Page 151 - FINAL COMPENDIUM 2020-2021 27.2.2022

AN INVESTIGATION INTO THE DENTAL NERVE INNERVATION OF GAS 1 MUTANT MICE

Yim KM, Cobourne M, Seppala M

Introduction: Gas1 (Growth arrest-specific gene 1) encodes a transmembrane protein
that together with other co-receptors, Cdo (cell-adhesion-molecule-related/
downregulated by oncogenes) and Boc (Brother of Cdo), can either facilitate or negatively
regulate Shh (Sonic hedgehog) signalling in a context- dependent manner. Gas1 loss-of-
function mice have been shown to develop with supernumerary teeth and their
development is preceded by prolonged Shh expression in normally regressing
rudimentary R2 epithelial swelling in the non-odontogenic diastema. Dental nerves play
an important role in tooth formation and their innervation is guided by distinct molecular
signalling pathways. The dental nerve innervation takes place in an extremely coordinated
manner and it has been shown that the timing of axon navigation, axon differentiation as
well as the patterning of the axons in the tooth germ mesenchymal tissue happens
concomitantly to the tooth morphogenesis demonstrating a strong association with
variable stages of tooth development. Objective: To compare the nerve innervation
between the control and Gas1-/- developing molars in terms of their nerve branching,
innervation to the target field and survival of the nerve axons. Materials and methods: A
total of 38 specimens were sectioned to include stages of early dental nerve innervation
that are crucial time point for supernumerary development (E12.5, E13.5 and E14.5).
Sections covering the maxillary and mandibular molar regions were chosen for
Haematoxylin and Eosin(H&E) staining and immunostaining. The primary antibody used
as a nerve market were either Rabbit Anti-beta III Tubulin antibody (TUJ1) or Mouse Anti-
beta III Tubulin antibody (TUJ1). Goat Anti-Mouse Alexa Fluor 488 or Donkey Anti-Rabbit
Fluor 568 were used as a secondary antibody marking the nerve with either green(anti-
mouse) or red(anti-rabbit) in immunostaining. Results: Quantitative comparison of E14.5
control and Gas 1-/-nerve axons were done by comparing 10 chosen sections (four of
them containing both maxilla and mandible). These 10 chosen sections (3 maxillary
control, 4 maxillary mutants, 4 mandibular control and 3 mandibular mutants) were
chosen based on the quality of the image for qualitative analysis to assess the branching
morphogenesis of the developing dental nerve axons in the presumptive follicular field.
Cross sections of the axons were calculated twice in a dark room to enable better visibility
and blinding was done. The average value indicated that that there was no remarkable
difference in the number of the axonal cross sections between the maxillary control (10.7)
and mutants (8.25) nor mandibular control (11.4) and mutants (12.5). Conclusion: This
study revealed comparable nerve innervation between control and Gas1-/- mice and
therefore other molecular and cellular mechanisms are more likely to play a part in the
aetiology of supernumerary development in Gas1-/- mice.

Thesis dissertation towards the requirement for the Master of Science in Orthodontics Programme, King’s College London,
2020

Dr Yim Kah Mun Prof. Martyn Cobourne
Orthodontic Unit Dr Maisa Seppala
Jalan Abdul Samad Dental Clinic King’s College London
Johor Bahru, Johor United Kingdom

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