Branch Mode Selection during Early Lung Development

Many organs of higher organisms, such as the vascular system, lung, kidney, pancreas, liver and glands, are heavily branched structures. The branching process during lung development has been studied in great detail and is remarkably stereotyped. The branched tree is generated by the sequential, non-random use of three geometrically simple modes of branching (domain branching, planar and orthogonal bifurcation). While many regulatory components and local interactions have been defined an integrated understanding of the regulatory network that controls the branching process is lacking. We have developed a deterministic, spatio-temporal differential-equation based model of the core signaling network that governs lung branching morphogenesis. The model focuses on the two key signaling factors that have been identified in experiments, fibroblast growth factor (FGF10) and sonic hedgehog (SHH) as well as the SHH receptor patched (Ptc). We show that the reported biochemical interactions give rise to a Schnakenberg-type Turing patterning mechanisms that allows us to reproduce experimental observations in wildtype and mutant mice. The kinetic parameters as well as the domain shape are based on experimental data where available. The developed model is robust to small absolute and large relative changes in the parameter values. At the same time there is a strong regulatory potential in that the switching between branching modes can be achieved by targeted changes in the parameter values. We note that the sequence of different branching events may also be the result of different growth speeds: fast growth triggers lateral branching while slow growth favours bifurcations in our model. We conclude that the FGF10-SHH-Ptc1 module is sufficient to generate pattern that correspond to the observed branching modesComment: Initially published at PLoS Comput Bio

Evaluation of appendicitis risk prediction models in adults with suspected appendicitis

Background Appendicitis is the most common general surgical emergency worldwide, but its diagnosis remains challenging. The aim of this study was to determine whether existing risk prediction models can reliably identify patients presenting to hospital in the UK with acute right iliac fossa (RIF) pain who are at low risk of appendicitis. Methods A systematic search was completed to identify all existing appendicitis risk prediction models. Models were validated using UK data from an international prospective cohort study that captured consecutive patients aged 16–45 years presenting to hospital with acute RIF in March to June 2017. The main outcome was best achievable model specificity (proportion of patients who did not have appendicitis correctly classified as low risk) whilst maintaining a failure rate below 5 per cent (proportion of patients identified as low risk who actually had appendicitis). Results Some 5345 patients across 154 UK hospitals were identified, of which two‐thirds (3613 of 5345, 67·6 per cent) were women. Women were more than twice as likely to undergo surgery with removal of a histologically normal appendix (272 of 964, 28·2 per cent) than men (120 of 993, 12·1 per cent) (relative risk 2·33, 95 per cent c.i. 1·92 to 2·84; P < 0·001). Of 15 validated risk prediction models, the Adult Appendicitis Score performed best (cut‐off score 8 or less, specificity 63·1 per cent, failure rate 3·7 per cent). The Appendicitis Inflammatory Response Score performed best for men (cut‐off score 2 or less, specificity 24·7 per cent, failure rate 2·4 per cent). Conclusion Women in the UK had a disproportionate risk of admission without surgical intervention and had high rates of normal appendicectomy. Risk prediction models to support shared decision‐making by identifying adults in the UK at low risk of appendicitis were identified

Immobilisation of bovine enterokinase and application of the immobilised enzyme in fusion protein cleavage

Kubitzki T, Noll T, Luetz S. Immobilisation of bovine enterokinase and application of the immobilised enzyme in fusion protein cleavage. BIOPROCESS AND BIOSYSTEMS ENGINEERING. 2008;31(3):173-182.Two immobilisation methods for enterokinase were developed, which yielded high remaining activities for the cleavage of the fusion protein MUC1-IgG Fc. Different carrier materials were compared regarding remaining enzyme activity and storage stability. Immobilisation procedures involving support material activation using glutardialdehyde were found to result in low remaining activities. Applying less aggressive activation procedures, remaining activities of similar to 60% were received when immobilising enterokinase on either Estapor paramagnetic microspheres or hexamethylamino Sepabeads(R). In case of hexamethylamino Sepabeads(R) we were able to increase the half-life time 4.3-fold at 23 degrees C and 3.8-fold at 4 degrees C compared to the free enzyme at the same temperatures. By immobilising the biocatalyst the downstream process is simplified allowing the easy removal of the enzyme from the reaction mixture. The immobilised enterokinase cleaves the fusion protein MUC1-IgG Fc in at least two repeated batches, proving the efficiency of the immobilisation method and the reusability of the biocatalyst

Expression analysis of Shh signaling members in early stages of chick lung development

Lung organogenesis is guided by epithelial-mesenchymal interactions that coordinate cellular events responsible for the formation of the respiratory system. Several signaling pathways have been implicated in this process; among them, sonic hedgehog (Shh) signaling has emerged as a crucial regulator of branching morphogenesis in the mammalian lung. Canonical Shh signaling requires the presence of patched (Ptch) and smoothened (Smo) transmembrane receptors in order to induce the activation of glioblastoma (Gli) zinc finger transcription factors that are the true effectors of the pathway. Signal transduction is finely regulated by Ptch1, Gli, and Hhip (hedgehog-interacting protein). The present work characterizes, for the first time, the expression pattern of shh, ptch1, smo, gli1, and hhip in early stages of the embryonic chick lung. In situ hybridization studies revealed that these genes are expressed in the same cellular compartments as their mammalian counterparts, although their proximo-distal distribution is slightly changed. Moreover, the molecular interactions between fibroblast growth factor (FGF) and Shh signaling pathway were assessed, in vitro, by grafting beads soaked in SU5402 (an FGF receptor inhibitor). In the chick lung, Shh signaling seems to have some features that are species specific since shh is not a downstream target of FGF signaling. Nonetheless and despite the observed differences, these findings suggest a role for Shh signaling in the epithelial-mesenchymal interactions that control chick lung morphogenesis.This project was funded by: project ON.2 SR&TD Integrated Program (NORTE-07-0124-FEDER-000017) co-funded by Programa Operacional Regional do Norte (ON.2- O Novo Norte), Quadro de Referência Estratégico Nacional (QREN), through Fundo Europeu de Desenvolvimento Regional (FEDER); FEDER funds through the Operational Programme Competitiveness Factors – COMPETE; National Funds through FCT - Foundation for Science and Technology under the project POCI-01-0145-FEDER-007038. RSM was supported by the ON.2 SR&TD Integrated Program (N-01- 01-01-24-01-07), ref: UMINHO/BPD/31/2013