Differential subcellular localization and activity of kelch repeat proteins KLHDC1 and KLHDC2.

We have previously identified and characterized human KLHDC2/HCLP-1, a kelch repeat protein that interacts with and inhibits transcription factor LZIP. In this study, we identified and characterized a paralog of KLHDC2 called KLHDC1. KLHDC1 and KLHDC2 share about 50% identity at the level of amino acid sequence and both gene loci localize to human chromosome 14q21.3. This cluster of KLHDC1 and KLHDC2 genes is highly conserved in vertebrates ranging from pufferfish to human. Both genes are expressed highly in skeletal muscle, but weakly in various other tissues. While KLHDC2 was predominantly found in the nucleus, KLHDC1 is a cytoplasmic protein. Neither KLHDC1 nor KLHDC2 binds to actin. In addition, KLHDC1 was unable to inhibit LZIP/CREB3-mediated transcriptional activation. Thus, KLHDC1 and KLHDC2 have differential localization and activity in cultured mammalian cells.postprin

Analytical method for magnetic field calculation in a low-speed permanent-magnet harmonic machine

Magnetic-gearing effect has become increasingly attractive when designing direct-drive low-speed permanent-magnet machines. The machines derived from the magnetic-gearing effect can be termed as harmonic machines. Unlike the conventional types, harmonic machines rely on the field harmonics to achieve energy conversion and transmission. The detailed knowledge of the field distributions in the air gap is vitally important for predicting and optimizing its performance. In this paper, we present an analytical approach to calculate the magnetic field distribution in a low-speed permanent-magnet harmonic machine. A series-slot model which is composed of a group of partial differential equations concerning the scalar magnetic potential is built up. Then, the field solutions are obtained by using the method of separating variables and analyzing the field boundary conditions. Finally, the flux densities are derived from the scalar magnetic potentials. All the results agree well with those obtained from the finite element method. © 2011 IEEE.published_or_final_versio

Analytical method for magnetic field calculation in a low-speed permanent-magnet harmonic machine

Magnetic-gearing effect has become increasingly attractive when designing direct-drive low-speed permanent-magnet machines. The machines derived from the magnetic-gearing effect can be termed as harmonic machines. Unlike the conventional types, harmonic machines rely on the field harmonics to achieve energy conversion and transmission. The detailed knowledge of the field distributions in the air gap is vitally important for predicting and optimizing its performance. In this paper, we present an analytical approach to calculate the magnetic field distribution in a low-speed permanent-magnet harmonic machine. A series-slot model which is composed of a group of partial differential equations concerning the scalar magnetic potential is built up. Then, the field solutions are obtained by using the method of separating variables and analyzing the field boundary conditions. Finally, the flux densities are derived from the scalar magnetic potentials. All the results agree well with those obtained from the finite element method. © 2011 IEEE.published_or_final_versio

Do frailty and comorbidity indices improve risk prediction of 28-day ED reattendance? Reanalysis of an ED discharge nomogram for older people

Background: In older people, quantification of risk of reattendance after emergency department (ED) discharge is important to provide adequate post ED discharge care in the community to appropriately targeted patients at risk. Methods: We reanalysed data from a prospective observational study, previously used for derivation of a nomogram for stratifying people aged 65 and older at risk for ED reattendance. We investigated the potential effect of comorbidity load and frailty by adding the Charlson or Elixhauser comorbidity index and a ten-item frailty measure from our data to develop four new nomograms. Model I and model F built on the original nomogram by including the frailty measure with and without the addition of the Charlson comorbidity score; model E adapted for efficiency in the time-constrained environment of ED was without the frailty measure; and model P manually constructed in a purposeful stepwise manner and including only statistically significant variables. Areas under the ROC curve of models were compared. The primary outcome was any ED reattendance within 28 days of discharge. Results: Data from 1357 patients were used. The point estimate of the respective areas under ROC were 0.63 (O), 0.63 (I), 0.68 (E), 0.71 (P) and 0.63 (F). Conclusion: Addition of a comorbidity index to our previous model improves stratifying elderly at risk of ED reattendance. Our frailty measure did not demonstrate any additional predictive benefit

Preparation of exosomes for siRNA delivery to cancer cells

Extracellular vesicles, in particular exosomes, have recently gained interest as novel drug delivery vectors due to their biological origin, abundance, and intrinsic capability in intercellular delivery of various biomolecules. This work establishes an isolation protocol to achieve high yield and high purity of exosomes for siRNA delivery. Human Embryonic Kidney cells (HEK-293 cells) are cultured in bioreactor flasks and the culture supernatant (hereon referred to as conditioned medium) is harvested on a weekly basis to allow for enrichment of HEK-293 exosomes. The conditioned medium (CM) is pre-cleared of dead cells and cellular debris by differential centrifugation and is subjected to ultracentrifugation onto a sucrose cushion followed by a washing step, to collect the exosomes. Isolated HEK-293 exosomes are characterized for yield, morphology and exosomal marker expression by nanoparticle tracking analysis, protein quantification, electron microscopy and flow cytometry, respectively. Small interfering RNA (siRNA), fluorescently labeled with Atto655, is loaded into exosomes by electroporation and excess siRNA is removed by gel filtration. Cell uptake in PANC-1 cancer cells, after 24 h incubation at 37 °C, is confirmed by flow cytometry. HEK-293 exosomes are 107.0 ± 8.2 nm in diameter. The exosome yield and particle-to-protein ratio (P:P) ratio are 6.99 ± 0.22 × 1012 particle/mL and 8.3 ± 1.7 × 1010 particle/µg, respectively. The encapsulation efficiency of siRNA in exosomes is ~ 10-20%. Forty percent of the cells show positive signals for Atto655 at 24 h post-incubation. In conclusion, exosome isolation by ultracentrifugation onto sucrose cushion offers a combination of good yield and purity. siRNA could be successfully loaded into exosomes by electroporation and subsequently delivered into cancer cells in vitro. This protocol offers a standard procedure for developing siRNA-loaded exosomes for efficient delivery to cancer cells

Simultaneous dual-band optical coherence tomography for endoscopic applications

published_or_final_versio

Systematic review:genetic associations for prognostic factors of urinary bladder cancer

Introduction: Many germline associations have been reported for urinary bladder cancer (UBC) outcomes and prognostic characteristics. It is unclear whether there are overlapping genetic patterns for various prognostic endpoints. We aimed to review contemporary literature on genetic associations with UBC prognostic outcomes and to identify potential overlap in reported genes. Methods: EMBASE, MEDLINE, and PubMed databases were queried for relevant articles in English language without date restrictions. The initial search identified 1346 articles. After exclusions, 112 studies have been summarized. Cumulatively, 316 single-nucleotide polymorphisms (SNPs) were reported across prognostic outcomes (recurrence, progression, death) and characteristics (tumor stage, grade, size, age, risk group). There were considerable differences between studied outcomes in the context of genetic associations. The most commonly reported SNPs were located in OGG1, TP53, and MDM2. For outcomes with the highest number of reported associations (ie, recurrence and death), functional enrichment annotation yields different terms, potentially indicating separate biological mechanisms. Conclusions: Our study suggests that all UBC prognostic outcomes may have different biological origins with limited overlap. Further validation of these observations is essential to target a phenotype that could best predict patient outcome and advance current management practices

Vaccinia-related kinase 1 promotes hepatocellular carcinoma by controlling the levels of cell cycle regulators associated with G1/S transition

We identified the specific role of vaccinia-related kinase 1 (VRK1) in the progression of hepatocellular carcinoma (HCC) and evaluated its therapeutic and prognostic potential. VRK1 levels were significantly higher in HCC cell lines than a normal hepatic cell line, and were higher in HCC than non-tumor tissue. VRK1 knockdown inhibited the proliferation of SK-Hep1, SH-J1 and Hep3B cells; moreover, depletion of VRK1 suppressed HCC tumor growth in vivo. We also showed that VRK1 knockdown increased the number of G1 arrested cells by decreasing cyclin D1 and p-Rb while upregulating p21 and p27, and that VRK1 depletion downregulated phosphorylation of CREB, a transcription factor regulating CCND1. Additionally, we found that luteolin, a VRK1 inhibitor, suppressed HCC growth in vitro and in vivo, and that the aberrant VRK1 expression correlated with poor prognostic features of HCC. High levels of VRK1 were associated with shorter overall and disease-free survival and higher recurrence rates. Taken together, our findings suggest VRK1 may act as a tumor promoter by controlling the level of cell cycle regulators associated with G1/S transition and could potentially serve as a therapeutic target and/or prognostic biomarker for HCC.1110Ysciescopu