Importance of multiple loading scenarios for the identification of material coefficients of the human cornea

A mechanical balance between intraocular pressure and tissue stiffness defines the refractive shape of the human cornea. More and more daily surgical procedures modify that shape to achieve vision correction, which increases the demand for a profound understanding of the tissue mechanics. The wide variety of published mechanical properties foreshadows the difficulty of this task. The aim of this study is to show that such problems may arise from using the inverse method for fitting material models with multiple coefficients to a limited number (usually one) of experimental data. Using multiple sets of experimental data for the fitting process is proposed as a possible solution.     

Design and synthesis of 4-substituted-8-(2-phenyl-cyclohexyl)-2,8-diaza-spiro[4.5]decan-1-one as a novel class of GlyT1 inhibitors: achieving selectivity against the mu opioid and nociceptin/orphanin FQ peptide (NOP) receptors

A novel class of 4-substituted-8-(2-phenyl-cyclohexyl)-2,8-diaza-spiro[4.5]decan-1-ones have been discovered and developed as potent and selective GlyT1 inhibitors. The molecules are devoid of activity at the GlyT2 isoform and display excellent selectivities against the mu opioid receptor as well as the nociceptin/orphanin FQ peptide (NOP) receptor. A novel, straightforward and efficient synthetic strategy for the assembly of the target molecules is also presented.     

A novel role for retinoids in patterning the avian forebrain during presomite stages

Retinoids, and in particular retinoic acid (RA), are known to induce posterior fates in neural tissue. However, alterations in retinoid signalling dramatically affect anterior development. Previous reports have demonstrated a late role for retinoids in patterning craniofacial and forebrain structures, but an earlier role in anterior patterning is not well understood. We show that enzymes involved in synthesizing retinoids are expressed in the avian hypoblast and in tissues directly involved in head patterning, such as anterior definitive endoderm and prechordal mesendoderm. We found that in the vitamin A-deficient (VAD) quail model, which lacks biologically active RA from the first stages of development, anterior endodermal markers such as Bmp2, Bmp7, Hex and the Wnt antagonist crescent are affected during early gastrulation. Furthermore, prechordal mesendodermal and prospective ventral telencephalic markers are expanded posteriorly, Shh expression in the axial mesoderm is reduced, and Bmp2 and Bmp7 are abnormally expressed in the ventral midline of the neural tube. At early somite stages, VAD embryos have increased cell death in ventral neuroectoderm and foregut endoderm, but normal cranial neural crest production, whereas at later stages extensive apoptosis occurs in head mesenchyme and ventral neuroectoderm. As a result, VAD embryos end up with a single and reduced telencephalic vesicle and an abnormally patterned diencephalon. Therefore, we propose that retinoids have a dual role in patterning the anterior forebrain during development. During early gastrulation, RA acts in anterior endodermal cells to modulate the anteroposterior (AP) positional identity of prechordal mesendodermal inductive signals to the overlying neuroectoderm. Later on, at neural pore closure, RA is required for patterning of the mesenchyme of the frontonasal process and the forebrain by modulating signalling molecules involved in craniofacial morphogenesis.     

Making and repairing the mammalian brain--in vitro production of dopaminergic neurons

Midbrain dopamine (DA) neurons play an essential role in modulating motor control, and their degeneration is the hallmark feature of Parkinson's disease (PD). In vitro production of DA neurons provides insight into the mechanisms that control cell fate choice, and offers an alternative to the use of fetal tissue for experimental cell replacement in PD. Here we will review the advantages and disadvantages of the various renewable cell sources and protocols tested, and discuss their relevance for basic studies and for cell therapy.     

Int-B13, an Unusual Site-Specific Recombinase of the Bacteriophage P4 Integrase Family, Is Responsible for Chromosomal Insertion of the 105-Kilobase clc Element of Pseudomonas sp. Strain B13

Pseudomonas sp. strain B13 carries the clcRABDE genes encoding chlorocatechol-degradative enzymes on the self-transmissible 105-kb clc element. The element integrates site and orientation specifically into the chromosomes of various bacterial recipients, with a glycine tRNA structural gene (glyV) as the integration site. We report here the localization and nucleotide sequence of the integrase gene and the activity of the integrase gene product in mediating site-specific integration. The integrase gene (int-B13) was located near the right end of the clc element. It consisted of an open reading frame (ORF) of maximally 1,971 bp with a coding capacity for 657 amino acids (aa). The full-length protein (74 kDa) was observed upon overexpression and sodium dodecyl sulfate-polyacrylamide gel electrophoresis separation. The N-terminal 430 aa of the predicted Int-B13 protein had substantial similarity to integrases from bacteriophages of the P4 family, but Int-B13 was much larger than P4-type integrases. The C-terminal 220 aa of Int-B13 were homologous to an ORF flanking a gene cluster for naphthalene degradation in Pseudomonas aeruginosa PaK1. Similar to the bacteriophages R73 and P4, the clc element integrates into the 3′ end of the target tRNA gene. This target site was characterized from four different recipient strains into which the clc element integrated, showing sequence specificity of the integration. In Pseudomonas sp. strain B13, a circular form of the clc element, which carries an 18-bp DNA sequence identical to the 3′-end portion of glyV as part of its attachment site (attP), could be detected. Upon chromosomal integration of the clc element into a bacterial attachment site (attB), a functional glyV was reconstructed at the right end of the element. The integration process could be demonstrated in RecA-deficient Escherichia coli with two recombinant plasmids, one carrying the int-B13 gene and the attP site and the other carrying the attB site of Pseudomonas putida F1.     

Iodotyrosine deiodination in the normal and acutely TSH-stimulated thyroid.

The deiodination of L-MIT-125I was measured in rat thyroid homogenates and slices before and after acute TSH stimulation. Slices and homogenates were incubated with identical concentrations of tissue and substrate in the presence and absence of NADPH. 1 USP unit TSH added in vitro to thyroid slices failed to stimulate deiodination; a single in vivo ip injection of 3 USP units TSH was also unable to raise deiodinating activity. In contrast to TSH, NADPH added to homogenates and slices enhanced deiodination significantly. However, several arguments, including a review of the literature, strongly militate against the hypothesis of an increased intracellular concentration of the coenzyme NADPH being the prerequisite to enhanced deiodination. The results suggest that deiodinase activity in acutely stimulated thyroids is not limited by the intracellular concentration of the enzyme itself nor by the availability of co-enzyme. Therefore, the increased iodide release induced by acute TSH stimulation is a mere consequence of the enhanced thyroglobulin proteolysis and does not require higher enzyme concentration. It will be shown subsequently that a different conclusion must be drawn in experiments with chronic TSH stimulation.