The Morphological Metrology Study Of Intraepidermal Nerve Fiber In Healthy Human

Skin biopsy with measurement of intraepidermal nerve fiber density is a sensitive measure of small fiber injury, which has been used only for about 15 years. Evaluating intraepidermal nerve fiber density (IENFD) by using of skin biopsy has a number of advantages over conventional measures in the study of small-fiber sensory neuropathies. Because this technique is sensitive, specific, simple, inexpensive, painless, minimally invasive and easily repeatable at the same sites or at different time, it holds promise as a clinical trial endpoint measure. But up to now, there have been no such report on morphology and quantitation of IENFs in our country.71 biopsy samples surgical procedures involving 71 patients of different ages (3-80 years) were mainly analyzed in our experiment. The biopsy samples were divided into five groups based on anatomical site: upper arm, elbow, wrist, upper leg and distal leg. A biopsy sample from finger pulp was also involved. 24 biopsy samples from wrist were divided into four groups based on age, each group had 6 examples, 18 biopsy samples from distal leg were also divided into three groups as before. Specimens were fixed routinely in formalin and thereafter embedded in paraffin. Ten-micrometer paraffin thick sections were incubated with rabbit polyclonal antibodies to human PGP9.5. Using light microscopy, we observed the morphological character of IENFs, and at the same time completed the quantitation of epidermal innervation. At last, we analyzed effect of age and anatomic site on IENFD by statistical analysis, and discuss the reference value scope of IENFD.The result of immunohistochemical staining suggested that, the normal human skin had abundant epidermal nerve fibers, subepidermal nerve plexuses,dermal nerve fibers and Meissner's corpuscles (finger pulp) immunoreactive to PGP 9.5. Individual epidermal nerve fibers ascended perpendicularly in the epidermis after arising from the subepidermal nerve plexuses, which paralleled the junction between the epidermis and the dermis, and finally terminated at the stratum granulosum or the lower portion of the stratum corneum, the outermost layer of the epidermis. The epidermal innervation of every skin biopsy was determined in all subjects. The results were expressed as mean±SD density. The comparison of epidermal innervation between the healthy control groups on the basis of age(range: 8~58 years) in wrist did not show any significant age-related effect on IENFD change(P=0.830), The comparison of epidermal innervation between the healthy control groups on the basis of age(range: 3~68 years) in distal leg did not show any significant age-related effect on IENFD change(P=0.478); The density of the intraepidermal nerves varied in the different body areas(P = 0.001). There were no statistically significant differences between the innervation of upper arm, elbow and upper leg. Conversely, the upper arm and upper leg were significantly more densely innervated compared with the wrist and distal leg. We determined that the density of intraepidermal fibers in normal controls was 62.87±15.25 fibers/mm~2 at the distal part of the leg, which was lower than that of Finnish.From the above results, we can draw the conclutions as follow: (1) Values of the skin biopsy technique in clinical application were very high, because it was convenient to draw the materials, simple to operate, inexpensive, and had quantitative index. It may be an outcome measure in peripheral neuropathy trials; (2) Normal means and ranges for the density of epidermal nerve fibers in a reference population have been established. The density of intraepidermal fibersin normal controls was 62.87±15.25 fibers/mm2 at the distal part of the leg in China. Density of IENF below the lower 5th percentile was considered abnormal. The 5th percentile of IENFD in distal leg was 40.55 fibers/mm2; (3) Epidermal innervation of skin at wrist or at distal leg did not change with age. The density of the intraepidermal nerves varied in the different body areas,there was a decreasing gradient of epidermal innervation from proximate limbs to distal end; (4) Ethnicity may effect on IENFD, the density of intraepidermal fibers of Chinese at distal leg was lower than that of Finnish.