Molecular basis of craniosynostis
One in 2500 children are born with craniosynostosis, a devastating
medical disorder where the bones of the skull fuse prematurely,
resulting in abnormal skull development, visual and neurological
problems and mental impairment. There is a clear need to develop
adjunct therapies to minimize the need for repeated invasive
cranial surgery and enable proper skull and brain growth.
The underlying causes of the majority of craniosynostoses
are not known and to develop new therapies we need to know
what the mechanisms are and how they act on skull growth.
Two molecules we have discovered from our studies in children
with craniosynostosis have been a major focus of our research
in the past year. These are a protein known as retinol binding
protein 4 (RBP4) that binds vitamin A, an important vitamin
for bone growth, and a proteoglycan called glypican which
regulates signalling by growth factors.
In collaboration with surgeons in the Australian Craniofacial
Unit we have continued to recruit children undergoing surgical
treatment for craniosynostosis to our study. Notably, Senior
Craniofacial Consultant and Associate Professor, Peter Anderson,
remains an invaluable collaborator.
In studying the function of RBP4 and glypican we have optimised
methods for their detection in human and mouse cells using
antibodies. We have identified where they are located and
have used flow cytometry to analyse the distribution of some
of them on the cell surface. To complement the protein studies
we have optimised the in situ hybridization technique to define
where the genes are active in tissue. To date, our studies
have focussed on gene expression in embryonic limbs where
we have shown that glypicans are expressed in several different
tissue compartments and that other molecules are either specific
to the cartilage forming regions or the bone forming regions.
We are now adapting this method to study the patterns of gene
expression in cranial sutures and bone formation in the skull.
In our studies of gene function, where we hope to gain insight
into function by overproducing or inhibiting gene activity,
we have continued to refine our lentiviral vectors for gene
delivery and are now developing vectors to study the effects
of gene knockdown. Importantly, we have also used Dual Luciferase
assay technology to show that glypicans can modulate the action
of growth factors in cultured human suture cells, indicating
that these are potentially very important molecules for skull
growth. A collaboration with Prof Jorge Filmus, University
of Toronto, Canada, has been important in obtaining molecular
reagents and advice for our studies on glypicans.
We are also expanding our use of mouse models in the study
of skull growth and craniosynostosis and have imported RBP4
gene knockout mice from Dr Loredana Quadro, Rutgers University,
USA, for our study of RBP4 function. Other strains of gene
knockout mice are also being imported, and by cross-breeding
them to produce new models we hope to obtain insight into
the role these molecules play in skull growth and craniosynostosis.
Antibody detection of glypican (white stain) in cultured
human suture cells
Mouse model of the human Crouzon craniosynostosis syndrome
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