism underlying failure of closure 1. In fish, the cilia motility is dependent on heparan sulfate chains, but this has not been studied in mammals. The present study identified a novel cellular localisation of cohesin/proteoglycan protein Smc3 and its GAG chains. Both Smc3 and CS-E are expressed in the midbody, primary and motile cilia. For the first time, this study showed the nuclear expression of CS chains in mouse embryo. The coordinated movement of Smc3 and CS-E chains during cytokinesis and ciliogenesis suggests conserved role of this protein in mouse cilia and cytokinetic apparatus

Vangl2 disruption alters the biomechanics of late spinal neurulation leading to spina bifida in mouse embryos

Human mutations in the planar cell polarity component VANGL2 are associated with the neural tube defect spina bifida. Homozygous Vangl2 mutation in mice prevents initiation of neural tube closure, precluding analysis of its subsequent roles in neurulation. Spinal neurulation involves rostral-to-caudal ‘zippering’ until completion of closure is imminent, when a caudal-to-rostral closure point, ‘Closure 5’, arises at the caudal-most extremity of the posterior neuropore (PNP). Here, we used Grhl3Cre to delete Vangl2 in the surface ectoderm (SE) throughout neurulation and in an increasing proportion of PNP neuroepithelial cells at late neurulation stages. This deletion impaired PNP closure after the ∼25-somite stage and resulted in caudal spina bifida in 67% of Grhl3Cre/+Vangl2Fl/Fl embryos. In the dorsal SE, Vangl2 deletion diminished rostrocaudal cell body orientation, but not directional polarisation of cell divisions. In the PNP, Vangl2 disruption diminished mediolateral polarisation of apical neuroepithelial F-actin profiles and resulted in eversion of the caudal PNP. This eversion prevented elevation of the caudal PNP neural folds, which in control embryos is associated with formation of Closure 5 around the 25-somite stage. Closure 5 formation in control embryos is associated with a reduction in mechanical stress withstood at the main zippering point, as inferred from the magnitude of neural fold separation following zippering point laser ablation. This stress accommodation did not happen in Vangl2-disrupted embryos. Thus, disruption of Vangl2-dependent planar-polarised processes in the PNP neuroepithelium and SE preclude zippering point biomechanical accommodation associated with Closure 5 formation at the completion of PNP closure