Ryuichi Shinjo

Keratan Sulfate Restricts Neural Plasticity after Spinal Cord Injury

Chondroitin sulfate (CS) proteoglycans are strong inhibitors of structural rearrangement after injuries of the adult CNS. In addition to CS chains, keratan sulfate (KS) chains are also covalently attached to some proteoglycans. CS and KS sometimes share the same core protein, but exist as independent sugar chains. However, the biological significance of KS remains elusive. Here, we addressed the question of whether KS is involved in plasticity after spinal cord injury. Keratanase II (K-II) specifically degraded KS, i.e., not CS, in vivo. This enzyme digestion promoted the recovery of motor and sensory function after spinal cord injury in rats. Consistent with this, axonal regeneration/sprouting was enhanced in K-II-treated rats. K-II and the CS-degrading enzyme chondroitinase ABC exerted comparable effects in vivo and in vitro. However, these two enzymes worked neither additively nor synergistically. These data and further in vitro studies involving artificial proteoglycans (KS/CS-albumin) and heat-denatured or reduced/alkylated proteoglycans suggested that all three components of the proteoglycan moiety, i.e., the core protein, CS chains, and KS chains, were required for the inhibitory activity of proteoglycans. We conclude that KS is essential for, and has an impact comparable to that of CS on, postinjury plasticity. Our study also established that KS and CS are independent requirements for the proteoglycan-mediated inhibition of axonal regeneration/sprouting.

ADAMTS-13 is produced by glial cells and upregulated after spinal cord injury

ADAMTS-13, a member of the family of disintegrins and metalloproteinases with thrombospondin motifs, is produced primarily in the liver, particularly by hepatic stellate cells. This metalloproteinase cleaves von Willebrand factor multimers and thereby regulates blood coagulation. Here, we investigated the expression of ADAMTS-13 in the central nervous system. ADAMTS-13 mRNA was expressed in cultured astrocytes and microglia but not in neurons. The protein production of ADAMTS-13 was also detected in these cultured glial cells. Furthermore, we found that the expression of ADAMTS-13 was significantly increased in the rat spinal cord after injury. Supporting the in vivo data, ADAMTS-13 protein was detected in GFAP- and CD11b-positive glial cells in injured spinal cord. Consistent with this, the proteolytic activity of ADAMTS-13 was increased after spinal cord injury. Our data suggest that ADAMTS-13 may have a critical role in the central nervous system, particularly after neuronal injuries.

Midkine overcomes neurite outgrowth inhibition of chondroitin sulfate proteoglycan without glial activation and promotes functional recovery after spinal cord injury

Injuries in the mammalian central nervous system induce a variety of factors which promote or inhibit neuronal axon regeneration/sprouting. However, the inhibitory activities are much stronger, and indeed are the major obstacle to functional recovery. Chondroitin sulfate proteoglycans (CSPGs) are produced by activated glial cells, and are among the strongest inhibitors. Here, we investigated the role of the growth factor midkine (MK), which binds to CSPGs, in neuronal injury. MK expression was induced by spinal cord injury, and was mainly produced by activated astrocytes. A prolonged culture of neurons also produced MK. MK not only enhanced neurite outgrowth on the substratum coated with poly-l-lysine, but also overcame the neurite growth inhibition by the CSPG substratum. Moreover, we found that MK activated neither astrocytes nor microglia as evaluated by morphological changes and cell proliferation or nitric oxide production. These properties would be advantageous for the treatment of neuronal injuries in vivo. Therefore, we next explored the therapeutic effect of MK in a rat spinal cord injury model. MK or vehicle was administered intrathecally for 2 weeks using an osmotic pump after spinal cord contusion injury. Rats treated with MK showed significantly better functional recovery after 5 weeks. These results suggest that MK may offer a potent alternative for the treatment of neuronal injuries without activating glial cells.

Ponte Osteotomy During Dekyphosis for Indirect Posterior Decompression with Ossification of Posterior Longitudinal Ligament of the Thoracic Spine.

Study Design: Retrospective clinical study. Purpose: To investigate the outcomes after indirect posterior decompression and dekyphosis using multilevel Ponte osteotomies for ossification of the posterior longitudinal ligament (OPLL) of the thoracic spine. Summary of Background Data: There are no previous reports on the use of Ponte osteotomy to treat thoracic OPLL. Methods: The subjects were 10 patients with an average age at surgery of 47 years, who underwent indirect posterior decompression and dekyphosis using multilevel Ponte osteotomies at our institute. Minimum follow-up period was 2 years, and averaged 2 year 6 months. Using radiographs and CT images, we investigated fusion range, preoperative and postoperative Cobb angles of thoracic fusion levels, intraoperative ultrasonography, and clinical results. Results: The mean fusion area was 9.8 vertebraes, with average laminectomy of 7.3 laminas. The mean preoperative thoracic kyphosis of fusion levels on standing radiograph measured 35 degrees and was changed to 21 degrees after surgery. The mean number of Ponte osteotomies was 3 levels. The mean preoperative and postoperative (at the 1 y follow-up) JOA scores were 3.5 and 7.5 points, respectively, and the recovery rate was 56%. On intraoperative ultrasonography, 7 of the cases were included in the floating (+) and 3 in the floating (−) groups, and the recovery rates were 66.0% and 33.4%, respectively. Conclusions: “The Ponte procedure for indirect spinal cord decompression” is a novel concept used for the first time with thoracic OPLL in our study, and we consider it a useful method to achieve more effectively dekyphosis and indirect spinal cord decompression if there is not the spinal cord free from OPLL on intraoperative ultrasonography after only laminectomies.

Intradural disc herniation: Radiographic findings and surgical results with a literature review

OBJECTIVE To report a series of four cases of intradural disc herniation (IDH) with a review of the literature. SUMMARY OF BACKGROUND DATA IDH is a rare type of disc herniation. Preoperative diagnosis is difficult and IDH is only confirmed during surgery in most cases. Here, we describe four cases of IDH, including three with lumbar hernia and one with thoracic hernia. METHODS A retrospective chart review, surgical database query, and review of radiology reports are presented for each case, along with a literature review of IDH. RESULTS Two of the four patients had a history of surgery at the same spinal level. Ring enhancement in gadolinium-enhanced MRI, an air image in computed tomography, and complete block in myelography were observed in the series. Surgery was performed with a transdural approach in all patients. One patient underwent transforaminal lumbar interbody fusion after postoperative recurrence. Three patients with lumbar involvement had nerve root symptoms preoperatively, but showed symptomatic improvement in the early postoperative period. In contrast, the patient with thoracic involvement had preoperative muscle weakness due to myelopathy symptoms, and had residual symptoms after surgery. CONCLUSIONS IDH is a rare disease and characteristic imaging findings can be useful for diagnosis. Intraoperative findings lead to a definitive diagnosis in many cases and recognition of the pathological characteristics of IDH is important.

Evaluation of the effect of tranilast on rats with spinal cord injury

BACKGROUND Glial and fibrotic scars inhibit neural regeneration after spinal cord injury (SCI). N-[3,4-dimethoxycinnamoyl]-anthranilic acid (tranilast) inhibits transforming growth factor β, alleviates allergic reactions, and decreases hypertrophic skin scars. We evaluated its ability to improve motor function and inhibit the spread of tissue damage in rats with SCI. METHODS Rats with SCI were divided into groups that received tranilast (30 mg/[kg · day]) by intravenous administration (group IV), tranilast (200mg/[kg · day]) by oral administration (group OR), and saline injections (control). Motor functions were assessed by determining Basso, Beattie, and Bresnahan (BBB) scores and %grip tests for 8 weeks after SCI. Histological evaluation of ionized calcium binding adaptor molecule 1 (Iba1) at 1 week after SCI and glial fibrillary acidic protein (GFAP), fibronectin, and chondroitin sulfate (CS) at week 8 was performed. RESULTS Motor function recovery, BBB score, and the %grip test were significantly higher in the tranilast-treated groups than in the control group. At week 1 after SCI, inflammatory-cell invasion was more severe and Iba1 expression was significantly higher in the control group. At week 8, although the number of GFAP-positive cells increased greatly from the impaction site to the proximal and distal sites in the control group, these cells were confined around a cavity in the tranilast-treated groups. GFAP distribution coincided with that of fibronectin. Anti-CS antibody level in the tranilast-treated groups was significantly lower than that in the control group. CONCLUSIONS Tranilast inhibits inflammation in the acute phase of SCI and reduces glial and fibrotic scars and could present a new method for treating SCI.

Arundic acid (ONO-2506) inhibits secondary injury and improves motor function in rats with spinal cord injury

BACKGROUND Arundic acid (ONO-2506) inhibits the production and release of S100 protein from astrocytes. While numerous studies have assessed the effect of ONO-2506 in the diseased brain, to the best of our knowledge, no study has examined the effect of ONO-2506 in spinal cord injury (SCI). In this study, we administered ONO-2506 to rats with SCI in order to evaluate its effectiveness in improving motor function and protecting against histological injury. METHODS All rats underwent laminectomy with SCI at the 10th thoracic vertebra. Rats were divided into 3 groups that received different concentrations of ONO-2506 as follows: 10 mg/kg (Group I) and 20 mg/kg (Group II). The third group (control group) was administered only saline. ONO-2506 or saline was administered by intravenous injection for a week after SCI. Recovery of motor function was assessed by determining the Basso, Beattie, and Bresnahan (BBB) scores and using the %grip test. Using immunohistochemistry, S100 protein and glial fibrillary acidic protein expression was assessed at week 12 post SCI. RESULTS The BBB score of Group II was significantly better than that of the control group. At week 12 post SCI, the %grip was 43.0% in Group II and 20.3% in Group I. The score for the %grip test was greater for Group II than for the control group (7.0%); thus, motor function improvement appeared to be dose dependent. Regarding immunostaining evaluation, S100 protein staining was lower in Group II compared to the control group, and the astrocytic morphology resembled that of normal spinal cord sections. The SCI lesion expanded from the injured site to both proximal and distal sites in the control group and in Group I. However, despite the presence of cavitation, secondary expansion of the SCI lesion was prevented in Group II as a result of inhibition of S100 protein. CONCLUSIONS Administration of ONO-2506 (20 mg/kg) improves motor function and inhibits expansion of secondary injury in SCI rats.

Radiologic evaluation after posterior instrumented surgery for thoracic ossification of the posterior longitudinal ligament: union between rostral and caudal ossifications.

Study Design: Retrospective clinical study. Objective: To investigate, using multislice CT images, how thoracic ossification of the posterior longitudinal ligament (OPLL) changes with time after thoracic posterior fusion surgery. Summary of Background Data: Few studies have evaluated thoracic OPLL preoperatively and post using computed tomography (CT). Methods: The subjects included 19 patients (7 men and 12 women) with an average age at surgery of 52 years (38-66 y) who underwent indirect posterior decompression with corrective fusion and instrumentation at our institute. Minimum follow-up period was 1 year, and averaged 3 years 10 months (12–120 mo). Using CT images, we investigated fusion range, preoperative and postoperative Cobb angles of thoracic fusion levels, intraoperative and postoperative blood loss, operative time, hyperintense areas on preoperative MRI of thoracic spine and thickness of the OPLL on the reconstructed sagittal, multislice CT images taken before the operation and at 3 months, 6 months and 1 year after surgery. The basic fusion area was 3 vertebrae above and below the OPLL lesion. Results: The mean operative time was 7 hours and 48 min (4 h 39 min–10 h 28 min), and blood loss was 1631 mL (160–11,731 mL). Intramedullary signal intensity change on magnetic resonance images was observed at the most severe ossification area in 18 patients. Interestingly, the rostral and caudal ossification regions of the OPLLs, as seen on sagittal CT images, were discontinuous across the disk space in all patients. Postoperatively, the discontinuous segments connected in all patients without progression of OPLL thickness by 5.1 months on average. Conclusions: All patients needing surgery had discontinuity across the disk space between the rostral and caudal ossified lesions as seen on CT. This discontinuity was considered to be the main reason for the myelopathy because a high-intensity area on magnetic resonance imaging was seen in 18 of 19 patients at the same level. Rigid fixation with instrumentation may allow the discontinuous segments to connect in patients without a concomitant thickening of the OPLL.

Characteristics and surgical results of the distal type of cervical spondylotic amyotrophy.

OBJECT Cervical spondylosis that causes upper-extremity muscle atrophy without gait disturbance is called cervical spondylotic amyotrophy (CSA). The distal type of CSA is characterized by weakness of the hand muscles. In this retrospective analysis, the authors describe the clinical features of the distal type of CSA and evaluate the results of surgical treatment. METHODS The authors performed a retrospective review of 17 consecutive cases involving 16 men and 1 woman (mean age 56.3 years) who underwent surgical treatment for the distal type of CSA. The condition was diagnosed on the basis of cervical spondylosis in the presence of muscle impairment of the upper extremity (intrinsic muscle and/or finger extension muscles) without gait disturbance, and the presence of a compressive lesion involving the anterior horn of the spinal cord, the nerve root at the foramen, or both sites as seen on axial and sagittal views of MRI or CT myelography. The authors assessed spinal cord or nerve root impingement by MRI or CT myelography and evaluated surgical outcomes. RESULTS The preoperative duration of symptoms averaged 11.8 months. There were 14 patients with impingement of the anterior horn of the spinal cord and 3 patients with both anterior horn and nerve root impingement. Twelve patients were treated with laminoplasty (plus foraminotomy in 1 case), 3 patients were treated with anterior cervical discectomy and fusion, and 2 patients were treated with posterior spinal fixation. The mean manual muscle testing grade was 2.4 (range 1-4) preoperatively and 3.4 (range 1-5) postoperatively. The surgical results were excellent in 7 patients, good in 2, and fair in 8. CONCLUSIONS Most of the patients in this series of cases of the distal type of CSA suffered from impingement of the anterior horn of the spinal cord, and surgical outcome was fair in about half of the cases.

Keratan sulfate expression is associated with activation of a subpopulation of microglia/macrophages in Wallerian degeneration.

Wallerian degeneration is a fundamental process of axonal degeneration distal to the injury site. Although axonal degeneration itself is accomplished in a few days, the subsequent process of removing debris, including myelin debris, in the central nervous system takes more time. Since this debris is a potent inhibitor of axonal regeneration, the removal process is critical for functional recovery after neuronal injuries. Although it is known that microglia/macrophages are involved in this process, the underlying mechanisms are not fully understood. Here, we found that keratan sulfate (KS) expression was induced far from the injury site after spinal cord injury. A hemilateral section of the spinal cord at the third cervical level induced KS expression in a restricted area of the ipsilateral column at the first lumbar level 1 week after injury. This localized KS expression lasted for at least 1 month after injury. The KS signal was merged with a portion of Iba1-positive cells, suggesting that a subpopulation of microglia/macrophages expressed KS. KS-positive cells expressed CD68 and CD86, but not CD206 or arginase 1, suggesting that these microglia/macrophages were in an activated state probably polarized to M1. Our study has explored for the first time the relation between KS expression and activation of microglia/macrophages in Wallerian degeneration.