Tetsuya Kiyotani

Peripheral nerve regeneration across an 80-mm gap bridged by a polyglycolic acid (PGA)-collagen tube filled with laminin-coated collagen fibers : a histological and electrophysiological evaluation of regenerated nerves

We evaluated peripheral nerve regeneration across an 80-mm gap using a novel artificial nerve conduit. The conduit was made of a polyglycolic acid (PGA)-collagen tube filled with laminin-coated collagen fibers. Twelve beagle dogs underwent implantation of the nerve conduit across an 80-mm gap in the left peroneal nerve. In four other dogs used as negative controls, the nerve was resected and left unconnected. Histological observation showed that numerous unmyelinated and myelinated nerve fibers, all smaller in diameter and with a thinner myelin sheath than normal nerve fibers, regrew through and beyond the gap 12 months after implantation. The distribution of the regenerated axonal diameters was different from that of the normal axonal diameters. Compound muscle action potentials, motor evoked potentials, and somatosensory evoked potentials were recorded in most animals 3 months after implantation. Peak amplitudes and latencies recovered gradually, which indicating the functional establishment of the nerve connection with the target organs. In addition to the ordinary electrophysiological recoveries, potentials with distinct latencies originating from Aalpha, Adelta and C fibers became distinguishable at the 6th lumbar vertebra following stimulation of the peroneal nerve distal to the gap 12 months after implantation. The pattern of walking without load was restored to almost normal 10-12 months after implantation. Neither electrophysiological nor histological restoration was obtained in the controls. Our nerve conduit can guide peripheral nerve elongation and lead to favorable functional recovery across a wider nerve gap than previously reported artificial nerve conduits.

NERVE REGENERATION ACROSS A 25-MM GAP BRIDGED BY A POLYGLYCOLIC ACID-COLLAGEN TUBE: A HISTOLOGICAL AND ELECTROPHYSIOLOGICAL EVALUATION OF REGENERATED NERVES

In the study reported here we have examined the nerve regeneration that occurs over a 25-mm gap using a novel biodegradable nerve guide tube. The tube was a composite of polyglycolic acid (PGA) mesh coated with collagen which was filled with neurotrophic factors. The left sciatic nerve of ten adult cats was dissected. The stumps were connected by the tube, and fixed gap. Histological examinations carried out 4-16 months after implantation of the tube revealed regeneration of well vascularized nerve tissue. Regeneration of both myelinated, unmyelinated axons and Schwann cells was confirmed by electron microscopy 5 months after surgery. Following injection of horseradish peroxidase (HRP) into a site peripheral to the regenerated segment of the sciatic nerves, motoneurons in the ventral horn of the spinal cord, afferent terminals in the medial portion of the dorsal column of the medulla oblongata, and sensory afferent nerve terminals in the dorsal horn of the spinal cord were labelled. Electrophysiological examinations revealed restoration of evoked electromyograms and sensory evoked potentials (SEPs) recorded from the cerebral cortex as well as the spinal cord. We also found that some of the regenerated motor axons exhibited branching in the regenerated segments. In two cases, a single motoneuronal axon from the regenerated side projected to both flexors and extensors, simultaneously. Our results indicate that the PGA-collagen composite tube is a promising tool for use as a nerve guide tube in peripheral nerve regeneration.

The experimental replacement of a cervical esophageal segment with an artificial prosthesis with the use of collagen matrix and a silicone stent

OBJECTIVE Attempts have been made to replace esophageal defects with a variety of artificial materials. However, because of the artificial nature of the materials, problems such as infection, leakage, stricture, or dislocation could not be avoided. Therefore we have designed a new type of artificial esophagus that is gradually replaced by host tissue. METHODS Our artificial esophagus was a two-layered tube consisting of a collagen sponge matrix and an inner silicone stent. We used it to replace 5 cm esophageal segmental defects in 43 dogs, and the inner silicone stent was removed endoscopically at weekly intervals from 2 to 4 weeks. RESULTS In the 27 dogs from which the silicone stent was removed at 2 or 3 weeks, constriction of the regenerated esophagus progressed and the dogs became unable to swallow within 6 months. In the 16 dogs from which the silicone stent was removed at 4 weeks, highly regenerated esophageal tissue successfully replaced the defect, leaving no foreign body in the host. Moreover, the regenerated esophagi had stratified flattened epithelia, striated muscle tissue composed of an inner circular and an outer longitudinal muscle layer, and esophageal glands. CONCLUSIONS Even in mature adult higher mammals, esophageal high-order structures can be regenerated provided that an adequate three-dimensional extracellular structure is put in place for a sufficient period.

Intrathoracic esophageal replacement in the dog with the use of an artificial esophagus composed of a collagen sponge with a double-layered silicone tube

OBJECTIVES Intrathoracic esophageal replacement with an artificial esophagus is considered difficult. We attempted to replace the intrathoracic esophagus with an artificial esophagus composed of a collagen sponge with a double-layered silicone tube and examined the state of host tissue regeneration. METHODS A 5-cm long gap was created in the intrathoracic esophagus in 9 dogs and repaired by interposition of our prosthesis. The dogs were fed only by intravenous hyperalimentation for 28 days. The silicone tube was removed at 29 days after the operation, and oral feeding was reintroduced. RESULTS One dog was put to death at each of the following times: 1, 2, 3, 3, 6, 12, and 24 months after the operation. One dog is still surviving without problems after more than 26 months. One dog died of malnutrition at 10 months. In all dogs, the host regenerated tissue had replaced the resulting gap at the time of silicone tube removal. The mucosa had fully regenerated within 3 months and the glands within 12 months. The process of stenosis and shrinkage was complete within 3 months and did not advance thereafter. The lamina muscularis mucosae were observed as islets of smooth muscle within 12 months. Although the skeletal muscle regenerated close to the anastomoses, it did not extend to the middle of the regenerated esophagus even after 24 months. CONCLUSIONS Use of a collagen sponge with a double-layered silicone tube was shown to be feasible even in the thorax and to allow the regenerated host tissue, consisting of the mucosa, glands, and lamina muscularis mucosae, to replace the esophageal gap.

Porous-Type Tracheal Prosthesis Sealed With Collagen Sponge

BACKGROUND Reconstruction of a long section of the trachea is clinically problematic. Tracheal reconstructions using prostheses have met with limited success due to local infection, hemorrhage, luminal stenosis and prosthesis dislocation. METHODS We have designed a porous type of tracheal prosthesis in which the mesh is sealed with collagen sponge. We used this prosthesis (50 mm in length) to reconstruct the cervical trachea in 10 mongrel dogs and evaluated its efficacy. RESULTS One dog died due to an accident with anesthesia at 6 weeks and 1 of suffocation at 10 weeks. The other 8 dogs had an uneventful postoperative course until they were killed between 6 and 24 months after implantation. At sacrifice, all the prostheses had become completely incorporated into the host. Microscopic examination revealed advanced formation of a new epithelial lining in 1 dog at 6 months, and a confluent epithelial lining was observed in another dog at 12 months. Central stenosis was not significant in any of the animals. CONCLUSIONS This tracheal prosthesis gives good results in canine tracheal reconstruction, and appears very promising for the clinical repair of tracheal defects.

Intrathoracic tracheal reconstruction with a collagen-conjugated prosthesis: Evaluation of the efficacy of omental wrapping

Reconstructions of the intrathoracic trachea in 24 dogs were done with the use of 50 mm long collagen-conjugated tracheal prostheses. Omental wrapping was also done in 14 of the dogs (omentopexy group) to evaluate the efficacy of this option in comparison with results in the other 10 dogs (control group). All 24 dogs had uneventful postoperative courses and were killed at 4 weeks or 3, 6, or 12 months after the operation. Better epithelialization and fewer complications, such as mesh exposure and luminal stenosis, were observed in the omentopexy group than in the control group. Angiography and analysis of regenerated blood vessels revealed that vessel ingrowth had started within 4 weeks and that vessel formation reached its maximal point within 6 to 12 months in the omentopexy group. In contrast, revascularization of the subepithelial region in the control group was poor even after 3 months, and vessel formation continued for as long as 12 months. The differences between the two groups were considered to be mainly a result of the speed of blood vessel ingrowth into the regenerated mucosa. We conclude that our prosthesis can be used safely for intrathoracic tracheal reconstruction and that omental wrapping is a useful supplementary method that reduces the occurrence of complications.

A new type of surgical adhesive made from porcine collagen and polyglutamic acid

We have developed a new adhesive for surgical use. The new adhesive is made of three components: porcine collagen, poly(L-glutamic acid) and water-soluble carbodiimides (WSC). The optimum concentration of each component was determined by measuring the time required for gel formation in experiments in vitro. Using these optimum concentrations, we applied the adhesive to wounds made on rats. A conventional fibrin glue was used as a control. Measurement of tensile strength and histological examination were performed 5, 7, 10, and 14 days after the operation. The tensile strength of wounds treated with 2.5 mg/mL collagen glue was not significantly different from that of wounds treated with fibrin glue except at 7 days after the operation (p < 0.05 by Students t-test). Histological examination revealed that the speed of cell infiltration into, and absorption of 2.5 mg/mL collagen glue was slower than for fibrin glue, but faster than for 5.0 mg/mL collagen glue. One of the important advantages of our collagen glue is that the absorption rate of it can be controlled by the collagen concentration. Therefore, it seems to be adequate for sealing air leakage from the lung, which takes a relatively long period for recovery. Moreover it does not contain human serum, and, hence, it requires no blood donation and can be obtained with low cost.

Use of a newly developed artificial nerve conduit to assist peripheral nerve regeneration across a long gap in dogs.

There is now considerable evidence that peripheral nerves have the potential to regenerate if an appropriate microenvironment is provided. However, there are only a few reports of the successful use of artificial nerve conduits to repair major nerve defects more than 30 mm in length. In this study, we examined nerve regeneration across a long gap in the dog peroneal nerve using a novel artificial nerve conduit developed by our group. The conduit consists of a polyglycolic acid (PGA) collagen tube filled with laminin coated collagen fibers. In 12 dogs, the nerve conduit was implanted across an 80 mm gap in the left peroneal nerve. Three months after surgery, compound muscle action potentials (CMAPs) and somatosensory evoked potentials (SEPs) were detected. Evaluation of locomotor function revealed obvious limping for up to 3 months, but no marked difficulty in walking by 6 months. Microscopic observation of the regenerated nerve segment at 12 months showed numerous myelinated nerve fibers, which were smaller in diameter and enclosed in a thinner myelin sheath than normal axons. These results suggest that our artificial nerve conduit has potential usefulness in enhancing peripheral nerve regeneration, even across large gaps.

Experimental Study of Nerve Regeneration in a Biodegradable Tube Made From Collagen and Polyglycolic Acid

The authors developed a novel bioabsorbable nerve conduit to induce nerve regeneration across long gaps. It is a composite tube made of polyglycolic acid mesh coated with collagen. Seven cats underwent 25 mm resection of the left sciatic nerve. The proximal and distal nerve stumps were inserted into the tube and fixed with sutures to bridge the 25 mm gap between them. Histologic examination 4 months after implantation of the tube showed regeneration of nerve tissue structure, including myelinated axons and Schwann cells, and somatosensory evoked potentials and electromyograms demonstrated functional recovery of the regenerated nerves. Horseradish peroxidase staining revealed restoration of anterograde and retrograde axonal transport. These results indicate that this polyglycolic acid-collagen composite tube is promising as a nerve conduit that provides adequate nerve regeneration.

FACIAL NERVE REPAIR USING A COLLAGEN CONDUIT IN CATS

We evaluated facial nerve regeneration using a collagen tube as a nerve conduit in five cats. In three 5 mm of the facial nerve were resected, a collagen tube was implanted, and a 5 mm segment of the opposite facial nerve was resected, reversed 180 degrees, and sutured back as an autologous nerve graft. In one a collagen tube was implanted on one side, and in the remaining one a 5 mm nerve segment was reversed. Histological, electrophysiological, and horseradish peroxidase labelling examinations were carried out 4-24 weeks postoperatively. Histological study showed that the nerve was well vascularised and regenerated. Electrophysiological examination confirmed the recovery of evoked electromyograms through to the regenerated axons. Horseradish peroxidase examination also confirmed restoration of the whole facial nerve. The collagen tube is an efficient nerve conduit.