Pigmentation of the oral mucosa by PCB poisoning in Yusho patients.

OBJECTIVES: Toxins, such as PCBs, dramatically affect patients even decades after exposure. Although 40 years have passed since the accidental poisoning with polychlorinated biphenyl (PCB) in Western Japan in 1968, high concentrations of PCBs are still detected in the serum of the "Yusho" (oil disease) patients. In this study, an epidemiological examination was carried out to reveal the prevalence of the oral pigmentation and blood concentrations of PCBs and polychlorinated quaterphenyl (PCQ) in Yusho victims. DESIGN: We performed a group examination of patients (Yusho victims) from 2004 to 2006, including 72 Yusho victims and 15 control subjects. The oral examination was performed by two oral and maxillofacial surgeons. The serum concentrations of PCB and PCQ were determined using gas chromatography; blood samples from Yusho victims were analyzed for PCB and PCQ by saponification in 1M NaOH ethanol solution, extraction with n-hexane column chromatography on silica gel, and then gas chromatography with electron capture detection. RESULTS: The mean Yusho victim's serum PCB and PCQ concentrations were 3.3ppb and 0.9ppb, respectively. In controls, these were 0.7ppb and 0ppb, respectively. Oral pigmentation was observed in 24 out of 72 Yusho patients. In controls, oral pigmentation was observed in one out of 15 persons. Oral pigmentation was most frequently observed in the buccal mucosa, followed by gingival mucosa. The blood concentration of PCB in Yusho patients with oral pigmentations was significantly higher than that in Yusho patients without oral pigmentation. CONCLUSION: These results indicated that PCB-related compounds may be responsible for the higher prevalence of oral pigmentation in Yusho victims, even though a long time has passed since the Yusho poisoning accident.

Immunohistochemical study of VEGF expression in oral squamous cell carcinomas: correlation with the mTOR-HIF-1α pathway.

The aim of this study was to clarify the relationship between vascular endothelial growth factor (VEGF) expression and clinicopathological factors in oral squamous cell carcinoma (OSCC). We also examined the correlation between the VEGF expression and the mammalian target of rapamycin (mTOR)-hypoxia inducible factor-1α (HIF-1α) pathway. Formalin-fixed paraffin-embedded tissues from 120 OSCC cases and 10 samples of normal mucosa were stained immunohistochemically for VEGF-A, VEGF-C, p-mTOR and HIF-1α proteins. VEGF-A and VEGF-C protein expression was detected in 76 out of 120 (63%) and 81 of 120 (67.5%) OSCCs, respectively, and their expression was significantly higher in primary OSCC than in normal oral mucosa. VEGF-A expression was significantly associated with the tumor stage and age. VEGF-C expression was significantly associated with the cancer cell invasion. The cases with combined p-mTOR+/HIF-1α(+)/VEGF-A(+) expression had a significantly higher tumor stage and invasion grade, and combined p-mTOR+/HIF-1α(+)/VEGF-C(+) expression was significantly associated with tumor stage, regional lymph node metastasis and invasion grade. In a survival analysis, no obvious correlation was observed with any of the immunohistochemical results. This study indicated that the mTOR-HIF-1α-VEGF pathway affects the progression of OSCC, and inhibition of this pathway may be useful for the treatment of OSCC.

Expression of thymidylate synthase and dihydropyrimidine dehydrogenase in primary oral squamous cell carcinoma and corresponding metastases in cervical lymph nodes: association with the metastasis suppressor CD82.

Thymidylate synthase (TS) and dihydropyrimidine dehydrogenase (DPD) are 5-fluorouracil (5-FU) metabolizing enzymes and are involved in the sensitivity of carcinoma patients to 5-FU. Although 5-FU is often used for the treatment of oral carcinoma, there has not been any investigation into the expression of these enzymes in metastatic lymph nodes or of their roles in the effectiveness of 5-FU in treating lymph node-metastatic cancer. Oral squamous cell carcinoma (OSCC) often metastasizes to the lymph nodes, and these enzymes may be significant in the survival of patients with this disease. This study investigated the expression of TS and DPD in cervical lymph node metastases and its relationship with primary OSCC, as well as the interaction between these enzymes and Kangai 1(KAI1/CD82) which is a metastasis suppressor protein. Surgical specimens from 20 cases of OSCC with lymph node metastasis, 20 cases of OSCC without lymph node metastasis, and 10 cases of normal mucosa were examined by immunohistochemistry. The relationship between TS and DPD expression and clinicopathological data was analyzed. TS and DPD proteins were overexpressed in primary OSCC compared to that in normal mucosa. TS expression of the primary oral cancer cells in the group with lymph node metastasis was higher than that of those without. DPD expression did not significantly correlate with the occurrence of lymph node metastasis, nor was it different between primary oral cancer cells and cervical metastases. CD82 expression was significantly reduced in lymph node metastases. These findings indicate that TS and CD82 may be of great value in assessing lymph node metastasis of OSCC, and could be taken as new targets for therapy of metastatic OSCC.

Efficacy and toxicity of S-1 plus cisplatin combination neoadjuvant chemotherapy in patients with oral cancer.

S-1 is a newly developed oral fluoropyrimidine derivative that is now widely used as a chemotherapeutic agent in the treatment of various carcinomas. This study was performed to assess the efficacy and safety profile of the combination of S-1 and cisplatin(S-1/CDDP)in patients with oral cancer as neo-adjuvant chemotherapy. We reviewed our experience of 12 patients diagnosed with oral carcinoma, who were treated with S-1/CDDP. S-1 was administered orally at a dose of 50mg twice a day for 21 consecutive days, followed by a 14-day rest period. CDDP(60mg/m2)in 500 mL physiological saline was administered by intravenous drip as a 120-min infusion on day 8, together with standard premedications and hydration. Seven partial responders were obtained. The median follow-up duration was 54. 8 months, and all patients were alive excluding one case. This regimen was well tolerated, with only one case of grade 3 thrombocytopenia, and no grade 4 patient. No treatment-related death was observed. Moreover, we evaluated immunohistochemical expressions of thymidylate synthase (TS), dihydropyrimidine dehydrogenase(DPD), and orotate phosphoribosyl transferase(OPRT)which are associated with chemosensitivity to 5-FU-based therapies. We investigated the relation between the immunohistochemical score and clinicopathological factors, however we could not clarify the relationship between the efficacy of chemotherapy and results of immunohistochemistry.

Lingual metastasis as an initial presentation of renal cell carcinoma.

Distant metastasis of malignant neoplasm to the oral soft tissue is extremely rare. We report a case of renal cell carcinoma (RCC) metastasizing to the tongue. A 47-year-old man visited our hospital with chief complaint of a lump on the middle third of the dorsum of his tongue and the lesion fell off from the tongue. Although histopathological diagnosis of the mass was granuloma teleangiectaticum, similar nodule reappeared in the same area 2 weeks later. The second lesion was composed of granuloma teleangiectaticum and aggregation of neoplastic clear cells in ductal arrangement. The clear cells were immunohistochemically positive for EMA and CD10. The abdominal CT scan revealed a 5.5 cm mass in the left kidney, suggesting RCC. Thus, the lingual lesion was consistent with metastatic RCC. There has been no recurrence for 2 years after the radical nephrectomy and local excision of the tongue.

Fabrication of an integrated holographic imaging element for a three-dimensional eye-gaze detection system.

We evaluate the imaging characteristics of an integrated optical imaging element that is used to obtain images from opposite directions in one imaging sensor for a three-dimensional eye-gaze detection system. The element consists of a transmission-type holographic imaging element, a reflection-type holographic imaging element, and a noise reduction filter. In the evaluation of the imaging characteristics, modulation transfer functions of both the reflection-type and the transmission-type holographic imaging elements are evaluated. Results indicate that both holographic imaging elements have enough resolution, even under white-light illumination conditions, for eye-gaze detection. We also demonstrate the simultaneous detection of images by an artificial eye and objects by using the integrated element under white light or sunlight.

[Immunohistochemical and clinicopathological study of thymidylate synthase, dihydropyrimidine dehydrogenase, and orotate phosphoribosyl transferase expression in oral cancer patients responding to UFT].

Thymidylate synthase(TS), dihydropyrimidine dehydrogenase(DPD), and orotate phosphoribosyl transferase(OPRT)are initial key enzymes in the 5-fluorouracil(5-FU)metabolic pathway. In this study, we investigated clinicopathological and immunohistochemical expressions of TS, DPD, and OPRT in oral cancer patients who showed a complete response(CR)to UFT. We also evaluated patients showing a partial response(PR)and stable disease(SD)following UFT. The numbers of CR, PR, and SD cases were 3, 5, and 10, respectively. Pathologically, all CR and PR cases were the well-differentiated type, and 5 out of 10 SD cases were of the moderately or poorly-differentiated type. Three out of the 5 cases of moderately or poorlydifferentiated type were DPD-negative. Most cases of CR and PR were DPD-positive. OPRT expression showed no difference with the UFT response. We suggest that UFT affects high DPD patients with the well-differentiated type, but may not influence low DPD patients with the moderately or poorly-differentiated type.

Overexpression of cortactin increases invasion potential in oral squamous cell carcinoma.

Cortactin, an F-actin binding protein, stabilizes F-actin networks and promotes actin polymerization by activating the Arp2/3 complex. Overexpression of cortactin has been reported in several human cancers. Cortactin stimulates cell migration, invasion, and experimental metastasis. However, the underlying mechanism is not still understood. In the present study, we therefore evaluated the possibility that cortactin could be appropriate as a molecular target for cancer gene therapy. In 70 primary oral squamous cell carcinomas and 10 normal oral mucosal specimens, cortactin expression was evaluated by immunological analyses, and the correlations of the overexpression of cortactin with clinicopathologic factors were evaluated. Overexpression of cortactin was detected in 32 of 70 oral squamous cell carcinomas; significantly more frequently than in normal oral mucosa. Cortactin overexpression was more frequent in higher grade cancers according to T classification, N classifications, and invasive pattern. Moreover, RNAi-mediated decrease in cortactin expression reduced invasion. Downregulation of cortactin expression increased the expression levels of E-cadherin, ß-catenin, and EpCAM. The siRNA of cortactin also reduced PTHrP expression via EGF signaling. These results consistently indicate that the overexpression of cortactin is strongly associated with an aggressive phenotype of oral squamous cell carcinoma. In conclusion, we propose that cortactin could be a potential molecular target of gene therapy by RNAi targeting in oral squamous cell carcinoma.

Comparative evaluation of the breaking strength of a simple mobile mandibular advancement splint.

Mandibular advancement splints (MASs) are used to advance the mandible forward in patients with sleep-disordered breathing. The conventional rigid MAS restricts the movement of the mandible, and this immobility sometimes produces discomfort, including temporomandibular disorders. A simple method for fabricating a mobile MAS was devised, using a connector made from a polyethylene toothed belt, with the intention of making the MAS more comfortable. The experimental connector is easily constructed, inexpensive, and small enough for use as an intraoral MAS. To evaluate durability, the axial and diagonal tensile breaking strengths of the MAS, using high- or low-density polyethylene (HDPE or LDPE) lateral toothed belts, were compared with those of a conventional mobile MAS (Silensor). The values were compared by factorial analysis of variances and post hoc Scheffe's S multiple comparison intervals, with the value of statistical significance set at alpha = 0.05. In addition, the experimental LDPE connectors were clinically tested in 30 patients (23 males and 7 females aged 19-71 years) and evaluated. Compared with the Silensor, the experimental MAS exhibited sufficient breaking strength, especially when a diagonal tensile load was applied to mimic mandibular lateral translation. When examining the clinical evaluation between 3 and 4 months after insertion, no damage or failure was observed. The experimental connecting system may have clinical applications. To make the connector stronger for clinical use, HDPE should be used.

Akt regulates skeletal development through GSK3, mTOR, and FoxOs.

Although Akt plays key roles in various cellular processes, the functions of Akt and Akt downstream signaling pathways in the cellular processes of skeletal development remain to be clarified. By analyzing transgenic embryos that expressed constitutively active Akt (myrAkt) or dominant-negative Akt in chondrocytes, we found that Akt positively regulated the four processes of chondrocyte maturation, chondrocyte proliferation, cartilage matrix production, and cell growth in skeletal development. As phosphorylation of GSK3beta, S6K, and FoxO3a was enhanced in the growth plates of myrAkt transgenic mice, we examined the Akt downstream signaling pathways by organ culture. The Akt-mTOR pathway was responsible for positive regulation of the four cellular processes. The Akt-FoxO pathway enhanced chondrocyte proliferation but inhibited chondrocyte maturation and cartilage matrix production, while the Akt-GSK3 pathway negatively regulated three of the cellular processes in limb skeletons but not in vertebrae due to less GSK3 expression in vertebrae. These findings indicate that Akt positively regulates the cellular processes of skeletal growth and endochondral ossification, that the Akt-mTOR, Akt-FoxO, and Akt-GSK3 pathways positively or negatively regulate the cellular processes, and that Akt exerts its function in skeletal development by tuning the three pathways in a manner dependent on the skeletal part.

Single nucleotide polymorphism that accompanies a missense mutation (Gln488His) impedes the dimerization of Hsp90.

A single nucleotide polymorphism (SNP) that causes a missense mutation of highly conserved Gln488 to His of the alpha isoform of the 90-kDa heat shock protein (Hsp90alpha) molecular chaperone is observed in Caucasians. The mutated Hsp90alpha severely reduced the growth of yeast cells. To investigate this molecular mechanism, we examined the domain-domain interactions of human Hsp90alpha by using bacterial 2-hybrid system. Hsp90alpha was expressed as a full-length form, N-terminal domain (residues 1-400), or middle (residues 401-617) plus C-terminal (residues 618-732) domains (MC domain/amino acids 401-732). The Gln488His substitution in MC domain did not affect the intra-molecular interaction with N-terminal domain, whereas the dimeric interaction-mediated by the inter-molecular interaction between MC domains was decreased to 32%. Gln488Ala caused a similar change, whereas Gln488Thr, which exceptionally occurs in mitochondrial Hsp90 paralog, fully maintained the dimeric interaction. Therefore, the SNP causing Gln488His mutation could abrogate the Hsp90 function due to reduced dimerization.

Ribonucleotide reductase small subunit p53R2 promotes oral cancer invasion via the E-cadherin/beta-catenin pathway.

The p53-inducible p53R2 gene has been isolated and shown to play a crucial role in DNA repair and synthesis after DNA damage. Moreover, the expression and activity of p53R2 has been reported to be associated with the anticancer agent resistance of human cancer cells. Previously, we reported that the presence of p53R2 expression was a predictive factor for regional lymph node metastasis in oral squamous cell carcinoma; however, the mechanism of cancer metastasis by p53R2 expression is still unclear. In the present study, we analyzed the correlation of p53R2 expression with cancer invasion in vitro. Three human oral cancer cell lines (SAS, HSC-3 and Ca9-22) were cultured, and the invasive potential of these cancer cells was evaluated using Matrigel invasion assay. To investigate the effect of p53R2 on cancer invasion, the down-regulation of p53R2 was examined by small interfering RNA (siRNA). Moreover, we examined the intracellular localization of cell adhesion molecules (E-cadherin and beta-catenin) in subcellular extractions of cancer cells by immunoblotting. The proteolytic activity of matrix metalloproteinases (MMPs) was assessed by gelatin zymography. Down-regulation of p53R2 significantly enhanced the invasion potential (p<0.01), and enhanced nuclear translocation of beta-catenin with loss of total cellular E-cadherin expression in p53 mutant cancer cells, but not in p53 wild-type cancer cells. These changes in the invasion index by p53R2 siRNA transfection were not accompanied by alterations in MMP activity and expression. These results suggested that the expression of p53R2 could be associated with the invasion of cancer cells, and indicated that p53R2 might promote cancer invasion via the E-cadherin/beta-catenin pathway without the alteration of MMP activity.

Substitution of only two residues of human Hsp90alpha causes impeded dimerization of Hsp90beta.

Two isoforms of the 90-kDa heat-shock protein (Hsp90), i.e., Hsp90alpha and Hsp90beta, are expressed in the cytosol of mammalian cells. Although Hsp90 predominantly exists as a dimer, the dimer-forming potential of the beta isoform of human and mouse Hsp90 is less than that of the alpha isoform. The 16 amino acid substitutions located in the 561-685 amino acid region of the C-terminal dimerization domain should be responsible for this impeded dimerization of Hsp90beta (Nemoto T, Ohara-Nemoto Y, Ota M, Takagi T, Yokoyama K. Eur J Biochem 233: 1-8, 1995). The present study was performed to define the amino acid substitutions that cause the impeded dimerization of Hsp90beta. Bacterial two-hybrid analysis revealed that among the 16 amino acids, the conversion from Ala(558) of Hsp90beta to Thr(566) of Hsp90alpha and that from Met(621) of Hsp90beta to Ala(629) of Hsp90alpha most efficiently reversed the dimeric interaction, and that the inverse changes from those of Hsp90alpha to Hsp90beta primarily explained the impeded dimerization of Hsp90beta We conclude that taken together, the conversion of Thr(566) and Ala(629) of Hsp90alpha to Ala(558) and Met(621) is primarily responsible for impeded dimerization of Hsp90beta.

Clinicopathologic significance of EpCAM expression in squamous cell carcinoma of the tongue and its possibility as a potential target for tongue cancer gene therapy.

Epithelial adhesion molecule (EpCAM) is a transmembrane glycoprotein involved in intercellular adhesion. In particular, EpCAM appears to be overexpressed by the majority of human epithelial carcinomas, including colorectal, breast, head and neck, and hepatic carcinomas. We therefore hypothesized that EpCAM would be a good molecular target for cancer gene therapy. EpCAM protein expression in 48 primary tongue cancers and 10 normal oral mucosa was evaluated using anti-EpCAM immunohistochemistry, and correlation was examined with the clinicopathologic factors. In four human tongue cancer cell lines (SAS, HSC-2, OSC19 and OSC20), we investigated EpCAM expression by reverse transcription-polymerase chain reaction (RT-PCR). The invasive potential of cancer cells was evaluated using Matrigel invasion assay. Moreover, the effect of EpCAM inhibition was analyzed using RNA interference (RNAi). EpCAM overexpression was detected in 30 of 48 tongue cancers (62.5%), and was significantly higher in primary squamous cell carcinoma (SCC) of the tongue than in normal oral mucosa. The expression of EpCAM was significantly associated with tumor size, regional lymph node metastasis, histological differentiation and invasion pattern. Cancer cell lines with higher EpCAM expression had more invasive potential. Moreover, RNAi-mediated EpCAM reduction decreased the invasion potential and proliferation activity. These results indicated that the overexpression of EpCAM was correlated with a more aggressive phenotype of tongue cancer. Moreover, we suggested that EpCAM could be a molecular target, and that RNAi targeting EpCAM could be useful for tongue cancer gene therapy.