Evaluation of anatomical and histological characteristics of human peripheral nerves: as an effort to develop an efficient allogeneic nerve graft.

Management of peripheral nerve defects is a complicated problem in clinical contexts. Autologous nerve grafting, a gold standard for surgical treatment, has been well known to have several limitations, such as donor site morbidity, a limited amount of available donor tissue, and size mismatches. Acellular nerve allografts (ANAs) have been developed as an alternative and have been applied clinically with favorable outcomes. However, because of the limited availability of commercialized ANAs due to supplier-related issues and high costs, efforts continue to produce alternative sources for ANAs. The present study evaluated the anatomical and histological characteristics of human peripheral nerves using 25 donated human cadavers. The length, diameter, and branching points of various peripheral nerves (median, ulnar, tibial, lateral femoral cutaneous, saphenous, and sural nerves) in both the upper and lower extremities were evaluated. The cross-sectional area (CSA), ratio of fascicular area, and numbers of fascicles were also evaluated via histologic analysis. CSA, the ratio of fascicular area, and the number of fascicles were analyzed statistically in correlation with demographic data (age, sex, height, weight, BMI). The mean length of all evaluated nerves ranged from 17.1 to 41.4 cm, and the mean diameter of all evaluated nerves ranged from 1.2 to 4.9 mm. Multiple regression analysis revealed correlations between the ratio of fascicular area and sex (p = 0.005) and BMI (p = 0.024) (R2 = 0.051). The results of the present study will be helpful in selecting necessary nerve allograft sources while considering the characteristics of each nerve in the upper and lower extremities during ANAs production.

Comparison of two different decellularization methods for processed nerve allograft.

The use of processed nerve allografts as an alternative to autologous nerve grafts, the gold standard treatment for peripheral nerve defects, is increasing. However, it is not widely used in Korea due to cost and insurance issues. Moreover, the main detergent used in the conventional Hudson method is unavailable. Therefore, a new nerve allograft decellularization process is needed. We aimed to compare the traditional Hudson method with a novel decellularization process that may remove cellular content more efficiently while preserving the extracellular matrix (ECM) structure using low concentration sodium dodecyl sulfate (SDS) and nuclease. After each decellularization process, DNA content was measured in nerve tissue. Masson's trichrome staining and scanning electron microscopy were performed to determine the state of preservation of the ECM. A significantly greater amount of DNA content was removed in the novel method, and the ECM structure was preserved in both methods. For the in vivo study, a 15-mm long sciatic nerve defect was created in two groups of Sprague-Dawley rats, and processed nerve allografts decellularized using the Hudson or novel method were transplanted. Functional and histological recovery results were measured 12 weeks post-transplantation. Ankle contracture angle, maximal isometric tetanic force of the tibialis anterior (TA), and the TA mass were compared between the groups, as well as the percent neural tissue (100 × neural area/intrafascicular area). There was no significant difference in functional and histological nerve recovery between the methods. The novel method is appropriate for developing a processed nerve allograft.

Effectiveness and Biocompatibility of Decellularized Nerve Graft Using an In Vivo Rat Sciatic Nerve Model.

BACKGROUND: Decellularized nerve allografting is one of promising treatment options for nerve defect. As an effort to develop more efficient nerve graft, recently we have developed a new decellularization method for nerve allograft. The aim of this study was to evaluate the effectiveness and biocompatibility of nerve graft decellularized by our newly developed method. METHODS: Forty-eight inbred male Lewis rats were divided into two groups, Group I (autograft group, n = 25), Group II (decellularized isograft group, n = 23). Decellularized nerve grafts were prepared with our newly developed methods using amphoteric detergent and nuclease treatment. Serum cytokine level measurements at 0, 2, and 4 weeks and histologic evaluation for inflammatory cell infiltration at 6 and 16 weeks after nerve graft. RESULTS: There was no significant difference in mean maximum isometric tetanic force and weight of tibialis anterior muscle or ankle angle at toe-off phase between two groups at 6 and 16 weeks survival time points (p > 0.05). There was no inflammatory cell infiltration in either group and histomorphometric assessments of 6- and 16-week specimens of the isograft group did not differ from those in the autograft group with regard to number of fascicle, cross sectional area, fascicle area ratio, and number of regenerated nerve cells. CONCLUSION: Based on inflammatory reaction, axonal regeneration, and functional outcomes, our newly developed decellularized nerve grafts were fairly biocompatible and had comparable effectiveness to autografts for nerve regeneration, which suggested it would be suitable for nerve reconstruction as an alternative to autograft.

The anatomy of the saphenous and sural nerves as a source of processed nerve allografts.

As an alternative to autologous nerve donors, acellular nerve allografts (ANAs) have been studied in many experiments. There have been numerous studies on processing ANAs and various studies on the clinical applications of ANA, but there have not been many studies on sources of ANAs. The purposes of the present study were to evaluate the course of the saphenous and sural nerves in human cadavers and help harvest auto- or allografts for clinical implications. Eighteen lower extremities of 16 fresh cadavers were dissected. For the saphenous nerve and sural nerve, the distances between each branch and the diameters at the midpoint between each branch were measured. In the saphenous nerve, the mean length between each branch ranged from 7.2 to 28.6 cm, and the midpoint diameter ranged from 1.4 to 3.2 mm. In the sural nerve, the mean length between each branch ranged from 17.4 to 21 cm, and the midpoint diameter ranged from 2.3 to 2.8 mm. The present study demonstrates the length of the saphenous and sural nerve without branches with diameters larger than 1 mm. With regard for the clinical implications of allografts, the harvest of a selective nerve length with a large enough diameter could be possible based on the data presented in the present study.

Comparison of cytokine expression and ultrastructural alterations in fresh-frozen and dried electron beam-irradiated human amniotic membrane and chorion.

The purpose of the current study was to compare the effects of drying and fresh-freezing on human amniotic membrane (HAM) and amnion/chorion membrane (HACM) in terms of histological and structural characteristics and cytokine levels. HAM and HACM samples, obtained from six placentae, were investigated. HAM and HACM were dried, electron beam-irradiated (dehydration group; d-HAM/d-HACM), or fresh-frozen (freezing group; f-HAM/f-HACM). Luminex assay was used to assay the levels of 15 cytokines. The ultrastructural characteristics of HAM and HACM were evaluated using light and transmission electron microscopies. Total cytokine contents did not show the statistical difference between dehydration and fresh-freezing process. Significantly higher levels of total cytokines were observed in HACM than in HAM. Epidermal growth factor (EGF) level was significantly higher in d-HAM than in the other samples. The levels of most of the other growth factors were higher in HACM than in HAM, but there was no statistical difference between the dehydration process and the fresh-freezing process. The levels of the cytokines, other than the growth factors, were higher in HACM than in HAM, and higher concentrations of cytokines were observed in the freezing group than in the dehydration group. Histological examination revealed that the dehydration group had thinner tissues than the freezing group, but the structural stability, including the basement membrane, did not differ between the two groups. Microscopic structures such as microvilli and nuclei were well-preserved in the freezing group, based on the results of the transmission electron microscopy. Our dehydration process maintained the histological structure of HAM/HACM and a variety of growth factors and cytokines were identified. Especially, the HAM, processed with the dehydration method, had a higher EGF level than that processed with the fresh-freezing method. Therefore, dehydration method can be used to effectively promote wound repair.