Isolation and characterisation of the chick orthologue of the Opitz syndrome gene, Mid1, supports a conserved role in vertebrate development

© UBC PressThe X-linked form of Opitz syndrome (OS) is caused by loss of function of the microtubule-associated MID1 protein. The phenotype of OS includes defects along the central body axis, namely hypertelorism, cleft lip and palate, hypospadias and cardiac structural anomalies. Here we describe the isolation and characterisation of full-length cDNA clones representing the chick Mid1 gene and the detailed profile of its expression in stage 7 to 28 chick embryos. Consistent with the remarkable sequence conservation of MID1 between human and chick was the good correlation of the pattern of cMid1 expression with the tissues affected in OS. In stage 10 embryos, transcripts were concentrated in the head mesenchyme which includes migratory neural crest cells. However, the incomplete overlap with a neural crest marker, Sox10, suggests that Mid1 is a marker for somitomeric mesoderm and potentially for a subset of neural crest cells. Consistent with this, cMid1 expression was also detected at later stages in neural crest-derived facial mesenchyme, in the myotome and in the condensing muscle blocks of the limb. Expression of cMid1 was observed in the neural epithelium of the forebrain beginning at stage 7 with increased signal in presumptive rhombomeres 2/3. By stage 15, expression is highest in the diencephalon. Other areas with high expression are certain facial epithelia and the midgut that will give rise to the oesophagus and trachea. These data indicate that Mid1 plays an evolutionarily conserved developmental function in vertebrates that may involve effects on cellular proliferation, tissue interactions and morphogenesis.Joy M. Richman, Katherine K. Fu, Liza L. Cox, Jane P. Sibbons and Timothy C. Co

ASKAP and MeerKAT surveys of the magellanic clouds

The Magellanic Clouds are a stepping stone from the overwhelming detail of the Milky Way in which we are immersed, to the global characteristics of galaxies both in the nearby and distant universe. They are interacting, gas-rich dwarf galaxies of sub-solar metallicity, not unlike the building blocks that assembled the large galaxies that dominate groups and clusters, and representative of the conditions at the height of cosmic star formation. The Square Kilometre Array (SKA) can make huge strides in understanding galactic metabolism and the ecological processes that govern star formation, by observations of the Magellanic Clouds and other, nearby Magellanic-type irregular galaxies. Two programmes with SKA Pathfinders attempt to pave the way: the approved Galactic ASKAP Spectral Line Survey (GASKAP) includes a deep survey in H I and OH of the Magellanic Clouds, whilst MagiKAT is proposed to perform more detailed studies of selected regions within the Magellanic Clouds - also including Faraday rotation measurements and observations at higher frequencies. These surveys also close the gap with the revolutionizing surveys at far-IR wavelengths with the Spitzer Space Telescope and Herschel Space Observatory

Synthesis of C-2-symmetric aza- and azaoxa-macrocyclic ligands derived from (1R,2R)-1,2-diaminocyclohexane and their applications in catalysis

Investigations have been undertaken into the synthesis of chiral derivatives of 1,4,7-triazacyclononane and 1,7-diaza-4-oxacyclononane that incorporate the C-2-symmetric (R,R)-1,2-diaminocyclohexane backbone. The target ligands 17a and 17b have been prepared as their hydrochloride salts, the latter of which has been characterized by single crystal X-ray diffraction revealing an extensively hydrogen bonded polymeric network in the solid state. These investigations have shown that the formation of the fused bicyclic system in these ligands via standard Richman-Atkins macrocyclisation conditions is extremely difficult, particularly for the intermediate 16a, when three tosyl amide nitrogen atoms must be accommodated in the macrocyclic ring. In addition to these target ligands the unexpected piperazine 15 as well as the novel binucleating ligand 19 have also been prepared. Preliminary investigation into the coordination chemistry of 17b resulted in the formation of the copper(II) complex [Cu(17b)Cl-2] in which the copper centre exists in the expected square-pyramidal geometry with the two chloride ions and the nitrogen donors occupying the equatorial positions and with the oxygen donor apically situated. The complex has been screened for activity and found to be a potent catalyst for two hetero-Diels-Alder reactions. The first aza-Diels-Alder reaction of imine 20 with Danishefsky's diene 21 proceeds to yield the cycloadduct 22 in 94% yield. The second nitroso-Diels-Alder reaction relies on the in situ oxidation of hydroxylamine 23 to dienophile 24, catalysed by the complex in the presence of tert-butyl hydroperoxide, which is then trapped as cycloadduct 25 by cyclohexadiene in 69% yield

Characterisation of porcine dermis scaffolds decellularised using a novel non-enzymatic method for biomedical applications

Off-the-shelf availability of tissue-engineered skin constructs, tailored by different combinations of reagents to produce a highly preserved biological matrix is often the only means to help patients suffering skin damage. This study assessed the effect of five different decellularisation methods on porcine dermal scaffolds with regard to matrix composition, biomechanical strength, and cytotoxicity using an in vitro biocompatibility assay. Results demonstrated that four out of the five tested decellularisation protocols were efficient in producing acellular scaffolds. Nevertheless, decellularisation method using osmotic shock without enzymatic digestion showed to be efficient not only in removing cellular material and debris from dermal scaffolds but was also beneficial in the preservation of extracellular matrix components (glycosaminoglycans and collagen). Histological assessment revealed that the dermal architecture of coarse collagen bundles was preserved. Examinations by scanning electron microscopy and transmission electron microscopy showed that the arrangement and ultrastructure of collagen fibrils in the scaffolds were retained following non-enzymatic method of decellularisation and also after collagen crosslinking using genipin. Moreover, this decellularised scaffold was not only shown to be biologically compatible when co-cultured with bone marrow-derived mesenchymal stem cells and fibroblasts, but also stimulated the cells to release trophic factors essential for tissue regeneration

Development and Characterization of a Porcine Liver Scaffold

The growing number of patients requiring liver transplantation for chronic liver disease cannot be currently met due to a shortage in donor tissue. As such, alternative tissue engineering approaches combining the use of acellular biological scaffolds and different cell populations (hepatic or progenitor) are being explored to augment the demand for functional organs. Our goal was to produce a clinically relevant sized scaffold from a sustainable source within 24 h, while preserving the extracellular matrix (ECM) to facilitate cell repopulation at a later stage. Whole porcine livers underwent perfusion decellularization via the hepatic artery and hepatic portal vein using a combination of saponin, sodium deoxycholate, and deionized water washes resulting in an acellular scaffold with an intact vasculature and preserved ECM. Molecular and immunohistochemical analysis (collagen I and IV and laminin) showed complete removal of any DNA material, together with excellent retention of glycosaminoglycans and collagen. Fourier-transform infrared spectroscopy (FTIR) analysis showed both absence of nuclear material and removal of any detergent residue, which was successfully achieved after additional ethanol gradient washes. Samples of the decellularized scaffold were assessed for cytotoxicity by seeding with porcine adipose-derived mesenchymal stem cells in vitro, these cells over a 10-day period showed attachment and proliferation. Perfusion of the vascular tree with contrast media followed by computed tomography (CT) imaging showed an intact vascular network. In vivo implantation of whole intact nonseeded livers, into a porcine model (as auxiliary graft) showed uniform perfusion macroscopically and histologically. Using this method, it is possible to create an acellular, clinically sized, liver scaffold with intact vasculature in less than 24 h