Cholesterol Down-Regulates BK Channels Stably Expressed in HEK 293 Cells

Cholesterol is one of the major lipid components of the plasma membrane in mammalian cells and is involved in the regulation of a number of ion channels. The present study investigates how large conductance Ca²⁺-activated K⁺ (BK) channels are regulated by membrane cholesterol in BK-HEK 293 cells expressing both the α-subunit hKCa1.1 and the auxiliary β1-subunit or in hKCa1.1-HEK 293 cells expressing only the α-subunit hKCa1.1 using approaches of electrophysiology, molecular biology, and immunocytochemistry. Membrane cholesterol was depleted in these cells with methyl-β-cyclodextrin (MβCD), and enriched with cholesterol-saturated MβCD (MβCD-cholesterol) or low-density lipoprotein (LDL). We found that BK current density was decreased by cholesterol enrichment in BK-HEK 293 cells, with a reduced expression of KCa1.1 protein, but not the β1-subunit protein. This effect was fully countered by the proteasome inhibitor lactacystin or the lysosome function inhibitor bafilomycin A1. Interestingly, in hKCa1.1-HEK 293 cells, the current density was not affected by cholesterol enrichment, but directly decreased by MβCD, suggesting that the down-regulation of BK channels by cholesterol depends on the auxiliary β1-subunit. The reduced KCa1.1 channel protein expression was also observed in cultured human coronary artery smooth muscle cells with cholesterol enrichment using MβCD-cholesterol or LDL. These results demonstrate the novel information that cholesterol down-regulates BK channels by reducing KCa1.1 protein expression via increasing the channel protein degradation, and the effect is dependent on the auxiliary β1-subunit.     

Triterpenoid saponins from Eryngium yuccifolium ‘Kershaw Blue’

Phytochemical investigation on the root of Eryngium yuccifolium ‘Kershaw Blue’ resulted in the isolation and identification of two new polyhydroxyoleanene saponins, named eryngioside M and eryngioside N, together with 15 known triterpenoid saponins eryngiosides A-L, 21β-angeloyloxy-3β-[β-d-glucopyranosyl-(1 → 2)]-[β-d-xylopyranosyl-(1 → 3)]-β-d-glucuronopyranosyloxyolean-12-ene-15α,16α,22α,28-tetrol, saniculasaponin III, and saniculasaponin II. Their structures were established by extensive spectroscopic and chemical analyses. Eryngioside M and saniculasaponin II showed week cytotoxicity against human non-small cell lung tumor cells (A549) with GI₅₀ values of 37.5 ± 1.59 μM and 35.5 ± 1.11 μM, respectively.     

Genome-Wide Association Analysis with Gray Matter Volume as a Quantitative Phenotype in First-Episode Treatment-Na?ve Patients with Schizophrenia

Reduced Gray matter (GM) volume is a core feature of schizophrenia. Mapping genes that is associated with the heritable disease-related phenotypes may be conducive to elucidate the pathogenesis of schizophrenia. This study aims to identify the common genetic variants that underlie the deficits of GM volume in schizophrenia. High-resolution T1 images and whole genome genotyping data were obtained from 74 first-episode treatment-naïve patients with schizophrenia and 51 healthy controls in the Mental Health Centre of the West China Hospital, Sichuan University. All participants were scanned using a 3T MR imaging system and were genotyped using the HumanHap660 Bead Array. Reduced GM volumes in three brain areas including left hOC3v in the collateral sulcus of visual cortex (hOC3vL), left cerebellar vermis lobule 10 (vermisL10) and right cerebellar vermis lobule 10 (vermisR10) were found in patients with schizophrenia. There was a group by genotype interaction when genotypes from genome-wide scan were subsequently considered in the case-control analyses. SNPs from three genes or chromosomal regions (TBXAS1, PIK3C2G and HS3ST5) were identified to predict the changes of GM volume in hOC3vL, vermisL10 and vermisR10. These results also highlighted the usefulness of endophenotype in exploring the pathogenesis of neuropsychiatric diseases such as schizophrenia although further independent replication studies are needed in the future.     

Genome-Scale Screening of Drug-Target Associations Relevant to Ki Using a Chemogenomics Approach

The identification of interactions between drugs and target proteins plays a key role in genomic drug discovery. In the present study, the quantitative binding affinities of drug-target pairs are differentiated as a measurement to define whether a drug interacts with a protein or not, and then a chemogenomics framework using an unbiased set of general integrated features and random forest (RF) is employed to construct a predictive model which can accurately classify drug-target pairs. The predictability of the model is further investigated and validated by several independent validation sets. The built model is used to predict drug-target associations, some of which were confirmed by comparing experimental data from public biological resources. A drug-target interaction network with high confidence drug-target pairs was also reconstructed. This network provides further insight for the action of drugs and targets. Finally, a web-based server called PreDPI-K_i was developed to predict drug-target interactions for drug discovery. In addition to providing a high-confidence list of drug-target associations for subsequent experimental investigation guidance, these results also contribute to the understanding of drug-target interactions. We can also see that quantitative information of drug-target associations could greatly promote the development of more accurate models. The PreDPI-K_i server is freely available via: http://sdd.whu.edu.cn/dpiki.     

Autophagic Protein Beclin 1 Serves as an Independent Positive Prognostic Biomarker for Non-Small Cell Lung Cancer

Beclin 1, a key regulator of autophagy, has been found to be aberrantly expressed in a variety of human malignancies. Herein, we employed immunohistochemistry (IHC) to detect the protein expression of Beclin 1 in non-small cell lung cancer (NSCLC) and paired normal adjacent lung tissues, and analyzed its clinicopathological/prognostic significance in NSCLC. Receiver operating characteristic (ROC) curve analysis was utilized to determine a cutoff point (>2 VS. ≤2) for Beclin 1 expression in a training set (n = 105). For validation, the ROC-derived cutoff value was subjected to analysis of the association of Beclin 1 with patients’ clinical characteristics and outcome in a testing set (n = 111) and the overall patient cohort (n = 216). Our data showed that Beclin 1 was significantly lower in NSCLC tissues compared with the adjacent normal tissues, negatively associating with tumor recurrence rate (65.8% VS 32.3%; p < 0.001). In the testing set and the overall patient cohort, low expression of Beclin 1 showed significantly inferior overall survival (OS) (p < 0.001) and progression-free survival (PFS) (p < 0.001) compared to high expression of Beclin 1. In the testing set and the overall patient cohort, the median duration of OS for patients with high and low expression of Beclin 1 was 108 VS. 24.5 months (p < 0.001) and 108 VS. 28 months (p < 0.001), respectively. Furthermore, low expression of Beclin 1 was also a poor prognostic factor within each stage of NSCLC patients. Multivariate analysis identified that Beclin 1 was an independent prognostic factor for NSCLC. Our findings in the present study provided evidence that Beclin 1 may thus emerge as an independent prognostic biomarker in this tumor entity in the future.     

A Field Evaluation of an External and Neutrally Buoyant Acoustic Transmitter for Juvenile Salmon: Implications for Estimating Hydroturbine Passage Survival

Turbine-passed fish are exposed to rapid decreases in pressure which can cause barotrauma. The presence of an implanted telemetry tag increases the likelihood of injury or death from exposure to pressure changes, thus potentially biasing studies evaluating survival of turbine-passed fish. Therefore, a neutrally buoyant externally attached tag was developed to eliminate this bias in turbine passage studies. This new tag was designed not to add excess mass in water or take up space in the coelom, having an effective tag burden of zero with the goal of reducing pressure related biases to turbine survival studies. To determine if this new tag affects fish performance or susceptibility to predation, it was evaluated in the field relative to internally implanted acoustic transmitters (JSATS; Juvenile Salmon Acoustic Telemetry System) used widely for survival studies of juvenile salmonids. Survival and travel time through the study reach was compared between fish with either tag type in an area of high predation in the Snake and Columbia rivers, Washington. An additional group of fish affixed with neutrally-buoyant dummy external tags were implanted with passive integrated transponder (PIT) tags and recovered further downstream to assess external tag retention and injury. There were no significant differences in survival to the first detection site, 12 river kilometers (rkm) downstream of release. Travel times were also similar between groups. Conversely, externally-tagged fish had reduced survival (or elevated tag loss) to the second detection site, 65 rkm downstream. In addition, the retention study revealed that tag loss was first observed in fish recaptured approximately 9 days after release. Results suggest that this new tag may be viable for short term (<8 days) single-dam turbine-passage studies and under these situations, may alleviate the turbine passage-related bias encountered when using internal tags, however further research is needed to confirm this.     

Correction of Diabetic Erectile Dysfunction with Adipose Derived Stem Cells Modified with the Vascular Endothelial Growth Factor Gene in a Rodent Diabetic Model

The aim of this study was to determine whether adipose derived stem cells (ADSCs) expressing vascular endothelial growth factor (VEGF) gene can improve endothelial function, recover the impaired VEGF signaling pathway and enhance smooth muscle contents in a rat diabetic erectile dysfunction (DED) model. DED rats were induced via intraperitoneal injection of streptozotocin (40 mg/kg), and then screened by apomorphine (100 µg/kg). Five groups were used (n = 12/group)–Group 1 (G1): intracavernous injection of lentivirus-VEGF; G2: ADSCs injection; G3: VEGF-expressing ADSCs injection; G4: Phosphate buffered saline injection; G1–G4 were DED rats; G5: normal rats. The mean arterial pressure (MAP) and intracavernosal pressure (ICP) were measured at days 7 and 28 after the injections. The components of the VEGF system, endothelial, smooth muscle, pericytes markers in cavernoursal tissue were assessed. On day 28 after injection, the group with intracavernosum injection of ADSCs expressing VEGF displayed more efficiently and significantly raised ICP and ICP/MAP (p<0.01) than those with ADSCs or lentivirus-VEGF injection. Western blot and immunofluorescent analysis demonstrated that improved erectile function by ADSCs-VEGF was associated with increased expression of endothelial markers (VEGF, VEGF R1, VEGF R2, eNOS, CD31 and vWF), smooth muscle markers (a-actin and smoothelin), and pericyte markers (CD146 and NG2). ADSCs expressing VEGF produced a therapeutic effect and restored erectile function in diabetic rats by enhancing VEGF-stimulated endothelial function and increasing the contents of smooth muscle and pericytes.     

Integrated Meta-QTL and Genome-Wide Association Study Analyses Reveal Candidate Genes for Maize Yield

Journal of Plant Growth Regulation - Crop yield is of a quantitative nature. Two complementary approaches linkage mapping and genome-wide association study (GWAS) have been employed for the...