Kenichi Amagasaki

Quantification of thrombospondin-1 secretion and expression of αvβ3 and α3β1 integrins and syndecan-1 as cell-surface receptors for thrombospondin-1 in malignant glioma cells

Malignant glioma cells secrete thrombospondin-1 (TSP-1) which participates in the motility of glioma cells, and binds to cell surface αvβ3 and α3β1 integrins, and syndecan-1. This study evaluated the amount of TSP-1 secretion from malignant glioma cells, and the expression of αvβ3 and α3β1 integrins, and syndecan-1. The amounts of TSP-1 in the supernatants from 10 malignant glioma cell lines and eight non-glioma malignant tumor cell lines were measured by enzyme-linked immunosorbent assay. Expression of αvβ3 and α3β1 integrins, and syndecan-1 were examined by flow cytometry. The amounts of TSP-1 secreted by malignant glioma cells were 43 to 2431 ng/1 × 106 cells/24 h (mean ± SD=626 ± 792). Seven of 10 glioma cell lines secreted more than 100 ng of TSP-1 and three of these cell lines secreted more than 1 μg. Seven of eight non-glioma cell lines secreted less than 100 0ng of TSP-1. All glioma cell lines expressed α3β1 integrin and syndecan-1, and seven of 10 glioma cell lines expressed αvβ3 integrin. Treatment of the glioma cell lines with TGF-β2 did not change the expression of αvβ3 integrin. These results suggest that malignant glioma cells secrete high levels of TSP-1, which may be important in the migration of glioma cells via interactions with αvβ3 and α3β1 integrins, and syndecan-1.

Antisense-mediated reduction in thrombospondin-1 expression reduces cell motility in malignant glioma cells

Thrombospondin‐1 (TSP‐1) is a multifunctional matrix protein implicated in cancer cell adhesion, migration, invasion, inhibition of angiogenesis and activation of latent transforming growth factor‐β. The involvement of TSP‐1 in the motility of malignant glioma cells was investigated by transfection of TSP‐1 complementary deoxyribonucleic acid (cDNA) sense and antisense expression vectors into the glioblastoma cell line T98G‐G7 that secretes high amounts of TSP‐1. TSP‐1 production in the 3 antisense cDNA‐transfected clones was significantly reduced to 51%, 43% and 47% compared to the host T98G‐G7 cells. Motility of the 3 clones was evaluated by invasion assay and compared to the motility of host T98G‐G7 cells and 2 sense‐transfected clones. Migration of cells was significantly reduced in the 3 antisense‐transfected clones with reduced TSP‐1 production to 56%, 61% and 43% compared to the host T98G‐G7 cells. The host T98G‐G7 and another TSP‐1‐secreting A172 and YMG5 glioblastoma cells were also treated with a synthetic peptide, WSHWSPWSSCSVTCG, which includes 3 consecutive sequences of the adhesion sites in the TSP‐1 molecule and with a control peptide. The synthetic peptide significantly inhibited the migration of T98G‐G7 and A172 cells in a dose‐related manner. Maximum inhibition of migration was achieved by 100 μg/ml of the peptide and the reduction of cell motility compared to untreated cells was 34.6 % and 53.9 %, respectively. On the other hand, the inhibition of migration by the peptide was minimal in YMG5 cells, which secretes a smaller amount of TSP‐1 than T98G‐G7 and A172 cells. These results suggest that TSP‐1 secreted by malignant glioma cells is involved in the motility of glioma cells.

Treatment of ruptured anterior circulation aneurysm based on computerized tomography angiography: surgical results and indications for additional digital subtraction angiography.

The purpose of this study is to determine whether or to what extent computerized tomography angiography (CTA) can replace digital subtraction angiography (DSA) for ruptured anterior circulation cerebral aneurysms. We reviewed our patients in two different periods and evaluated in what cases we should perform additional DSA. In the first period, both CTA and DSA were routinely performed except on patients that were suitable for emergent surgery with CTA only. In the second period, only CTA was performed routinely as a preoperative workup and if further information was necessary, additional DSA was performed. A total of 106 (52 in the first period and 54 in the second period) patients were included. Characteristics of the patients and outcome at three months were similar in the two periods. In the second period, ten patients underwent additional DSA. Our results suggest that most of cases with ruptured anterior circulation aneurysms can be successfully treated on the basis of CTA alone.

Recurrence of medulloblastoma 19 years after the initial diagnosis

Abstract A 25-year-old woman presented with recurrent medulloblastoma. She had been diagnosed with cerebellar medulloblastoma and treated for it at the age of 5 years. The new tumor was partially resected and chemotherapy was begun. However, a disseminated tumor of the spine was discovered and, despite radiotherapy, the patient became comatose and died. Histological examination of the tumor specimen showed characteristics similar to those of the first tumor. Late recurrence of medulloblastoma is rare, and a latency period of 19 years is the longest reported. This case violates Collins’ rule and exceeds other proposed periods of risk. Extension of the period of follow-up in patients with medulloblastoma is advocated.

Establishment and partial characterization of five malignant glioma cell lines

Five malignant glioma cell lines (YMG1, 2, 3, 4, and 5) were established from surgical specimens obtained from patients with glioblastoma or anaplastic astrocytoma, and these lines were partially characterized. Three glioma cell lines (YMG1, 3, and 5) were weakly positive for GFAP by Western blot analysis and two cell lines were negative. S‐100 protein was positive in all glioma cell lines. The expression of p53, p16, p15, cyclin‐dependent kinase 4 (CDK4), and EGF receptor (EGFR) proteins was examined by Western blotting. YMG1 and 2 cell lines showed accumulation of p53 protein and loss of p16 and p15 expression. YMG3 and 4 showed accumulation of p53 protein and expression of p16 and p15 proteins. YMG5 revealed weak expression of p53 protein, suggesting wild‐type p53, and loss of p16 and p15 expression. All cell lines expressed various levels of CDK4 protein. YMG1, 2, and 3 showed higher EGFR protein expression and YMG4 and 5 showed lower EGFR expression compared to U251 glioblastoma cells, which express high levels of EGFR. Fluorescence in situ hybridization analysis for EGFR gene expression did not show any amplification in the glioma cell lines. Immunohistochemical studies revealed that the patterns of p53 and EGFR expressions in the original tumor tissues were mostly correlated with those in the malignant glioma cell lines. These results suggest that the characteristics of p53 and EGFR expression in the malignant glioma cell lines were passed over from the original tumor tissues. These newly established malignant glioma cell lines can be used for further analysis of the mechanisms of tumor growth and progression.

Cell density regulates thrombospondin-1 production in malignant glioma cells

Thrombospondin-1 (TSP-1) is a multifunctional matrix protein implicated in cancer cell adhesion, migration, and invasion, inhibition of angiogenesis, and activation of latent transforming growth factor-β (TGF-β). The effect of cell density was investigated on the production of TSP-1, basic fibroblast growth factor (bFGF), and vascular endothelial growth factor (VEGF) by two glioblastoma cell lines. The effect of TGF-β was also examined. The amount of intracellular TSP-1 protein decreased significantly as the cell density increased in cultures of both T98G and A172 cells. The amount of intracellular TSP-1 was highest in sparse tumor cell cultures and lowest in densely confluent tumor cell cultures. The maximum reduction of TSP-1 protein production was 56.8% and 44.6% in T98G and A172 cells, respectively. The cell density did not affect the production of bFGF or VEGF. TGF-β2 treatment did not affect the production of TSP-1, bFGF, or VEGF proteins. Treatment with excess TGF-β2 resulted in a slight but significant decrease (22%; P < 0.02) of TGF-β2 production by A172 cells, but not by T98G cells. The present results indicate that the production of TSP-1 protein is regulated by cell density of glioblastoma cells, while that of angiogenic factors is not affected by tumor cell density. This suggests that high tumor cell density may tilt the angiogenic balance in favor of angiogenesis.

Current usage of three-dimensional computed tomography angiography for the diagnosis and treatment of ruptured cerebral aneurysms

Our previous study suggested that 3D-CT angiography could replace digital subtraction (DS) angiography in most cases of ruptured cerebral aneurysms, especially in the anterior circulation. This study reviewed our further experience. One hundred and fifty patients with ruptured cerebral aneurysms were treated between November 1998 and March 2002. Only 3D-CT angiography was used for the preoperative work-up study in patients with anterior circulation aneurysms, unless the attending neurosurgeons agreed that DS angiography was required. Both 3D-CT angiography and DS angiography were performed in patients with posterior circulation aneurysms, except for recent cases that were possibly treated with 3D-CT angiography alone. One hundred sixteen (84%) of 138 patients with ruptured anterior circulation aneurysms underwent surgical treatment, but additional DS angiography was required in 22 cases (16%). Only two recent patients were treated surgically with 3D-CT angiography alone in 12 patients with posterior circulation aneurysms. Most patients with ruptured anterior circulation aneurysms could be treated successfully after 3D-CT angiography alone. However, additional DS angiography is still necessary in atypical cases. 3D-CT angiography may be limited to complementary use in patients with ruptured posterior circulation aneurysms.

Lower cranial nerve palsy after the infrafloccular approach in microvascular decompression for hemifacial spasm

Background: The infrafloccular approach was introduced as a variation in microvascular decompression (MVD) for hemifacial spasm. However, the rate of postoperative lower cranial nerve (CN) palsy can be high. This study investigated the surgical factors in relation to the occurrence of postoperative lower CN palsy. Methods: The case records of 103 patients who underwent MVD were reviewed. Dissection around the lower CNs to approach the root exit zone of CN VII was divided into two steps – incision of the rhomboid lip at the root of the lower CNs and separation of CN IX and flocculus/choroid plexus. The correlations of these steps and other characteristics to the occurrence of lower CN palsy were analyzed. Results: Ten of the 103 patients suffered from postoperative transient lower CN palsy. The rhomboid lip was incised in 30 cases (29.1%), separation of CN IX and flocculus or choroid plexus was necessary in 24 cases (23.3%), and both steps were required in 7 cases (6.8%). The steps showed no correlation with postoperative lower CN palsy. Posterior inferior cerebellar artery (PICA) as the offending vessel was significantly correlated with postoperative lower CN palsy (P < 0.05). Conclusions: Our study showed that the offending PICA was the only significant factor for postoperative lower CN palsy. Therefore, correct dissection around the lower CNs, particularly for complicated PICA, is necessary to reduce the risk of postoperative lower CN palsy.

Trigeminal neuralgia caused by a trigeminocerebellar artery

This 31-year-old woman presented with typical right trigeminal neuralgia caused by a trigeminocerebellar artery, manifesting as pain uncontrollable with medical treatment. Preoperative neuroimaging studies demonstrated that the offending artery had almost encircled the right trigeminal nerve. This finding was confirmed intraoperatively, and decompression was completed. The neuralgia resolved after the surgery; the patient had slight transient hypesthesia, which fully resolved within the 1st month after surgery. The neuroimaging and intraoperative findings showed that the offending artery directly branched from the upper part of the basilar artery and, after encircling and supplying tiny branches to the nerve root, maintained its diameter and coursed toward the rostral direction of the cerebellum, which indicated that the artery supplied both the trigeminal nerve and the cerebellum. The offending artery was identified as the trigeminocerebellar artery. This case of trigeminal neuralgia caused by a trigeminocerebellar artery indicates that this variant is important for a better understanding of the vasculature of the trigeminal nerve root.

Fibrin glue injection into the hematoma cavity for refractory chronic subdural hematoma: A case report

Background: Repeat burr hole irrigation and drainage has been effective in most cases of recurrent chronic subdural hematoma (CSDH), however, refractory cases require further procedures or other interventions. Case Description: An 85-year-old male presented with left CSDH, which recurred five times. The hematoma was irrigated and drained through a left frontal burr hole during the first to third surgery and through a left parietal burr hole during the fourth and fifth surgery. The hematoma had no septation and was well-evacuated during each surgery. Antiplatelet therapy for preventing ischemic heart disease was stopped after the second surgery, the hematoma cavity was irrigated with artificial cerebrospinal fluid at the third surgery, and the direction of the drainage tube was changed to reduce the postoperative subdural air collection at the fourth surgery. However, none of these interventions was effective. He was successfully treated by fibrin glue injection into the hematoma cavity after the fifth surgery. Conclusion: This procedure may be effective for refractory CSDH in elderly patients.