Standard deviation of the distances between consecutive epidermal nerve fibers as a parameter to study their distribution in human skin

Abstract

Evaluation of epidermal nerve fiber (ENF) density by skin biopsy is nowadays considered as the gold standard for the diagnosis of small fiber neuropathies. ENFs show in normal subjects quite a regular distribution between the keratinocytes while in patients with small fiber neuropathies we can find, besides a loss of ENFs, evident anomalies of their distribution. Different patterns of nerve fiber dispersion can express different pathophysiological processes. The aim of this study was to define a tool capable of describing the distribution of ENF in the epidermis. For this purpose, we measured the distance between subsequent epidermal fibers on skin samples of healthy and diabetic subjects. We calculated the standard deviation of the interfiber length as a parameter (the dispersion index – DI) to evaluate the variability of the distance between consecutive ENFs. We selected thigh skin samples from five healthy subjects (age range 30–50 years) and five age and sex matched diabetic patients. Samples were processed using immunohistochemical techniques. Sections were PGP 9.5 and Collagen IV double stained to show nerve fibers and basement membrane. One randomly selected section for each subject was acquired for its entire length (usually seven 20x confocal images obtained from a z‐stack of 16 two micron optical sections) using a confocal microscope (CARV, Atto Biosciences, Rockville MD, USA). We measured the distance between consecutive ENFs along the entire epidermal length, using dedicated software (ScionImage, Scion Corporation Frederick, Maryland USA). In diabetic patients the DI resulted significantly higher than in normal subjects. We suggest that DI along with the ENF density can be useful to better characterize epidermal innervation.