Yoshihito Matsumoto

Matrix metalloproteinase-2 and -9 expression in astrocytic tumors.

In this study, we investigated whether the expression of matrix metalloproteinase (MMP)-2 and MMP-9 correlated with invasiveness, proliferative potential, or prognosis in astrocytic tumors. Thirty-seven astrocytic tumors (8 diffuse astrocytomas, 15 anaplastic astrocytomas, and 14 glioblastomas) and three gliomatosis cerebri were investigated immunohistochemically. The invasive glioma group included three cases of gliomatosis cerebri and two of glioblastoma associated with cerebrospinal fluid dissemination. The expression of MMP-2 and MMP-9 was evaluated by assigning an immunohistochemical (IHC) score defined as the sum of expression frequency and intensity. mRNA expression patterns for the MMPs were also evaluated in a reverse transcription-polymerase chain reaction assay. Neither the MMP-2 nor MMP-9 IHC score was related to histological malignancy. The MMP-2 IHC score of the invasive glioma group was significantly higher than those of other kinds of astrocytic tumors. However, the MMP-9 IHC score did not correlate with dissemination among astrocytic tumors. An inverse correlation was observed between the MIB-1 labeling index and the IHC scores of MMP-2, but it was not significant. A Kaplan-Meyer survival analysis revealed no significant relationship between the survival rate and MMP-2 or MMP-9 expression. Our study showed that MMP-2 expression, but not MMP-9 expression, may be associated with invasion in astrocytic tumors.

Matrix Metalloproteinase-2 and Matrix Metalloproteinase-9 Expressions Correlate with the Recurrence of Intracranial Meningiomas

AbstractBackground: We studied whether or not the expressions of MMP-2 and MMP-9 were correlated with the proliferative potential or recurrence of intracranial meningiomas. Methods: The expressions of MMP-2 and MMP-9 of 56 meningioma tissues were examined and scored by immunohistochemistry (IHC). Expressions of the mRNA of each MMP were also examined by reverse transcriptase polymerase chain reaction (RT-PCR) assay in 15 cases. The relationships between IHC score and each of age, sex, tumor location, histology, MIB-1 labeling index (LI), and recurrence or regrowth rate were studied. Results: Nine of 56 cases showed recurrence or regrowth. IHC revealed that MMP-2 was highly expressed in 13 of 56 cases, whereas MMP-9 was highly expressed in 22 of 56 cases. Among the 15 cases analyzed by RT-PCR assay, MMP-2 and MMP-9 were expressed in 9 and 13 cases, respectively. Based on the Kaplan–Meier curve, the high expressions of MMPs were related to recurrence/regrowth (MMP-2: p < 0.001; MMP-9: p < 0.05). No significant relationship was observed between the expressions of MMPs and either age, sex, tumor location, or MIB-1 LI. Conclusions: Our study indicated that MMP-2 and MMP-9 expressions are prognostic factors predicting the recurrence of meningioma, independent of proliferative potential.

The use of mild hypothermia for patients with severe vasospasm: a preliminary report

The purpose of this study was to determine the effect of mild hypothermia on cerebral ischaemia due to severe vasospasm, which was refractory to medical and intravascular treatments and to assess the brain protection of this treatment in patients who underwent delayed aneurysm clipping after presenting with ischaemic neurological deficits. Mild hypothermia (32-34 degrees C of brain temperature) was employed in two groups: (1) Patients (Hunt and Kosnik grades I to II) who showed progressive neurological deficits due to vasospasm and did not respond to conventional therapy (Group 1) and (2) Patients who received delayed aneurysm clipping after presenting with ischaemic neurological deficits due to vasospasm (Group 2). Seven of 8 patients in both Groups showed a favorable outcome with mild hypothermia (good recovery in 5 and moderate disability in two patients). Mild hypothermia is considered to be effective on critical cerebral ischaemia due to vasospasm even after failure to response the conventional therapies and to provide brain protection in delayed aneurysm clipping.

Matrix metalloproteinase-2 expression correlates with cavernous sinus invasion in pituitary adenomas

UNLABELLED The aim of this study was to evaluate the relationship between expression of matrix metalloproteinase-2 (MMP-2) and cavernous sinus invasion in pituitary adenoma. METHOD Tissue samples from 54 pituitary adenomas were examined for expression of MMP-2 protein by immunohistochemistry. In sixteen tumors, the expression of MMP-2 mRNA was also examined by RT-PCR. Immunostaining was semiquantatively scored based on intensity (0-3) and distribution (0-3). RESULTS There were 34 women and 20 men, with a mean age of 49.9 years (range 18-76). There were 12 tumors with cavernous sinus invasion, and 42 noninvasive cases. The MMP-2 score of pituitary adenomas with cavernous sinus invasion (3.9 +/- 0.5) was significantly higher than those without invasion (2.3 +/- 0.2; P < 0.01). There was no difference in MMP-2 score between macroadenomas (3.0 +/- 0.3) and microadenomas (2.1 +/- 0.4; P > 0.05), and also, no difference between the functioning adenomas (2.8 +/- 0.3) and non-functioning adenomas (2.8 +/- 0.3; P > 0.05). We found no correlation between the MMP-2 score and the Ki-67 labeling index (r2 = -0.05; P = 0.72). MMP-2 mRNA expression was also intense in invasive pituitary adenomas and was significantly higher in invasive pituitary adenomas than those without invasion (68.2 +/- 15.3; 21.8 +/- 8.2; P < 0.05). CONCLUSION This study suggests that MMP-2 may be associated with aggressiveness and invasion in pituitary adenoma but is not related to tumor size or secretory function. MMP-2 may be a useful tool for assessing the invasive potential.

Treatment of cold injury-induced brain edema with a nonspecific matrix metalloproteinase inhibitor MMI270 in rats.

Blood-brain barrier (BBB) disruption is a critical event leading to vasogenic brain edema and secondary brain damage after cold injury-induced brain trauma. Matrix metalloproteinases (MMPs), a family of proteolytic enzymes which degrade the extracellular matrix, are implicated in BBB disruption in this model. The purpose of this study was to examine the effects of MMI270 (N-hydroxy-2(R)-[(4-methoxysulfony) (3-picolyl)-amino]-3-metylbutaneamide hydrochloride monohydrate), a synthetic nonspecific MMP inhibitor, on cold injury-induced brain edema in rats. Cold injury was induced by applying a copper probe cooled with liquid nitrogen on the parietal skull for 30 sec in 38 rats. Treatment with MMI270, a bolus injection at a dose of 30 mg/kg, was started immediately after the induction of cold injury and was continued for 24 h at a dose of 40 mg/kg/day using an intraperitoneal osmotic minipump (n = 7). In the untreated control group (n = 7), rats were administered a vehicle and implanted with a vehicle-containing osmotic pump. Two percent Evans Blue (EB) in saline (1 mL/kg) was administrated intravenously immediately after the cold injury in another group of rats, six of which were untreated and six of which were treated with MMI270 at the above dose. At 24 h after the cold injury, the brain water content and the BBB permeability to EB were determined. To assess the protective effect of MMI270 on secondary brain lesion after the cold injury, the MMI270-treated rats received a bolus injection at a dose of 30 mg/kg, followed by a continuous administration of MMI270 for 7 days at a dose of 40 mg/kg/day using an osmotic minipump (n = 6). In the untreated control group (n = 6), the rats were administered the vehicle and implanted with a vehicle-containing osmotic pump. At 7 days after cold injury, the secondary brain lesion was assessed using hematoxylin and eosin (H-E) staining. Compared with the untreated control group, treatment with MMI270 significantly reduced the brain water content in the ipsilateral core and intermediate areas (p < 0.05 and p < 0.01) and protected the BBB integrity to EB in the ipsilateral core area (p < 0.05) at 24 h after the cold injury. The secondary lesion was significantly smaller in the MMI270-treated animals compared with the untreated animals (p < 0.05) a 7 days after the cold injury. O kur results indicate that treatment with MMI270 in rats exhibits protection in acute brain edema formation and secondary brain damage by attenuating the BBB permeability after cold injury.

Determinants of drug response in camptothecin-11-resistant glioma cell lines

SummaryCamptothecin-11 (CPT-11) is a new derivative of camptothecin, a plant alkaloid antitumor agent. Previous studies indicated that antitumor activity of CPT-11 was mediated through interaction of the drug with its target enzyme, DNA topoisomerase I (topo I).To elucidate the mechanisms of CPT-11 resistance, we have characterized glioma cell lines (T98G/CPT-11, C6/CPT-11) selected from the wild types (T98G. C6) for acquired resistance to CPT-11. T98G/CPT-11 and C6/CPT-11 cells demonstrated 5.4- and 7.3-fold increases, respectively, in resistance to CPT-11.Total glutathione S-transferase (GST) and GST-p activities were similar in CPT-11-sensitive and -resistant cells. No difference in intracellular accumulation of CPT-11 was observed between CPT-11-resistant and parental cells, indicating that an alteration in the uptake was not responsible for resistance. In addition, CPT-11-resistant cell lines showed no change in the total activity of Topo I, indicating an alteration in total Topo I activity was not responsible for resistance.In contrast, significantly increased intracellular glutathione (GSH) levels were found in T98G/CPT-11 and C6/CPT-11 cells (4.3- and 2.1-fold). Furthermore, Topo I samples from T98G/CPT-11 and C6/CPT-11 cells were at least 4- and 2-fold more resistant to the inhibitory effect of the CPT-11 on the relaxation activity of Topo I than were Topo I samples from their respective parent lines. The resistance of the enzyme itself to the effects of CPT-11 may be responsible for the resistance to CPT-11. Thus, at least two distinct mechanisms have been selected for the CPT-11-resistant cells.

Technetium-99m Sestamibi Single Photon Emission Computed Tomography Findings Correlated With P-glycoprotein Expression, Encoded by the Multidrug Resistance Gene-1 Messenger Ribonucleic Acid, in Intracranial Meningiomas

The present study evaluated whether technetium-99m sestamibi (99mTc-MIBI) single photon emission computed tomography (SPECT) characteristics of intracranial meningioma are correlated with the histological malignancy, proliferative potential, and P-glycoprotein (Pgp) expression, encoded by the multidrug resistance gene-1 (MDR-1) messenger ribonucleic acid (mRNA). Twenty-one patients with intracranial meningiomas, including 17 benign and four nonbenign meningiomas, underwent 99mTc-MIBI SPECT imaging at 15 minutes (early) and 3 hours (delayed) after injection. The tumor-to-normal pituitary gland ratio was calculated on both early (ER) and delayed (DR) images. Retention index (RI) was calculated using the following formula: (DR - ER)/ER x 100%. Meningioma specimens were examined by immunohistochemistry using anti-Pgp and MIB-1 monoclonal antibody. MDR-1 mRNA expression was also investigated using reverse transcription-polymerase chain reaction assay. 99mTc-MIBI was highly accumulated and retained in the tumors. 99mTc-MIBI SPECT findings were not related to MIB-1 labeling index. 99mTc-MIBI SPECT RI of the Pgp-positive group (-9.12 +/- 22.27%) was significantly lower than that of the Pgp-negative group (28.79 +/- 22.80%) (p = 0.0016). No significant difference was seen in ER and DR between the positive and negative groups. These results show that 99mTc-MIBI may not be useful for determining proliferative potential and histological malignancy, but could predict anticancer drug resistance related to the expression of MDR-1 mRNA and its gene product Pgp in patients with intracranial meningiomas.

Incidence of Mutation and Deletion in Topoisomerase IIα mRNA of Etoposide and mAMSA–resistant Cell Lines

The efficacy of all chemotherapeutic agents is limited by the occurrence of drug resistance. To further understand resistance to topoisomerase II inhibitors, 50 sublines were isolated as single clones from parental cells by exposure to VP–16 (etoposide) or mAMSA (m–amsacrine). Subsequently, a population of cells from each subline was exposed to three–fold higher drug concentrations allowing 16 stable sublines to be established at higher extracellular drug concentration. Finally, 66 sublines were picked up. The frequency and nature of mutations in the topoisomerase II gene in the drug–selected cell lines were evaluated. In order to screen a large number of cell lines, an RNAse protection assay was developed and mismatches were observed in 13.6% of resistant cell lines (12% of resistant cell lines exposed to lower drug concentrations and 18.8% of resistant cell lines exposed to higher drug concentrations). Some of these mutations are located in vital regions of topoisomerase II (phosphorylation sites in the C–terminal or N–terminal, and nuclear localizing signal of topoisomerase II). Our findings suggest that mutations of topoisomerase II gene are an important and frequent mechanism of resistance to topoisomerase II inhibitors.

Expression of Drug Resistance Genes in VP-16 and mAMSA-selected Human Carcinoma Cells

The cell lines described in the present study were isolated as part of an effort to understand resistance to topoisomerase (topo) II inhibitors. To that end, 50 sublines were isolated from four human breast cancer cell lines, i.e., MCF‐7, T47D, MDA‐MB‐231, and ZR‐75B. As an initial step, a concentration that would be lethal to the majority of cells (IC99) was selected for both VP‐16 and mAMSA, for each cell line. The identification of an increasing number of putative drug resistance‐related proteins provided the opportunity to examine expression of the corresponding genes in the selected cell lines. Northern blot analysis revealed different responses to the selecting agents in the different cell lines. Previous studies examining expression of multidrug resistance (MDR)‐l in resistant cell lines had found undetectable levels in all cells. In the ZR‐75B sublines, increased expression of MDR‐associated protein (MRP) and canalicular multispecific organic anion transporter (cMOAT) was observed, and when the relative levels of overexpression were compared, a high correlation was found. In contrast, increased expression of MRP was observed in some of the MDA‐MB‐231 sublines, without a concomitant increase in cMOAT expression. Finally, in both T47D and MCF‐7 sublines, increased expression of cMOAT or MRP was observed infrequently, and where it occurred, was of a much smaller magnitude. In the analysis of expression of MRP, the highest levels were found in the ZR‐75B and MDA‐MB‐231 sublines, with lower levels in the MCF‐7 and T47D clones. Similarly, differences in the expression of topo IIα were observed among the sublines. Although the differences in expression appear to depend on the parental cell line from which the resistant sublines were derived, a strong correlation was observed between the expression of MRP and the levels of topo IIα. Cell lines with low levels of MRP had lower levels of topo IIα, while those with high levels of MRP maintained higher levels of topo IIα. While a reduced topo IIα level was common, there did not appear to be a compensating increase in the expression of topo IIβ or topo I or casein kinase (CK) IIα in any of the cell lines. While the possibility that such compensation could occur has been discussed and even reported in some cell lines, such an adaptation was not observed in the present study, suggesting that it is not common.

A comparison of intraarterial carboplatin and ACNU for the treatment of gliomas

BACKGROUND Intraarterial chemotherapy with carboplatin for malignant gliomas has been tried recently, but its therapeutic efficacy and toxicity have not yet been elucidated. METHODS We treated patients with malignant glioma by intraarterial chemotherapy using carboplatin, and compared the efficacy as well as the side effects with intraarterial ACNU. RESULTS Twenty patients were treated with carboplatin (300 mg/m2) and 22 patients were treated with ACNU (80-200 mg/m2). Response (complete remission+partial response) rate for carboplatin was 12.5% compared to 45% for ACNU. Despite higher response rate for ACNU, the difference in the survival curves of the two groups was not significant. Three patients who were treated with high dose (150-200 mg/m2) of ACNU developed hemiparesis and aphasia. Seven patients treated with carboplatin developed 10 incidences of neurotoxicities; two hemiparesis, one aphasia, one blindness, one visual field disturbance, three convulsions, and two developed incidences of disturbances of consciousness. CONCLUSIONS Intraarterial carboplatin was not superior to intraarterial ACNU in achieving remissions, and showed much greater tendency to produce neurotoxicities.