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.

99mTc-MIBI and 201Tl SPET in the detection of recurrent brain tumours after radiation therapy.

The purpose of this study was to evaluate whether Tc-hexakis-2-methoxyisobutylisonitrile ( Tc-MIBI) or Tl single photon emission tomography (SPET) could detect recurrent tumours in patients with previous radiation therapy for brain tumours. Dual SPET with Tc-MIBI and Tl was performed in 21 patients suspected of having recurrent brain tumours. SPET images were acquired 15 min (early) and 2 h (delayed) after injection. The ratio of the average counts for the region of interest in the lesion area and its mirror image in normal brain tissue was obtained. Early and delayed ratios were calculated. On the basis of histological and/or clinical findings, the final diagnosis was considered as recurrent tumours in 15 patients and radiation necrosis in six. Both ratios using Tc-MIBI and Tl were significantly higher in recurrent tumours than in radiation necrosis. Based on a cut-off of 5.89 of the early ratio using Tc-MIBI to distinguish between recurrent tumours and radiation necrosis, the accuracy was 90%. Based on a cut-off of 6.77 of the delayed ratio using Tc-MIBI, the accuracy was 86%. The corresponding values using cut-offs of 2.40 and 1.85 with Tl were 90% and 86%, respectively. However, within recurrent tumours, both ratios for Tc-MIBI were significantly higher than those for Tl. Early Tc-MIBI SPET may be especially useful for the detection of recurrent tumours in patients who have previously undergone radiation therapy for brain tumours.

Amygdaloid projections to ventromedial striatal subterritories in the primate.

The ventral striatum is the part of the striatum associated with reward and goal-directed behaviors, which are mediated in part by inputs from the amygdala. The ventral striatum is divided into 'shell' and 'core' subterritories which have different connectional, histochemical and pharmacological properties. Behavioral studies also indicate that subterritories of the ventral striatum are differentially involved in specific goal-directed behaviors. The amygdala is a heterogeneous structure which has multiple nuclei involved in processing emotional information. While the existence of an amygdalostriatal pathway has long been established, the relationship between amygdaloid afferents and specific subterritories of the ventral striatum is not known. In this study we operationally defined the ventromedial striatum as the region receiving cortical inputs primarily from the orbital and medial prefrontal cortex. We placed retrograde tracer injections into subregions of the ventromedial striatum of macaques monkeys to determine the relative contribution of specific amygdaloid inputs to each region. Calbindin-D28k immunostaining was used to further define the shell subterritory of the ventromedial striatum. Based on these definitions, the amygdala innervates the entire ventromedial striatum, and has few to no inputs to the central striatum. The basal and accessory basal nuclei are the major source of input to the ventromedial striatum, innervating both the shell and ventromedial striatum outside the shell. However, a restricted portion of the dorsomedial shell receives few basal nucleus inputs. Afferent inputs from the basal nucleus subdivisions are arranged such that the parvicellular subdivision projects mainly to the ventral shell and core, and the magnocellular subdivision targets the ventral shell and ventromedial putamen. In contrast, the intermediate subdivision of the basal nucleus projects broadly across the ventromedial striatum avoiding only the dorsomedial shell. The shell has a specific set of connections derived from the medial part of the central nucleus and periamygdaloid cortex. Within the shell, the dorsomedial region is distinguished by additional inputs from the medial nucleus. The ventromedial caudate nucleus forms a unique transition zone with the shell, based on inputs from the periamygdaloid cortex. Together, these results indicate that subterritories of the ventromedial striatum are differentially modulated by amygdaloid nuclei which play roles in processing olfactory, visual/gustatory, multimodal sensory, and 'drive'-related stimuli.

Incidence of mutation and deletion in topoisomerase II alpha 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.

[Chemotherapy for gliomas based on the expression levels of drug resistant genes].

Drug resistance, which often occurs during chemotherapy, is still a great obstacle to the success of human malignancy treatment. Among many possible mechanisms of drug resistance (biological, biochemical, kinetic or pharmacological), both typical and atypical multidrug-resistance (MDR) have been extensively studied. We picked up MDR-1, MXR, MRP1, MRP2, TopoII alpha, MGMT, and GST-pi as drug-resistant gene, based on experimental data and previous reports. Expression of these genes were measured in 14 malignant glioma specimens by reverse transcription polymerase chain reaction assay. We chose anticancer drugs for each patient, based on results of drug resistant gene expression to acquire good response to drugs. Though our follow-up periods are not long enough to analyze the results of our chemotherapy, 78% (7/9) of our glioma patients who were treated with our chemotherapy are free from tumor progression. The assays, which measure the expression of drug resistant genes, are necessary to allow rapid detection of the drug-sensitivity to chemotherapy in malignant glioma patients.

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)-1 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 IIalpha 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 IIalpha. Cell lines with low levels of MRP had lower levels of topo IIalpha, while those with high levels of MRP maintained higher levels of topo IIalpha. While a reduced topo IIalpha level was common, there did not appear to be a compensating increase in the expression of topo IIbeta or topo I or casein kinase (CK) IIalpha 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.

Hypophosphorylation of topoisomerase IIalpha in etoposide (VP-16)-resistant human carcinoma cell lines associated with carboxy-terminal truncation.

Topoisomerase IIalpha is a target for many chemotherapeutic agents in clinical use. To define mechanisms of resistance and regions crucial for the function of topoisomerase IIalpha, drug-resistant cell lines have been isolated following exposure to topoisomerase II poisons. Two resistant sublines, T47D-VP and MCF-7-VP, were isolated from human carcinoma cell lines following exposure to 300 or 500 ng / ml etoposide (VP-16). Cytotoxicity studies confirmed resistance to etoposide and other topoisomerase II poisons. KCl-sodium dodecyl sulfate (K-SDS) precipitation assays using intact cells showed reduced DNA-topoisomerase II complex formation following VP-16 or amsacrine (m-AMSA). RNAse protection analysis identified a deletion of 200 base pairs in the topoisomerase IIalpha cDNA of T47D-VP and rising dbl quote, left (low)AA insertion" in the topoisomerase IIalpha cDNA of MCF-7-VP. Reduced topoisomerase IIalpha mRNA and protein levels were observed in both cell lines. It was somewhat surprising to find that nuclear extracts from T47D-VP and MCF-7-VP cells had comparable topoisomerase II activity to that of parental cells. Analysis of the extent of phosphorylation demonstrated that topoisomerase IIalpha from the resistant cells was relatively hypophosphorylated compared to that of parental cells. In these cell lines, hypophosphorylation secondary to loss of a portion of the C-terminal domain of topoisomerase IIalpha mediated the restored activity, despite a fall in topoisomerase IIalpha mRNA and protein, and this resulted in cross resistance to topoisomerase II poisons.

The efficacy and safety of transvenous embolisation in the treatment of intracranial dural arteriovenous fistulas.

To evaluate the role of transvenous embolisation including its efficacy and safety in the treatment of intracranial dural arteriovenous fistulas (DAVFs), we retrospectively analysed seven cases of intracranial DAVFs treated with transvenous embolisation in combination with arterial embolisation. Four DAVFs were in the cavernous sinus, two in the transverse-sigmoid sinus, and one in the inferior petrosal sinus. The transarterial and transvenous embolic agents included fibred platinum coils (FPC) and interlocking detachable coils (IDC). In all patients, the transarterial embolisation alone had failed to cure the DAVFs. After the combined transvenous embolisation, the anatomical cure was proven in five patients, and all patients were clinically cured. There were no complications in any patient. In conclusion, the transvenous embolisation is a useful and safe approach in the management of intracranial DAVFs.

Comparison of 99Tcm-MIBI with 201Tl chloride SPET in patients with malignant brain tumours.

The purpose of this study was to evaluate the usefulness of 99Tcm-MIBI accumulation for the differentiation of histological diagnosis of malignant brain tumours in comparison with the findings obtained using 201Tl chloride. A total of 25 patients with malignant brain tumours were investigated. The histological categories of tumours included glioblastoma multiforme (n = 5), anaplastic astrocytoma (n = 4), malignant lymphoma (n = 5), and metastatic tumour (n = 11). Simultaneous dual single photon emission tomography (SPET) images with 99Tcm-MIBI and 201Tl were acquired 15 min (early) and 2 h (delayed) after injection, and the early ratio, delayed ratio and retention index were measured. The new indices 201Tl/99Tcm-MIBI ratios and 201Tl/99Tcm-MIBI retention index were also calculated. With respect to the histological type, a higher retention index using 99Tcm-MIBI was noted in glioblastoma multiforme compared with metastatic tumour. Higher values of both ratios using 201Tl were noted in glioblastoma multiforme compared to metastatic tumour. The value of the delayed ratio obtained using 201Tl was higher in glioblastoma multiforme than in anaplastic astrocytoma, and the value was also higher in malignant lymphoma than in metastatic tumour. The 201Tl/99Tcm-MIBI early ratio of glioblastoma multiforme was significantly higher than that of metastatic brain tumour. The 201Tl/99Tcm-MIBI retention index of malignant lymphoma was significantly higher than that of glioblastoma multiforme. In the histological type of tumour, 99Tcm-MIBI is not superior to 201Tl, but the combined indices using 201Tl/99Tcm-MIBI may add new information about differential diagnosis.

Treatment of arteriovenous malformation of the brain--preliminary experience.

With the availability of new techniques, such as intravascular embolisation and radiosurgery, the therapeutic approach to arteriovenous malformations (AVMs) of the brain has recently been modified. The present study reports the authors, experiences in treating AVMs over the past 13 years. Spetzler-Martin grading of AVMs was I and II in 19 cases, III in 12, IV in 5 and V in 1 case. Four therapeutic regimens were utilised: surgical resection alone, embolisation and resection, and radiosurgery alone or after surgical resection. Generally, for low-grade AVMs (Spetzler-Martin grades I, II and III), the therapeutic choice was surgical resection in 27 cases, in combination with pre-operative embolisation in two of these patients. Two cases received radiotherapy only and one case received radiosurgery after embolisation, while one case was treated conservatively. Of the five cases of grade IV, four required surgical treatment, whereas the fifth case was treated conservatively. Favourable results (good recovery and moderate disability) were obtained in 96% of the low-grade AVMs as compared with the high-grade AVMs (66%) that had a poor outcome (due to primary brain damage resulting from haemorrhage at the onset in three cases and due to postoperative re-bleeding in one case). This report summarises preliminary experience in treating intracranial AVMs by surgical resection, intravascular embolisation and radiotherapy. Good therapeutic results can be expected by combining these therapeutic modalities.

Expression of DNA topoisomerase I, IIalpha, and IIbeta in human brain tumors.

We investigated the expression of DNA topoisomerase I (Topo I), IIalpha (Topo IIalpha), and IIbeta (Topo IIbeta) mRNA using reverse transcription-polymerase chain reaction (RT-PCR) assay in 31 human brain tumors, and examined the relationship between DNA topoisomerase mRNA expression and Topo IIalpha and MIB-1 positive index (PI) as a cell proliferation marker. Topo IIalpha mRNA was expressed in 11 of 31 cases, and Topo I and IIbeta were each expressed in 18 of 31 cases. A significant correlation was seen between the MIB-1 PI and Topo IIalpha PI (P < 0.001). The cases with overexpression of Topo IIalpha mRNA had significantly high MIB-1 and Topo IIalpha PI (P < 0.0001). There was no significant correlation between Topo I and IIbeta mRNA expression and MIB-1 PI. We concluded that it was useful as a cell proliferation marker to analyze the expression of Topo IIalpha mRNA using RT-PCR in human brain tumors.

Visualization of the motor activation area using SPECT in neurosurgical patients with lesions near the central sulcus.

UNLABELLED: The purpose of this study was to visualize the motor area related to finger movement and a fist-making task using SPECT in patients with lesions near the central sulcus. METHODS: Eleven patients (9 with a brain tumor, 1 with cerebral infarction, and 1 with an arteriovenous malformation) were investigated. The first intravenous injection of 99mTc-ethyl cysteinate dimer (ECD) for the motor activation SPECT images was administered 2 min after completion of the fist-making task with the hand contralateral to the brain lesion. The movement was stopped 2 min after injection, and activation SPECT was performed. After the scan, the second dose of 99mTc-ECD was injected into resting patients, and a second set of SPECT images was acquired. The first set of images was subtracted from the second set to obtain control images. Regions of interest were set bilaterally on the sensorimotor hand area; the supplementary motor area; the frontal, temporal, and occipital lobes; and the cerebellar hemispheres. The results of activation SPECT were expressed as positive or negative for a high-count area, and the regional percentage change for activation images relative to resting images was calculated. RESULTS: Visual assessment of activation images was positive in 9 patients for the sensorimotor hand area and 7 patients for the supplementary motor area. The regional percentage change between activation and resting images for the high-count areas was 19.7% for the sensorimotor hand area and 18.2% for the supplementary motor area. Both values were significantly higher than those for other areas (P<0.05). CONCLUSION: Motor activation SPECT using a 99mTc-ECD split-dose method is easy to perform and may be helpful for presurgical visualization and identification of the sensorimotor hand area or the supplementary motor area.

Protective effect of mild hypothermia on symptomatic vasospasm: a preliminary report.

Mild hypothermia (32-34 degrees C of brain temperature) was used for brain protection in patients with progressive ischemic neurological deficits associated with severe cerebral vasospasm and who did not respond to medical treatment or intravascular angioplasty. Results showed that 2 of 3 patients in Hunt & Kosnik grade I to III and 2 patients who underwent delayed operation on day 5 and 9 each and had ischemic neurological deficits made good recovery with this treatment. Favourable outcome was obtained in 4 of 9 patients in grade IV and V. Mild hypothermia is thought to provide brain protection in critical ischemia due to severe cerebral vasospasm and can lengthen therapeutic time to employ angioplasty and intraarterial Papaverin infusion.

DNA topoisomerase IIalpha protein and mRNA expression in intracranial meningiomas.

We examined the expression of DNA topoisomerase IIalpha (Topo IIalpha) immunohistochemically using a monoclonal antibody and compared its proliferative potential [MIB-1 labeling index (LI)] and recurrence to verify the possible influence of Topo IIalpha on the progress of meningiomas. The reverse transcription-polymerase chain reaction (RT-PCR) assay was also performed to evaluate the expression of Topo IIalpha mRNA. Formalin-fixed, paraffin-embedded tissue sections of 52 meningiomas (18 meningothelial types, 16 fibrous types, 4 transitional types, 4 psammomatous types, 1 angiomatous type, 1 secretory type, 5 atypical types, and 3 anaplastic types) were used for immunostaining. The Topo IIalpha labeling index (LI) was 1.4 +/- 1.9% (mean +/- SE) in benign meningiomas and 4.5 +/- 1.6% in atypical or anaplastic meningiomas, representing significant differences between them (P < 0.0001). RT-PCR assay revealed that Topo IIalpha mRNA expression was associated with Topo IIalpha LI. A significant correlation was seen between Topo IIalpha LI and MIB-1 LI (r = 0.517; P < 0.01). Recurrence was significantly more frequent in patients with more than 1.5% of Topo IIalpha LI than in those with 1.5% or less (P < 0.005). In conclusion, Topo IIalpha protein and mRNA expression correlated with clinical malignancy and the potential for predicting the regrowth of meningiomas.

Proliferative potential of malignant glioma cells before and after interstitial brachytherapy.

The viability of tumor cells in radionecrotic tissue after interstitial brachytherapy (BRTX) was evaluated using immunohistochemical markers of proliferative potential in primary and recurrent tumors. Tumor specimens from 30 patients with malignant gliomas (14 anaplastic astrocytomas, 16 glioblastomas) taken before and after BRTX were examined using MIB-1 monoclonal antibody. Histological examination of specimens obtained by craniotomy or stereotactic biopsy after BRTX revealed tumor recurrence in 18 patients and radionecrosis in 12 patients including two with pure radionecrosis and 10 with a mixture of both tumor and radionecrosis. The MIB-1 index of the tumors with radionecrosis was 7.6 +/- 5.5%, and that of the primary tumors was 17.0 +/- 11.2%, showing a significant difference (p < 0.05). There was no significant difference between the MIB-1 index of the primary tumors with local recurrence after BRTX and the primary tumors which underwent radionecrosis. Although morphologically viable tumor cells were found in the radionecrotic tissue, BRTX causes a reduction in the proliferative potential of these tumor cells.

[A clinical analysis of patients with primary intracranial germinomas: the relationship between proliferative potential and recurrence].

We analyzed clinically 30 germinomas (22 pure germinomas and 8 germinomas with syncytiotrophoblastic giant cell (STGC) and also investigated the proliferative potential of 18 germinomas immunohistochemically using MIB-1 monoclonal antibody in formalin-fixed paraffin-embedded sections. The majority of patients responded favorably and completely to the treatment. Seven patients (four had germinomas with STGC and the other three had pure germinomas) suffered recurrence after a complete response to radiotherapy and/or chemotherapy. The patients with germinomas with STGC experienced recurrence significantly more often than those with pure germinomas (P < 0.01). Three of the patients died. The MIB-1 indexes of the germinomas ranged from 18% to 80%. The average MIB-1 index of pure germinomas is 58.5 +/- 17.3%, that of germinomas with STGC is 45.3 +/- 17.1%. There was no significant difference between the two groups. The MIB-1 indexes of 3 recurrent cases were 64.3%, 51.9%, and 20.8%, respectively. There were no significant differences in the MIB-1 indexes between nonrecurrent cases and recurrent ones.

Increased phosphorylation of DNA topoisomerase II in etoposide resistant mutants of human glioma cell line.

The efficacy of the epipodophyllotoxins VP-16 and VM-26 is limited by the occurrence of drug resistance in the tumor cell population. Cellular insensitivity to drugs that stabilize the cleavable complex is frequently expressed as multidrug resistance (MDR). In some cell lines, overexpression of MDR-1/P-glycoprotein or the multidrug resistance associated protein (MRP) has been demonstrated and implicated as the mechanism of resistance. Typically, these cells have reduced drug accumulation, secondary to increased drug efflux. In other cell lines, an atypical MDR phenotype has been identified, with the predominant mechanism of resistance shown to be qualitative and/or quantitative changes in the levels and activity of topoisomerase II. For VP-16, increased expression of MDR-1 or MRP and alterations in topoisomerase II have been shown to confer tolerance. To further understand resistance to VP-16, T98G-VP(1000) was initially isolated as a single clone from parental cell, T98G, by exposure to VP-16. Subsequently, a population of cells from this subline was exposed to three-fold higher drug concentration allowing stable sublines to be established at higher extracellular drug concentration. Characterization of the resistant sublines demonstrates the adaptation that occurs with advancing drug concentrations during in vitro selections. Reduced topoisomerase II mRNA and protein levels were observed in the initial isolate. This reduction was accompanied by a decrease in topoisomerase II activity and cellular growth rate and was associated with 47-fold resistance to topoisomerase II poisons. With advancing resistance, MRP expression increased, with increased VP-16 efflux and reduced accumulation. This adaptation allowed for partial restoration of topoisomerase II activity secondary to increased expression and hyperphosphorylation, with a resultant increase in growth rate. In this cell line, hyperphosphorylation coincided with increased casein kinase II mRNA protein levels, without increased PKC protein levels, suggesting a role for this kinase in the acquired hyperphosphorylation. In this cell line, hyperphosphorylation mediated the increased activity despite a fall in topoisomerase II protein levels secondary to an acquired 615 bp deletion in one topoisomerase II allele, which resulted in reduced protein levels. In this subline, high levels of resistance were attained as a result of synergism between the reduced topoisomerase II levels and MRP overexpression. These studies demonstrate how cellular adaptation to increasing drug pressure occurs and how more than one mechanism can contribute to the resistant phenotype when increasing selecting pressure is applied. Reduced expression of topoisomerase II is sufficient to confer substantial resistance early in the selection process, with synergy from additional mechanisms helping to confer high levels of resistance.

[Incidence of mutation and deletion in topoisomerase II mRNA of etoposide and m-AMSA resistant cell lines].

The efficacy of all chemotherapeutic agents is limited by the occurrence of drug resistance. To further understand resistance to topoisomerase (topo) II inhibitors, 50 sublines were isolated as single clones from parental cells by exposure to ETP or m-AMSA. 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. The frequency and nature of mutations in topo II in the drug selected cell lines have been evaluated. In order to screen a large number of cell lines, an RNase protection assay was developed. Fragments covering the entire coding sequence of topo II was isolated after PCR amplification and subcloned in pGEM3Z vector. Using this approach, mismatches was 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). Our findings suggest that mutations of topo II gene seem to be an important and frequent mechanism of resistance to topo II inhibitors.

Proliferative potential and apoptosis in rat glioma cell lines after hyperthermia.

The proliferative potential of cultured rat glioma cells (C6 and 9L) was evaluated after hyperthermia using immunohistochemical staining with bromodeoxyuridine (BrdU) and Ki-67 monoclonal antibodies. Apoptosis was assessed by in situ end-labeling of deoxyribonucleic acid breaks. Both BrdU and Ki-67 labeling indexes decreased with increasing hyperthermia time. The decrease of the Ki-67 labeling index was not as great as that of the BrdU labeling index. The number of apoptotic cells increased with time after hyperthermia. These results indicate that the antitumor effect of hyperthermia may reflect the induction of apoptosis in the cells within the cell cycle, and the resultant reduction of the proliferative potential of surviving cells, especially in the S phase.