Taihoh Shibata

Steroid hormones protect spinal cord neurons from glutamate toxicity

The effects of steroid hormones on glutamate neurotoxicity were examined in cultured spinal cord neurons. The extent of neuronal damage, produced by glutamate exposure for 15 min, was estimated based on the activity of lactate dehydrogenase released from degenerated neurons to the media during 24 h of post-exposure incubation. This damage was dependent on the glutamate concentrations used. The addition of dexamethasone, a synthetic steroid, in post-exposure media remarkably reduced the extent of damage in a dose-dependent manner. The half effective concentration for the steroid was approximately 0.7 microM, which was in the range of pharmacological concentration. Dexamethasone was effective even when it was added 2 h after glutamate exposure. Some endogenous steroid hormones--aldosterone, progesterone and testosterone--also showed similar neuroprotective effects. However, cholesterol, a precursor of these steroid hormones, had no effect on glutamate neurotoxicity. This direct protective effect on neurons against glutamate neurotoxicity may explain, at least partly, the mechanisms of beneficial effects of steroid hormones on in vivo spinal cord injury.

Outcomes of surgical treatment for cervical myelopathy in patients more than 75 years of age.

STUDY DESIGN Retrospective study on the results of surgical treatment of compressive cervical myelopathy in patients more than 75 years of age. OBJECTIVES To investigate clinical features and surgical outcomes of compressive cervical myelopathy in aged patients and to discuss the role of surgical treatment. SUMMARY OF BACKGROUND DATA There are few data focused on the outcomes of surgery in patients with cervical myelopathy who are more than 75 years of age. METHODS Seventeen patients with compressive cervical myelopathy who underwent surgery were reviewed. The average age at the time of surgery was 77.2 years. Posterior decompression in 15 patients and anterior decompression in 2 patients were performed. Neurologic deficits before and after surgery were assessed using a scoring system proposed by the Japanese Orthopaedic Association (JOA score). Independence of daily living was evaluated. Radiologic features were examined with radiographs and magnetic resonance imaging. Clinical results were compared with those of patients less than 65 years old as a control. RESULTS The preoperative mean JOA score was 6.1, the postoperative maximum JOA scores averaged 11.4, and the recovery rate was 48.4%. These were significantly inferior to scores in those less than 65 years of age. All seven of the patients who could not walk even with aids before surgery became independent in daily activities after surgery. At the final follow-up, the mean JOA score had decreased to 10.7 and the recovery rate to 39.1%. Five of nine patients whose follow-up periods were more than 5 years showed decreases in JOA score, although all patients were still ambulatory. CONCLUSIONS Surgical decompression for cervical myelopathy appears to be beneficial, even in patients more than 75 years of age, in improving neurologic function and ability to engage in activities of daily living.

Morphologic Differences of the Vascular Buds in the Vertebral Endplate: Scanning Electron Microscopic Study

Study Design Vascular buds in rabbit vertebral endplates were examined by scanning electron microscopy of corrosion casts. Objectives To examine morphologic differences between vascular buds in two regions of the vertebral endplate (inner anular and nucleus pulposar). Summary of Background Data Vascular buds are specific structures present at the vertebral endplate that are important as nourishing channels. There is a significant difference in permeability between the lateral portion (inner anular) and the central portion (nucleus pulposar) of the endplate, the latter usually being permeable and the former being impermeable. Morphologic differences between vascular buds in the two regions have not been investigated previously. Methods Eight 20-week-old rabbits were used. Vascular buds in rabbit vertebral endplates were examined by scanning electron microscopy of corrosion casts. Results The vascular buds in the region of the inner anulus form simple loops, but those in the area near the nucleus pulposus exhibit swollen and complex coil-like loops. Although they differ structurally, the average number of vascular buds per area does not vary between the two regions. Conclusions We suggest that the morphologic difference between the vascular buds in the two regions (inner anular and nucleus pulposar) plays a principal role in permeability at the endplate.

Reconstruction surgery for patients with musculoskeletal tumor, using a pasteurized autogenous bone graft.

BackgroundThe pasteurized autogenous bone graft (PABG) is a new method to reuse resected and diseased autogenous bones after heat treatment at a comparatively low temperature (60°C–65°C).MethodsThe subjects of this study were ten patients with musculoskeletal tumor who underwent surgery with a PABG in the 6 years between 1995 and 2000.ResultsThe pasteurized bone developed into bone union in all patients, except for the elderly patients who required repeat surgery. There were no infected patients. The PABG was performed by three different types of reconstruction, a segmental method, an intercalary method, and a combination method with an artificial joint as a spacer. No local recurrence of tumor the pasteurized bone was observed from in any patient.ConclusionThe PABG appears to be a comparatively easy, safe, inexpensive, and effective reconstruction method for musculoskeletal tumors.

Existence of Lipoprotein Lipase in Human Sarcomas and Carcinomas

Aqueous extracts of acetone/ether powders of surgically obtained specimens of human tumors hydrolyzed 3H‐labeled triolein in a dose‐dependent manner. The lipolytic activity in these extracts was inhibited by anti‐lipoprotein lipase (LPL) IgG dose‐dependently, 25 μg of anti‐LPL IgG causing 95% inhibition of the activity. Thus, LPL accounts for most of the lipolytic activity in extracts of acetone/ether powders of the tumors. All sarcomas and carcinomas examined contained LPL activity. Western blotting showed that they gave a band corresponding to that of human adipose tissue LPL (Mr=57,000). Immunocytochemical studies showed that LPL was present in cultured human osteosarcoma cells and distributed throughout the cells. We determined the proliferating cell nuclear antigen (PCNA)‐labeling index as an indicator of the proliferative activity of tumor cells and measured LPL activity in extracts of tumors in areas corresponding to those used for determining the PCNA‐labeling index. In malignant fibrous histiocytomas, the PCNA‐labeling index in area a, which corresponds to the subcapsular region, was higher than that in area b, which corresponds to the central region. The LPL activity in area a was 10 times that in area b. In rectal cancer, the index in area c, which corresponds to the subserosal region, was higher than that in area d, which corresponds to the submucosal region. The LPL activity in area c was 1.9 times that in area d. These findings indicate heterogeneity in the distributions of LPL activity within tumors and higher levels of LPL activity in tumors that are proliferating actively.

Atlanto-occipital hypermobility in subjects with Down's syndrome

Study Design The upper cervical spines of 57 subjects with Downs syndrome were retrospectively examined, with special attention to atlanto-occipital mobility. Objective To examine the magnitude of atlanto-occipital mobility and its clinical significance in subjects with Downs syndrome. Summary of Background Data Atlanto-occipital translation of more than 1 mm in adults implies instability. However, the normal value in children with Downs syndrome has not been established, and the value In Downs syndrome has not been evaluated based on a comparison between subjects with Downs syndrome and control subjects. Methods Measurements were made by Wiesel and Rothmans method in 38 subjects with Downs syndrome and 34 control subjects. Results Atlanto-occipital translation in the Downs syndrome group ranged from 0–6.4 mm (mean, 2.3 mm), whereas in the control group it ranged from 0–2.1 mm (mean, 0.61 mm). The difference was statistically significant. Of the 38 subjects with Downs syndrome, 37 were asymptomatic. Conclusion The magnitude of atlanto-occipital translation, as expected, apparently was greater in subjects with Downs syndrome than in control subjects. Although the possibility of neurologic complications should be considered whenever unusually high atlanto-occipital mobility is seen, a majority of the subjects with Downs syndrome were asymptomatic.

Protection by diphenyliodonium against glutamate neurotoxicity due to blocking ofN-methyl-d-aspartate receptors

The protective effect of diphenyliodonium, known as an inhibitor of flavin enzymes including nitric oxide synthases, was examined against the neurotoxicity of excitatory amino acids on cultured spinal neurons of the rat. Diphenyliodonium reduced the neuronal damage induced by 15-min exposure to glutamate or N-methyl-D-aspartate in a dose-dependent manner; half effective concentrations (EC50) were about 3 microM for both. Protection was only observed when diphenyliodonium was added into the exposure medium. Diphenyliodonium showed no effect on the toxicity induced by 24 h exposure to non-N-methyl-D-aspartate receptor agonists. Using a microfluorometry technique with Fura 2, we observed that diphenyliodonium reversibly inhibited the N-methyl-D-aspartate-evoked intracellular Ca2+ elevation. The amount of 45Ca2+ influx induced by N-methyl-D-aspartate was also inhibited by diphenyliodonium in a dose-dependent manner; EC50 was about 3 microM. Furthermore, we examined the effect of diphenyliodonium on an opening activity of the N-methyl-D-aspartate receptors estimated by binding of dizocilpine maleate to membrane fractions from whole brain of adult rat and from cultured spinal neurons. Diphenyliodonium inhibited the binding of dizocilpine maleate dose-dependently; EC50 was 5-8 microM. These results suggest that diphenyliodonium is a new antagonist to the N-methyl-D-aspartate receptors and that diphenyliodonium protects neurons against glutamate toxicity due to a direct blocking of the Ca2+ influx. This conclusion is supported by the similarity of the stereochemical structures predicted by computer between diphenyliodonium and dizocilpine maleate.

Neurotoxicity of acromelic acid in cultured neurons from rat spinal cord

Acromelic acid A, which contains the kainic acid structure in its molecule, is known to cause selective damage of interneurons in the rat lower spinal cord. In the present study, the potent neurotoxicity of acromelic acid A was demonstrated in cultured rat spinal neurons in terms of the activity of lactate dehydrogenase that was released from degenerated neurons into the culture medium. Acromelic acid A increased the lactate dehydrogenase activity in time- and concentration-dependent manners, and its EC50 was about 2.5 microM, which was much lower than that of kainic acid (70 microM) and (RS)-alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (EC50; 11 microM). The maximum level of lactate dehydrogenase released by acromelic acid A was quite similar to that by kainic acid, but was about twice the level produced by (RS)-alpha-3-hydroxy-5-methyl-4-isoxazolepropionic acid. Exposure to acromelic acid A caused release of L-glutamate from the cells into the medium; however, the concentration of L-glutamate released was far below the level for inducing the neurotoxic effects. The neurotoxicity of 10 microM acromelic acid A was almost completely inhibited by 30 microM 6-nitro-7-sulphamoylbenzo(F)quinoxaline-2,3-dione and 6-cyano-7-nitroquinoxaline-2,3-dione, potent antagonists for non-N-methyl-D-aspartate receptors, but was partly (35%) reduced by 30 microM dizocilpine maleate. In cultured hippocampal neurons, the toxicity of acromelic acid A was significantly less effective (EC50: 18 microM) than that in spinal neurons, whereas the toxicity of kainic acid was almost the same in both neurons. These results suggest that acromelic acid A directly activates non-N-methyl-D-aspartate receptors on the cultured spinal neurons to induce neuronal death. A new type of non-N-methyl-D-aspartate receptors which is specific to acromelic acid A is suggested to be present at least in spinal neurons.

Transient increase of cyclic AMP induced by glutamate in cultured neurons from rat spinal cord.

Abstract: We demonstrated that glutamate increased the cyclic AMP level in cultured neurons from rat spinal cord. A bath application of glutamate (300 µM) elicited a rapid increase of the cyclic AMP concentration reaching a level three times as high as the basal level in ∼3 min, and its content then decreased to the control level in 15 min. The increase was not observed in a Ca2+‐free medium and was inhibited by an antagonist of NMDA receptors or a voltage‐sensitive Ca2+ channel blocker. Preincubation with W7 also inhibited the glutamate‐evoked cyclic AMP increase. NMDA, aspartate, and high‐K+ conditions also induced a cyclic AMP increase; however, a decreasing phase did not follow. The decreasing phase was observed when (2S,1′S,2′S)‐2‐(carboxycyclopropyl)‐glycine, a potent agonist for metabotropic glutamate receptors, was combined with NMDA. These results suggest that the cyclic AMP increase is mediated by a Ca2+ influx via both NMDA receptors and voltage‐sensitive Ca2+ channels followed by an activation of the Ca2+/calmodulin system, and the decreasing phase observed in the case of glutamate exposure is due to the activation of the metabotropic glutamate receptors.

Three-dimensional structure of the vascular network in normal and immobilized muscles of the rat☆☆☆

OBJECTIVE To examine whether there are three-dimensional changes in capillaries of rat soleus muscle during mechanical limb immobilization. DESIGN Scanning electron microscopy, with the microcorrosion cast technique, was used to examine vascular networks in immobilized Wistar rat soleus muscles from 4 to 12 weeks after limb immobilization and to compare these networks with those in the control muscle. INTERVENTION Immobilization of the soleus muscle was achieved by unilateral pinning of the ankle joint in full plantar flexion. RESULTS The wavelike structure of the vascular network in the control muscles changed into a straight configuration in 8- and 12-week immobilized muscles.