The aim of this study was to estimate dermal nerve fiber length (DNFL) using a stereological sampling technique in comparison with a previously reported manual estimation. DNFL was analyzed in skin punch biopsy specimens from 24 healthy volunteers and 18 patients with small fiber neuropathy (SFN) using global spatial sampling that yields unbiased and reliable length estimation. The estimation was carried out in 50-μm biopsy sections after immunostaining with anti-protein gene product (PGP) 9.5 antibodies. The length of the PGP9.5-positive dermal nerves from the dermal-epidermal junction and 200 μm down was measured (DNFL mm(-2) ). Results were compared with our previously reported manual method. Patients showed a significantly (p <0.0001) lower DNFL (105 mm(-2) ± 6.4 SD) than healthy subjects (246 mm(-2) ± 8.39 SD). Moderate correlation with age was observed for both healthy subjects (Pearson's r = -0.33) and patients (Pearson's r = -0.59). A significant (p <0.001) correlation between global spatial sampling and manual estimation was observed in both patients and healthy subjects (Pearson's r = 0.62 and 0.61, respectively). These findings provide further evidence on the reliability of dermal nerve morphometry in human skin and strengthen the hypothesis that dermal nerve fibers undergo significant degeneration in SFN.
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