Chimeric Investigations into the Diamide Binding Site on the Lepidopteran Ryanodine Receptor

Alterations to amino acid residues G4946 and I4790, associated with resistance to diamide insecticides, suggests a location of diamide interaction within the pVSD voltage sensor-like domain of the insect ryanodine receptor (RyR). To further delineate the interaction site(s), targeted alterations were made within the same pVSD region on the diamondback moth (Plutella xylostella) RyR channel. The editing of five amino acid positions to match those found in the diamide insensitive skeletal RyR1 of humans (hRyR1) in order to generate a human−Plutella chimeric construct showed that these alterations strongly reduce diamide efficacy when introduced in combination but cause only minor reductions when introduced individually. It is concluded that the sites of diamide interaction on insect RyRs lie proximal to the voltage sensor-like domain of the RyR and that the main site of interaction is at residues K4700, Y4701, I4790 and S4919 in the S1 to S4 transmembrane domains

Diamide insecticide resistance in transgenic Drosophila and Sf9-cells expressing a full-length diamondback moth ryanodine receptor carrying an I4790M mutation

BACKGROUNDResistance to diamide insecticides in Lepidoptera is known to be caused primarily by amino acid changes on the ryanodine receptor (RyR). Recently, two new target site mutations, G4946V and I4790M, have emerged in populations of diamondback moth, Plutella xylostella, as well as in other lepidopteran species, and both mutations have been shown empirically to decrease diamide efficacy. Here, we quantify the impact of the I4790M mutation on diamide activation of the receptor, as compared to alterations at the G4946 locus.RESULTSI4790M when introduced into P. xylostella RyR expressed in an insect-derived Sf9 cell line was found to mediate just a ten-fold reduction in chlorantraniliprole efficacy (compared to 104- and 146-fold reductions for the G4946E and G4946V variants, respectively), whilst in the field its presence is associated with a ≥150-fold reduction. I4790M-mediated resistance to flubendiamide was estimated to be >24-fold. When the entire coding sequence of P. xylostella RyR was integrated into Drosophila melanogaster, the I4790M variant conferred ~4.4-fold resistance to chlorantraniliprole and 22-fold resistance to flubendiamide in the 3rd instar larvae, confirming that it imparts only a moderate level of resistance to diamide insecticides. Although the I4790M substitution appears to bear no fitness costs in terms of the flies' reproductive capacity, when assessed in a noncompetitive environment, it does, however, have potentially major impacts on mobility at both the larval and adult stages.CONCLUSIONSI4790M imparts only a moderate level of resistance to diamide insecticides and potentially confers significant fitness costs to the insect

Free fatty acids link metabolism and regulation of the insulin-sensitizing fibroblast growth factor-21

OBJECTIVE—Fibroblast growth factor (FGF)-21 improves insulin sensitivity and lipid metabolism in obese or diabetic animal models, while human studies revealed increased FGF-21 levels in obesity and type 2 diabetes. Given that FGF-21 has been suggested to be a peroxisome proliferator–activator receptor (PPAR) –dependent regulator of fasting metabolism, we hypothesized that free fatty acids (FFAs), natural agonists of PPAR, might modify FGF-21 levels. RESEARCH DESIGN AND METHODS—The effect of fatty acids on FGF-21 was investigated in vitro in HepG2 cells. Within a randomized controlled trial, the effects of elevated FFAs were studied in 21 healthy subjects (13 women and 8 men). Within a clinical trial including 17 individuals, the effect of insulin was analyzed using an hyperinsulinemic-euglycemic clamp and the effect of PPAR activation was studied subsequently in a rosiglitazone treatment trial over 8 weeks. RESULTS—Oleate and linoleate increased FGF-21 expression and secretion in a PPAR-dependent fashion, as demonstrated by small-interfering RNA–induced PPAR knockdown, while palmitate had no effect. In vivo, lipid infusion induced an increase of circulating FGF-21 in humans, and a strong correlation between the change in FGF-21 levels and the change in FFAs was observed. An artificial hyperinsulinemia, which was induced to delineate the potential interaction between elevated FFAs and hyperinsulinemia, revealed that hyperinsulinemia also increased FGF-21 levels in vivo, while rosiglitazone treatment had no effect. CONCLUSIONS—The results presented here offer a mechanism explaining the induction of the metabolic regulator FGF-21 in the fasting situation but also in type 2 diabetes and obesity

Development and implementation of blood pressure screening and referral guidelines for German community pharmacists

Involvement of community pharmacists in the detection and control of hypertension improves patient care. However, current European or North‐American guidelines do not provide specific guidance how to implement collaboration between pharmacists and physicians, especially when and how to refer patients with undetected or uncontrolled hypertension to a physician. The German Society of Cardiology and the ABDA – Federal Union of German Associations of Pharmacists developed and tested referral recommendations for community pharmacists, embedded in two guideline worksheets. The project included a guideline‐directed blood pressure (BP) measurement and recommendations when patients should be referred to their physician. A “red flag” referral within 4 weeks was recommended when SBP was >140 mm Hg or DBP >90 mm Hg (for subjects 160 mm Hg or >90 mm Hg (≥80 years) in undetected individuals, or >130 mm Hg or >80 mm Hg (140 mm Hg or >80 mm Hg (≥65 years) in treated patients. BP was measured in 187 individuals (86 with known hypertension, mean [±SD] age 62 ± 15 years, 64% female, and 101 without known hypertension, 47 ± 16 years, 75% female) from 17 community pharmacies. In patients with hypertension, poorly controlled BP was detected in 55% (n = 47) and were referred. A total of 16/101 subjects without a history of hypertension were referred to their physician because of uncontrolled BP. Structured BP testing in pharmacies identified a significant number of subjects with undetected/undiagnosed hypertension and patients with poorly controlled BP. Community pharmacists could play a significant role in collaboration with physicians to improve the management of hypertension

Angiotensin II type 1-receptor activating antibodies in renal-allograft rejection (authors reply inN Engl J Med. 2005 May 12;352(19):2027-8)

BACKGROUND: Antibodies against HLA antigens cause refractory allograft rejection with vasculopathy in some, but not all, patients. METHODS: We studied 33 kidney-transplant recipients who had refractory vascular rejection. Thirteen had donor-specific anti-HLA antibodies, whereas 20 did not Malignant hypertension was present in 16 of the patients without anti-HLA antibodies, 4 of whom had seizures. The remaining 17 patients had no malignant hypertension. We hypothesized that activating antibodies targeting the angiotensin II type 1 (AT1) receptor might be involved. RESULTS: Activating IgG antibodies targeting the AT1 receptor were detected in serum from all 16 patients with malignant hypertension and without anti-HLA antibodies, but in no other patients. These receptor-activating antibodies are subclass IgG1 and IgG3 antibodies that bind to two different epitopes on the second extracellular loop of the AT1 receptor. Tissue factor expression was increased in renal-biopsy specimens from patients with these antibodies. In vitro stimulation of vascular cells with an AT1-receptor-activating antibody induced phosphorylation of ERK 1/2 kinase and increased the DNA binding activity of the transcription factors activator protein 1 (AP-1) and nuclear factor-κB. The AT1 antagonist losartan blocked agonistic AT1-receptor antibody-mediated effects, and passive antibody transfer induced vasculopathy and hypertension in a rat kidney-transplantation model. CONCLUSIONS: A non-HLA, AT1-receptor-mediated pathway may contribute to refractory vascular rejection, and affected patients might benefit from removal of AT 1-receptor antibodies or from pharmacologic blockade of AT 1 receptors

An altered plasma lipidome-phenome network characterizes heart failure with preserved ejection fraction

AIMS: Heart failure with preserved ejection fraction (HFpEF) is a multifactorial, multisystemic syndrome that involves alterations in lipid metabolism. This study aimed to test whether distinct plasma lipid profiles or lipid entities or both are associated with clinical and functional echocardiographic parameters in HFpEF. METHODS AND RESULTS: We examined the human plasma lipidome in HFpEF patients (n = 18) with left ventricular ejection fraction =50% and N-terminal pro-brain natriuretic peptide (NT-proBNP) >125 pg/mL and control subjects (n = 12) using mass spectrometry-based shotgun lipidomics. The cohort included 8 women and 22 men with average age of 67.8 ± 8.6 SD. The control and disease groups were not significantly different with respect to age, body mass index, systolic and diastolic blood pressure, and waist-to-hip ratio. The disease group experienced more fatigue (P < 0.001), had more often coronary artery disease (P = 0.04), and received more medications (beta-blockers, P < 0.001). The disease group had significantly different levels of HFpEF-relevant parameters, including NT-proBNP (P < 0.001), left ventricular mass index (P = 0.005), left atrial volume index (P = 0.001), and left ventricular filling index (P < 0.001), and lower left ventricular end-diastolic diameter (P = 0.014), with no difference in left ventricular ejection fraction. Significant differences in lipid profiles between HFpEF patients and controls could not be detected, including no significant differences in abundance of circulating lipids binned by carbon chain length or by double bonds, nor at the level of individual lipid species. However, there was a striking correlation between selected lipids with smoking status that was independent of disease status, as well as between specific lipids and hyperlipidaemia [with corresponding significance of either false discovery rate (FDR) <0.1 or FDR < 0.01]. In an exploratory network analysis of correlations, we observed significantly stronger correlations within the HFpEF group between individual lipids from the cholesterol ester and phosphatidylcholine (PC) classes and clinical/echocardiographic parameters such as left atrial volume index, left ventricular end-diastolic diameters, and heart rate (FDR < 0.1). In contrast, the control group showed significantly stronger negative correlations (FDR < 0.1) between individual species from the PC and sphingomyelin classes and left ventricular mass index or systolic blood pressure. CONCLUSIONS: We did not find significant direct associations between plasma lipidomic parameters and HFpEF and therefore could not conclude that any specific lipids are biomarkers of HFpEF. The validation in larger cohort is needed to confidently conclude the absence of first-order associations

Search for transient optical counterparts to high-energy IceCube neutrinos with Pan-STARRS1

In order to identify the sources of the observed diffuse high-energy neutrino flux, it is crucial to discover their electromagnetic counterparts. IceCube began releasing alerts for single high-energy ($E > 60$ TeV) neutrino detections with sky localisation regions of order 1 deg radius in 2016. We used Pan-STARRS1 to follow-up five of these alerts during 2016-2017 to search for any optical transients that may be related to the neutrinos. Typically 10-20 faint ($m < 22.5$ mag) extragalactic transients are found within the Pan-STARRS1 footprints and are generally consistent with being unrelated field supernovae (SNe) and AGN. We looked for unusual properties of the detected transients, such as temporal coincidence of explosion epoch with the IceCube timestamp. We found only one transient that had properties worthy of a specific follow-up. In the Pan-STARRS1 imaging for IceCube-160427A (probability to be of astrophysical origin of $\sim$50 %), we found a SN PS16cgx, located at 10.0' from the nominal IceCube direction. Spectroscopic observations of PS16cgx showed that it was an H-poor SN at z = 0.2895. The spectra and light curve resemble some high-energy Type Ic SNe, raising the possibility of a jet driven SN with an explosion epoch temporally coincident with the neutrino detection. However, distinguishing Type Ia and Type Ic SNe at this redshift is notoriously difficult. Based on all available data we conclude that the transient is more likely to be a Type Ia with relatively weak SiII absorption and a fairly normal rest-frame r-band light curve. If, as predicted, there is no high-energy neutrino emission from Type Ia SNe, then PS16cgx must be a random coincidence, and unrelated to the IceCube-160427A. We find no other plausible optical transient for any of the five IceCube events observed down to a 5$\sigma$ limiting magnitude of $m \sim 22$ mag, between 1 day and 25 days after detection.Comment: 20 pages, 6 figures, accepted to A&