cAMP/PKA Pathways and S56 Phosphorylation Are Involved in AA/PGE2-Induced Increases in rNaV1.4 Current

Arachidonic acid (AA) and its metabolites are important second messengers for ion channel modulation. The effects of extracellular application of AA and its non-metabolized analogue on muscle rNa_V1.4 Na⁺ current has been studied, but little is known about the effects of intracellular application of AA on this channel isoform. Here, we report that intracellular application of AA significantly augmented the rNa_V1.4 current peak without modulating the steady-state activation and inactivation properties of the rNa_V1.4 channel. These results differed from the effects of extracellular application of AA on rNa_V1.4 current. The effects of intracellular AA were mimicked by prostaglandin E₂ but not eicosatetraynoic acid (ETYA), the non-metabolized analogue of AA, and were eliminated by treatment with cyclooxygenase inhibitors, flufenamic acid, or indomethacin. AA/PGE₂-induced activation of rNa_V1.4 channels was mimicked by a cAMP analogue (db-cAMP) and eliminated by a PKA inhibitor, PKAi. Furthermore, inhibition of EP2 and EP4 (PGE₂ receptors) with AH6809 and AH23848 reduced the intracellular AA/PGE₂-induced increase of rNa_V1.4 current. Two mutated channels, rNa_V1.4S56A and rNa_V1.4T21A, were designed to investigate the role of predicted phosphorylation sites in the AA/PGE₂–mediated regulation of rNa_V1.4 currents. In rNa_V1.4S56A, the effects of intracellular db-cAMP, AA, and PGE₂ were significantly reduced. The results of the present study suggest that intracellular AA augments rNa_V1.4 current by PGE₂/EP receptor-mediated activation of the cAMP/PKA pathway, and that the S56 residue on the channel protein is important for this process.     

Familial Clarification of Saucrosmylidae stat. nov. and New Saucrosmylids from Daohugou,China (Insecta,Neuroptera)

BackgoundSaucrosmylids are characterized by the typically large body size, complicated venation and diverse wing markings, which were only discovered in Middle Jurassic of Daohugou, Ningcheng county, Inner Mongolia, China.Conclusions/SignificanceThe intriguing group represents a particular lineage of Neuroptera in the Mesozoic Era. The familial status of Saucrosmylidae was firstly advanced that clarified the former incorrect citation and use of the family name. As an extinct clade, many species of the saucrosmylids were erected just based on a single fore- or hindwing, and it should be realized that providing more stable characters is necessary when describing new lacewing taxa just based on an isolated hindwing. It is vital for the systematics of Saucrosmylidae.     

Effect of Carotene and Lycopene on the Risk of Prostate Cancer: A Systematic Review and Dose-Response Meta-Analysis of Observational Studies

Background

Many epidemiologic studies have investigated the association between carotenoids intake and risk of Prostate cancer (PCa). However, results have been inconclusive.

Methods

We conducted a systematic review and dose-response meta-analysis of dietary intake or blood concentrations of carotenoids in relation to PCa risk. We summarized the data from 34 eligible studies (10 cohort, 11 nested case-control and 13 case-control studies) and estimated summary Risk Ratios (RRs) and 95% confidence intervals (CIs) using random-effects models.

Results

Neither dietary β-carotene intake nor its blood levels was associated with reduced PCa risk. Dietary α-carotene intake and lycopene consumption (both dietary intake and its blood levels) were all associated with reduced risk of PCa (RR for dietary α-carotene intake: 0.87, 95%CI: 0.76–0.99; RR for dietary lycopene intake: 0.86, 95%CI: 0.75–0.98; RR for blood lycopene levels: 0.81, 95%CI: 0.69–0.96). However, neither blood α-carotene levels nor blood lycopene levels could reduce the risk of advanced PCa. Dose-response analysis indicated that risk of PCa was reduced by 2% per 0.2mg/day (95%CI: 0.96–0.99) increment of dietary α-carotene intake or 3% per 1mg/day (95%CI: 0.94–0.99) increment of dietary lycopene intake.

Conclusions

α-carotene and lycopene, but not β-carotene, were inversely associated with the risk of PCa. However, both α-carotene and lycopene could not lower the risk of advanced PCa.     

Drug in Adhesive Patch of Zolmitriptan: Formulation and In vitro /In vivo Correlation

The objective of the present study was to develop transdermal patch for zolmitriptan, determine its in vivo absorption using the rabbit skin. Solvent evaporation technique prepared zolmitriptan patch was settled in two-chamber diffusion cell combined with excised rabbit abdomen skin for permeation study. A sufficient cumulative penetration amount of zolmitriptan (258.5 ± 26.9 μg/cm² in 24 h) was achieved by the formulation of 4% zolmitriptan, 10% Azone, and adhesive of DURO-TAK® 87–4098. Pharmacokinetic parameters were determined via i.v. and transdermal administrations using animal model of rabbit. The results revealed that the absolute bioavailability was about 63%. Zolmitriptan could be detected with drug level of 88 ± 51 ng/mL after transdermal administration of 15 min. The in vivo absorption curve obtained by deconvolution approach using WinNonlin® program was correlated well with the in vitro permeation curve, the correlation coefficient R is 0.84, and the result indicated that in vitro skin permeation experiments were useful to predict the in vivo performance. In addition, little skin irritation was found in the irritation study. As a conclusion, the optimized zolmitriptan transdermal patches could effectively deliver adequate drug into systemic circulation in short time without producing any irritation phenomenon and worth to be developed.KEY WORDS: chemical enhancer, drug-in-adhesive patch, in vitro/in vivo correlation, pharmacokinetic, zolmitriptan     

Outer Hair Cell Lateral Wall Structure Constrains the Mobility of Plasma Membrane Proteins

Nature’s fastest motors are the cochlear outer hair cells (OHCs). These sensory cells use a membrane protein, Slc26a5 (prestin), to generate mechanical force at high frequencies, which is essential for explaining the exquisite hearing sensitivity of mammalian ears. Previous studies suggest that Slc26a5 continuously diffuses within the membrane, but how can a freely moving motor protein effectively convey forces critical for hearing? To provide direct evidence in OHCs for freely moving Slc26a5 molecules, we created a knockin mouse where Slc26a5 is fused with YFP. These mice and four other strains expressing fluorescently labeled membrane proteins were used to examine their lateral diffusion in the OHC lateral wall. All five proteins showed minimal diffusion, but did move after pharmacological disruption of membrane-associated structures with a cholesterol-depleting agent and salicylate. Thus, our results demonstrate that OHC lateral wall structure constrains the mobility of plasma membrane proteins and that the integrity of such membrane-associated structures are critical for Slc26a5’s active and structural roles. The structural constraint of membrane proteins may exemplify convergent evolution of cellular motors across species. Our findings also suggest a possible mechanism for disorders of cholesterol metabolism with hearing loss such as Niemann-Pick Type C diseases.     

Arabidopsis COP1 SUPPRESSOR 2 Represses COP1 E3 Ubiquitin Ligase Activity through Their Coiled-Coil Domains Association

CONSTITUTIVE PHOTOMORPHOGENIC 1 (COP1) functions as an E3 ubiquitin ligase and mediates a variety of developmental processes in Arabidopsis by targeting a number of key regulators for ubiquitination and degradation. Here, we identify a novel COP1 interacting protein, COP1 SUPPRESSOR 2 (CSU2). Loss of function mutations in CSU2 suppress the constitutive photomorphogenic phenotype of cop1-6 in darkness. CSU2 directly interacts with COP1 via their coiled-coil domains and is recruited by COP1 into nuclear speckles in living plant cells. Furthermore, CSU2 inhibits COP1 E3 ubiquitin ligase activity in vitro, and represses COP1 mediated turnover of HY5 in cell-free extracts. We propose that in csu2 cop1-6 mutants, the lack of CSU2’s repression of COP1 allows the low level of COP1 to exhibit higher activity that is sufficient to prevent accumulation of HY5 in the dark, thus restoring the etiolated phenotype. In addition, CSU2 is required for primary root development under normal light growth condition.     

Identification of differentially methylated markers among cytogenetic risk groups of acute myeloid leukemia

Aberrant DNA methylation is known to occur in cancer, including hematological malignancies such as acute myeloid leukemia (AML). However, less is known about whether specific methylation profiles characterize specific subcategories of AML. We examined this issue by using comprehensive high-throughput array-based relative methylation analysis (CHARM) to compare methylation profiles among patients in different AML cytogenetic risk groups. We found distinct profiles in each group, with the high-risk group showing overall increased methylation compared with low- and mid-risk groups. The differentially methylated regions (DMRs) distinguishing cytogenetic risk groups of AML were enriched in the CpG island shores. Specific risk-group associated DMRs were located near genes previously known to play a role in AML or other malignancies, such as MN1, UHRF1, HOXB3, and HOXB4, as well as TRIM71, the function of which in cancer is not well characterized. These findings were verified by quantitative bisulfite pyrosequencing and by comparison with results available at the TCGA cancer genome browser. To explore the potential biological significance of the observed methylation changes, we correlated our findings with gene expression data available through the TCGA database. The results showed that decreased methylation at HOXB3 and HOXB4 was associated with increased gene expression of both HOXB genes specific to the mid-risk AML, while increased DNA methylation at DCC distinctive to the high-risk AML was associated with increased gene expression. Our results suggest that the differential impact of cytogenetic changes on AML prognosis may, in part, be mediated by changes in methylation.     

Immuno‐modification of enhancing stem cells targeting for myocardial repair

Despite the controversy in mechanism, rodent and clinical studies have demonstrated beneficial effects of stem/progenitor cell therapy after myocardial infarction (MI). In a rat ischaemic reperfusion MI model, we investigated the effects of immunomodification of CD 34⁺ cells on heart function and myocardial conduction. Bispecific antibody (BiAb), consisting of an anti‐myosin light chain antibody and anti‐CD45 antibody, injected intravenously was used to direct human CD34⁺ cells to injured myocardium. Results were compared to echocardiography guided intramyocardial (IM) injection of CD34⁺ cells and PBS injected intravenously. Treatment was administered 2 days post MI. Echocardiography was performed at 5 weeks and 3 months which demonstrated LV dilatation prevention and fractional shortening improvement in both the BiAb and IM injection approaches, with BiAb achieving better results. Histological analyses demonstrated a decrease in infarct size and increase in arteriogenesis in both BiAb and IM injection. Electrophysiological properties were studied 5 weeks after treatments by optical mapping. Conduction velocity (CV), action potential duration (APD) and rise time were significantly altered in the MI area. The BiAb treated group demonstrated a more normalized activation pattern of conduction and normalization of CV at shorter pacing cycle lengths. The ventricular tachycardia inducibility was lowest in the BiAb treatment group. Intravenous administration of BiAb offers an effective means of stem cell delivery for myocardial repair post‐acute MI. Such non‐invasive approach was shown to offer a distinct advantage to more invasive direct IM delivery.