Notch1 inactivation promotes invasion and metastasis of nasopharyngeal carcinoma cells partly through Slug activation.

Nasopharyngeal carcinoma (NPC) is a rare form of the head and neck cancer of the epithelial lining of the nasopharynx and exhibits the highest metastatic rate among head and neck cancers. Underlying mechanisms of metastasis remain largely unknown. Here, we explored whether Notch1 affects the invasion and metastasis of NPC cells. In vitro migration and invasion capacities were evaluated after the knockdown of Notch1 expression in NPC cells. To investigate the role of Notch1 in in vivo metastasis, we examined the metastatic ability to the lungs following administration of cancer cells via mouse tail vein. The expression of epithelial-mesenchymal transition (EMT) markers associated with Notch1-mediated metastasis was investigated, and their roles in metastasis and relationship with Notch1 expression were investigated. Suppression of Notch1 expression increased the ability of NPC cells to invade Matrigel in vitro. Knockdown of Notch1 expression in NPC cells resulted in extensive lung metastasis in a mouse model and increased the mRNA expression of Slug in NPC cells. Slug-specific RNA interference resulted in the loss of the metastatic and invasion capacities in Notch1-suppressed NPC cells. These findings show that Notch1 has a significant suppressive role in the regulation of metastasis in NPCs, suggestive of its prudent use in clinical trials.

Cyclin-dependent kinase 7 is a therapeutic target in high-grade glioma.

High-grade glioma (HGG) is an incurable brain cancer. The transcriptomes of cells within HGG tumors are highly heterogeneous. This renders the tumors unresponsive or able to adapt to therapeutics targeted at single pathways, thereby causing treatment failure. To overcome this, we focused on cyclin-dependent kinase 7 (CDK7), a ubiquitously expressed molecule involved in two major drivers of HGG pathogenesis: cell cycle progression and RNA polymerase-II-based transcription. We tested the activity of THZ1, an irreversible CDK7 inhibitor, on patient-derived primary HGG cell lines and ex vivo HGG patient tissue slices, using proliferation assays, microarray analysis, high-resolution respirometry, cell cycle analysis and in vivo tumor orthografts. The cellular processes affected by CDK7 inhibition were analyzed by reverse transcriptase-quantitative PCR, western blot, flow cytometry and immunofluorescence. THZ1 perturbed the transcriptome and disabled CDK activation, leading to cell cycle arrest at G2 and DNA damage. THZ1 halted transcription of the nuclear-encoded mitochondrial ribosomal genes, reducing mitochondrial translation and oxidative respiration. It also inhibited the expression of receptor tyrosine kinases such as epidermal growth factor receptor (EGFR) and platelet-derived growth factor receptor-α (PDGFR-α), reducing signaling flux through the AKT, extracellular-signal-regulated kinase 1/2 (ERK1/2) and signal transducer and activator of transcription 3 (STAT3) downstream pathways. Finally, THZ1 disrupted nucleolar, Cajal body and nuclear speckle formation, resulting in reduced cytosolic translation and malfunction of the spliceosome and thus leading to aberrant mRNA processing. These findings indicate that CDK7 is crucial for gliomagenesis, validate CDK7 as a therapeutic target and provide new insight into the cellular processes that are affected by THZ1 and induce antitumor activity.

A randomised controlled trial comparing meat-based with human cadaveric models for teaching ultrasound-guided regional anaesthesia.

The aim of this prospective, blinded, randomised controlled study was to compare novices' acquisition of the technical skills of ultrasound-guided regional anaesthesia using either a meat phantom model or fresh-frozen human cadavers. The primary outcome was the time taken to successfully perform an ultrasound-guided sciatic nerve block on a cadaver; secondary outcomes were the cumulative score of errors, and best image quality of the sciatic nerve achieved. After training, the median (IQR [range]) time taken to perform the block was 311(164-390 [68-600]) s in the meat model trained group and 210 (174-354 [85-600]) s in the fresh-frozen cadaver trained group (p = 0.24). Participants made a median (IQR [range]) of 18 (14-33 [8-55]) and 15 (12-22 [8-44]) errors in the two groups respectively (p = 0.39). The image quality score was also not different, with a median (IQR [range]) of 62.5 (59.4-65.6 [25.0-100.0])% vs 62.5 (62.5-75.0 [25.0-87.5])% respectively (p = 0.58). The training and deliberate feedback improved all participants' block performance, the median (IQR [range]) times being 310 (206-532 [110-600]) s before and 240 (174-354 [85-600]) s after training (p = 0.02). We conclude that novices taught ultrasound scanning and needle guidance skills using an inexpensive and easily constructed meat model perform similarly to those trained on a cadaveric model.

Tracking metastatic breast cancer: the future of biology in biosensors.

Circulating tumour cells associated with breast cancer (brCTCs) represent cells that have the capability to establish aggressive secondary metastatic tumours. The isolation and characterization of CTCs from blood in a single device is the future of oncology diagnosis and treatment. The methods of enrichment of CTCs have primarily utilized simple biological interactions with bimodal reporting with biased high purity and low numbers or low purity and high background. In this review, we will discuss the advances in microfluidics that has allowed the use of more complex selection criteria and biological methods to identify CTC populations. We will also discuss a potential new method of selection based on the response of the oncogenic DNA repair pathways within brCTCs. This method would allow insight into not only the oncogenic signalling at play but the chemoresistance mechanisms that could guide future therapeutic intervention at any stage of disease progression.

Design and validation of the Regional Anaesthesia Procedural Skills Assessment Tool.

The aim of this study was to create and evaluate the validity, reliability and feasibility of the Regional Anaesthesia Procedural Skills tool, designed for the assessment of all peripheral and neuraxial blocks using all nerve localisation techniques. The first phase was construction of a 25-item checklist by five regional anaesthesia experts using a Delphi process. This checklist was combined with a global rating scale to create the tool. In the second phase, initial validation by 10 independent anaesthetists using a test-retest methodology was successful (Cohen kappa ≥ 0.70 for inter-rater agreement, scores between test to retest, paired t-test, p > 0.12). In the third phase, 70 clinical videos of trainees were scored by three blinded international assessors. The RAPS tool exhibited face validity (p < 0.026), construct validity (p < 0.001), feasibility (mean time to score < 3.9 min), and overall reliability (intraclass correlation coefficient 0.80 (95% CI 0.67-0.88)). The Regional Anaesthesia Procedural Skills tool used in this study is a valid and reliable assessment tool to score the performance of trainees for regional anaesthesia.

The LIM-only transcription factor LMO2 determines tumorigenic and angiogenic traits in glioma stem cells.

Glioblastomas (GBMs) maintain their cellular heterogeneity with glioma stem cells (GSCs) producing a variety of tumor cell types. Here we interrogated the oncogenic roles of Lim domain only 2 (LMO2) in GBM and GSCs in mice and human. High expression of LMO2 was found in human patient-derived GSCs compared with the differentiated progeny cells. LMO2 is required for GSC proliferation both in vitro and in vivo, as shRNA-mediated LMO2 silencing attenuated tumor growth derived from human GSCs. Further, LMO2 is sufficient to induce stem cell characteristics (stemness) in mouse premalignant astrocytes, as forced LMO2 expression facilitated in vitro and in vivo growth of astrocytes derived from Ink4a/Arf null mice and acquisition of GSC phenotypes. A subset of mouse and human GSCs converted into vascular endothelial-like tumor cells both in vitro and in vivo, which phenotype was attenuated by LMO2 silencing and promoted by LMO2 overexpression. Mechanistically, the action of LMO2 for induction of glioma stemness is mediated by transcriptional regulation of Jagged1 resulting in activation of the Notch pathway, whereas LMO2 directly occupies the promoter regions of the VE-cadherin gene for a gain of endothelial cellular phenotype. Subsequently, selective ablation of human GSC-derived VE-cadherin-expressing cells attenuated vascular formation in mouse intracranial tumors, thereby significantly prolonging mouse survival. Clinically, LMO2 expression was elevated in GBM tissues and inversely correlated with prognosis of GBM patients. Taken together, our findings describe novel dual roles of LMO2 to induce tumorigenesis and angiogenesis, and provide potential therapeutic targets in GBMs.

Oct4 is a critical regulator of stemness in head and neck squamous carcinoma cells.

Cancer stem cells (CSCs) have been suggested as responsible for the initiation and progression of cancers. Octamer-binding transcription factor 4 (Oct4) is an important regulator of embryonic stem cell fate. Here, we investigated whether Oct4 regulates stemness of head and neck squamous carcinoma (HNSC) CSCs. Our study showed that ectopic expression of Oct4 promotes tumor growth through cyclin E activation, increases chemoresistance through ABCC6 expression and enhances tumor invasion through slug expression. Also, Oct4 dedifferentiates differentiated HNSC cells to CSC-like cells. Furthermore, Oct4(high) HNSC CSCs have more stem cell-like traits compared with Oct4(low) cells, such as self-renewal, stem cell markers' expression, chemoresistance, invasion capacity and xenograft tumorigeneity in vitro and in vivo. In addition, knockdown of Oct4 led to markedly lower HNSC CSC stemness. Finally, there was a significant correlation between Oct4 expression and survival of 119 HNSC patients. Collectively, these data suggest that Oct4 may be a critical regulator of HNSC CSCs and its targeting may be potentially valuable in the treatment of HNSC CSCs.

SOX2 regulates self-renewal and tumorigenicity of stem-like cells of head and neck squamous cell carcinoma.

BACKGROUND: Head and neck squamous cell carcinomas (HNSCCs) display cellular heterogeneity and contain cancer stem cells (CSCs). Sex-determining region Y [SRY]-box (SOX)2 is an important regulator of embryonic stem cell fate and is aberrantly expressed in several types of human tumours. Nonetheless, the role of SOX2 in HNSCC remains unclear. METHODS: We created cells ectopically expressing SOX2 from previously established HNSCC cells and examined the cell proliferation, self-renewal capacity, and chemoresistance of these cells compared with control cells. In addition, we knocked down SOX2 in primary spheres obtained from HNSCC tumour tissue and assessed the attenuation of stemness-associated traits in these cells in vitro and in vivo. Furthermore, we examined the clinical relevance of SOX2 expression in HNSCC patients. RESULTS: SOX2 is aberrantly expressed in primary tissue of HNSCC patients but not in healthy tissue. SOX2 expression correlated with tumour recurrence and poor prognosis of HNSCC patients. Ectopic expression of SOX2 induced cell proliferation via cyclin B1 expression and stemness-associated features, such as self-renewal and chemoresistance. In addition, a knockdown of SOX2 in HNSCC CSCs attenuated their self-renewal capacity, chemoresistance (through ABCG2 suppression), invasion capacity (via snail downregulation), and in vivo tumorigenicity. CONCLUSIONS: These results suggest that SOX2 may have important roles in the 'stemness' and progression of HNSCC. Targeting SOX2-positive tumour cells (CSCs) could be a new therapeutic strategy in HNSCCs.

Forced arterial suction thrombectomy with the penumbra reperfusion catheter in acute basilar artery occlusion: a retrospective comparison study in 2 Korean university hospitals.

BACKGROUND AND PURPOSE: A performance of forced arterial suction thrombectomy was not reported for the treatment of acute basilar artery occlusion. This study compared revascularization performance between intra-arterial fibrinolytic treatment and forced arterial suction thrombectomy with a Penumbra reperfusion catheter in patients with acute basilar artery occlusion. MATERIALS AND METHODS: Fifty-seven patients with acute basilar artery occlusion were treated with intra-arterial fibrinolysis (n = 25) or forced arterial suction thrombectomy (n = 32). Baseline characteristics, successful revascularization rate, and clinical outcomes were compared between the groups. RESULTS: Baseline characteristics, the frequency of patients receiving intravenous recombinant tissue plasminogen activator, and mean time interval between symptom onset and femoral puncture did not differ between groups. The forced arterial suction thrombectomy group had a shorter procedure duration (75.5 minutes versus 113.3 minutes, P = .016) and higher successful revascularization rate (88% versus 60%, P = .017) than the fibrinolysis group. Fair outcome, indicated by a modified Rankin Scale 0-3, at 3 months was achieved in 34% of patients undergoing forced arterial suction thrombectomy and 8% of patients undergoing fibrinolysis (P = .019), and the mortality rate was significantly higher in the fibrinolysis group (25% versus 68%, P = .001). Multiple logistic regression analysis identified the forced arterial suction thrombectomy method as an independent predictor of fair outcome with adjustment for age, sex, initial NIHSS score, and the use of intravenous recombinant tissue plasminogen activator (odds ratio, 7.768; 95% CI, 1.246-48.416; P = .028). CONCLUSIONS: In acute basilar artery occlusion, forced arterial suction thrombectomy demonstrated a higher revascularization rate and improved clinical outcome compared with traditional intra-arterial fibrinolysis. Further clinical trials with the newer Penumbra catheter are warranted.

A randomised controlled trial of ultrasound-assisted spinal anaesthesia.

Pre-procedural ultrasound scanning has been used to facilitate spinal anaesthesia in patients with difficult anatomical landmarks and shown to improve first-attempt success rates in some studies. We studied whether pre-procedural ultrasound scanning improved first-attempt success rate and decreased time taken for the procedure in the general adult population. In this prospective, randomised controlled trial, 170 American Society of Anesthesiologists 1 to 3 patients aged between 21 and 80 years were recruited. Informed consent was obtained. Patients were randomised into two groups, ultrasound-guided identification of landmarks (Ultrasound Group) and manual palpation of landmarks (Manual Palpation Group). The primary outcome was first-attempt success rate and secondary outcomes were time taken to perform procedure, number of needle redirections, patient satisfaction and complications. The first-attempt success rate was 64% in the Ultrasound Group and 52% in the Manual Palpation Group (P=0.16). Time taken for procedure was shorter in the Ultrasound Group compared to the Manual Palpation Group (2.9±3.6 minutes versus 3.9±3.7 min, P= 0.007). Patient satisfaction was higher in the Ultrasound Group. There were no differences in complications. As there was no statistically significant difference in first-attempt success rates between the two groups, existing evidence for routine pre-procedural scanning for all patients is inadequate. The current use of pre-procedural ultrasound scanning will probably be limited to selected patients where spinal anaesthesia may be technically challenging with conventional methods.

Isolation and characterization of antifungal compound from Lactobacillus plantarum KCC-10 from forage silage with potential beneficial properties.

AIMS: The purpose of this study was to isolate, identify and characterize an antifungal compound from Lactobacillus plantarum KCC-10 from forage silage with potential beneficial properties. METHODS AND RESULTS: The 16S rRNA gene-based phylogenetic affiliation was determined using bioinformatic tools and identified as Lactobacillus sp. KCC-10 with 100% sequence similarity to L. plantarum. The antifungal substances were extracted with ethyl acetate from spent medium in which Lactobacillus sp. KCC-10 was cultivated. Antifungal activity was assessed using the broth microdilution technique. The compounds were obtained by eluting the crude extract with various concentrations of solvents followed by chromatographic purification. Based on the infrared, (13) C nuclear magnetic resonance (NMR) and (1) H NMR spectral data, the compound was identified as a phenolic-related antibiotic. The minimum inhibitory concentration of the compound against Aspergillus clavatus, A. oryzae, Botrytis elliptica and Scytalidium vaccinii was 2.5 mg ml(-1) and that against A. fumigatus, A. niger and S. fusca was 5.0 mg ml(-1) , respectively. In addition, Lactobacillus sp. KCC-10 was highly sensitive towards oxgall (0.3%) but grew well in the presence of sodium taurocholate (0.3%). An antimicrobial susceptibility pattern was an intrinsic feature of this strain; thus, consumption does not represent a health risk to humans or animals. CONCLUSION: Novel L. plantarum KCC-10 with antifungal and potential probiotic properties was characterized for use in animal food. SIGNIFICANCE AND IMPACT OF THE STUDY: This study revealed that L. plantarum KCC-10 exhibited good antifungal activity similar to that of probiotic Lactobacillus strains.

Comparison of the Incidence of parenchymal hematoma and poor outcome in patients with carotid terminus occlusion treated with intra-arterial urokinase alone or with combined IV rtPA and intra-arterial urokinase.

BACKGROUND AND PURPOSE: Patients with acute CTO generally have a poor prognosis, despite IV or IA thrombolytic treatment. The goal of this study was to analyze the results of patients with CTO who had IA urokinase treatment with or without initial IV rtPA based on a bridging protocol. MATERIALS AND METHODS: Sixteen consecutive patients with acute ischemic stroke due to CTO who had combined IV and IA or a single IA thrombolytic treatment were enrolled. The baseline characteristics and prognosis were described. The patients who did and did not develop a PH shortly after treatment were compared. RESULTS: The mean age was 66.4 years, and the median initial NIHSS score was 17. The median dose of IA urokinase was 320,000 U, and recanalization (TICI grade II-III) was achieved in 12 patients (75%). However, 5 patients died and 10 patients had poor prognosis with mRS 5-6 at discharge. Six patients (37.5%) with a PH had a higher NIHSS score 1 day after treatment (26.7 versus 13.6, P = .002), and they had more frequent mortality (66.7% versus 10.0%, P = .018) and worse prognosis (mRS 5-6; 100% versus 40%, P = .016) at discharge than patients without PH. CONCLUSIONS: Patients with CTO who received IA urokinase treatment based on a bridging protocol had a poor prognosis. The development of PH might affect this outcome.

A novel role for hSMG-1 in stress granule formation.

hSMG-1 is a member of the phosphoinositide 3 kinase-like kinase (PIKK) family with established roles in nonsense-mediated decay (NMD) of mRNA containing premature termination codons and in genotoxic stress responses to DNA damage. We report here a novel role for hSMG-1 in cytoplasmic stress granule (SG) formation. Exposure of cells to stress causing agents led to the localization of hSMG-1 to SG, identified by colocalization with TIA-1, G3BP1, and eIF4G. hSMG-1 small interfering RNA and the PIKK inhibitor wortmannin prevented formation of a subset of SG, while specific inhibitors of ATM, DNA-PK(cs), or mTOR had no effect. Exposure of cells to H(2)O(2) and sodium arsenite induced (S/T)Q phosphorylation of proteins. While Upf2 and Upf1, an essential substrate for hSMG-1 in NMD, were present in SG, NMD-specific Upf1 phosphorylation was not detected in SG, indicating hSMG-1's role in SG is separate from classical NMD. Thus, SG formation appears more complex than originally envisaged and hSMG-1 plays a central role in this process.

Ministry of Health clinical practice guidelines: management of gambling disorders.

The Ministry of Health (MOH) has published clinical practice guidelines on Management of Gambling Disorders to provide doctors and patients in Singapore with evidence-based guidance on the management of gambling disorders. This article reproduces the introduction and executive summary (with recommendations from the guidelines) from the MOH clinical practice guidelines on Management of Gambling Disorders for the information of readers of the Singapore Medical Journal. Chapters and page numbers mentioned in the reproduced extract refer to the full text of the guidelines, which are available from the Ministry of Health website (http://www.moh.gov.sg/mohcorp/publications.aspx?id=26136). The recommendations should be used with reference to the full text of the guidelines. Following this article are multiple choice questions based on the full text of the guidelines.

Aptasensors: a review.

Aptamers are a promising class of agents for biomolecules detection due to their small size, chemical stability and cost effectiveness over conventional bioreceptors such as antibodies. Recent advances in micro/nano-fabrication and biotechnology have driven active participation of engineers and molecular biologists in the development of aptasensors. This review examines aptasensors from a developer standpoint discussing surface immobilization techniques and mechanisms used to detect biomolecular interactions in the context of biotechnology and nanomedicine. The factors that affect accuracy, sensitivity and stability of aptasensors are also addressed.

CD137 enhances monocyte-ICAM-1 interactions in an E-selectin-dependent manner under flow conditions.

Expression of CD137, a member of the tumor necrosis factor receptor family, is inducible on vascular endothelial cells by proinflammatory stimuli. Its ligand is expressed as a transmembrane protein on the surface of monocytes, and transmits activating signals into monocytes (reverse signaling) inducing monocyte migration. These findings led to the hypothesis that CD137 expression on activated endothelial cells facilitates recruitment of monocytes into inflammatory tissues including atherosclerotic lesions. Data from this study demonstrate that CD137 expression is inducible on endothelial cells by TNF stimulation. Recombinant CD137 protein and CD137 expressed on activated endothelial cells enhance ICAM-1-mediated adhesion of monocytes under defined flow conditions in vitro. CD137 seems not to play a role in tethering of monocytes since this activity is completely E-selectin-dependent. In addition, LFA-1 affinity and clustering on monocytic cells is enhanced by CD137. In summary, CD137 expression is induced on vascular endothelial cells by proinflammatory mediators and strengthens ICAM-1 and LFA-1-mediated adhesion of monocytes. These data support a role for CD137 in the recruitment of monocytes to inflammatory tissues.

Pharmacokinetics and bioequivalence evaluation of gliclazide/metformin combination tablet and equivalent doses of gliclazide and metformin in healthy Korean subjects.

OBJECTIVE: To evaluate the bioequivalence of a gliclazide/metformin combination tablet (at dose of 80/500 mg) with co-administration of metformin (500 mg) and gliclazide (80 mg) as individual tablets in healthy male Korean volunteers. SUBJECTS, MATERIALS AND METHODS: The study was conducted as an open-label, randomized, 2-period crossover design in 32 healthy male Korean volunteers who received a combination tablet of gliclazide/metformin at a dose of 80/500 mg or co-administration of gliclazide and metformin as individual tablets in each study period. There was a 7-day washout period between doses. Serum concentrations of gliclazide and metformin up to 32 hours after administration were determined using a validated HPLC method with UV detection. The pharmacokinetic parameters such as AUC0-t (the area under the curve from zero to the time), AUC0- yen (the area under the curve from zero to infinity), Cmax (maximum serum concentration), tmax (time to reach Cmax) and t1/2 (terminal half-life), were analyzed by non-compartmental analysis. Analysis of variance (ANOVA) was carried out using logarithmically transformed AUC0-t, AUC0- yen and Cmax, and untransformed tmax. In addition, blood glucose concentration was also logarithmically transformed and analyzed. Tolerability and safety profiles were also investigated. RESULTS: There were no significant differences between the single combination tablet and the individual tablets in AUC0-t, AUC0- yen, Cmax and blood glucose concentration. The point estimates (90% confidence intervals) for AUC0-t, AUC0- yen and Cmax were 1.0293 (0.9476 - 1.1178), 1.0253 (0.9185 - 1.1443) and 1.0425 (0.9986 - 1.0883) for gliclazide, and 0.9887 (0.9137 - 1.0697), 0.9915 (0.9189 - 1.0697) and 0.9882 (0.9295 - 1.0505) for metformin, respectively, satisfying the bioequivalence criteria of 80 - 125% as proposed by the US FDA and the Korean legislation. Significant F test values were found between the subjects and subject nested sequence (SEQ) for AUC0-t and Cmax, indicating substantial inter-subject variation in the pharmacokinetics of gliclazide and metformin. However, a SEQ effect in the two-way crossover design did not impair the bioequivalence conclusion. No statistically significant differences were found for tmax and blood glucose concentration between two treatments. CONCLUSION: The combination tablet of gliclazide/metformin is bioequivalent to co-administration of individual tablets. As a result, the combination tablets are regarded therapeutically equivalent and exchangeable to the co-administration of individual tablets in clinical practice. Moreover, the combination tablets are expected to improve convenience and adherence to prescribed therapy and to contribute to better blood glucose control for patients with Type 2 diabetes.

Case report. Tuberculous cervical lymphadenitis mimicking metastatic lymph nodes from papillary thyroid carcinoma.

We report two unusual cases of tuberculous lymphadenitis mimicking metastatic lymph nodes from papillary thyroid carcinoma (PTC). Pre-operative ultrasonography of the cervical nodes suggested a metastasis with cystic necrosis and calcification in PTC patients, but permanent pathology revealed tuberculosis lymphadenitis after neck dissection. In cases suspicious for metastatic cervical nodes in patients with PTC, fine-needle aspiration cytology may be indicated for the differential diagnosis of tuberculosis lymphadenitis, especially in those who have experienced tuberculosis in the past.

Acute renal failure following detergent ingestion.

A 42-year-old woman with a history of depression and epilepsy ingested two types of household detergent and developed gastrointestinal symptoms, and subsequently acute renal failure. Coingestants included nontoxic quantities of paracetamol and therapeutic doses of sodium valproate and fluoxetine. The patient developed acute renal failure, and also had fever and unilateral ear inflammation. The acute renal failure resolved four days later. Patients presenting with detergent poisoning are typically screened and treated for gastrointestinal and respiratory toxicity. We discuss the mechanism of development of acute renal failure in our patient, review the literature linking detergent poisoning and nephrotoxicity, and propose a direct relationship between detergent poisoning and acute renal failure.