Intradural avulsion of the individual nerve roots

Transcription

1 J Neurosurg 119: , 2013 AANS, 2013 Contralateral L-6 nerve root transfer to repair lumbosacral plexus root avulsion: experimental study in rhesus monkeys Laboratory investigation Haodong Lin, M.D., Ph.D., Aimin Chen, M.D., Ph.D., and Chunlin Hou, M.D. Department of Orthopedic Surgery, Changzheng Hospital, The Second Military Medical University, Shanghai, People s Republic of China Object. Nerve transfer is used for brachial plexus injuries but has rarely been applied to repairs in the lower extremities. The aim of this study was to evaluate the feasibility and effectiveness of using the contralateral L-6 nerve root to repair lumbosacral plexus root avulsions. Methods. Eighteen rhesus monkeys were randomized into 3 groups. In the experimental group, the left L4 7 and S-1 nerve roots were avulsed and the right L-6 nerve root was transferred to the left inferior gluteal nerve and the sciatic nerve branch innervating the hamstrings. In the control group, the left L4 7 and S-1 nerve roots were avulsed and nerve transfer was not performed. In the sham operation group, the animals underwent a procedure that did not involve nerve avulsion and nerve transfer. Functional outcomes were measured by electrophysiological study, muscle mass investigation, and histological study. Results. The mean amplitudes of the compound muscle action potentials from the gluteus maximus and biceps femoris in the experimental group were higher than those in the control group but lower than those in the sham group (p < 0.05). The muscle mass and myofiber cross-sectional area of these muscles were heavier and larger than those in the control group (p < 0.05). The number of myelinated nerve fibers of the inferior gluteal nerve and the branch of the sciatic nerve innervating the hamstrings in the control group was significantly smaller than the number in the experimental and sham groups (p < 0.01). Conclusions. In this animal model, the contralateral L-6 (analogous to S-1 in humans) nerve root can be used to repair lumbosacral plexus root avulsion. ( Key Words lumbosacral plexus nerve roots nerve transfer rhesus monkey peripheral nerve Intradural avulsion of the individual nerve roots that form the brachial plexus was initially described in 1911 by Frazier and colleagues. 7 Brachial plexus avulsion and the resulting myelographic picture is now a well-known condition. However, it was not until 1960 that Finney and Wulfman first reported on intradural lumbosacral nerve root avulsion. 6 Recent reports suggest that this condition is frequently overlooked. Avulsion of the intradural lumbosacral nerve root occurs more frequently than previously suspected. 1 Nerve transfer is a valuable surgical technique in peripheral nerve reconstruction, especially in brachial plexus injuries. 2,3 In reality, the technique has been used as a salvage procedure when no proximal nerve source is available. Although many types of nerve transfers have been reported for the upper extremity, 8,13,14 little attention has been given to this option for repairs in the lower extremity. 4,10 We hypothesized that the surgical Abbreviation used in this paper: CMAP = compound muscle action potential. strategies currently used for brachial plexus repair could be adopted for the treatment of intradural lumbosacral nerve root avulsion. In our previous experiments, we have confirmed that division of the L-6 nerve root in the monkey sacral plexus, which is analogous to the S-1 root in humans, did not permanently affect limb function. 12 This paper describes a preliminary study whose aim was to investigate whether the L-6 nerve root could be used as a donor nerve to repair lumbosacral plexus root avulsion in a monkey model. Methods Animal Preparation and Surgical Procedures A total of 18 healthy rhesus monkeys of either sex, each weighing 3500 to 4500 g, were randomly divided This article contains some figures that are displayed in color on line but in black-and-white in the print edition. 714

2 Contralateral L-6 nerve root transfer into 3 groups. All procedures were approved by The Second Military Medical University Animal Care and Use Committee, and the animals were treated in accordance with the Animal Care Guidelines of the National Bureau of Health. Prior to surgery, all animals were observed for normal lower-extremity function and the absence of any previous injury. In preparation for surgery, the animals were anesthetized with an intravenous injection of ketamine (50 mg/kg body weight) combined with valium (1.5 mg/kg body weight). A longitudinal incision was made that was centered at L-3, L-4, L-5, L-6, and L-7. The skin, subcutaneous tissue, and deep fascia were incised to expose the spinal and the articular processes. An L3 7 hemilaminectomy was performed, and the dura mater was then opened and anatomical landmarks used to identify the L3 7 ventral roots and dorsal roots on the left side. In the experimental group (n = 8), these 5 roots were individually avulsed and separated from the spinal cord surface by applying a constant traction with a pair of fine jeweler s forceps along the course of each root. The right L-6 nerve root was severed extradurally, and palliative nerve transfers were performed using the contralateral L-6 nerve root. The inferior gluteal nerve and a branch to the hamstring muscles were chosen as the common target for neurotization. The sciatic and the gluteal nerves were reached by detaching the gluteus maximus laterally. Two nerve grafts from the sural nerve of approximately 10 cm in length were used as the nerve grafts. One end of the nerve grafts was connected to the proximal stump of the right L-6 nerve root using a 10-0 absorbable suture. The distal end of one nerve graft was sutured to the inferior gluteal nerve and the other was sutured to the branch of the sciatic nerve innervating the hamstrings (Fig. 1). The wound was closed in layers. In the control group (n = 5), the animals underwent lumbar hemilaminectomy and root injury but nerve transfer was not performed. In the sham operation group (n = 5), the animals underwent lumbar hemilaminectomy and opening of the dura, but without root injury and nerve transfer. In all cases during the surgery, a total dose of U of penicillin was administered by intravenous drip, and U/day of penicillin was administered via an intramuscular injection for 3 days postoperatively. The monkeys remained in a standard environment at the animal department for the postsurgical period. Six months postsurgery, the surviving rhesus monkeys were examined. Evaluation of Nerve Regeneration Electrophysiological Study. Six months after surgery, the animals were prepared after induction of general anesthesia. The previous operative site was explored, and the nerve repair sites were inspected for integrity. A 4-channel electrophysiology apparatus (Esaote Phasis, Esaote Biomedica) was applied to record the CMAP of the gluteus maximus muscle and the biceps femoris. The electrophysiological setup involved 2 reference electrodes: a recording electrode in the gluteus maximus muscle or biceps femoris and a ground electrode positioned subcutaneously at an adjacent region. The stimulating electrode was placed on Fig. 1. The surgical procedure in the experimental group. The contralateral L-6 nerve root was transferred to the inferior gluteal nerve and the branch of the sciatic nerve innervating the hamstrings. BSNIH = branch of the sciatic nerve innervating the hamstrings; IGN = inferior gluteal nerve. the inferior gluteal nerve or the branch of the sciatic nerve innervating the hamstrings just distal to the nerve repairs. Nerve stimulation (a 0.2-msec, 1-mA rectangular pulse wave) and recordings were performed using a PowerLab 4SP digital data acquisition system (Keypoint 3.02). Digitized data were stored in a personal computer, and peak amplitudes of the CMAP were calculated. Muscle Tetanus Contraction Force Measurement. After the electrodiagnostic study, the biceps femoris contraction force was assessed using a force-displacement transducer (FT03 Force Displacement Transducers, Grass Instruments) and computerized recording software. Initially, the resting muscle length was determined. The distal tendon of the biceps femoris was then divided at its distal insertion and attached to the force transducer. The length of the muscle was approximately equal to its resting length in situ for tension adjustment. To avoid motion artifact during measurements, the hip and knee joints were immobilized on the operating platform. Stimulating current was applied using a bipolar platinum electrode distal to the repaired side. The threshold stimulus was defined as the stimulus that was required to produce an observable muscle twitch. Stimulation of the branch of the sciatic nerve innervating the hamstrings was performed for activation of the biceps femoris muscle at different thresholds (1 10 times the threshold), at different volt- 715

3 H. Lin, A. Chen, and C. Hou ages (range V), and different frequencies (range Hz). Under different stimuli, the maximal tetanic strength was determined with 1 V and 60 Hz, and recorded as grams/weight. The mean maximal isometric muscle contraction of the repeated muscle contraction forces (5 times with a pulse duration of 1.0 msec) was recorded. The measurements were analyzed and recorded using the MacLab System (ADInstruments, Inc.). Neuromorphometry. Nerve specimens were obtained from the inferior gluteal nerve and the branch of the sciatic nerve innervating the hamstrings (3 to 5 mm in length at the point 2 mm distal to the nerve repaired site) and were fixed in 2.5% glutaraldehyde and postfixed in 2% osmium tetroxide. Each nerve was embedded in 100% Epon. Transverse sections of 1-mm thickness were made from the nerve to obtain sections at successive 1-mm intervals. In both the control group and the sham operation group, similar specimens were also taken of comparable length and at a similar location. The axon count was used to evaluate axonal regeneration. The first 15 sections were selected. Every third section was stained. A total of 5 sections were analyzed to determine the total number of myelinated axons. Five fields were randomly chosen under 400 magnification. All the myelinated axons in the chosen fields were counted. To determine the total number of axons, each histological section was photographed using a digital camera (Panasonic WV-CP410) coupled with an optical microscope (model DWLB2, Leica Microsystem), and Image Analysis Software (Leica FW4000) was used for the neuromorphometric analysis. The average of the number of axons counted in these fields was calculated. The investigator who assessed all the outcomes was blinded to the experimental group assignment. Muscle Mass and the Cross-Sectional Area of Muscle Fiber. After harvesting the nerve, we removed the gluteus maximus muscle and the biceps femoris from the surgical side of the animal. The surface of the muscles was gently blotted with absorbent paper to remove any blood or serum. The muscle was then promptly weighed using an Analytical Balance (with a verification scale interval of 1 mg; R200D, Sartorius). Muscle samples were cut from the midbelly of the harvested muscle and fixed in buffered 4% paraformaldehyde. The muscle samples were subsequently washed in water, dehydrated in a graded ethanol series, cleared in xylene, embedded in paraffin, and cut into 5-mm-thick transverse sections. Four sections were acquired from every specimen. They were subjected to H & E staining before photographs were taken with a DFC 300FX color digital camera (Leica). For each section, photographs were taken from 5 random fields and analyzed with a Leica QWin software package (Q550 IW Image Analysis System, Leica Imaging Systems Ltd.) to measure the cross-sectional area of muscle fibers. Statistical Analysis Results were expressed as the mean ± SD, and SPSS 11.0 software (SPSS Inc.) was used for data analysis. Oneway analysis of variance was used to determine significant differences between the groups. Differences were considered significant at p < Results General Observations All of the animals in the control group and the sham operation group survived for the duration of the experiment without wound infection. Two monkeys died of postanesthesia pneumonia, and therefore the 6 remaining live monkeys constituted the experimental group. No obvious atrophy of the gluteus maximus muscle and the biceps femoris was seen in the sham operation group. However, obvious atrophy of the gluteus maximus muscle and the biceps femoris was seen in all animals in both the control group and the experimental group 6 months after surgery. In the experimental group, all monkeys were able to abduct the hip joint and flex the knee joint. This means that, in the affected limb, the strength of their gluteus maximus and biceps femoris had been significantly improved to Grade M3/5 or better (according to the Medical Research Council grading scale). By contrast, the values for the control group were shown to be Grade M0/5 without an exception. Electrophysiology Compound muscle action potentials were recorded from the gluteus maximus muscle when the inferior gluteal nerve was stimulated and from the biceps femoris when the branch of the sciatic nerve innervating the hamstrings was stimulated. In the experimental group, the mean CMAP measured from the gluteus maximus muscle and the biceps femoris was 6.62 ± 1.54 mv and 7.06 ± 1.82 mv, respectively. In the sham operation group, the mean CMAP measured from the gluteus maximus muscle and the biceps femoris was 8.56 ± 2.04 mv and 8.86 ± 1.95 mv, respectively. The sham operation group demonstrated a significantly higher amplitude than did the experimental group (p < 0.01). In the control group, a CMAP was not detected when the same stimulus was applied to the inferior gluteal nerve and the branch of the sciatic nerve innervating the hamstrings (Fig. 2). Muscle Tetanus Contraction Force Measurement In the experimental group, the mean contractile force of the biceps femoris was ± g. In the sham operation group, the mean contractile force was ± g. The difference between both groups was statistically significant (p < 0.01), showing a greater contractile force in the sham operation group. In the control group, the mean contractile force was 0 g, which was smaller than that of both the experimental and the sham operation groups. Neuromorphometry In the inferior gluteal nerve and the branch of the sciatic nerve innervating the hamstrings segment distal to the repair side, well-myelinated nerve fibers were seen in both the experimental group and the sham operation group. The mean number of regenerative myelinated nerve fibers in the segment distal to the repair site of the inferior gluteal nerve in the experimental group was ± 74.6 and ± 86.4, respectively. The absolute mean regenerative numbers in the sham operation group were ± 70.4 and ± 92.5, respectively, which 716

4 Contralateral L-6 nerve root transfer ris weight in the experimental group was ± g and ± g, respectively. The mean gluteus maximus muscle and biceps femoris weight in the sham operation group was ± g and ± g, respectively, which was significantly heavier than that in the experimental group (p < 0.05). In the control group, the mean gluteus maximus and biceps femoris mass was ± g and ± g, respectively, which was significantly lighter than those in the experimental and sham operation groups (p < 0.01). Cross-Sectional Area of Muscle Fiber The cross-sectional area of the muscle fibers from the gluteus maximus muscle and the biceps femoris in the experimental group was ± μm2 and ± μm2, respectively, which was significantly smaller than that in the sham operation group ( ± μm2 and ± μm2, respectively) (p < 0.05). The cross-sectional area of the muscle fibers from the gluteus maximus muscle and the biceps femoris in the control group was ± μm2 and ± μm2, respectively, which was significantly smaller than that in the sham operation group and the experimental group (p < 0.01) (Fig. 4). Discussion Fig. 2. Compound muscle action potentials of the biceps femoris were recorded 6 months after surgery. The CMAP amplitude of the biceps femoris muscles in the experimental group was lower than that in the sham operation group but was higher than that in the control group: experimental group (A); control group (B); and sham operation group (C). were significantly greater than those of the experimental group (p < 0.05). In the control group, the mean number of regenerative nerve fibers was 0 (Fig. 3). Muscle Mass The mean gluteus maximus muscle and biceps femo- Nerve transfer has proved to be an important addition to the techniques available for the repair of brachial plexus lesions but has rarely been applied to the lower extremities.4,10 Avulsion of the intradural lumbar nerve root occurs more frequently than was previously suspected.1 Huittinen9 recently reported the results of 42 medicolegal autopsies performed following industrial, domestic, or traffic accidents, in which fracture of the pelvis occurred. He had previously observed that 46% of patients with double vertical pelvic fractures had signs of persistent neural damage, predominantly in the lumbosacral segments, on follow-up examination. In the autopsy series, all 42 cadavers underwent complete dissection of the sacrum, posterior wall of the pelvis, piriformis muscle, sacral plexus, lumbosacral trunk, and anterior rami of the lumbar and sacral spine nerves. In 15 cases, the intradural and intraforaminal course of the spinal nerves concerned were exposed by laminectomy and dural opening. Rupture of the roots of the cauda equina was recorded in 6 autopsy cases. Fig. 3. Cross-section of the inferior gluteal nerve stumps harvested 6 months after surgery from animals in the experimental group (A), control group (B), and sham operation group (C). The number of myelinated nerve fibers of the inferior gluteal nerve in the experimental group (A) was significantly more than that of the control group (B) but less than that of the sham operation group (C). Toluidine blue, original magnification

5 H. Lin, A. Chen, and C. Hou Fig. 4. Cross-section of the biceps femoris harvested 6 months after the surgery from animals in the experimental group (A), control group (B), and sham operation group (C). The myofiber cross-sectional area of the biceps femoris in the experimental group (A) was smaller than that in the sham operation group (C) but larger than that in the control group (B). H & E, original magnification 400. In cases in which the proximal stump of the nerve or the root is difficult or impossible to retrieve, palliative procedures can be performed. One study showed that, with the help of nerve grafts, a connection between the lower intercostal nerves and the lumbar plexus provided some functional return to the lower extremities.11 However, the number of axons that the intercostal nerves contained was much lower than that of the lumbosacral nerve roots. Therefore, the intercostal nerves cannot provide the adequate donor axons that are necessary for the effective reconstruction of lower-limb function. 5,10 Thus, the search for more effective donor nerves continues. In our previous experiments, we have confirmed that the division of a single L-6 nerve root of the sacral plexus in rhesus monkeys (homologous to S-1 in humans) does not have a harmful effect on lower-limb function and could potentially act as a donor nerve for the treatment of lumbosacral plexus root avulsion.12 The current study sought to determine whether the L-6 nerve root could be used as a donor nerve to repair lumbosacral plexus root avulsion in a monkey model. The most basic requirements for standing and walking are the stability and flexion of the hips and knees. These key functions depend on sufficient power in proximal muscles, including the quadriceps muscles, hamstrings, and the gluteal muscles. Unlike the upper extremity, in which hand function is crucial, it is still possible to stand and walk with impaired intrinsic muscle function or even with loss of sensitivity in the foot. In the latter case, however, if function is regenerated in a few proximal key muscles of the leg, the patient may show dramatic improvement, resulting in, for example, being able to stand and walk independently after having been previously wheelchair bound. Thus, recovering these key functions is critical. In the present study, we transferred the contralateral L-6 to repair the inferior gluteal nerve and the branch of the sciatic nerve innervating the hamstrings, in an attempt to recover the function of the gluteus maximus muscle and the biceps femoris muscle. The results showed that, although the mean amplitude of the CMAPs from the gluteus maximus and the biceps femoris muscles in the experimental group was lower than that of the sham operation group, it was higher than that of the control group. The muscle mass and myofiber cross-sectional area of the muscles in the experimental group were heavier and larger than those in the control group. The number of myelinated nerve fibers 718 of the inferior gluteal nerve and the branch of the sciatic nerve innervating the hamstrings in the experimental group was significantly greater than that in the control group. The results demonstrated that the gluteus maximus muscle and the biceps femoris regained significant innervation. The present study suggests that L-6 (analogous to S-1 in humans) from the healthy, uninjured side may be considered as a potential donor nerve for transfer. Conclusions Using the contralateral L-6 nerve root as a donor nerve to repair lumbosacral plexus root avulsion was feasible and effective in this monkey model. Therefore, the L-6 nerve root, which corresponds to the S-1 nerve root in humans, can be used as a donor nerve to repair lumbosacral plexus root avulsion. However, further studies are needed to assess the safety and feasibility of this approach, because there may be differences between humans and monkeys that could not be assessed in the current study. Disclosure This study was supported by Key Project of Shanghai Science and Technology Commission (grant no ), the Shanghai Rising-Star Program (grant no. 11QA ), and Project of Talents Cultivation of Shanghai Municipal Health Bureau (grant no. XYQ ). Author contributions to the study and manuscript preparation include the following. Conception and design: Hou. Acquisition of data: Lin. Analysis and interpretation of data: Chen. Drafting the article: Lin. Critically revising the article: all authors. Reviewed submitted version of manuscript: all authors. Approved the final version of the manuscript on behalf of all authors: Hou. Statistical analysis: Chen. References 1. Alexandre A, Corò L, Azuelos A: Microsurgical treatment of lumbosacral plexus injuries. 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6 Contralateral L-6 nerve root transfer turator nerve transfer as an option for femoral nerve repair: case report. Neurosurgery 66 (6 Suppl Operative):375, Chin CH, Chew KC: Lumbosacral nerve root avulsion. Injury 28: , Finney LA, Wulfman WA: Traumatic intradural lumbar nerve root avulsion with associated traction injury to the common peroneal nerve. Am J Roentgenol Radium Ther Nucl Med 84: , Frazier CH, Penn G, Skillern JR: Supraclavicular subcutaneous lesions of the brachial plexus not associated with skeletal injuries with the report of a case of avulsion of the anterior and posterior spinal roots. JAMA LVII: , Goubier JN, Teboul F, Papadogeorgou E: Nerve transfers in children with traumatic partial brachial plexus injuries. Microsurgery 28: , Huittinen VM: Lumbosacral nerve injury in fracture of the pelvis. A postmortem radiographic and patho-anatomical study. Acta Chir Scand Suppl 429:3 43, Kolawole TM, Hawass ND, Shaheen MA, Badr AH, Rahman NU: Lumbosacral plexus avulsion injury: clinical, myelographic and computerized tomographic features. J Trauma 28: , Lang EM, Borges J, Carlstedt T: Surgical treatment of lumbosacral plexus injuries. J Neurosurg Spine 1:64 71, Lin H, Xu Z, Liu Y, Chen A, Hou C: The effect of severing L6 nerve root of the sacral plexus on lower extremity function: an experimental study in rhesus monkeys. Neurosurgery 70: , Siqueira MG, Martins RS: Phrenic nerve transfer in the restoration of elbow flexion in brachial plexus avulsion injuries: how effective and safe is it? Neurosurgery 65 (4 Suppl): A125 A131, Sulaiman OA, Kim DD, Burkett C, Kline DG: Nerve transfer surgery for adult brachial plexus injury: a 10-year experience at Louisiana State University. Neurosurgery 65 (4 Suppl): A55 A62, 2009 Manuscript submitted June 19, Accepted May 6, Please include this information when citing this paper: published online June 7, 2013; DOI: / JNS Address correspondence to: Chunlin Hou, M.D., Department of Orthopedic Surgery, Changzheng Hospital, The Second Military Medical University, Fengyang Road 415, Shanghai , People s Republic of China. chunlin_hou@yahoo.com.cn. 719