Cellular mechanisms of facial primordium growth

Lead Research Organisation: King's College London
Department Name: Dental Institute School Office

Abstract

Cleft palate and cleft lip are not only disfiguring birth defects, they also cause serious problems for children affected, ranging from speech difficulties to shortened lifespan. Management and treament are stressful and costly. Facial clefting happens when bulges in the sides of the developing embryo s face fail to meet or fuse. These bulges normally make the upper lip and the roof of the mouth. This project will look at how cells making the mouse face (which develops in the same way) multiply, rearrange themselves and change shape normally and in mouse strains in which clefting happens. Individual cells will be tracked and monitored to build up a three-dimensional picture of what the cells do. This will provide a map of what should happen, and therefore help pinpoint what goes wrong when clefting happens. Understanding how the face develops normally and what goes wrong with the cells in facial tissue when clefting occurs will help future diagnosis (for example, finding mothers at risk) and treatments (potentially re-growing what is missing).

Technical Summary

Normal development of the early face and palate depends upon controlled elongation and spatial co-ordination of the early embryonic processes that build these structures. Disruption of these mechanisms can lead to clefting of the oro-facial region, which is the commonest form of craniofacial anomaly. The directional orientation of embryonic tissue can take place through a number of cellular mechanisms, which include terminally localised and orientated cell division, changes in cell shape and cell rearrangement. Whilst an increasing number of mutant mouse models have demonstrated the importance of normal cell proliferation, particularly during palatogenesis, there is currently little known about the relative contributions of these different processes during development of the early face and palate; in particular, the relationship between cell proliferation and tissue elongation. We aim to identify and quantify in detail the cellular events underlying tissue elongation in the developing mouse embryo. Proliferation and programmed cell death will be mapped in three-dimensions using wild type embryos to provide a hitherto unavailable baseline of these cell activities during facial development. In addition, detailed lineage-tracing studies will be carried out to provide a picture of cellular lineage growth and extension using diI-labelling of murine facial explants in vitro and in vivo clonal-labelling using Tamoxifen-inducible conditional reporter mice. Finally, directional cellular rearrangement and shape change will be analysed in developing murine facial processes using time-lapse live imaging of diI-labelled explants and immunofluoresent labelling. The proposed detailed and systematic analysis of tissue growth and elongation in the facial processes will allow progress in understanding how genetic mutation can lead to cellular disruption during the complex process of facial morphogenesis.
 
Description MRC PhD Studentship
Amount £50,000 (GBP)
Organisation Medical Research Council (MRC) 
Sector Public
Country United Kingdom
Start 10/2010 
End 09/2013
 
Description Project Grant - Epithelial bending
Amount £435,467 (GBP)
Funding ID BB/L002965/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 03/2014 
End 02/2017
 
Description Project Grant - Palate Patterning
Amount £478,391 (GBP)
Funding ID BB/J009105/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 06/2012 
End 05/2015
 
Title Image analysis for cell-to-tissue phenotyping 
Description We have developed a software pipeline adapting freeware image analysis tools (ImageJ/Fiji) to provide a whole-cell-population capture of growth and directional processes that identifies explicitly the cellular differences between normal and abnormal tissue growth, applicable to fixed material and, for example, to definining the cell-behaviour lesion in birth defect models 
Type Of Material Physiological assessment or outcome measure 
Year Produced 2013 
Provided To Others? Yes  
Impact This is the basis of the collaboration with Yang Chai, listed under "collaborations" and will be the basis of collaborations with other groups. 
 
Description Consortium to analyse cellular facial phenotypes 
Organisation Cornell University
Department Department of Cell and Developmental Biology
Country United States 
Sector Academic/University 
PI Contribution We have developed novel image analysis methods for examinining cell size, shape, spacing and proliferation in fixed tissue specimens
Collaborator Contribution Our collaborators have developed genetic knockouts with cleft palate phenotypes that we can analyse
Impact Work is in progress
Start Year 2012
 
Description Consortium to analyse cellular facial phenotypes 
Organisation University of Southern California
Department Herman Ostrow School of Dentistry
Country United States 
Sector Academic/University 
PI Contribution We have developed novel image analysis methods for examinining cell size, shape, spacing and proliferation in fixed tissue specimens
Collaborator Contribution Our collaborators have developed genetic knockouts with cleft palate phenotypes that we can analyse
Impact Work is in progress
Start Year 2012
 
Description Davidson Imaging Morphogenesis Collaboration 
Organisation University of Pittsburgh
Department Department of Bioengineering
Country United States 
Sector Academic/University 
PI Contribution Significant intellectual input in generating the framework for and writing the article published and in devising and writing grant applications
Collaborator Contribution This collaboration led directly to the publication in Nature Cell Biology, is ongoing and is expected to lead to at least one further publication from this grant.
Impact Publication 17762892
Start Year 2006
 
Description Jaeger Network Inference collaboration 
Organisation Centre for Genomic Regulation (CRG)
Country Spain 
Sector Academic/University 
PI Contribution We identified the mouse palate epithelium as a Turing morphogen patterning system with iterative gene expression regulation and proposed a project to infer the underlying gene network using the partner's methodology.
Collaborator Contribution Dr. Johannes Jaeger developed a method of inferring transcriptional network structure from dense gene expression analysis by in situ measurement. He helped construct a proposal to apply this method for the first time to a mammalian tissue, namely the oral palate epithelium that we had studied
Impact Grant proposal to the BBSRC funded from June 2012 for three years. Has led to significant coverage in the media on our research on animal patterning
Start Year 2011
 
Description Yang Chai USC image-based cellular phenotyping 
Organisation University of Southern California
Department Centre for Craniofacial Molecular Biology
Country United States 
Sector Academic/University 
PI Contribution We have demonstrated the applicability of a new image-based method for measuring the cellular basis of malformation phenotypes such as cleft palate and used it to compare normal and cleft mutant mice to understand the aetiology of this birth defect
Collaborator Contribution Yang Chai and team have included our method as a component of a major grant application to the NIH FaceBase Consortium for understanding the molecular genetic basis of facial phenotypes
Impact Grant application pending. I will be a subcontractor to the grant if it is awarded.
Start Year 2012
 
Description Career presentation for Inspiring Futures (careers teacher organisation) hosted at Wellcome Trust 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Schools
Results and Impact I made a presentation and participated in workshop discussions with a group of secondary school teachers involved in giving career advice
Year(s) Of Engagement Activity 2015
 
Description Newspaper interviews (February 2012) 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Media (as a channel to the public)
Results and Impact King's College London press release and short video prompted several press inquiries including from the Daily Mail, Daily Telegraph and Frankfurter Allgemeiner Zeitung

Newpaper articles published in Daily Mail, Daily Telegraph, Frankfurter Allgemeiner Zeitung and on-line articles and reports in Huffington Post, Smithsonian science blog
Year(s) Of Engagement Activity 2012
 
Description Participation in I'm A Scientist, Get Me Out of Here! 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? Yes
Geographic Reach National
Primary Audience Schools
Results and Impact Numerous children in schools across the country (13, 14 years old) asked questions about science careers and motivation for this career track and specific questions about biology, chemistry, etc.

See http://imascientist.org.uk/
Year(s) Of Engagement Activity 2011
 
Description Radio interview (Albany, New York) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Type Of Presentation Keynote/Invited Speaker
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact Contributed to the series "The Academic Minute", a short-form radio broadcast by a National Public Radio affiliate in upstate New York.

Recording will be broadcast at some point in early 2013
Year(s) Of Engagement Activity 2012
 
Description School visits (Careers), London 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact Visits to several different secondary and primary schools in classes and smaller and larger groups describing the science we do and the career pathways in science

influenced several school students to pursue science-based A-levels and degree options
Year(s) Of Engagement Activity 2006,2007,2008,2009,2010,2011,2012,2013,2014,2015,2017
 
Description Science Live/European Researchers Night at Natural History Museum 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? Yes
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact More than 3,000 members of the public attended the event as a whole, an open evening at the Natural History Museum, with about 800 scientists including NHM staff and invited others, including myself. I was at the "Science Bar" available to members of the public wanting to meet or question a real scientist about any issue relating to science. Each year of this activity, I spoke to between 10 and 20 members of the public in detail over the course of the evening.

Talked at length with a range of members of the public many of whom were thirsty for science knowledge having somehow missed out at school. One individual said that she had learned more talking to me that evening than in the whole of her school career!
Year(s) Of Engagement Activity 2012,2013,2014,2015,2016
 
Description Science-Humanities Workshop 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? Yes
Type Of Presentation Workshop Facilitator
Geographic Reach Regional
Primary Audience Other academic audiences (collaborators, peers etc.)
Results and Impact Some 50 academics bridging the science-humanities divide participated and discussed a range of issues and proposals for actual and potential synergies

"Scoping Study" report published. See www.reading.ac.uk/cultivating-common-ground
Year(s) Of Engagement Activity 2012