Gene delivery to rat and human Schwann cells and nerve segments: a comparison of AAV 1-9 and lentiviral vectors.

Schwann cells (SCs) in an injured peripheral nerve form pathways for regenerating axons. Although these cells initially support regeneration, SCs lose their pro-regenerative properties following a prolonged period of denervation. Gene transfer to SC can enhance their therapeutic potential. In this article, we compared adeno-associated viral (AAV) vectors based on serotypes 1-9 for their capability to transduce cultured primary rat and human SCs and nerve segments. AAV1 is the best serotype to transduce rat SCs, whereas AAV2 and AAV6 performed equally well in human SCs. Transduction of monolayers of cultured rat and human SCs did not accurately predict the transduction efficiency in nerve segments. Rat nerve segments could be genetically modified equally well by a set of four AAV vectors (AAV1, AAV5, AAV7, AAV9), whereas AAV2 was superior in human nerve segments. The current experiments were undertaken as a first step towards future clinical implementation of ex vivo AAV-based gene therapy in surgical nerve repair. The transduction of rat and human SCs and nerve segments by entirely different AAV serotypes, as documented here, highlights one of the challenges of translating gene therapy from experimental animals to human patients.

Developing a potentially immunologically inert tetracycline-regulatable viral vector for gene therapy in the peripheral nerve.

Viral vector-mediated gene transfer of neurotrophic factors is an emerging and promising strategy to promote the regeneration of injured peripheral nerves. Unfortunately, the chronic exposure to neurotrophic factors results in local trapping of regenerating axons or other unwanted side effects. Therefore, tight control of therapeutic gene expression is required. The tetracycline/doxycycline-inducible system is considered to be one of the most promising systems for regulating heterologous gene expression. However, an immune response directed against the transactivator protein rtTA hampers further translational studies. Immunogenic proteins fused with the Gly-Ala repeat of the Epstein-Barr virus Nuclear Antigen-1 protein have been shown to successfully evade the immune system. In this article, we used this strategy to demonstrate that a chimeric transactivator, created by fusing the Gly-Ala repeat with rtTA and embedded in a lentiviral vector (i) retained its transactivator function in vitro, in muscle explants, and in vivo following injection into the rat peripheral nerve, (ii) exhibited a reduced leaky expression, and (iii) had an immune-evasive advantage over rtTA as shown in a novel bioassay for human antigen presentation. The current findings are an important step toward creating a clinically applicable potentially immune-evasive tetracycline-regulatable viral vector system.

Reestablishment of the Smile after Hypoglossal-Facial Nerve Transfer: What Can We Learn?

Objective The aim of this study was to assess the ability to smile following a hypoglossal-facial nerve transfer (N12-N7). Design This is a retrospective chart review. Setting National tertiary referral center for skull base pathology. Participants Seventeen patients. Main Outcome Measures The ability to smile following an N12-N7 transfer was assessed by five medical doctors on photographs of the whole face and frontal, orbital, and oral segments. The (segmented) photographs were scored for the symmetry, asymmetry, and correct or incorrect assessment of the affected side. Results Seventeen patients were analyzed by 5 assessors providing 85 assessments. The whole face at rest was judged symmetrical in 26% of the cases and mildly asymmetrical in 56%. Frontal, orbital, and oral segments were symmetrical in 63, 20, and 35%, respectively. The affected side was correctly identified in 76%. When smiling, the whole face was symmetrical in 6% and mildly asymmetric in 59%. The affected side was correctly identified in 94%. The frontal, orbital, and oral segments during smiling were symmetrical in 67, 15, and 6%, respectively. The affected side of the frontal, orbital, and buccal facial segments during smiling was correctly identified in 89, 89, and 96%, respectively. Interobserver variability with Fleiss' kappa analysis showed that the strength of the agreement during smile of the total face was good (0.771) Conclusion Following an N12-N7 transfer, a good facial symmetry at rest can be achieved. During smiling, almost all patients showed asymmetry of the face, which was predominantly determined by the orbital and oral segments. To improve the ability to smile after an N12-N7 transfer, additional procedures are needed.

Correlating birthweight with neurological severity of obstetric brachial plexus lesions.

OBJECTIVE: To investigate the nature and extent of neurosurgically treated obstetric plexus lesions with obstetric and neonatal precedents. DESIGN: Retrospective analysis of prospectively collected data. SETTING: Leiden, the Netherlands. POPULATION: A 9-year cohort of infants (n = 206) neurosurgically treated for obstetric brachial plexus lesion at a tertiary referral centre for nerve lesions. METHOD: Obstetric and neonatal data (parity, diabetic status, pregnancy gestation, mode of cephalic delivery and birthweight) were collected using a standardised protocol and correlated to neurological severity of the brachial plexus lesion. MAIN OUTCOME MEASURE: Neurological severity of the brachial plexus lesion. RESULTS: Nulliparous women delivered significantly lower birthweight newborns (P = 0.016), injuries in those infants were associated with the least severe injury classification. The most prominent association in ordinal logistic regression was between neurological injury severity and larger birthweight (P < 0.001). CONCLUSIONS: Birthweight is correlated with neurological severity of the injury in a group of infants experiencing brachial plexus injury resulting from cephalic vaginal delivery.

[Surgical treatment of nerve lesions]. / Operatieve behandeling van zenuwletsels.

A peripheral nerve lesion may result in loss of motor and sensory function and pain. Nerve surgery may lead to function recovery or contribute to pain reduction. Different surgical techniques are used depending on the specific type of lesion. Common techniques consist of coapting, grafting, moving or burying the nerve. The outcome depends on the interval between the time the lesion occurred and the time of repair as well as the patient's age. Here we present three case studies of patients with a different type of nerve lesion and its consequences. We describe the specific surgical intervention performed to treat the nerve lesion. Immediate repair of the nerve should be performed when the patient has an open wound. In closed lesions, the decision to repair a damaged nerve depends on the severity of the trauma, the nerve(s) involved, the location and the age of the patient. In general, repair should be performed as soon as possible.

Nerve Surgery to Treat Intractable Genitofemoral Neuropathic Pain following Laparoscopic Live Kidney Donation.

To date live laparoscopic donor nephrectomies (LLDN) are frequently performed. The most common complications entail bleeding, wound infection, and incisional hernia. Here we discuss a 50-year-old patient with a severe less known complication, namely, postoperative persistent neuropathic pain in the scrotum and left upper leg. Satisfactory pain control could not be obtained in 3 years of postoperative pain treatment which consisted of neuroleptic drugs, blocks of the L1/L2 dorsal roots with local anaesthetics, and pulsed radiofrequency lesioning. Exploratory laparoscopy was performed to assess the aspect of the genitofemoral nerve (GFN). A hemoclip used for the closure of the ureter at the time of nephrectomy was found in close relation to the GFN. The clip was removed and the GFN was subsequently cut proximal to the side of this clip. Soon after surgery the patient was completely pain-free and could return to his normal activities. Surgery should be considered in case of GFN neuropathic pain following LLDN.

Pores in synthetic nerve conduits are beneficial to regeneration.

Current opinion holds that pores in synthetic nerve guides facilitate nerve regeneration. Solid factual support for this opinion, however, is absent; most of the relevant studies assessed only morphological parameters and results have been contradictory. To evaluate the effect of pores, the rat sciatic nerve was either autografted or grafted with nonporous, macroporous (10-230 mum), and microporous (1-10 microm) biodegradable epsilon-caprolactone grafts. Twelve weeks later, the grafted nerves were resected, and the electrophysiological properties were determined in vitro. Subsequently midgraft-level sections were inspected, and peroneal nerve sections were evaluated morphometrically. Finally, the gastrocnemic and tibial muscle morphometrical properties were quantified. The microporous nerve graft performed much better than the nonporous and macroporous grafts with respect to most parameters: it was bridged by a free floating bundle that contained myelinated nerve fibers, there were more nerve fibers present distal to the graft, the electrophysiological response rate was higher, and the decrease in muscle cross-sectional area was markedly smaller. Hence, the present study demonstrates the beneficial effect of synthetic nerve guide pores on nerve regeneration, although with the caveat that not pores per se, but only small (1-10 microm) pores were effective.

Perfusion magnetic resonance imaging provides additional information as compared to anatomical imaging for decision-making in vestibular schwannoma.

OBJECTIVE: The added value of perfusion MRI for decision-making in vestibular schwannoma (VS) patients is unknown. MRI offers two perfusion methods: the first employing contrast agent (dynamic susceptibility contrast (DSC)-MRI) that provides information on cerebral blood volume (CBV) and cerebral blood flow (CBF), the second by magnetic labeling of blood (arterial spin labeling (ASL)-MRI), providing CBF-images. The goal of the current study is to investigate whether DSC and ASL perfusion MRI provides complimentary information to current anatomical imaging in treatment selection process of VS. METHODS: Nine patients with growing VS with extrameatal diameter >9 mm were included (>2 mm/year and 20% volume expansion/year) and one patient with 23 mm extrameatal VS without growth. DSC and ASL perfusion MRI were obtained on 3 T MRI. Perfusion in VS was scored as hyperintense, hypointense or isointense compared to the contralateral region. RESULTS: Seven patients showed hyperintense signal on DSC and ASL sequences. Three patients showed iso- or hypointense signal on at least one perfusion map (1 patient hypointense on both DSC-MRI and ASL; 1 patient isointense on DSC-CBF; 1 patient isointense on ASL). All patients showed enhancement on post-contrast T1 anatomical scan. CONCLUSION: Perfusion MR provides additional information compared to anatomical imaging for decision-making in VS.

Reduced functional connectivity within the primary motor cortex of patients with brachial plexus injury.

This study aims at the effects of traumatic brachial plexus lesion with root avulsions (BPA) upon the organization of the primary motor cortex (M1). Nine right-handed patients with a right BPA in whom an intercostal to musculocutaneous (ICN-MC) nerve transfer was performed had post-operative resting state fMRI scanning. The analysis of empirical functional correlations between neighboring voxels revealed faster correlation decay as a function of distance in the M1 region corresponding to the arm in BPA patients as compared to the control group. No differences between the two groups were found in the face area. We also investigated whether such larger decay in patients could be attributed to a gray matter diminution in M1. Structural imaging analysis showed no difference in gray matter density between groups. Our findings suggest that the faster decay in neighboring functional correlations without significant gray matter diminution in BPA patients could be related to a reduced activity in intrinsic horizontal connections in M1 responsible for upper limb motor synergies.

Neuroexcitatory plasma amino acids are elevated in migraine.

To investigate the role of glutamic (Glu) and aspartic acid (Asp) in migraine, we measured the plasma amino acids in migraine patients with and without aura, between and during attacks, and compared the profiles with the plasma amino acid profiles of tension headache patients and healthy controls. Between attacks, migraineurs (notably with aura) had substantially higher plasma Glu and Asp levels than did controls and tension headache patients. In addition, patients with migraine without aura showed low plasma histidine levels. During migraine attacks, Glu (and to a lesser extent Asp) levels were even further increased. The results suggest a defective cellular reuptake mechanism for Glu and Asp in migraineurs, and we hypothesize a similar defect at the neuronal/glial cell level, predisposing the brain of migraineurs to develop spreading depression.

Cortical excitability of the biceps muscle after intercostal-to-musculocutaneous nerve transfer.

OBJECTIVE: Restoration of volitional control over elbow flexion has been demonstrated in patients who have undergone intercostal-to-musculocutaneous nerve transfer. We investigated the cortical area involved in the control over elbow flexion after intercostal-to-musculocutaneous nerve transfer. METHODS: Maps of magnetically excitable cortical areas of the affected arms of five patients were compared with maps of their healthy arms and maps of both arms of four healthy control subjects. The intercostal cortical area was also studied, requiring needle electromyography mapping (n = 1). RESULTS: The cortical areas of affected arms were smaller and less excitable than those of healthy arms. The locations of these areas could not be distinguished from that of the normal cortical biceps area but seemed to differ from that of the intercostal cortical area. CONCLUSION: The existence of a biceps-like cortical area related to the reinnervated muscle can be explained in two ways. Interneurons from the original biceps area might excite the cortical neurons controlling the intercostal muscles. Alternatively, corticospinal neurons of the original biceps area may project directly onto spinal intercostal motor neurons. Cerebral plasticity does occur in intercostal-to-musculocutaneous nerve transfers and may be crucial for their clinical success.

Evaluation of intercostal to musculocutaneous nerve transfer in reconstructive brachial plexus surgery.

OBJECT: Direct coaptation of intercostal nerves (ICNs) to the musculocutaneous (MC) nerve was performed to restore elbow flexion in 25 patients with brachial plexus root avulsions. METHODS: Seventy-five ICNs were transected as close as possible to the sternum to obtain sufficient length and then tunneled to the axilla and coapted to the MC nerve. Direct coaptation was achieved in 95% of ICNs, and functional elbow flexion was regained in 64% of the patients. The results were compared with several reported transfer techniques in which either an ICN or other donor nerves were used. CONCLUSIONS: Direct coaptation was equally effective and more straightforward than transfers involving interposition of grafts. The use of alternative donors such as the accessory nerve carries inherent disadvantages compared with the use of ICNs, and the results are not substantially better. Direct ICN-MC nerve transfer is a valuable reconstructive procedure.

Changing central nervous system control following intercostal nerve transfer.

OBJECT: The goal of this study was to find which central nervous system (CNS) pathways are involved in volitional control over reinnervated biceps or pectoral muscles. METHODS: Intercostal nerves (ICNs) were coapted to the musculocutaneous nerve (MCN) or the medial pectoral nerve (MPN) in 23 patients with root avulsions of the brachial plexus to restore biceps or pectoral muscle function. The facilitatory effects of respiration and voluntary contraction on cortical motor-evoked potentials of biceps or pectoral muscles were used to study CNS control over the reinnervated muscles. The time course of the facilitatory effect of respiration and voluntary contraction differed significantly. In the end stage of nerve regeneration, the facilitatory effect of voluntary contraction was significantly larger than that of respiration, indicating that the CNS control network over the muscle comes to resemble that of the recipient nerve (MCN or MPN) rather than that of the donor nerve (ICN). CONCLUSIONS: The strengthening of previously subthreshold synaptic connections in a CNS network connecting ICN to MCN or MPN neurons may underlie changing excitability.

Initial report on the limited value of hypoglossal nerve transfer to treat brachial plexus root avulsions.

OBJECT: Hypoglossal nerve (12th cranial nerve) transfer was performed to treat the sequelae of brachial plexus root avulsion in 12 adults and two infants, and the patients were followed to assess the effectiveness of the surgery. METHODS: The 12th cranial nerve was transected at the base of the tongue, and a sural nerve graft was used to bridge the gap between the donor (12th) and recipient nerves: C-5 spinal, axillary, suprascapular, or musculocutaneous nerve. The mean graft length in adult patients was 15.75 +/- 5.5 cm (+/- standard deviation, median 14.5 cm) and in the two infants the graft lengths were 7 and 8 cm, respectively. After a mean postoperative interval of 1138 +/- 254 days, electromyographic examination of the target muscles showed tongue movement-related activity in all patients. Muscle force strength measured according to the Medical Research Council's guidelines, was Grade 3 or higher in 21% of patients. Contraction, however, could only be attained by tongue movements, and volitional control was not achieved. CONCLUSIONS: Although recovery of muscle strength was obtained by 12th cranial nerve transfer, the functional gain remained virtually nonexistent because central control was missing.

Correlation between histopathological findings in C-5 and C-6 nerve stumps and motor recovery following nerve grafting for repair of brachial plexus injury.

OBJECT: Proximal spinal nerve stumps were used as donor sites for grafts to repair brachial plexus traction lesions. The quality of the stumps was assessed histologically, and its correlation with the strength attained in the target muscle was studied. METHODS: Four histopathological parameters in frozen tissue sections of 31 C-5 or C-6 nerve stumps were examined by a neuropathologist. The total quantity of myelin was compared with normal values. Also, thick myelinated fibers, fibrosis, and misdirected axons were assessed. Stumps embedded in plastic were used in a morphometric study of myelinated fiber profiles. The fiber density, mean size, and size distribution in five donor stumps were determined; three normal C-5 spinal nerves obtained at autopsy served as controls. Finally, the relative area occupied by fiber profiles and interspace was computed. Linear regression was used as a multivariate analysis, adjusting the outcome of surgical repair for effects of age, interval between trauma and surgery, and graft length. Histopathological examination showed that the total quantity of myelin in donor stumps used for biceps muscle reinnervation was considerably reduced. On morphometric examination the fiber density did not differ significantly between stumps obtained in patients and control stumps obtained at autopsy. However, a significant reduction of the area occupied by myelinated fibers was measured: from 46% in controls to 13% in patients (p < 0.0001). Likewise, a significant reduction was found in the mean fiber size: from 7.4 microm in controls to 3.7 microm in patients (p < 0.0001). The relationship between the myelin quantity in the proximal stump and the grade of biceps muscle recovery was statistically significant (p = 0.02). From the 95% confidence interval it was concluded that the estimated effect of a mean increase of myelinated fibers by 25% almost equals an increase in recovery of one point on the Medical Research Council scale (grade range 0-5). CONCLUSIONS: Both histopathological and morphometric examination showed a reduction of the quantity of myelin in C-5 or C-6 stumps used as donor sites for grafts. The amount of myelin is significantly correlated with biceps muscle function after nerve grafting. Because it is possible to assess the quantity of myelin by intraoperative examination of frozen sections, this correlation is potentially useful in the decision-making process of whether to use stumps for grafting or to use nerve transfer procedures to restore biceps muscle function.

Surgical repair of brachial plexus injury.

This preliminary report surveys the authors' experience in the treatment of 66 patients with brachial plexus traction injuries. The results of nerve reconstructive surgery obtained in 44 patients with a follow-up of at least 2 years were disappointing; on average, 10% of upper limb function was added. The role of several factors involved in this type of injury and the major draw-backs of treatment are discussed in combination with future possibilities. Current techniques of neurotizations are presented in one case.

The dorsoscapular nerve in traumatic brachial plexus lesions.

The origin and course of the nerval innervation of the levator scapulae and rhomboid muscles was studied in four human cadavers. Special attention was given to surgical anatomy. The levator scapulae muscle receives two small segmental nerves from C3 and C4, respectively. The rhomboid muscle receives two segmental nerves, one from C4 and the other from C5. Moreover, in a series of 137 patients with a traumatic brachial plexus lesion, the linkage between levator scapulae and rhomboid muscle function impairment and site of the brachial plexus lesion was studied. Paresis of one or both of these muscles was noted in 13 cases. It was concluded that the rhomboid muscle may function on a single C4 nerve supply without any loss of strength. Arguments are put forward to support the relevance of rhomboid muscle testing in the assessment of brachial plexus lesions.

Respiration-related activity in the biceps brachii muscle after intercostal-musculocutaneous nerve transfer.

Needle EMG and polygraphic recordings of respiration and biceps activity were obtained in 7 patients who had undergone intercostal to musculocutaneous nerve transfer because of severe traumatic injury to the brachial plexus. EMG activity during expiration and inspiration was assessed during quiet breathing, deep breathing, and during sustained inspiration and expiration, and compared to the clinical strength of the biceps muscle. Biceps activity was consistently found during both expiration and inspiration, and in either case separately related to the flexion force levels. Initially this relationship appeared equally strong for both phases. With time, control over flexion changes from a completely respiratory-driven control to a conscious volitional control; however, involuntary effects of respiration always remained present in polygraphic recordings. Theoretical explanations for the change in control are put forward.

[Brachial plexus injury due to unhealed or wrongly healed clavicular fracture]. / Plexus-brachialisletsel door niet genezen of in afwijkende stand genezen claviculafracturen.

Two men, aged 65 and 27 years, had persistent dysesthesia of digits IV and V 18 and 8 months, respectively, after conservative treatment of dislocated comminuted clavicular fractures. In both patients, the fracture was found to have healed poorly. There was pseudoarthrosis, shortening and malrotation. Following surgical decompression of the neurogenic structures by elongation and internal fixation of the clavicle, motor and sensory functions recovered completely. The incidence of clavicular fractures is high, but complications are rare. Symptom recognition and timely treatment can prevent irreversible nerve damage.

Neonatal brachial plexus palsy with neurotmesis of C5 and avulsion of C6: supraclavicular reconstruction strategies and outcome.

BACKGROUND: Nerve reconstruction strategies for restoration of elbow flexion and shoulder function in patients with neonatal brachial plexus palsy with neurotmesis of C5 and avulsion of C6 are not well defined and the outcomes are unclear. METHODS: From 1990 to 2008, nerve surgery was performed in 421 patients with neonatal brachial plexus palsy. This study focused on thirty-four infants who had a neurotmetic lesion of C5 and avulsion or intraforaminal neurotmesis of C6, irrespective of C7. The C8 and T1 functions were intact. Intraplexal transfer of C6 to C5 with direct coaptation was preferred for restoration of elbow flexion. The suprascapular nerve was reconnected either by extra-intraplexal transfer of the accessory nerve or by grafting from C5 to restore shoulder function. Additional grafts were attached from C5 to the C5 contribution of the posterior division of the superior trunk when technically possible. RESULTS: Transfer of either the C6 anterior root filaments or the entire C6 nerve to C5 was performed in seventeen patients (group A) with direct coaptation in fifteen of them. Grafting from C5 to the anterior division of the superior trunk was performed in the remaining seventeen infants (group B). An accessory-to-suprascapular nerve transfer was applied in twenty-nine infants. The suprascapular nerve was reconnected in five patients by grafting from C5. It was possible to attach one, two, or three additional grafts from C5 to the posterior division of the superior trunk in twenty-one patients. All infants had biceps muscle recovery to a Medical Research Council (MRC) grade of ≥4, twenty-two (65%) of the thirty-four patients obtained Mallet grade-IV abduction, and eleven (32%) of the thirty-four obtained Mallet grade-IV external rotation. CONCLUSIONS: In patients with neonatal brachial plexus palsy who have neurotmesis of C5 and avulsion of C6, elbow flexion can be successfully restored with supraclavicular intraplexal reconstruction with use of C5 as the proximal outlet. However, shoulder function recovery following suprascapular nerve reinnervation and additional grafting from C5 to the posterior division of the superior trunk is less successful. LEVEL OF EVIDENCE: Therapeutic Level III. See Instructions for Authors for a complete description of levels of evidence.