ラット坐骨神経モデルにおけるヒト線維芽細胞を用いたscaffold-free Bio 3D conduitの末梢神経再生に対する有効性

京都大学0048新制・課程博士博士(医学)甲第21675号医博第4481号新制||医||1036(附属図書館)京都大学大学院医学研究科医学専攻(主査)教授 渡邉 大, 教授 林 康紀, 教授 井上 治久学位規則第4条第1項該当Doctor of Medical ScienceKyoto UniversityDFA

Opening Wedge Osteotomy for Valgus Deformity of the Little Finger after Proximal Phalangeal Fracture in Children: Two Case Reports

In the treatment of posttraumatic valgus deformity of the pediatric little finger, it is usually difficult to achieve accurate correction of angular and rotational deformity using closing wedge osteotomy. We report two cases of valgus deformity of the little finger (both 11-year-old female patients) successfully treated using opening wedge osteotomy followed by intramedullary semirigid fixation with a single Kirschner wire. A wire tip inserted from the retrocondylar fossa of the proximal phalangeal head was advanced along the radial side of the intramedullary cortex after gradual opening of the osteotomy site. If needed, further fine adjustment of the rotational alignment can be performed even after K-wire insertion. Postoperatively, the gap between the little and ring fingers in the fully extended and adducted position and the finger overlapping in the fully flexed position were completely resolved. The flexibility of the pediatric bone and sagittal clearance between the wire and the inner wall of the proximal phalangeal medullary cavity allow fine adjustment of the rotational alignment even after wire insertion

The efficacy of a scaffold-free Bio 3D conduit developed from human fibroblasts on peripheral nerve regeneration in a rat sciatic nerve model

バイオ3Dプリンターを用いた末梢神経損傷に対する神経再生技術の開発. 京都大学プレスリリース. 2017-02-27.バイオ3Dプリンタを用いた神経再生技術の開発に成功 --末梢神経損傷に対する三次元神経導管の医師主導治験を開始--. 京都大学プレスリリース. 2021-01-15.Background: Although autologous nerve grafting is the gold standard treatment of peripheral nerve injuries, several alternative methods have been developed, including nerve conduits that use supportive cells. However, the seeding efficacy and viability of supportive cells injected in nerve grafts remain unclear. Here, we focused on a novel completely biological, tissue-engineered, scaffold-free conduit. Methods: We developed six scaffold-free conduits from human normal dermal fibroblasts using a Bio 3D Printer. Twelve adult male rats with immune deficiency underwent mid-thigh-level transection of the right sciatic nerve. The resulting 5-mm nerve gap was bridged using 8-mm Bio 3D conduits (Bio 3D group, n = 6) and silicone tube (silicone group, n = 6). Several assessments were conducted to examine nerve regeneration eight weeks post-surgery. Results: Kinematic analysis revealed that the toe angle to the metatarsal bone at the final segment of the swing phase was significantly higher in the Bio 3D group than the silicone group (-35.78 ± 10.68 versus -62.48 ± 6.15, respectively; p < 0.01). Electrophysiological studies revealed significantly higher compound muscle action potential in the Bio 3D group than the silicone group (53.60 ± 26.36% versus 2.93 ± 1.84%; p < 0.01). Histological and morphological studies revealed neural cell expression in all regions of the regenerated nerves and the presence of many well-myelinated axons in the Bio 3D group. The wet muscle weight of the tibialis anterior muscle was significantly higher in the Bio 3D group than the silicone group (0.544 ± 0.063 versus 0.396 ± 0.031, respectively; p < 0.01). Conclusions: We confirmed that scaffold-free Bio 3D conduits composed entirely of fibroblast cells promote nerve regeneration in a rat sciatic nerve model

The Efficacy of a Scaffold-free Bio 3D Conduit Developed from Autologous Dermal Fibroblasts on Peripheral Nerve Regeneration in a Canine Ulnar Nerve Injury Model: A Preclinical Proof-of-Concept Study

Autologous nerve grafting is widely accepted as the gold standard treatment for segmental nerve defects. To overcome the inevitable disadvantages of the original method, alternative methods such as the tubulization technique have been developed. Several studies have investigated the characteristics of an ideal nerve conduit in terms of supportive cells, scaffolds, growth factors, and vascularity. Previously, we confirmed that biological scaffold-free conduits fabricated from human dermal fibroblasts promote nerve regeneration in a rat sciatic nerve injury model. The purpose of this study is to evaluate the feasibility of biological scaffold-free conduits composed of autologous dermal fibroblasts using a large-animal model. Six male beagle dogs were used in this study. Eight weeks before surgery, dermal fibroblasts were harvested from their groin skin and grown in culture. Bio 3D conduits were assembled from proliferating dermal fibroblasts using a Bio 3D printer. The ulnar nerve in each dog’s forelimb was exposed under general anesthesia and sharply cut to create a 5 mm interstump gap, which was bridged by the prepared 8 mm Bio 3D conduit. Ten weeks after surgery, nerve regeneration was investigated. Electrophysiological studies detected compound muscle action potentials (CMAPs) of the hypothenar muscles and motor nerve conduction velocity (MNCV) in all animals. Macroscopic observation showed regenerated ulnar nerves. Low-level hypothenar muscle atrophy was confirmed. Immunohistochemical, histological, and morphometric studies confirmed the existence of many myelinated axons through the Bio 3D conduit. No severe adverse event was reported. Hypothenar muscles were re-innervated by regenerated nerve fibers through the Bio 3D conduit. The scaffold-free Bio 3D conduit fabricated from autologous dermal fibroblasts is effective for nerve regeneration in a canine ulnar nerve injury model. This technology was feasible as a treatment for peripheral nerve injury and segmental nerve defects in a preclinical setting

〈Review〉Peripheral nerve storage and allotransplantation using green tea polyphenol in rats—its possibility for clinical application—Review of our works.

Background. We examined whether peripheral nerve segments immersed in polyphenol solution could be preserved and allotransplanted into rats without using immunosuppressants. Methods. Sciatic nerve segments, 20 mm long, were harvested from male Lewis or DA rats and transplanted into 15-mm sciatic nerve deficits in Lewis rats immediately, or after polyphenol treatment (immersion in 1 mg/mL polyphenol solution for one week and in Dulbecco\u27s modified Eagle\u27s medium for three weeks more), without using immunosuppressants. In the FA group, DA rat nerve segments were transplanted into Lewis rats immediately. In the PA group, DA rat nerve segments were transplanted into Lewis rats after polyphenol treatment. In the FI group, Lewis rat nerve segments were transplanted immediately into Lewis rats. In the PI group, Lewis rat nerve segments were transplanted into Lewis rats after polyphenol treatment. Nerve regeneration was assessed electrophysiologically and histomorphologically. Nerve segments from male Lewis or DA rats were also transplanted into female Lewis rats as in the PI, PA, FI, and FA groups. Genomic DNA was extracted from each transplanted nerve segment and subjected to polymerase chain reaction amplification specific for the sex-determining region of Y-chromosome and β-actin genes. Immunostaining for S-100 and glial fibrillary acidic protein, and Y-chromosome-specific in situ hybridization were also performed. Results. Nerve regeneration in the PI and PA groups was similar to that in the FI group and was significantly greater than that in the FA group. The ratios of the Sry/β-actin polymerase chain reaction products for PA, PI, FI, and FA groups were 0.144, 0.294, 0.615, and 0, respectively. Some of the S-100-positive and glial fibrillary acidic protein-positive cells displayed Sry signals in the PI and FI groups, but not in the FA group. Conclusion. Some Schwann cells survived for one month in polyphenol-treated nerve segments that were transplanted allogeneically without immunosuppressants

Immunohistochemistry of the mid portion of the regenerated nerve eight weeks after surgery in both groups.

<p>A-C: Longitudinal sections in the Bio 3D group. D-F: Longitudinal sections in the silicone group. G-I: Transverse sections in the Bio 3D group. J-L: Transverse sections in the silicone group. A-F: scale bar = 100 μm. G-L: scale bar = 500 μm.</p

Electrophysiological studies and wet muscle weight of the tibialis anterior muscle eight weeks after surgery.

<p>A: CMAP was significantly higher in the Bio 3D group than the silicone group (<i>p</i> < 0.01). B: Regarding the NCV, there was no significant difference among two groups. C: Wet muscle weight was significantly higher in the Bio 3D group than in the silicone group (<i>p</i> < 0.01). All values are expressed as the percentage of those from the left hind limb. Error bars represent standard deviations.</p