Correlation of Nav1.8 and Nav1.9 sodium channel expression with neuropathic pain in human subjects with lingual nerve neuromas

Abstract

Background: Voltage-gated sodium channels Nav1.8 and Nav1.9 are expressed preferentially in small diameter sensory neurons, and are thought to play a role in the generation of ectopic activity in neuronal cell bodies and/or their axons following peripheral nerve injury. The expression of Nav1.8 and Nav1.9 has been quantified in human lingual nerves that have been previously injured inadvertently during lower third molar removal, and any correlation between the expression of these ion channels and the presence or absence of dysaesthesia investigated.

Results: Immunohistochemical processing and quantitative image analysis revealed that Nav1.8 and Nav1.9 were expressed in human lingual nerve neuromas from patients with or without symptoms of dysaesthesia. The level of Nav1.8 expression was significantly higher in patients reporting pain compared with no pain, and a significant positive correlation was observed between levels of Nav1.8 expression and VAS scores for the symptom of tingling. No significant differences were recorded in the level of expression of Nav1.9 between patients with or without pain.

Conclusions: These results demonstrate that Nav1.8 and Nav1.9 are present in human lingual nerve neuromas, with significant correlations between the level of expression of Nav1.8 and symptoms of pain. These data provide further evidence that changes in expression of Nav1.8 are important in the development and/or maintenance of nerve injury-induced pain, and suggest that Nav1.8 may be a potential therapeutic target.

Figure 1

Nav1.8 and Nav1.9 expression in lingual neuromas. A and B. Nav1.8 and Nav1.9 immunolabelling was bright and appeared to be axon-specific, showing positive co-localisation with PGP9.5. Images showing immunofluorescent labelling in the lingual nerve neuromas for PGP9.5 (green) and Nav1.8 and Nav1.9 (red). The composite images show the expression of Nav1.8 and Nav1.9 within PGP9.5 labelled tissue (yellow). C. No positive staining for sodium channel subtypes was present where the primary antibodies were omitted. Images of lingual nerve neuromas following incubation with Nav1.8 antibody, Nav1.9 antibody, and secondary antibody alone (Control). Scale bar = 100 μm (A and C); 50 μm (B).

Figure 2

Nav1.8 is expressed in human lingual nerve neuromas. A. Nav1.8 immunolabelling appeared greater in lingual nerve neuromas from patients with symptoms of dysaesthesia. Images of lingual nerve neuromas from patients without or with symptoms of dysaesthesia, showing double labelling with the general neuronal marker PGP9.5 (green) and Nav1.8 (red). Scale bar, 100 μm. B. The mean percentage area (± SEM) of PGP9.5 labelled tissue containing Nav1.8 was significantly higher in lingual nerve neuroma specimens taken from patients with symptoms of pain (unpaired t-test p = 0.0087; no pain 10.52 ± 4.93%, pain 31.82 ± 4.54%).

Figure 3

Correlation of Nav1.8 and Nav1.9 with symptoms of pain, tingling and discomfort. A. Regression plots showing the relationship between the expression of Nav1.8 and VAS scores for pain, tingling and discomfort as reported by the patient. The level of Nav1.8 expression correlated positively with reported levels of tingling (Pearson’s Correlation r = 0.64, p = 0.02). B. Regression plots showing the relationship between the expression of Nav1.9 and VAS scores for pain, tingling and discomfort as reported by the patient, revealed there was no significant correlation between the degree of symptoms experienced and the level of Nav1.9 expression.

Figure 4

Nav1.9 is expressed in human lingual nerve neuromas. A. Nav1.9 immunolabelling appeared greater in lingual nerve neuromas from patients with symptoms of dysaesthesia. Images of lingual nerve neuromas from patients without or with symptoms of dysaesthesia, showing double labelling with the general neuronal marker PGP9.5 (green) and Nav1.9 (red). Scale bar, 100 μm. B. The mean percentage area (± SEM) of PGP9.5 labelled tissue containing Nav1.9 did not reveal any significant differences between the two groups of neuromas (unpaired t-test p = 0.27; no pain 9.16 ± 1.51%, pain 12.8 ± 2.38%).