Characterization of the Ac/Ds behaviour in transgenic tomato plants using plasmid rescue

We describe the use of plasmid rescue to facilitate studies on the behaviour of Ds and Ac elements in transgenic tomato plants. The rescue of Ds elements relies on the presence of a plasmid origin of replication and a marker gene selective in Escherichia coli within the element. The position within the genome of modified Ds elements, rescued both before and after transposition, is assigned to the RFLP map of tomato. Alternatively to the rescue of Ds elements equipped with plasmid sequences, Ac elements are rescued by virtue of plasmid sequences flanking the element. In this way, the consequences of the presence of an (active) Ac element on the DNA structure at the original site can be studied in detail. Analysis of a library of Ac elements, rescued from the genome of a primary transformant, shows that Ac elements are, infrequently, involved in the formation of deletions. In one case the deletion refers to a 174 bp genomic DNA sequence immediately flanking Ac. In another case, a 1878 bp internal Ac sequence is deleted

The endothelial glycocalyx prefers albumin for evoking shear stress-induced, nitric oxide-mediated coronary dilatation

Background: Shear stress induces coronary dilatation via production of nitric oxide ( NO). This should involve the endothelial glycocalyx ( EG). A greater effect was expected of albumin versus hydroxyethyl starch ( HES) perfusion, because albumin seals coronary leaks more effectively than HES in an EG-dependent way. Methods: Isolated hearts ( guinea pigs) were perfused at constant pressure with Krebs-Henseleit buffer augmented with 1/3 volume 5% human albumin or 6% HES ( 200/0.5 or 450/0.7). Coronary flow was also determined after EG digestion ( heparinase) and with nitro-L-arginine ( NO-L-Ag). Results: Coronary flow ( 9.50 +/- 1.09, 5.10 +/- 0.49, 4.87 +/- 1.19 and 4.15 +/- 0.09 ml/ min/ g for `albumin', `HES 200', `HES 450' and `control', respectively, n = 5-6) did not correlate with perfusate viscosity ( 0.83, 1.02, 1.24 and 0.77 cP, respectively). NO-L-Ag and heparinase diminished dilatation by albumin, but not additively. Alone NO-L-Ag suppressed coronary flow during infusion of HES 450. Electron microscopy revealed a coronary EG of 300 nm, reduced to 20 nm after heparinase. Cultured endothelial cells possessed an EG of 20 nm to begin with. Conclusions: Albumin induces greater endothelial shear stress than HES, despite lower viscosity, provided the EG contains negative groups. HES 450 causes some NO-mediated dilatation via even a rudimentary EG. Cultured endothelial cells express only a rudimentary glycocalyx, limiting their usefulness as a model system. Copyright (c) 2007 S. Karger AG, Basel

Web Search Query Volume Correlates With Prescription Volumes of Antidepressants and Antipsychotics in the Netherlands and United Kingdom: An Explorative Study

BACKGROUND: The significant increase in Internet availability has resulted in a rise in search queries on health-related topics. Previous research has demonstrated the potential for analyzing web search query volume for nonpsychotropic prescription drugs, while studies on psychotropic drugs remain scarce. The aims of this study were to expand upon this scarce knowledge by investigating the relationship between web search query volumes and prescription volumes of antidepressants and antipsychotics in the United Kingdom and the Netherlands and to gain insight in topics of concern, such as withdrawal symptoms and discontinuation. METHODS: Data were obtained for the United Kingdom and the Netherlands from January 2010 until January 2021. Prescription volume data for 5 antidepressants (paroxetine, fluoxetine, sertraline, citalopram, venlafaxine) and 5 antipsychotics (quetiapine, olanzapine, clozapine, aripiprazole, and risperidone) were obtained. Web search query volumes and data on related search queries of these substances were acquired from Google Trends. Descriptive statistics and Pearson correlation analyses were performed. RESULTS: A strong, positive, and statistically significant correlation between web search query volume and prescription volume was observed for most included substances in both the Netherlands and the United Kingdom. The search queries related to the included antidepressants and antipsychotics indicate important topics of concern for specific substances, such as withdrawal symptoms and discontinuation. CONCLUSIONS: Web search data from Google Trends could potentially be used as a proxy for prescribing trends of antidepressants and antipsychotics and to gain insight in topics of concern of users of these substances. These findings highlight the importance of providing reliable patient information, particularly regarding adverse effects, withdrawal, and discontinuation

Charge modification of the endothelial surface layer modulates the permeability barrier of isolated rat mesenteric small arteries

We hypothesized that modulation of the effective charge density of the endothelial surface layer ( ESL) results in altered arterial barrier properties to transport of anionic solutes. Rat mesenteric small arteries ( diameter similar to 190 mu m) were isolated, cannulated, perfused, and superfused with MOPS- buffered physiological salt solutions. MOPS- solutions were of normal ionic strength ( 162 mM, MOPS), low ionic strength ( 81 mM, LO- MOPS), or high ionic strength ( 323 mM, HI- MOPS), to modulate ESL charge density ( normal, high, or low ESL charge, respectively). Osmolarity of MOPS, LO- MOPS, and HI- MOPS was kept constant at 297 mosmol/ l, using additional glucose when necessary. Perfusate solutions were supplemented with 1% BSA. Arteries were cannulated with a double-barreled theta- pipet on the inlet side and a regular pipet on the outlet side. fter infusion of FITC- labeled dextran of 50 kDa ( FITC- Delta 50) and the endothelial membrane dye 1,1'- dioctadecyl- 3,3,3', 3'- tetramethylindocarbocyanine perchlorate, the dynamics of arterial dye filling were determined with confocal microscopy. ESL thickness, as determined from the initial exclusion zone for FITC- Delta 50 on the luminal endothelial surface, was 6.3 +/- 1.4 mu m for LO- MOPS, 2.7 +/- 1.0 mu m for MOPS, and 1.1 +/- 1.3 mu m for HI- MOPS. At low ionic strength, FITC- Delta 50 permeated into the ESL with a total ESL permeation time ( tau(ESL)) of 26 min, and at normal ionic strength with a tau(ESL) of 20 min. No apparent exclusion of FITC-Delta 50 from the ESL could be observed at high ionic strength. In conclusion, we demonstrate that the modulation of solvent ionic strength influences the thickness and barrier properties of the ES

Localization of the permeability barrier to solutes in isolated arteries by confocal microscopy

Endothelial cells are covered by a surface layer of membrane-associated proteoglycans, glycosaminoglycans, glycoproteins, glycolipids, and associated plasma proteins. This layer may limit transendothelial solute transport. We determined dimension and transport properties of this endothelial surface layer (ESL) in isolated arteries. Rat mesenteric small arteries (diameter similar to 150 mum) were isolated and cannulated with a double-barreled theta-pipette on the inlet side and a regular pipette on the outlet side. Dynamics and localization of intra-arterial fluorescence by FITC-labeled dextrans (FITC-Deltas) and the endothelial membrane dye DiI were determined with confocal microscopy. Large FITC-Delta (148 kDa) filled a core volume inside the arteries within 1 min but was excluded from a 2.6 +/- 0.5-mum-wide region on the luminal side of the endothelium during 30 min of dye perfusion. Medium FITC-Delta (50.7 kDa) slowly penetrated this ESL within 30 min but did not permeate into the arterial wall. Small FITC-Delta (4.4 kDa) quickly passed the ESL and accumulated in the arterial wall. Prolonged luminal fluorochrome illumination with a bright mercury lamp destroyed the similar to3-mum exclusion zone for FITC-Delta148 within a few minutes. This study demonstrates the presence of a thick ESL that contributes to the permeability barrier to solutes. The layer is sensitive to phototoxic stress, and its damage could form an early event in atherosclerosi