Simultaneous Recognition of Over- and Under-Expressed MicroRNAs Using Nanopore Decoding.

This paper describes a strategy for simultaneous recognition of over- and under-expressed microRNAs (miRNAs) using the method of signal classification-based nanopore decoding. MiRNA has attracted attention as a promising biomarker for cancer diagnosis owing to its cancer-type-specific expression patterns. While nanopore technology has emerged as a simple and label-free method to detect miRNAs and their expression patterns, recognizing patterns involving simultaneous over/under-expression is still challenging due to the inherent working principles. Here, inspired by the sequence design for DNA computation with nanopore decoding, we designed diagnostic DNA probes targeting two individual over/under-expressed miRNAs in the serum of oral squamous cell carcinoma. Through nanopore measurements, our designed probes exhibited characteristic current signals depending on the hybridized miRNA species, which were plotted on the scatter plot of duration versus current blocking ratio. The classified signals reflected the relative abundance of target miRNAs, thereby enabling successful pattern recognition of over/under-expressed miRNAs, even when using clinical samples. We believe that our method paves the way for miRNA-targeting simple diagnosis as a liquid biopsy.

Responsiveness of the Zurich Claudication Questionnaire, the Oswestry Disability Index, the Japanese Orthopaedic Association Back Pain Evaluation Questionnaire, the 8-Item Short Form Health Survey, and the Euroqol 5 dimensions 5 level in the assessment of patients with lumbar spinal stenosis.

PURPOSE: To assess the responsiveness of the Zurich Claudication Questionnaire (ZCQ), the Oswestry Disability Index, the Japanese Orthopaedic Association Back Pain Evaluation Questionnaire, the visual analog scale (VAS), the 8-Item Short Form Health Survey (SF-8), and the EuroQol 5 dimensions 5 level as methods of assessing outcomes of surgery for lumbar spinal stenosis. METHODS: We analyzed 218 patients who had undergone lumbar surgery for spinal stenosis and completed one year of follow-up. The internal responsiveness of each questionnaire and any domains was assessed by the effect size and standardized response mean. External responsiveness was assessed by the Spearman rank correlation coefficient and the receiver operating characteristics (ROC) curve. RESULTS: The most responsive assessments were "symptom severity" and "physical function" on the ZCQ, "walking ability" on the JOABPEQ, "leg pain" on the VAS, and "social function" on the JOABPEQ. The moderately responsive assessments were the physical component summary on the SF-8, the ODI, the EQ5D-5L, "low back pain" on the JOABPEQ, and "leg numbness" on the VAS. The least responsive assessments were "low back pain" on the VAS, "mental health" and "lumbar function" on the JOABPEQ, and the mental component summary on the SF-8. CONCLUSIONS: Because of its high responsiveness, "symptom severity" on the ZCQ is recommended as a primary tool for assessing outcome when designing prospective studies for lumbar spinal stenosis.

Transfection of T-Box Transcription Factor BRACHYURY and SOX2 Synergistically Promote Self-Renewal and Invasive Phenotype in Oral Cancer Cells.

Recent studies suggest that epithelial⁻mesenchymal transition (EMT) correlates with cancer metastasis. In addition, there is growing evidence of the association of EMT with cancer stem cells (CSCs). Recently, we showed that the T-box transcription factor BRACHYURY could be a strong regulator of EMT and the CSC phenotype, which were effectively suppressed by a BRACHYURY knockdown in an adenoid cystic carcinoma cell line. In this study, we further tested whether BRACHYURY is a regulator of cancer stemness by means of forced expression of BRACHYURY in oral cancer cell lines. BRACHYURY, SOX2, or both were stably transfected into oral carcinoma cell lines. We analysed these transfectants with respect to self-renewal phenotypes using a sphere-formation assay, and we assessed the expression levels of EMT markers and stem cell markers using real-time reverse transcription-polymerase chain reaction (RT-PCR). Cell migration and invasiveness in vitro were evaluated using a wound healing assay and a tumour cell dissemination assay, respectively. Forced expression of BRACHYURY or SOX2 slightly increased expression of EMT and stem cell markers and the self-renewal phenotype. The expression levels, however, were much lower compared to those of cancer stem cell-like cells. Forced co-expression of BRACHYURY and SOX2 strongly upregulated EMT and stem cell markers and the self-renewal phenotype. Cell migration and invasiveness in vitro were also remarkably enhanced. These synergistic effects increased expression levels of FIBRONECTIN, SNAIL, SLUG, ZEB1, and TGF-ß2. In particular, the effects on FIBRONECTIN and TGF-ß2 were significant. We found that BRACHYURY and SOX2 synergistically promote cancer stemness in oral cancer cells. This finding points to the importance of gene or protein networks associated with BRACHYURY and SOX2 in the development and maintenance of the CSC phenotype.

Efficient regulation of branching morphogenesis via fibroblast growth factor receptor 2c in early-stage embryonic mouse salivary glands.

Salivary gland (SG) defects have a wide range of health implications, including xerostomia, bacterial infections, and oral health issues. Branching morphogenesis is critical for SG development. A clear understanding of the mechanisms underlying this process will accelerate SG regeneration studies. Fibroblast growth factor receptor 2 (FGFR2) interacts with multiple fibroblast growth factors (FGFs), which promote development. FGFR2 consists of two isoforms, FGFR2b and FGFR2c. FGFR2b is critical for SG development, but little is known about the expression and function of FGFR2c. We investigated the expression of all FGFR family members in fetal SGs between embryonic day 12.5 (E12.5) and E18.5. Based on RT-PCR, we observed an increase in the expression of not only Fgfr2b, but also Fgfr2c in early-stage embryonic mouse SGs, suggesting that FGFR2c is related to SG development. The branch number decreased in response to exogenous FGF2 stimulation, and this effect was suppressed by a mouse anti-FGFR2c neutralizing antibody (NA) and siRNA targeting FGFR2c, whereas FGFR2b signaling was not inhibited. Moreover, the expression of marker genes related to EMT was induced by FGF2, and this expression was suppressed by the NA. These results suggested that branching morphogenesis in SGs is regulated by FGFR2c, in addition to FGFR2b. Interestingly, FGFR2c signaling also led to increased fgf10 expression, and this increase was suppressed by the NA. FGFR2c signaling regulates branching morphogenesis through the activation of FGFR2b signaling via increased FGF10 autocrine. These results provide new insight into the mechanisms by which crosstalk between FGFR2b and FGFR2c results in efficient branching morphogenesis.

Involvement of the T-box transcription factor Brachyury in early-stage embryonic mouse salivary gland.

The mouse submandibular gland (SMG) is important organ for embryonic development, and branching morphogenesis is regulated by many molecules containing transcription factors. Real-time reverse transcriptase polymerase chain reaction revealed that the expression of Brachyury increased in the SMG and peaked between E12.5-E13.5, concomitant with the early stage of branching morphogenesis. The expression of Brachyury in SMG rudiments between E12.5-E13.5 was confirmed by western blotting. In addition, fibronectin and Btbd7 (regulated by fibronectin), which are both essential for cleft formation, were expressed strongly during the same period. The Sox2 and Wnt3a, which regulate cell growth, were also expressed strongly during E12.5-E13.5. On the other hand, cleft formation and branching morphogenesis was suppressed by knockdown of Brachyury gene, suggesting that Brachyury plays a central role in regulating cell growth and cleft formation in early-stage embryonic mouse salivary gland development.

Circulating sclerostin and dickkopf-1 levels in ossification of the posterior longitudinal ligament of the spine.

Sclerostin and dickkopf-1(DKK1) are Wnt/ß-catenin signal antagonists that play an important role in bone formation. Ossification of the posterior longitudinal ligament (OPLL) of the spine is characterized by pathological ectopic ossification of the posterior longitudinal ligament and ankylosing spinal hyperostosis. The aims of this study were to evaluate serum sclerostin and DKK1 levels in persons with OPLL and to identify its relationship with bone metabolism and bone mass in persons with OPLL. This was a case-control study, and 78 patients with OPLL were compared with 39 age- and sex-matched volunteers without OPLL. We analyzed the relationship with calciotropic hormones, bone turnover markers, OPLL localization, number of ossified vertebrae, and bone mineral density of total hip (TH-BMD). Serum sclerostin levels in men with OPLL were significantly higher than in men in the control group (control group: mean = 45.3 pmol/L; OPLL group: mean = 75.7 pmol/L; P = 0.002). Age and sclerostin levels were positively correlated in men with OPLL (r = 0.43; P = 0.002). Serum sclerostin levels in men with OPLL had a positive correlation with TH-BMD Z-score (r = 0.511; P = 0.011, n = 30). There was a strong negative correlation between serum sclerostin levels and serum DKK1 levels in men with OPLL (r = -0.506; P < 0.001). Bone and mineral metabolism in OPLL differs between men and women. In men with OPLL, systemic secretion of sclerostin increases with advancing age and with higher bone mass. These two Wnt/ß-catenin signal antagonists have the opposite effect in persons with OPLL, and higher serum sclerostin levels are counterbalanced by underproduction of DKK1.

3D morphometric analysis of laminae and facet joints in patients with degenerative spondylolisthesis.

OBJECTIVE: To clarify the three-dimensional (3D) morphometric characteristics of the spine in patients with degenerative spondylolisthesis (DS). METHODS: 3D morphometric analyses of laminae and facets were performed and compared for a DS group, an age-matched spinal canal stenosis (LCS) group, and a control group of young persons without spinal disease. 3D facet sagittal angles (3D-FSAs), 3D facet axial angle (3D-FAAs), and 3D-FAA tropism at L3 and at L4 were measured by extracting the 3D inferior articular process. The 3D lamina inclination angles (3D-LIAs) of L3 and L4 were also measured by extracting the ventral surface of the laminae. RESULTS: The 3D-FSAs at L4 in the DS group were significantly higher than for the other groups, but the difference in 3D-FSAs at L3 was not statistically significant among the groups. The 3D-FAAs at L4 in the DS group were significantly lower than in the control group. There was no significant difference in other factors. CONCLUSIONS: 3D morphometric analysis clarified that DS is significantly correlated with horizontalization (higher 3D-FSA), but is not correlated with sagittalization (lower 3D-FAA) and tropism (3D-FAA tropism) of facet joints or horizontalization of laminae (3D-LIA). There were no morphometric characteristics at the cranial adjacent segment of DS.

Relation of omega-3 fatty acid and C-reactive protein to peripheral artery disease in patients with coronary artery disease.

Eicosapentaenoic acid (EPA), a member of the omega-3 polyunsaturated fatty acid family, prevents cardiovascular disease. C-reactive protein (CRP) is a marker of inflammation, which promotes atherosclerosis. The aim of this study was to investigate the relationship among EPA, CRP, and the prevalence of peripheral artery disease (PAD), which is a manifestation of systemic atherosclerosis. A cross-sectional study was performed on 238 patients with coronary artery disease (CAD). Blood EPA and CRP levels and ankle-brachial pressure indices were measured. Cut-off values for plasma EPA levels and serum CRP levels were determined using receiver operating characteristic (ROC) analysis. Patients with ABIs ≤0.9 were defined as having PAD. EPA levels were significantly lower and CRP levels were significantly higher in patients with PAD than in those without [48 (26-77) vs. 58 (41-83) µg/ml, p = 0.026 and 3.3 (0.64-14.0) vs. 0.70 (0.32, 2.4) mg/l, p = 0.004]. Multivariate analysis for PAD revealed that high CRP levels and low EPA levels were significant and independent predictors of PAD [odds ratio 3.1 (95 % CI 1.4-6.9), p = 0.006 and odds ratio 4.9 (95 % CI 1.5-9.7), p = 0.004]. Furthermore, to predict PAD, adding high CRP levels and low EPA levels to the established risk factors significantly improved the area under the ROC curves, from 0.66 to 0.78, of the PAD prediction model (p = 0.004). A significant relationship among EPA, CRP, and PAD was confirmed in patients with CAD.

Docetaxel administered through a novel lymphatic drug delivery system (LDDS) improved treatment outcomes for lymph node metastasis.

Recently, sentinel lymph nodes (LNs) have been recognized as a starting point of hematogenous metastasis; thus, an increase in the control rate of LN metastasis is expected to improve the survival rate. Although surgical treatment and radiation therapy are commonly used for the radical treatment of LNs, these treatments are associated with lymphedema, pain, and an extended hospital stay. In a recent mouse study, activation of metastatic tumors in distant organs was reported after removing LNs, with or without metastasis to the LNs. Thus, there is the necessity for cancer treatment that can replace LN removal. Here, we evaluated the treatment efficacy of lymphatic drug delivery system (LDDS) with osmotic pressure and viscosity escalated Docetaxel at the early stage of LN metastasis. MXH10/Mo/lpr mice were inoculated with mouse breast cancer cells into Subiliac LN to create the metastatic mouse model. Docetaxel was injected into mouse mammary carcinoma cells inoculated LN as a single shot (SS) or double shot (DS) to understand the therapeutic mechanism of a single shot or double shot intervention using an in vivo imaging system, histology, and qPCR. The results showed that the DS administration of docetaxel at 1,960 kPa (12 mPa∙s) had better therapeutic outcomes with increased complete response and improved survival with reduced adverse events. The results also revealed that administration of a DS of docetaxel enhances differentiation of T helper cells, and improves survival and therapeutic outcomes. From a safety perspective, LDDS-administered DS of low-concentration docetaxel without any other anticancer treatments to LNs a novel approach to cancer management of LN metastasis. We emphasize that LDDS is a groundbreaking method of delivering anticancer drugs specifically to cancer susceptible LNs and is designed to enhance the effectiveness of cancer treatment while minimizing side effects.

Postoperative Changes of Spinopelvic Sagittal Parameters After Cervical Laminoplasty for Cervical Spondylotic Myelopathy.

STUDY DESIGN: A retrospective analysis of prospectively collected data. OBJECTIVE: To investigate postoperative changes of spinopelvic sagittal parameters after laminoplasty for cervical spondylotic myelopathy (CSM) accompanying postoperative cervical kyphotic deformity or cervical regional sagittal imbalance. SUMMARY OF BACKGROUND DATA: To the best of our knowledge, no study has been reported concerning postoperative changes of spinopelvic sagittal parameters accompanying postoperative deterioration of cervical sagittal alignment or balance after cervical laminoplasty. METHODS: Forty-five CSM patients without preoperative cervical kyphosis who underwent laminoplasty were included. None of the 45 patients had a medical history of previous spine surgery, hip joint surgery, or knee joint surgery. The patients were divided into 2 groups (kyphosis and lordosis groups) according to postoperative C2-7 angle, and they were also divided into 2 other groups (imbalance and balance groups) according to postoperative C1-7 sagittal vertical axis. Postoperative changes (Δ) of T1 slope (T1S), thoracic kyphosis, thoracolumbar kyphosis (TLK), lumbar lordosis (LL), Pelvic tilt, and C7 sagittal vertical axis were measured comparing lateral radiographs of the whole spine in the standing position taken at 1 year postoperatively with those before surgery. RESULTS: Both T1S and TLK significantly decreased after cervical laminoplasty in the kyphosis group compared with the lordosis group. On the other hand, both T1S and TLK increased significantly, and LL significantly decreased after surgery in the imbalance group compared with the balance group. CONCLUSIONS: At 1 year after laminoplasty for CSM, both T1S and TLK significantly decreased accompanying postoperative cervical kyphotic deformity as a compensatory action for postoperative cervical kyphosis to maintain the global sagittal balance of the spine, whereas both T1S and TLK increased significantly, and LL significantly decreased accompanying postoperative cervical reginal sagittal imbalance which resulted in postoperative forward inclination of the whole spine.

Primary Oral Mixed Neuroendocrine-Non-neuroendocrine Neoplasm (MiNEN): A Rare Case Report and Review of the Literature.

Mixed neuroendocrine-non-neuroendocrine neoplasms (MiNENs) are rare tumors recently characterized by the presence of both neuroendocrine and non-neuroendocrine components within the same tumor tissue. Although MiNEN found their place in the WHO classification for various organs, this composite tumor in the head and neck region remains exceptionally rare. We present a case of primary oral MiNEN in a 64-year-old male located on the left side of lower gingiva. Biopsy raised suspicion of neuroendocrine carcinoma (NEC) and the patient underwent partial mandibulectomy. The resected specimen showed two distinct components of NEC and squamous cell carcinoma (SCC) with the confirmation of immunohistochemical markers. There has been no sign of recurrence nor metastasis 6 years after the surgery. In addition, we have conducted a review of published cases with potential relevance to this entity, resulting in five cases. The diverse terminology reinforces the need for a standardized classification system of oral/head and neck MiNENs.

Numb Chin Syndrome as the Initial Presentation of Mandibular Metastasis of Colorectal Cancer: A Case Report.

Numb chin syndrome (NCS) is hypesthesia of the mandible and lower lip caused by damage to the inferior alveolar or mandibular nerves, commonly due to dental treatment or osteomyelitis, but occasionally caused by malignant tumors. We report the case of a male in his 60s. He came to our hospital with a chief complaint of mandibular pain and paresthesia in the right side of the mental region. He had noticed swelling of the left mandible one month before the initial visit and strong hypesthesia of the right side of the mental region one week before the initial visit. Panoramic radiographs showed slight osteosclerosis of the left side mandible at the initial visit. Blood tests showed only a slight inflammatory reaction. The diagnosis of mandibular osteomyelitis and numb chin syndrome was made, and a contrast-enhanced CT scan was performed to investigate the possibility of neoplastic lesions, but no obvious cause was found. Osteosclerosis was minimal. A tissue biopsy was recommended, but the patient did not consent. Considering the possibility of NCS due to a hematologic disorder, the patient was referred to a hematologist, but no cause could be identified at the initial visit. With time, the markedly severe pain worsened, and the possibility of a neoplastic lesion was again suspected. Blood tests were performed, which revealed abnormally high levels of CA19 and CEA. He consulted a gastroenterologist, who found a tumor in the ileocecal region on contrast-enhanced CT, and multiple systemic metastases were found on a PET-CT scan the next day. Systemic chemotherapy was administered for multiple metastatic unresectable colorectal cancer (cT4N1aMc2 stage IVc).

Microvascular reconstruction for oral cancer in older adult patients: the impact of age on surgical outcomes.

OBJECTIVE: Flap complications continue to be a challenge in microsurgical reconstruction for older adults. We aimed to evaluate the impact of age on surgical outcomes after microvascular reconstruction. STUDY DESIGN: We retrospectively investigated 103 patients with oral squamous cell carcinoma who had undergone microvascular reconstruction surgery to compare microsurgical reconstruction, common postoperative complications, and flap success rates in geriatric (>75 years) and non-geriatric (<75 years) patients. We also evaluated differences based on the American Society of Anesthesiologists Physical Status score. RESULTS: We found no significant differences between the geriatric and non-geriatric groups in peri-operative, postoperative, or general complications. Conversely, we found that delirium and aspiration pneumonia were significantly more likely to occur in geriatric patients and that multiple medical complications were significantly more likely to occur in geriatric patients with a high American Society of Anesthesiologists score. CONCLUSION: Microvascular reconstruction can be performed effectively and without excessive complications in geriatric patients, and age should not be considered a contraindication for this procedure. Comorbidities play a stronger role in the prediction of adverse events.

Detection of oral cancer and oral potentially malignant disorders using artificial intelligence-based image analysis.

BACKGROUND: We aimed to construct an artificial intelligence-based model for detecting oral cancer and dysplastic leukoplakia using oral cavity images captured with a single-lens reflex camera. SUBJECTS AND METHODS: We used 1043 images of lesions from 424 patients with oral squamous cell carcinoma (OSCC), leukoplakia, and other oral mucosal diseases. An object detection model was constructed using a Single Shot Multibox Detector to detect oral diseases and their locations using images. The model was trained using 523 images of oral cancer, and its performance was evaluated using images of oral cancer (n = 66), leukoplakia (n = 49), and other oral diseases (n = 405). RESULTS: For the detection of only OSCC versus OSCC and leukoplakia, the model demonstrated a sensitivity of 93.9% versus 83.7%, a negative predictive value of 98.8% versus 94.5%, and a specificity of 81.2% versus 81.2%. CONCLUSIONS: Our proposed model is a potential diagnostic tool for oral diseases.

Identification of Neck Lymph Node Metastasis-Specific microRNA-Implication for Use in Monitoring or Prediction of Neck Lymph Node Metastasis.

MicroRNAs (miRNAs) have attracted attention as non-invasive cancer biomarkers in various cancers; however, they have not been adequately investigated in oral squamous cell carcinoma (OSCC). This study investigated the diagnostic performance of serum-derived miRNAs at initial diagnosis for primary neck lymph node metastasis and the predictive performance for late neck lymph node metastasis based on long-term (up to approximately 8 years) follow-up of patients with OSCC. The expression of miRNAs in 40 patients with OSCC was quantified using real-time PCR (qPCR), and a comprehensive statistical analysis of the correlation of miRNA expression for primary and late neck lymph node metastases was performed. For the diagnosis of primary neck lymph node metastases, miR-423 and miR-125 were accurate. The miRNA index for primary metastasis diagnosis (miR-PM) calculated by regression analysis showed high diagnostic accuracy. The miR-5100 was useful for predicting late neck lymph node metastases. The miRNA index for late metastasis prediction (miR-LM) calculated using regression analysis showed high prediction accuracy. MiRNAs were useful for diagnosing primary neck lymph node metastases in OSCC and predicting late neck lymph node metastases. It may help to consider individualized treatment, including follow-up, surgical methods, and postoperative management.

Are intratumoral microbiota involved in the progression of intraductal papillary mucinous neoplasms of the pancreas?

BACKGROUND: Microbiota have been reported to influence the development of various gastrointestinal neoplasms through the mechanism of sustained inflammation; however, few data are available regarding their influence on intraductal papillary mucinous neoplasms. The aim of this study was to assess the association between specific microbiota and the clinicopathologic characteristics of intraductal papillary mucinous neoplasms of the pancreas. METHODS: DNA was extracted from formalin-fixed, paraffin-embedded samples of 30 patients who underwent pancreatectomy for intraductal papillary mucinous neoplasm, and polymerase chain reaction was used to create sequence libraries using the primer set for the V3 and V4 region of 16S recombinant DNA. Filtered sequence reads were then processed into operational taxonomic units with a 97% identity threshold and the relative abundance of bacteria compared between the 2 groups using operational taxonomic units. RESULTS: There was a trend toward fewer Firmicutes and more Proteobacteria and Fusobacteria in the relative abundance of main duct operational taxonomic units than in branch duct operational taxonomic units. The relative abundances of Bacteroidetes (P < .01) and Fusobacteria (P = .04) were significantly higher in invasive intraductal papillary mucinous neoplasms than in noninvasive intraductal papillary mucinous neoplasms. The relative abundance of the intestinal type was significantly lower in Firmicutes than the relative abundance of the nonintestinal type (P = .04). Notably, main duct operational taxonomic units with the intestinal subtype were affected by increased proportions of Proteobacteria and Fusobacteria, and Fusobacteria were abundant in the intestinal type of invasive main duct operational taxonomic units. CONCLUSION: Intratumoral microbiota may be involved in the progression of operational taxonomic units.

Bimodal artificial intelligence using TabNet for differentiating spinal cord tumors-Integration of patient background information and images.

We proposed a bimodal artificial intelligence that integrates patient information with images to diagnose spinal cord tumors. Our model combines TabNet, a state-of-the-art deep learning model for tabular data for patient information, and a convolutional neural network for images. As training data, we collected 259 spinal tumor patients (158 for schwannoma and 101 for meningioma). We compared the performance of the image-only unimodal model, table-only unimodal model, bimodal model using a gradient-boosting decision tree, and bimodal model using TabNet. Our proposed bimodal model using TabNet performed best (area under the receiver-operating characteristic curve [AUROC]: 0.91) in the training data and significantly outperformed the physicians' performance. In the external validation using 62 cases from the other two facilities, our bimodal model showed an AUROC of 0.92, proving the robustness of the model. The bimodal analysis using TabNet was effective for differentiating spinal tumors.

The T-box transcription factor Brachyury regulates epithelial-mesenchymal transition in association with cancer stem-like cells in adenoid cystic carcinoma cells.

BACKGROUND: The high frequencies of recurrence and distant metastasis of adenoid cystic carcinoma (AdCC) emphasize the need to better understand the biological factors associated with these outcomes. To analyze the mechanisms of AdCC metastasis, we established the green fluorescence protein (GFP)-transfected subline ACCS-GFP from the AdCC parental cell line and the metastatic ACCS-M GFP line from an in vivo metastasis model. METHODS: Using these cell lines, we investigated the involvement of the epithelial-mesenchymal transition (EMT) and cancer stem cell (CSCs) in AdCC metastasis by real-time RT-PCR for EMT related genes and stem cell markers. Characteristics of CSCs were also analyzed by sphere-forming ability and tumorigenicity. Short hairpin RNA (shRNA) silencing of target gene was also performed. RESULTS: ACCS-M GFP demonstrated characteristics of EMT and additionally displayed sphere-forming ability and high expression of EMT-related genes (Snail, Twist1, Twist2, Slug, zinc finger E-box binding homeobox 1 and 2 [Zeb1 and Zeb2], glycogen synthase kinase 3 beta [Gsk3ß and transforming growth factor beta 2 [Tgf-ß2]), stem cell markers (Nodal, Lefty, Oct-4, Pax6, Rex1, and Nanog), and differentiation markers (sex determining region Y [Sox2], Brachyury, and alpha fetoprotein [Afp]). These observations suggest that ACCS-M GFP shows the characteristics of CSCs and CSCs may be involved in the EMT of AdCC. Surprisingly, shRNA silencing of the T-box transcription factor Brachyury (also a differentiation marker) resulted in downregulation of the EMT and stem cell markers. In addition, sphere-forming ability, EMT characteristics, and tumorigenicity were simultaneously lost. Brachyury expression in clinical samples of AdCC was extremely high and closely related to EMT. This finding suggests that regulation of EMT by Brachyury in clinical AdCC may parallel that observed in vitro in this study. CONCLUSIONS: The use of a single cell line is a limitation of this study. However, parallel data from in vitro and clinical samples suggest the possibility that EMT is directly linked to CSCs and that Brachyury is a regulator of EMT and CSCs.

Pharmacokinetic analysis based on dynamic contrast-enhanced MRI for evaluating tumor response to preoperative therapy for oral cancer.

PURPOSE: To evaluate whether a pharmacokinetic analysis is useful for monitoring the response of oral cancer to chemoradiotherapy (CRT). MATERIALS AND METHODS: Twenty-nine patients were included. They underwent dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) before and after CRT. The DCE-MRI data were analyzed using a Tofts and Kermode (TK) model. The histological evaluation of the effects of CRT was performed according to Ohboshi and Shimosato's classification. RESULTS: None of the pre-CRT parameters were significantly different between the responders and nonresponders. The post-CRT volume of the extravascular extracellular space (EES) per unit volume of tissue (v(e) ) of responders (0.397 ± 0.080) was higher than that of nonresponders (0.281 ± 0.076) (P = 0.01). The change of the v(e) between the pre- and post-CRT of the responders (0.154 ± 0.093) was larger than that of the nonresponders (0.033 ± 0.073) (P = 0.001). Therefore, the increase in the v(e) strongly suggested a good tumor response to CRT, which reflected an increase of the EES secondary to the destruction of the cancer nest. The changes in the volume transfer constant (K(trans) ) were significantly different between the responders and nonresponders (P = 0.018). CONCLUSION: Both the increase of the v(e) and the elevation of permeability (K(trans) ) were indicative of a good tumor response to CRT. The pharmacokinetic analysis had potential for monitoring the histopathological response to CRT.