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To analyze the effect of polychlorinated biphenyl (PCB) 118 on fish bone metabolism, we examined osteoclastic and osteoblastic activities, as well as plasma calcium levels, in the scales of PCB (118)-injected goldfish. In addition, effect of PCB (118) on osteoclasts and osteoblasts was investigated in vitro. Immature goldfish, in which the endogenous effects of sex steroids are negligible, were used. PCB (118) was solubilized in dimethyl sulfoxide at a concentration of 10 ppm. At 1 and 2 days after PCB (118) injection (100 ng/g body weight), both osteoclastic and osteoblastic activities, and plasma calcium levels were measured. In an in vitro study, then, both osteoclastic and osteoblastic activities as well as each marker mRNA expression were examined. At 2 days, scale osteoclastic activity in PCB (118)-injected goldfish increased significantly, while osteoblastic activity did not change significantly. Corresponding to osteoclastic activity, plasma calcium levels increased significantly at 2 days after PCB (118) administration. Osteoclastic activation also occurred in the marker enzyme activities and mRNA expressions in vitro. Thus, we conclude that PCB (118) disrupts bone metabolism in goldfish both in vivo and in vitro experiments. © 2012 The Author(s).

We recently demonstrated that prostaglandin E2 PG¿ increases osteoclastic activity and induces bone resorption in both in vitro and in vivo experiments using goldfish. In the fish reproductive period, the plasma calcium (Ca) level in female teleosts increases remarkably to make vitellogenin, which is a major component of egg protein and a Ca-binding protein. In this period, however, there is no reported relationship between PGE2 and Ca metabolism in fish. To clarify the Ca metabolism in fish reproduction, we examined plasma PGE2 and Ca levels and measured tartrate-resistant acid phosphatase (TRAP) activities as an indicator of osteoclastic activity in goldfish. Plasma PGE2 levels in the reproductive stage significantly increased as compared with those in non-reproductive stages. Also, both plasma Ca and TRAP increased in the reproductive stage. Significant positive correlations were recognized between plasma Ca and the gonad somatic index (r=0.81, p

We have developed an immortalized oral epithelial cell line, ROE2, from fetal transgenic rats harboring temperature-sensitive simian virus 40 large T-antigen gene. The cells grew continuously at either a permissive temperature of 33°C or an intermediate temperature of 37°C. At the nonpermissive temperature of 39°C, on the other hand, growth decreased significantly, and the Sub-G1 phase of the cell cycle increased, indicating that the cells undergo apoptosis at a nonpermissive temperature. Histological and immunocytochemical analyses revealed that ROE2 cells at 37°C had a stratified epithelial-like morphology and expressed cytokeratins Krt4 and Krt13, marker proteins for oral nonkeratinized epithelial cells. Global-scale comprehensive microarray analysis, coupled with bioinformatics tools, demonstrated a significant gene network that was obtained from the upregulated genes. The gene network contained 16 genes, including Cdkn1a, Fos, Krt13, and Prdm1, and was associated mainly with the biological process of skin development in the category of biological functions, organ development. These four genes were validated by quantitative real-time polymerase chain reaction, and the results were nearly consistent with the microarray data. It is therefore anticipated that this cell line will be useful as an in vitro model for studies such as physiological functions, as well as for gene expression in oral epithelial cells.

Heat stress induces complex cellular responses, and its detailed molecular mechanisms still remain to be clarified. The objective of this study was to investigate the molecular mechanisms underlying cellular responses to mild hyperthermia (MHT) in normal human fibroblastic (NHF) cells.Cells were treated with MHT (41°C, 30 min) and then cultured at 37°C. Gene expression was determined by the GeneChip® system and bioinformatics tools.Treatment of the NHF cell lines, Hs68 and OUMS-36, with MHT did not affect the cell viability or cell cycle. In contrast, many probe sets were differentially expressed by1.5-fold in both cell lines after MHT treatment. Of the 1,196 commonly and differentially expressed probe sets analysed by k-means clustering, three gene clusters, Up-I, Down-I and Down-II, were observed. Interestingly, two gene networks were obtained from the up-regulated genes in cluster Up-I. The gene network E contained DDIT3 and HSPA5 and was mainly associated with the biological process of endoplasmic reticulum stress, while the network S contained HBEGF and LIF and was associated with the biological process of cell survival. Eighteen genes were validated by quantitative real-time polymerase chain reaction, consistent with the microarray data, in four kinds of NHF cells.Common genes that were differentially expressed and/or acted within a gene network in response to MHT in NHF cells were identified. These findings provide the molecular basis for a further understanding of the mechanisms of the MHT response in NHF cells.

Ultrasound (US) has been shown to induce apoptosis and cell lysis in cancer cells. In this study, we report on the potential of using Sonazoid, a new echo-contrast agent, that is chemically more stable and US-resistant in combination with US in cancer therapy. The biological effects and their mechanisms in the presence or absence of ultrasonic exposure in vitro were investigated. In addition, the impact of the agent on the expression levels of genes responsive to US was studied using global-scale microarrays and computational gene expression analysis tools. Our results show that the combination led to enhanced cell killing in the presence of 1 MHz acoustic field. The apoptosis induction was shown to be mediated by the mitochondrial pathway. The occurrence of US-induced DNA damage was also observed. Despite these findings, the agent at concentrations similar to those clinically used can be considered as well tolerated. Furthermore, Sonazoid enhanced expression of genes that related to apoptosis and are responsive to US, although it alone had almost no effect. These results indicate the potential of Sonazoid for US contrast enhancement as well as the possibility of its use in US-aided therapies.

To determine the clinical and hisotological efficacy and toxicities of induction chemotherapy with docetaxel (DOC) and nedaplatin (CDGP) for oral squamous cell carcinoma (OSCC) in the preoperative setting. A total of 30 patients with locally advanced but operable OSCC were enrolled. Combination induction chemotherapy consisted of DOC 60 mg/m2 followed by CDGP 100 mg/m2. All patients received one cycle of chemotherapy. In the clinical assessment, ten patients achieved partial response for an overall response rate of 33.3% (95% CI, 16.4–50.2%). Histological assessment of surgical specimens showed an overall response rate of 56.6% (95% CI, 38.9–74.3%). Although severe neutropenia was observed in 90% of patients, only one patient (3.3%) experienced severe infection. Toxicities associated with this regimen did not interfere with planned radical surgery. A single cycle of preoperative combination chemotherapy with DOC and CDGP showed moderate histological activity with an acceptable safety profile for the planned radical surgery. Further studies testing more cycles before surgery might be more appropriate.

著者人数: 26名, 資料番号: PA0910135000

In this study, to better understand the molecular mechanism underlying cellular responses to mild hyperthermia, we investigated gene expression patterns and genetic networks in human myelomonocytic lymphoma U937 cells using high-density oligonucleotide microarrays and computational gene expression analysis tools. The cells were incubated at 41 degrees C for 30 min (mild hyperthermia treatment) and then at 37 degrees C for 0-6 h. Although the mild hyperthermia treatment of the cells did not induce apoptosis, significant increases in the protein expression levels of heat shock proteins (HSPs), namely, Hsp27, Hsp40 and Hsp70, were observed following the activation of heat shock factor-1. Of the 22,283 probe sets analyzed, 423 probe sets were up-regulated and 515 probe sets were down-regulated by >1.5-fold in the cells 3 h post-treatment. Computational gene network analysis demonstrated that the significant genetic network A that contained many HSPs such as DNAJB1, HSPA1A, and HSPA1B was associated with cellular function and maintenance, post-transcriptional modification, or protein folding. Moreover, the significant genetic network B whose core contained v-myc myelocytomatosis viral oncogene homolog (MYC) was associated with cell morphology, cell cycle, and cellular development. The expression levels of nine selected genes were comparable to those determined by microarray analysis with real-time quantitative PCR assay. The present results indicate that mild hyperthermia affects the expression of a large number of genes and provides additional novel insights into the molecular basis of mild hyperthermia in cells.

We describe a rare case of systemic vascular malformation not classified as belonging to vascular malformationsyndrome.A 37-year-old woman was referred to our clinic because of a painless swelling on the left side of the tongue.99mTc-HSAD scintigraphy showed multiple sites of abnormal pooling in the right parotid gland, the right massetermuscle, the dorsum, the right anterior region of the thigh, the left gluteal region, the left knee, and the lumbarregion. The patient was given a diagnosis of a non-syndoromic multiple vascular malformation because vascularmalformation syndrome was ruled out by the results of further systemic examinations. The lesion in the tonguewas extirpated and pathologically diagnosed to be a venous malformation. The lesion in the parotid gland andother systemic lesions have not increased in size 3 years 6 months after the tongue operation.

In osteoclastic activity during space flight as well as hind limb unloading by tail suspension, inconsistent results have been reported in an in vivo study. The bone matrix plays an important role in the response to physical stress. However, there is no suitable in vitro co-culture system of osteoblasts and osteoclasts including bone matrix. On the other hand, fish scale is a calcified tissue that contains osteoblasts, osteoclasts, and bone matrix, all of which are similar to those found in human bones. Recently, we developed a new in vitro model system using goldfish scale. This system can detect the activities of osteoclasts and osteoblasts with tartrate-resistant acid phosphatase and alkaline phosphatase as the respective markers and precisely analyze the co-relationship between osteoblasts and osteoclasts. Using this system, we analyzed the bone metabolism under various degrees of acceleration (0.5-, 1-, 2-, 4-, and 6-G) by vibration with a G-load apparatus. After loading for 5 and 10 min, the scales were incubated for 6 and 24 h. The osteoblastic and osteoclastic activities were then measured. The osteoblastic activities gradually increased corresponding to 1-G to 6-G acceleration. In addition, ER mRNA expression was the highest under 6-G acceleration. On the other hand, the osteoclastic activity decreased at 24 h of incubation under low acceleration (0.5- and 1-G). This change coincided with TRAP mRNA expression. Under 2-G acceleration, the strength of suppression in osteoclastic activity was the highest. The strength of the inhibitory action under 4- and 6-G acceleration was lower than that under 2-G acceleration. In our co-culture system, osteoblasts and osteoclasts in the scale sensitively responded to several degrees of acceleration. Therefore, we strongly believe that our in vitro co-culture system is useful for the analysis of bone metabolism under loading or unloading.

This report is of a 60-year-old woman with mucosa-associated lymphoid tissue lymphoma of the palate. A moderate dose of radiotherapy and anti-CD20 monoclonal antibody chemotherapy were administered after tumour resection. There was no sign of recurrence 14 months after treatment.

Basaloid squamous cell carcinoma was first reported by Wain et al. in 1986 and has recently been designated a variant of squamous cell carcinoma because of its high potential for distant metastasis. We report a case of basaloid squamous cell carcinoma arising in the floor of the mouth in a 74-year-old man. The proliferative activities of this case and another case previously reported by our department were compared with those of 21 cases of poorly differentiated squamous cell carcinoma. The cases of basaloid squamous cell carcinoma showed high reactivity on Ki-67 immunohistochemical staining. This finding reflects the high malignancy of this disease and suggests that Ki-67 immunohistochemical staining is useful for diagnosis. Radiotherapy was very effective against the primary lesion in our two cases, but both had lung metastasis. In our patient, lung lesions were controlled by chemotherapy, but further investigations are needed to establish standard treatment, including systemic chemotherapy.

We evaluated the prognostic significance of p34cdc2 expression in 50 tongue squamous cell carcinomas (SCC) using immunohistochemical methods. The p34cdc2 protein was expressed in 33 cases of 50 tumor tissues (66.0%), compared with 15 cases of 42 controlled epithelia (35.7%). The expression of p34cdc2 was significantly correlated with the histological grade of tongue carcinoma (P

OBJECTIVE This study was performed to clarify the usefulness of Dextran magnetite (DM) for the oral cancer hyperthermia. METHODS Tumors were induced in golden hamster tongue by 9,10-dimethyl 1-1,2-benzanthracene (DMBA) application. DM suspension was locally injected into the tumor-bearing tongue and tongues were heated up to 43.0-45.0 degrees C, by AC magnetic field of 500 kHz. RESULTS The average time taken for the temperature to rise to 43.0 degrees C or above was 162 s (n = 17) at the margin of the tumor and 420 s (n = 17) at the center of the tumor. According to the tumor volume, the time required for an increase in the central temperature of tumor to 43.0 degrees C tended to be prolonged. Both temperatures could be maintained at approximately 43.0-45.0 degrees C for 30 min. The inhibition of the growth of tongue carcinoma in the four-time heating group was significantly greater than in the control group (P < 0.01). Moreover, the survival rate was significantly higher in the heated groups than in the control group (P < 0.01). Histological examination revealed a brown uniform DM accumulation at the stroma in the margin of the tumors. Many of tumor cells disappeared at the site adjacent to this accumulation. CONCLUSION These results strongly suggest the usefulness of this local hyperthermic system in the oral region that is accessible to this treatment.

The possibility of dextran magnetite complex (DM) as a new hyperthermic material was examined in this study. DM suspension of 56 mg ml(-1) iron concentration was locally injected into the normal tongue of golden hamster. DM injected tongues were heated by 500 kHz alternating current (AC) magnetic field and its serial changes in temperature were recorded at 30-s intervals. The temperature of DM injected tongue was maintained at about 43.0-45.0 degrees C for 30 min by changing the AC magnetic field intensity. While temperature elevations of the contralateral tongue and the rectum were only of minor degree. In experiment on the extent of heating area, there was correlation between volume of black stain area and amount of the injected DM suspension (Y = - 18.1 + 1.94X, r = 0.931, P < 0.0001, n = 9 ). Histological examination after heating revealed brown uniform DM accumulation in the connective tissue between fibers of the tongue muscle. Except for vascular dilatations, no tissue damage was seen in the heated tongue. Thus, DM which has the possibility of selective and uniform heating in local hyperthermia might be useful for oral cancer therapy.

Thrombospondin-1 (TSP-1) is a 450 kd glycoprotein synthesized and incorporated into the extracellular matrix by numerous cell types and reported to suppress tumor growth and progression by its inhibition of angiogenesis. In order to clarify the biological role of TSP-1 and determine its clinicopathological significance in oral squamous cell carcinomas (SCCs), we identified TSP-1 protein expression in 54 oral SCCs by immunohistochemistry and correlated it with microvessel density (MVD), clinicopathological features and patient's survival. Thirty-two out of 54 carcinomas (59%) were identified as having a low level of TSP-1 expression (TSP-1-L), and 22/54 (41%) carcinomas identified as having a high level of TSP-1 expression (TSP-1-H). The MVD counts (mean±S.D.=9.0±4.9) in TSP-1-H tumors was significantly lower than that (mean±S.D.=12.7±4.4) in TSP-1-H tumors (P=0.0065). The level of TSP-1 expression was not correlated with T category and histologic grade, but inversely correlated with the pattern of tumor invasion (P=0.0136) and with lymph nodal status (P=0.0119). Furthermore, Kaplan–Meier analysis showed that the 5-year survival rate of TSP-1-H group was significantly higher than that of TSP-l-L group. Our results suggested that TSP-1 expression exerts an inhibitory effect on tumor vascularity, and that it has value in assessment of aggressiveness and prognosis of oral SCCs.

1980年4月から1997年3月までの17年間に, 富山医科薬科大学医学部歯科口腔外科において診断された109例の歯原性腫瘍について, 1992年のWHO分類に基づき臨床的検討を行い以下の結果を得た。1.組織型別症例数では, 集合性歯牙腫51例 (46.8%) , 次いでエナメル上皮腫35例 (32.1%) , 複雑性歯牙腫17例 (15.6%) であり, 残りの6例 (5.4%) は4例が良性腫瘍, 2例が悪性腫瘍であった。2.性別頻度では男女比1: 0.9であった。初診時年齢は6歳から71歳に分布しており, 平均年齢は26.7歳であった。症例全体の約70%が30歳以下であった。3.主訴のほとんどが腫脹 (35.8%) あるいは歯の萠出異常 (25.7%) であった。エナメル上皮腫では, 約70%が腫脹であった。4.発生部位では, 43例 (39.4%) が上顎, 66例 (60.6%) が下顎であった。エナメル上皮腫の下顎臼歯部, 集合性歯牙腫の上顎前歯部, 複雑性歯牙腫の下顎臼歯部における好発傾向が認められた。

Dextran magnetite complex (DM) is a colloidal suspension of subdomain particles that efficiently generates heat when exposed to an alternating current (AC) magnetic field. Inductive heating using DM has attracted attention as a new method that can overcome various problems associated with currently available techniques for local hyperthermia. This study was performed to clarify the usefulness of DM for oral cancer hyperthermia.DM suspension was locally injected into the normal tongue and the tumor-bearing tongue of golden hamsters. The tongues were heated to 43.0-45.0°C by exposure to an AC magnetic field of 500 kHz. The following results were obtained:(I) Experiment on heating of normal tonguea) When DM was injected into the tongue in concentrations of 112, 56, and 42mg (as Fe/ml), the tongue temperature reached 43.0°C at 60, 120, and 570 seconds after treatment, respectively.b) By injecting DM in a concertration of 56mg Fe/ml and changing the intensity of the AC magnetic field, the temperature in the tongue could be maintained at 43.0-45.0°C for 30 minutes. Furthermore, the temperature rise in the rectum, used as control, was only 0.4°C during this period.c) Histological examination revealed brown uniform DM accumulation in the connective tissue between fibers of the tongue muscle.(II) Experiment on heating of tumor-bearing tongued) The central and marginal temperature of the tumors reached 43.0°C at 180 and 540 seconds, respectively, and both temperatures were maintained at 43.0-45.0°C for 30 minutes.e) An increase in tumor volume was associated with prolongation of the time required for the central temperature of the tumor to rise to 43.0°C.f) Histological examination revealed the brown uniform accumulation of DM in the stroma at the tumor margin.These results suggest that this local hyperthermic system may be useful in the oral region, where the tumor can be directly observed and palpated.

In squamous cell carcinomas of the oral cavity, spinal accessory lymph node (SALN) metastasis develops infrequently, and the prognosis of the patients with SALN metastasis is extremely poor.In this paper, patients with SALN metastasis of squamous cell carcinomas of the oral cavity were studied retrospectively. The SALN metastasis was histopathologically confirmed in 9 (5.0%) of 179 patients who underwent radical or modified radical neck dissection. The primary sites of their tumors were the tongue in 6 patients, the lower gingiva in 2, and the buccal mucosa in 1. The risk of SALN metastasis was related to the tumor spread into the oropharynx and the differentiation of the tumor, but not related to the T stage. In the patients with SALN metastasis, the mean total number of involved nodes on the affected side of the neck was 9.4. All of these patients had metastases in multiple groups of neck lymph nodes, and the majority had metastases in the upper and middle jugular groups of nodes. The mean number of involved SALNs was 2.0, and the mean size of these nodes was 0.52 cm in diameter.The outcome of treatment in the patients with SALN metastasis was poor: 1 patient was alive with no evidence of disease, and 8 patients died of disease with an average duration of survival of 8.0 months after neck dissection.To improve the prognosis of patients with SALN metastasis, both postoperative irradiation to the neck and intensive adjuvant chemotherapy should be employed.

Multiple primary malignancies (MPMs) were found in 55 (9.3%) out of 591 patients with oral cancer treated between 1972 and 1994 at Chiba Cancer Center Hospital. There were 43 men and 12 women. The mean age of the patients with MPM was 63.4 years, with a range of 42 to 87 years. MPMs occurred frequently in patients with cancers of the upper gingiva (15.4%) and floor of the mouth (14.9%). Additional malignancies developed in the digestive tract (59.3%) and respiratory system (23.7%). Thirty malignant tumors arose before oral cancer and 29 after oral cancer, and 18 MPMs developed synchronously (within 1 year). Among 29 malignancies diagnosed during follow-up of oral cancer, 11 were asymptomatic, 16 symptomatic, and 2 unknown whether they were asymptomatic or symptomatic. In a few patients, oral malignancies were treated palliatively due to other MPMs. The prognosis of patients with MPMs was poor: 9 died of oral cancer, and 12 died of other subsequent cancers. During both treatment and follow-up of oral cancer, synchronous and subsequent primary malignancies should be borne in mind to facilitate early detection.

Using our original in vitro assay system with goldfish scales, we examined the direct effect of prostaglandin E₂ (PGE₂) on osteoclasts and osteoblasts in teleosts. In this assay system, we measured the activity of alkaline phosphatase (ALP) and tartrate-resistant acid phosphatase (TRAP) as respective indicators of each activity in osteoblasts and osteoclasts. ALP activity in scales significantly increased following treatment at high concentration of PGE₂(10⁻⁷ and 10⁻⁶ M) over 6 hrs of incubation. At 18 hrs of incubation, ALP activity also significantly increased in the PGE₂ (10⁻⁹ to 10⁻⁶ M)-treated scale. In the case of osteoclasts, TRAP activity tended to increase at 6 hrs of incubation, and then significantly increased at 18 hrs of incubation by PGE₂ (10(-7) to 10⁻⁶ M) treatment. At 18 hrs of incubation, the mRNA expression of osteoclastic markers (TRAP and cathepsin K) and receptor activator of the NF-κB ligand (RANKL), an activating factor of osteoclasts expressed in osteoblasts, increased in PGE₂ treated-scales. Thus, PGE₂ acts on osteoblasts, and then increases the osteoclastic activity in the scales of goldfish as it does in the bone of mammals. In an in vivo experiment, plasma calcium levels and scale TRAP and ALP activities in the PGE₂-injencted goldfish increased significantly. We conclude that, in teleosts, PGE₂ activates both osteoblasts and osteoclasts and participates in calcium metabolism.

Local hyperthermia (HT) for various types of malignant tumors has shown promising antitumor effects. To confirm the detailed molecular mechanism underlying cell death induced by HT, gene expression patterns and gene networks in human oral squamous cell carcinoma (OSCC) cells were examined using a combination of DNA microarray and bioinformatics tools. OSCC HSC-3 cells were treated with HT at 44˚C for 90 min or mild hyperthermia (MHT) at 42˚C for 90 min, followed by culturing at 37˚C for 0-24 h. Treatment of cells with HT prevented cell proliferation (62%) and induced cell death (17%), whereas these alterations were not observed in cells treated with MHT. Microarray analysis revealed substantial differences with respect to gene expression patterns and biological function for the two different hyperthermic treatments. Moreover, we identified the temperature-specific gene networks D and H that were obtained from significantly up-regulated genes in the HT and MHT conditions, respectively, using Ingenuity pathway analysis tools. Gene network D, which contains 14 genes such as ATF3, DUSP1 and JUN, was associated with relevant biological functions including cell death and cellular movement. Gene network H, which contains 13 genes such as BAG3, DNAJB1 and HSPA1B, was associated with cellular function and maintenance and cellular assembly and organization. These findings provide a basis for understanding the detailed molecular mechanisms of cell death elicited by HT in human OSCC cells.

Although cancer cells exposed to temperatures >42.5°C undergo cell death as the temperature rises, exposure of up to 42.5°C induces slight or no cytotoxicity. The temperature of 42.5°C is, therefore, well known to be the inflection point of hyperthermia. To better understand the molecular mechanisms underlying cellular responses to heat stress at temperatures higher and lower than the inflection point, we carried out global scale microarray and computational gene expression analyses. Human leukemia U937 cells were incubated at 42°C or 44°C for 15 min and cultured at 37°C for 0-6 h. Apoptosis accompanied by the activation of caspase-3 and DNA fragmentation was only observed in cells treated with heat stress at 44°C, but not at 42°C. Although a large number of genes were differentially expressed by a factor of 2.0 or greater, we found substantial differences with respect to the biological functions and gene networks of the genes differentially expressed at the two temperatures examined. Interestingly, we identified temperature-specific gene networks that were considered to be mainly associated with cell death or cellular compromise and cellular function and maintenance at 44°C or 42°C, respectively, by using the Ingenuity pathway analysis tools. These findings provide the molecular basis for a further understanding of the mechanisms of the biological changes that are responsive to heat stress in human lymphoma cells.

The adaptive response of bone to mechanical loading in teleosts is not well understood. We recently developed a new assay system using teleost scales, which consists of osteoblasts, osteoclasts, and bone matrix protein. In this system, alkaline phosphatase (ALP) and tartrate-resistant acid phosphatase (TRAP) were used as markers of osteoblasts and osteoclasts, respectively. Using this assay system, we examined the effects of mechanical loading on ALP and TRAP activity in goldfish scales. ALP activity in the scales was significantly elevated (p

To define the molecular mechanisms that mediate hyperthermia-induced apoptosis, we performed microarray and computational gene expression analyses. U937 cells, a human myelomonocytic lymphoma cell line, were treated with hyperthermia at 42 degrees C for 90 min and cultured at 37 degrees C. Apoptotic cells ( approximately 15%) were seen 6 h after hyperthermic treatment, and elevated expression of heat shock proteins (HSPs) including Hsp27, Hsp40, and Hsp70 was detected, following the activation of heat shock factor-1. Of the 54,675 probe sets analyzed, 1334 were upregulated and 4214 were downregulated by2.0-fold in the cells treated with hyperthermia. A non-hierarchical gene clustering algorithm, K-means clustering, demonstrated 10 gene clusters. The gene network U1 or U2 that was obtained from up-regulated genes in cluster I or IX contained HSPA1B, DNAJB1, HSPH1, and TXN or PML, LYN, and DUSP1, and were mainly associated with cellular compromise, and cellular function and maintenance or death, and cancer, respectively. In the decreased gene cluster II, the gene network D1 including CCNE1 and CEBPE was associated with the cell cycle and cellular growth and proliferation. These findings will provide a basis for understanding the detailed molecular mechanisms of apoptosis induced by hyperthermia at 42 degrees C in cells.

To clarify the detailed molecular mechanism underlying cellular responses to nonthermal low-intensity pulsed ultrasound (LIPUS), gene expression patterns and genetic networks in human lymphoma U937 cells were examined using global-scale microarrays and computational gene expression analysis tools. Six hours after LIPUS treatment (0.3W/cm(2) for 1min), apoptosis (14+/-3.8%, mean+/-SD) without no cell lysis was observed. Of 22,283 probe sets analyzed, LIPUS down-regulated 193 genes and up-regulated 201 genes by >1.5-fold. For down-regulated genes, the significant genetic network D was associated with cellular growth and proliferation, gene expression, or cellular development. For up-regulated genes, the significant genetic network U was associated with cellular movement, cell morphology, and cell death. The present results indicate that LIPUS affects the expression of many genes and will provide novel insight into the biomolecular mechanisms of LIPUS in therapeutic application for cancer therapy.

OBJECTIVE AND METHODS This study was performed to clarify the usefulness of inductive heating system for the new endodontic therapy. Dextran magnetite complex (DM) suspensions were injected into the root canal of a permanent tooth, and the tooth was heated up to about 55.0 degrees C by alternating-current magnetic field. RESULTS AND CONCLUSION The time until the temperature in the pulp cavity reached 55.0 degrees C was 328 +/- 26 s (mean +/- s.d., n = 8) in the 56 mg as Fe ml(-1) of DM concentration. The temperature in the pulp cavity could be maintained at 53.5-59.0 degrees C for 1200 s by changing the magnetic field intensity safely, while temperature elevations of the dental surface on the coronal and apical sides were 4.9 degrees and 3.7 degrees C, respectively. Thus, this inductive heating system, which has the possibility of selective heating, might be useful for eliminating residues of pulp as a new ablation therapy.

The effects of various free radicals derived from 6-formylpterin (6-FP), alpha-phenyl-tert-butyl nitrone (PBN) and 2,2'-azobis (2-amidinopropane) dihydrochloride (AAPH) combined with hyperthermia, on gene expression in similarly enhanced apoptosis of human lymphoma U937 cells were investigated using cDNA microarrays containing approximately 16,600 genes and computational gene expression analysis tools. When the cells were treated for 10 min at 44 degrees C (15% apoptosis level), 39 up-regulated and 3 down-regulated genes were identified. In the up-regulated genes, apoptosis- and unfolded protein response-associated genes were contained. The combined treatment with heat and either chemical enhanced apoptosis level (approximately 30%) and showed a chemical-specific gene expression pattern. Furthermore, the expression levels of selected genes were confirmed by a real-time quantitative PCR. The present results will provide a basis for further understanding the molecular mechanisms in enhancement of heat-induced apoptosis by different intracellular oxidative stress.

We describe a 46-year-old man with T 4 N 2 bM 0 squamous cell cancer of the floor of the mouth. After concomitant radiotherapy (51.2Gy) and chemotherapy (CBDCA 480mg), he underwent total glossolaryngectomy, bilateral neck dissection, and reconstruction using a free rectus abdominis flap. Postoperatively, an IVH catheter was placed, and antibiotics (CEZ) were given prophylactically. Candida albicans sepsis developed 3 weeks after the operation. Administration of FCZ 200 mg daily for 17 days improved sepsis; however, candida osteomyelitis developed in the ribs and lumbar vertebrae 4 months after the operation. Persistent osteomyelitis of the ribs required partial costectomy. Postoperatively, the patient received FCZ 200 mg daily for 1 month, followed by FCZ 100mg daily. Osteomyelitis of the ribs completely healed, but the lesions of the lumbar vertebrae were not controlled.

The enhancement of heat-induced apoptosis by 6-formylpterin, an intra-cellular generator of hydrogen peroxide (H2O2), was examined in human myelomonocytic lymphoma U937 cells. The cells were treated with either 6-formylpterin alone at a nontoxic concentration of 300 microM (37 degrees C), heat shock (44 degrees C per 20 min) alone or a combination of the two, then incubated at 37 degrees C for 6 h. Assessments of apoptosis, mitochondrial membrane potential and caspase-3 activation were performed by flow cytometry. Moreover, caspase-8 activation and changes in the intra-cellular Ca2+ concentration ([Ca2+]i) were examined. Bax, Bcl-2, Bcl-XL, Bid, cytochrome c and PKCd were detected by Western blotting. The induction of heat-induced apoptosis evaluated by morphological observation and DNA fragmentation were promoted by the addition of 6-formylpterin. Mitochondrial membrane potential was decreased and the activation of caspase-3 and -8 was enhanced in the cells treated with the combination. A decreased-expression of Bid was noted, although no significant changes in Bax, Bcl-2 and Bcl-XL expression were observed after the combined treatment. Furthermore, both the release of cytochrome c from mitochondria to cytosol and the translocation of PKCd from cytosol to mitochondria, which were induced by heat shock, were enhanced by the addition of 6-formylpterin. The number of cells with a higher [Ca2+]i was also increased by the addition of 6-formylpterin. These findings suggest that the increase in [Ca2+]i, the activation of the mitochondria-caspase dependent pathway and the translocation of PKCd to mitochondria play principal roles in the enhancement of heat-induced apoptosis by 6-FP.

To examine the direct effects of tributyltin acetate (TBTA) on osteoclasts and osteoblasts, teleost scale, which has both osteoclasts and osteoblasts and is similar to mammalian membrane bone, was used in the present study. The activities of tartrate-resistant acid phosphatase and alkaline-phosphatase, as respective indicators of activity in both cells, were used. In freshwater teleost (goldfish) and marine teleosts (nibbler and wrasse), the osteoclastic activity in the scales did not change as a result of TBTA treatment (10(-9) to 10(-5) M). However, the osteoblastic activity decreased in the goldfish, nibbler, and wrasse after 6 h of incubation. In goldfish, even 10(-10) M of TBTA significantly inhibited the osteoblastic activity. The inhibitory activity in goldfish was stronger than that in nibbler and wrasse. Therefore, details of the mechanism were examined using goldfish. The mRNA expressions of the estrogen receptor and insulin-like growth factor-I, which participate in osteoblastic growth and differentiation, decreased in the TBTA-treated scales. However, the mRNA expression of metallothionein (MT), a metal-binding protein that protects the organism from heavy metal, increased much less than those of cadmium and methyl-mercury. Furthermore, we showed that the plasma calcium and hypocalcemic hormone (calcitonin) level increased in goldfish kept in water containing TBTA (10(-10) and 10(-8) M). The current data are the first to demonstrate that, in teleosts, TBTA inhibits osteoblastic activity without affecting osteoclastic activity and disrupts the calcium metabolism, including the calcemic hormone, in goldfish.

Radiation-induced apoptosis and its possible enhancement in the presence of 6-formylpterin (6-FP), a metabolite of folic acid, were examined in human myelomonocytic lymphoma U937 cells. When cells were treated with 6-FP at a nontoxic concentration of 300 microM, and then exposed to X-rays at a dose of 10 Gy, significant enhancement of radiation-induced apoptosis as determined by nuclear morphological change, phosphatidylserine (PS) externalization and DNA fragmentation were observed. Flow cytometry for the detection of intracellular hydrogen peroxide (H2O2) revealed that 6-FP increased the formation of intracellular H2O2, which further increased when the cells were irradiated. Decrease of mitochondria trans-membrane potential (MMP), release of cytochrome c from mitochondria, and activation of caspase-3 were enhanced after the combined treatment. Remarkable activation of protein kinase C delta (PKC delta) and its translocation from cytosol to mitochondria were detected in combined treatment. Increase of intracellular Ca2+ concentrations ([Ca2+]i) was also observed, however, neither calpain I nor calpain II could inhibit the apoptosis. In addition, c-Jun NH2-terminal kinase (JNK) activation was not enhanced in the combined treatment. A protein involved in a caspase-independent apoptosis pathway, apoptosis inducing factor (AIF), remained unchanged even 3 h after treatment. These results indicate that intracellular H2O2 generated by 6-FP enhances radiation-induced apoptosis via the mitochondria-mediated caspase-dependent pathway, with the active involvement of PKC delta.

Summary A phase I study of combination neoadjuvant chemotherapy with docetaxel (DOC) and nadaplatin (CDGP) was conducted in patients with untreated, advanced but operable oral squamous cell carcinoma. DOC was administered (one-hour i.v. infusion) on day 1 followed by CDGP (one-hour i.v. infusion). The dose levels of DOC and CDGP tested were 60/70, 60/80, 60/90, 60/100, and 70/100 (mg/m2). Fifteen patients enrolled in this study and median age was 60 years. Dose-limiting toxicity (DLT) occurred in one of six patients at DOC dose of 60 mg/m2 and CDGP dose of 100 mg/m2. The DLT was diarrhea. Because one additional patient at this dose-level developed grade 4 neutropenia lasting for 4 days that approached DLT and because fear of severe hematological problems was predicted, further dose escalation was not performed. This combination chemotherapy is active and well tolerated and warrants a phase II study. We recommended 60 mg/m2 DOC and 100 mg/m2 CDGP for phase II study.

Summary Introduction A newly devised retractor for mandibular torus is presented. It has a half-spoon-shaped head and was applied to various cases of mandibular torus for its evaluation. Results It was found that the elevated mucoperiosteum flap was safely protected lingually by using this retractor. Various mandibular tori were completely resected without damaging the surrounding tissues.

金沢大学環日本海域環境研究センター生物多様性研究部門, AAims: We previously demonstrated that monohydroxylated polycyclic aromatic hydrocarbons (OHPAHs) bound to a human estrogen receptor (ER) by a yeast two-hybrid assay, but polycyclic aromatic hydrocarbons did not have a binding activity. Therefore, the direct effect of 3-hydroxybenz[a]anthracene (3-OHBaA) and 4-hydroxybenz[a]anthracene (4-OHBaA) on osteoclasts and osteoblasts in teleosts was examined. As a negative control, 1-hydroxypyrene (1-OHPy), which has no binding activity to human ER, was used. Main methods: The effect of OHPAHs on osteoclasts and osteoblasts was examined by an assay system using teleost scale as each marker: tartrate-resistant acid phosphatase for osteoclasts and alkaline phosphatase for osteoblasts. Changes in cathepsin K (an osteoclastic marker) and insulin-like growth factor-I (IGF-I) (an osteoblastic marker) mRNA expressions in 4-OHBaA-treated goldfish scales were examined by using a reverse transcription-polymerase chain reaction. Key findings: In both goldfish (a freshwater teleost) and wrasse (a marine teleost), the osteoclastic activity in the scales was significantly suppressed by 3-OHBaA and 4-OHBaA, although 1-OHPy did not affect the osteoclastic activity. In reference to osteoblasts, the osteoblastic activity decreased with both 3-OHBaA and 4-OHBaA and did not change with the 1-OHPy treatment. However, 17β-estradiol (E2) significantly increased both the osteoclastic and osteoblastic activities in the scales of both goldfish and wrasse. The mRNA expressions of both cathepsin K and IGF-I decreased in the 4-OHBaA-treated scales but increased in the E2-treated scales. Significance: The current data are the first to demonstrate that 3-OHBaA and 4-OHBaA inhibited both osteoclasts and osteoblasts and disrupted the bone metabolism in teleosts. © 2009 Elsevier Inc. All rights reserved.

doi:10.1016/j.bone.2009.01.346 440 Effect of low-intensity pulsed ultrasound on osteoblasts and osteoclasts of the zebrafish scales N. Suzuki, Y. Furusawa, I. Takasaki, Y. Tabuchi, K. Kitamura, S. Wada, T. Hori, T. Kondo, T. Nemoto, N. Shimizu, A. Hattori Noto Marine Laboratory, Kanazawa University, Noto-cho, Ishikawa, Japan Graduate School of Medical and Pharmaceutical Science, University of Toyama, Sugitani, Toyama, Japan Life Science Research Center, University of Toyama, Sugitani, Toyama, Japan Graduate School of Medical Science, Kanazawa University, Kanazawa, Ishikawa, Japan Faculty of Medicine, University of Toyama, Sugitani, Toyama, Japan Institute of Nature and Environmental Technology, Kanazawa University, Kanazawa, Ishikawa, Japan College of Liberal Arts and Sciences, Tokyo Medical and Dental University, Ichikawa, Chiba, Japan