Saliva as a medium to detect and measure biomarkers related to pain

Abstract

Saliva is often neglected as a body fluid of diagnostic or prognostic value, even though generally well accepted by the patients. This is due to lack of a standardized collection procedure. The aim of this study was to identify the ideal saliva collection technique and develop new sensitive methods to detect and analyse markers related to pain in healthy pain-free subjects. Plasma and five different saliva collection approached was evaluated during strictly controlled conditions. Levels of nerve growth factor (NGF), calcitonin gene-related peptide (CGRP) and brain derived neurotropic factor (BDNF) were determined using novel western blotting based technology. Glutamate and substance P (SP) was determined using commercial available methods. Several new isoforms were found for NGF, CGRP and BDNF in saliva. The isoform pattern showed significant variation in both expression and chemiluminescence levels between different collection methods. New sensitive methods to study pain related markers in saliva were developed in this study. Furthermore, we are first to demonstrate a correlation between the Glutamate concentration in stimulated whole saliva and blood. However, the fundamental conclusion drawn is the importance of consistency in the collection method.

Figure 1

Illustrative overview of the main salivary glands and different collection approaches used in the study. The colours are associated with the different collection methods explained in the diagrams.

Figure 2

Salivary and plasma NGF, CGRP, BDNF, glutamate and substance P concentration in 20 healthy individuals matched for age and gender (Fig. 2A–H). Large variations were observed between different collection methods. Several isoforms was detected for NGF, CGRP and BDNF. The isoform pattern showed significant variation in both expression and chemiluminescence levels between different collection methods (Friedman; P < 0.05). All stimulated saliva samples, whether chemically or mechanically stimulated, showed significant higher expression of total-NGF and total-CGRP compared to unstimulated saliva samples and plasma. Higher concentration of glutamate was found in stimulated whole saliva comparing to other salivary collection methods. However, the plasma levels of glutamate and substance P were significantly higher in comparison with the levels detected in saliva.