Autofluorescent Flavoprotein Imaging of Intraepidermal Nerve Fibers: a Pilot Study

The first aim of the current project is to test the precision of AFI in the epidermis of 10 healthy volunteers. For this purpose, a range of nociceptive electrical stimuli with increasing intensities (5Hz @ 0.5mA-1.0mA) and one innocuous control stimulus (2000Hz @1mA) will be delivered to the third finger of each subject. The outcome measure is AFI-intensity, which is the change in autofluorescence intensity compared to baseline (delta F/F). The standard deviation of AFI intensity will be the measure of precision. Pearson's correlation coefficient will be calculated between electrical stimulus intensities and AFI intensity. A linear correlation needs to be confirmed, since this is a general characteristic of AFI. A paired t-tests will be performed to compare AFI intensities following 5Hz @ 1mA stimulation and 2000Hz @ 1mA stimulation. Lidocaine/prilocaine cream will be applied to the fingertips of the subjects and the electrical stimuli will be repeated, to serve as a negative control experiment. A repeated-measures ANOVA will be performed to compare AFI intensities before and after application of lidocaine/prilocaine cream.

The second aim of our study is to validate AFI in experimentally induced small nerve fiber degeneration of the epidermis, by comparing AFI intensities in subjects before and one week after application of a 8% capsaicin patch to the third fingertip. A repeated-measures ANOVA will be performed to compare AFI intensities before and after capsaicin-induced small nerve fiber degeneration. Assuming that all subjects develop epidermal small fiber degeneration following the 8% capsaicin patch, a statistically significant difference in AFI intensity would serve as a proof-of-principle and would provide validity to autofluorescent flavoprotein imaging of epidermal nociceptor activity as a diagnostic test for SFN. Comparing the distributions of before and after capsaicin-induced small nerve fiber degeneration will lead to a probability estimation of having SFN based on the outcome measure, i.e. AFI intensity. In future research, false-positive and false-negative consequences will be evaluated, leading to cut-off values in patients suspected for SFN.