Patty Lee

Rabbit facial nerve regeneration in NGF‐containing silastic tubes

Previous reports suggest that exogenous nerve growth factor (NGF) enhanced nerve regeneration in rabbit facial nerves.1 Rabbit facial nerve regeneration in 10‐mm Silastic® tubes prefilled with NGF was compared to cytochrome C (Cyt. C), bridging an 8‐mm nerve gap. Three weeks following implantation, NGF‐treated regenerates exhibited a more mature fascicular organization and more extensive neovascularization than cytochrome‐C‐treated controls. Morphometric analysis at the midtube of 3‐ and 5‐week regenerates revealed no significant difference in the mean number of myelinated or unmyelinated axons between NGF‐ and cytochrome‐C‐treated implants. However, when the number of myelinated fibers in 5‐week regenerates were compared to their respective preoperative controls, NGF‐treated regenerates had recovered a significantly greater percentage of myelinated axons than cytochrome‐C‐treated implants (46% vs. 18%, respectively). In addition, NGF‐containing chambers reinnervated a higher percentage of myelinated axons in the distal transected neural stumps (49% vs. 34%).

Facial nerve regeneration through autologous nerve grafts: A clinical and experimental study

Human facial nerve regeneration was compared in two systems: autologous neural cable grafts (N = 56) and direct end‐to‐end anastomosis (N = 34). The data were analyzed based on the following criteria: facial symmetry and muscle tone at rest, degree of total facial reinnervation, voluntary motion, syn‐kinesis, and electrophysiologic testing. Facial tone and symmetry at rest, and electrophysiologic tests were similar, while voluntary motion and facial rein‐nervation were decreased and synkinesis increased in the autologous cable graft repairs. Electrophysiologic tests failed to distinguish between the two repair methods in successful regenerates. Neural cable grafts were associated with more mass movement and less fine precise emotive facial movement. Excellent cable graft repair results diminish with time because of continual progressive synkinesis, which may take up to 4 years to develop.

Laryngeal Reinnervation with the Hypoglossal Nerve I. Physiology, Histochemistry, Electromyography, and Retrograde Labeling in a Canine Model

This study was performed to determine whether the hypoglossal nerve (cranial nerve XII [XII]) would serve as a useful donor for laryngeal reinnervation by anastomosis to the recurrent laryngeal nerve (RLN). Twenty hemilarynges in 10 dogs were studied pro-spectively after XII-RLN anastomosis (group A; n = 5), split XII—RLN anastomosis (group B; n = 3), XII-RLN anastomosis with a 2-cm interposition graft (group C; n = 2), no treatment (group D; n = 5), RLN section (group E; n = 2), or ansa cervicalis—RLN anastomosis (group F; n = 3). Spontaneous activity was observed monthly by infraglottic examination through permanent tracheostomies and was recorded by electromyography. Laryngeal adductory pressure and induced phonation were obtained by stimulating the RLN while passing a pressure transducer balloon or humidified air through the glottis. At sacrifice, the laryngeal muscles were stained for adenosine triphosphatase to determine the ratio of type I to type II fibers. Retrograde labeling of the brain stem was performed with horseradish peroxidase. Infraglottic examination at 6 months showed a full range of adductory motion in groups A and B during the swallow reflex, comparable with that in group D. Groups C and F showed good bulk and tone, but little spontaneous motion. Group E remained paralyzed. Stimulation of the transferred nerves caused more activity in groups A and B than in the other groups; groups C and F partially adducted at high levels. The laryngeal adductory pressure responses of groups A and B were similar to those of group D. The XII-reinnervated larynges were capable of producing normal induced phonation. Retrograde labeling of the RLN showed that the reinnervating axons originated only in the hypoglossal nucleus. Electromyography of the reinnervated adductor muscles confirmed spontaneous activity in the dogs (awake). Histochemical analysis confirmed slow-to-fast transformation of both the posterior and lateral cricoarytenoid muscles, indicating that significant reinnervation occurred. We conclude that the hypoglossal nerve functions well as a donor for adductory reinnervation of the larynx.

Comparison of Rabbit Facial Nerve Regeneration in Nerve Growth Factor-Containing Silicone Tubes to that in Autologous Neural Grafts

Previous reports suggest that nerve growth factor (NGF) enhanced nerve regeneration in rabbit facial nerves. We compared rabbit facial nerve regeneration in 10-mm silicone tubes prefilled with NGF or cytochrome C (Cyt C), bridging an 8-mm nerve gap, to regeneration of 8-mm autologous nerve grafts. Three weeks following implantation, NGF-treated regenerates exhibited a more mature fascicular organization and more extensive neovascularization than Cyt C-treated controls. Morphometric analysis at the middle of the tube of 3- and 5-week regenerates revealed no significant difference in the mean number of myelinated or unmyelinated axons between NGF- and Cyt C-treated implants. However, when the numbers of myelinated fibers in 5-week regenerates were compared to those in their respective preoperative controls, NGF-treated regenerates had recovered a significantly greater percentage of myelinated axons than Cyt C-treated implants (46% versus 18%, respectively). The number of regenerating myelinated axons in the autologous nerve grafts at 5 weeks was significantly greater than the number of myelinated axons in the silicone tubes. However, in the nerve grafts the majority of the axons were found in the extrafascicular connective tissue (66%). The majority of these myelinated fibers did not find their way into the distal nerve stump. Thus, although the number of regenerating myelinated axons within the nerve grafts is greater than that of axons within silicone tube implants, functional recovery of autologous nerve graft repairs may not be superior to that of intubational repairs.

Axonal regeneration in severed peripheral facial nerve of the rabbit: relation of the number of axonal regenerates to behavioral and evoked muscle activity.

The minimum number of regenerating facial nerve myelinated motor axons that are required to innervate and activate the mimetic musculature is not known. We compare rabbit facial nerve regeneration following complete transectional injuries of the buccal division to the evoked and behavioral muscle activities of the quadratus labii superioris muscle of the rabbit in three experimental models: end-to-end direct anastomosis (N = 4), 8-mm autologous nerve grafts (N = 8), and 10-mm silicone chamber implants (N = 40). Data are presented as total numbers of regenerating myelinated axons that traverse the surgical repair and innervate the fascicles of the transected distal nerve stump, as well as the percentage of regenerating neurites, as compared to the preoperative normal controls. Five weeks after neural repair, direct end-to-end anastomosis regained more myelinated axons across the reconstructed defect (2,632 × 1,232; 67%) than silicone tube implants (2,006 × 445; 51%) or autologous cable graft repairs (1,660 × 1,169; 42%). However, only a small percentage of myelinated fibers innervated the intrafascicular region of the distal transected neural stump in direct anastomosis (948 × 168; 24%), silicone tube implants (670 × 275; 17%), or autologous nerve grafts (445 × 120; 12%) in rabbits that regained evoked and behavioral mimetic muscle activity. All rabbits with direct anastomosis and neural cable grafts regained motor activity, despite the fact that 66% of regenerating motor neurites in cable graft repairs and 54% in direct anastomosis were collateral sprouts that did not contribute to effective muscle activity. In 17 rabbits with neural regenerates within the silicone tube implants that did not regain mimetic activity, the mean number of regenerating myelinated motor axons across the defect was 504 × 419 (13%), and the mean number of axons that innervated the distal transected nerve stump fascicles was 277 × 128 (7%). Therefore, the minimal number of motor axons that is required to activate the quadratus labii superioris muscle is 12% of the original motor axon population of the normal buccal nerve division.

Rabbit Facial Nerve Regeneration in Autologous Nerve Grafts After Antecedent Injury

Objective The effect of incomplete antecedent injuries on subsequent facial nerve regeneration within cable graft repairs is not known. The purpose of this study is to compare facial nerve regeneration after an immediate and delayed neural cable graft repair.

Early stages of facial nerve regeneration through silicone chambers in the rabbit

The early stages of rabbit peripheral facial nerve regeneration [N = 16], over a 10‐mm transectional gap, were analyzed at 1, 3, and 5 weeks of buccal nerve entubation in silicone chambers prefilled with saline. Normal nerve pooled data were obtained in nine nerves. Chronologic morphologic and morphometic light and electron microscopic computer‐captured data reveal that the regeneration process can be subdivided into four stages: 1. The establishment of an acellular intergap matrix; 2. the ingrowth of mesodermally derived cells (macrophages, fibroblasts, endothelial cells, etc.) to form an intergap scaffolding; 3. the ingrowth of ectodermally derived cells (neurons, Schwanns cells, etc.) to reconstitute the transected peripheral nerve; and 4. neural maturation by progressive axonal enlargement, myelination, and compartmentalization. The first two stages occur during the first week, the third stage during the third week, and the fourth stage from the fifth week onwards of neural entubation.

Facial nerve regeneration through semipermeable chambers in the rabbit

Peripheral neural regeneration, over a 10‐mm transectional gap, was determined in 70 rabbit buccal divisions of the facial nerve using two entubational systems (semipermeable and impermeable silicone chambers) prefilled with three natural occurring media (serum, blood, and saline) during a 5‐week period. The number of myelinated axonal regenerates at the midchamber and at 2 mm in the distal transected neural stump were counted in each group and compared to pooled myelinated axonal counts in 9 normal rabbit buccal divisions of the facial nerve. Semipermeable porous chambers had an overall greater regeneration success rate (75% vs. 42.8%) and regained, on the average, a higher number of myelinated axons (51.4% vs. 26.1%) than silicone chamber regenerates. Semipermeable chambers prefilled with serum or blood had significantly higher regeneration success rates, myelinated axonal counts, and percentages of neural innervation of the distal transected neural stump. Both entubational systems produced similar axonal counts with intraluminal saline. The highest overall success rate (93.7%) and average number of myelinated axons per chamber (3072) were achieved in semipermeable chambers prefilled with serum. The greatest variability in myelinated axonal counts (0 to 3266 axons) and percentage of distal stump innervation (5.5% to 98.1%) was seen in silicone chambers filled with saline. The percentage of myelinated axons from the midchamber that innervated the distal stump was greater in semipermeable chambers with blood (73%) and serum (54%) than in silicone saline chambers (43%). On the average, the distal stumps from semipermeable chambers filled with serum (47%) and blood (33.5%) regained a higher percentage of normal myelinated axonal counts than silicone‐saline chambers (12.5%). These results suggest that both the construction of entubational chamber and the intraluminal medium can have significant influence on neurite regeneration. Semipermeable chambers prefilled with serum have a strong neurite‐promoting potential in peripheral neural regeneration of rabbit facial nerves.

Effects of Thrombin and Protease Nexin–1 on Peripheral Nerve Regeneration

Thrombin and serine protease inhibitors such as protease nexin–1 (PN-1) have been implicated in neurite outgrowth activity. We compared rat sciatic nerve regeneration in 10-mm silicone tubes, bridging an 8-mm nerve gap, that were prefilled with thrombin (1.5 IU) and PN-1 (50 μg/mL or 1 mg/mL) to those filled with saline solution (control). Neural regeneration and fibrinoid matrix progression (deposition of extracellular matrix) were analyzed at 1, 4, 17, and 21 days after silicone tube implantation. At 1 and 4 days after implantation, thrombin reduced fibrinoid matrix length propagation from both the proximal and distal transected nerve stumps, but PN-1 and saline did not interfere with matrix progression (p <.05). Seventeen days after implantation, the number of silicone tubes containing myelinated neuronal regenerates at the mid-tube region was 1 of 6 for thrombin, 6 of 9 for PN-1, and 6 of 10 for saline solution. Twenty-one days after implantation, 11 of 11 tubes with saline solution, 9 of 11 with PN-1 at 1 mg/mL, and 7 of 9 with PN-1 at 50 μg/mL had myelinated neural regenerates in the mid—silicone tube region, while only 2 of 9 thrombin-containing silicone tubes contained myelinated axons. There was no statistically significant difference in myelinated neurite regenerates at 17 and 21 days after implantation among silicone tubes prefilled with saline solution and PN-1 (50 μg/mL or 1 mg/mL). Thrombin interfered with matrix progression and significantly reduced the number of myelinated neurite regenerates (p =.01). The PN-1 and saline solutions did not inhibit matrix progression or affect the number of myelinated axonal regenerates (p =.92).

Laryngeal chemodenervation: effects of injection site, dose, and volume.

A canine model was used to measure changes in laryngeal adductory pressure (LAP) following injections of vecuronium bromide, a short-acting neuromuscular blocking agent. At a constant volume, LAP was inversely related to the dose (and concentration) of vecuronium injected. At a constant dose (0.05 mg), LAP did not vary significantly over a wide range of injection volumes, from 0.05 to 0.50 mL. At a constant dose and volume, the site of injection was varied among the anterior, middle, and posterior vocal fold, the interarytenoid region, and the anterior contralateral vocal fold. Reduction in LAP was greatest (p < .05) for the posterior vocal fold injection site (78% reduction); less reduction was seen for the middle (54%) and anterior (52%) vocal fold and interarytenoid (43%) injection sites. These results have implications for laryngeal botulinum toxin injections, which are discussed.