Objectives:
Nerve growth factor (NGF) and its receptors have been shown to increase following traumatic injury. The aim of the study was to examine the changes that occur in the periodontal ligament of orthodontically traumatised teeth after localised injection of anti-NGF.

Design:
Orthodontic NiTi pullcoil springs were ligated to the right maxillary first molar and maxillary incisors of 17, eight week old, male Sprague-Dawley rats which were divided into two groups. The first group were injected with anti-NGF palatal to the right maxillary first molar. The second group were injected with saline in the same location to serve as a control. The left maxillary molars from each animal served as internal controls. Animals from the two groups were sacrificed at 7 and 14 days. Animals from a third matched group were neither injected nor had an orthodontic spring attached and were sacrificed to serve as an additional untreated control group. 25 µm horizontal frozen sections were obtained using a cryostat at -20°C and mounted on the slides to allow direct comparison between untreated and experimental sides of the mesiopalatal root periodontal ligament circumference. Immunohistochemical tissue labelling was performed, involving the avidin-biotin peroxidase technique and rabbit anti-calcitonin gene-related peptide (anti-CGRP) rat serum as the primary antibody. Photomicrographs were analysed using Image J software with a colour deconvolution plug-in to compare and quantify the staining intensity of the immunolabelled tissue. Data was compared between each of the groups using the Neuman-Keul paired test on Graphpad Prism 5 software.

Results:
A semi-quantitative comparison of the immunohistochemical CGRP staining showed a statistically significant (p<0.05) reduction in staining intensity in the anti-NGF injected group when compared with the saline injected group for both day 7 and day 14 groups. There was a statistically significant (p<0.05) reduction in staining intensity between the anti-NGF injected (group 1), the uninjected (group 2) and untreated control animals (group 3).

Conclusion:
Data suggest that the injection of anti-NGF reduces NGF tissue levels and prevents the re-innervation by CGRP-positive fibres in the periodontal ligament of rat molars subjected to orthodontic tooth movement.