The bloodless rat: a new model for macromolecular transport studies across lung endothelium

1982 ◽  
Vol 242 (5) ◽  
pp. H890-H899
Author(s):  
E. E. Schneeberger ◽  
B. A. Neary

To examine the effect of circulating proteins on the passage of intravenously injected native ferritin across pulmonary capillary endothelium, rats were exchange transfused with FC-43 fluorocarbon emulsion (FCE) under ether anesthesia. Protein concentration was reduced to less than 1.1 mg/ml by exchange transfusion, followed by FCE containing 15, 30, or 60 mg/ml of lyophilized rat serum protein (LRSP). Two minutes after ferritin injection lungs were prepared for ultrastructural morphometry. The diameter and numerical density of vesicles remained unchanged under all experimental conditions; however, at 0.6 mg/ml of circulating protein there was a 5- and 10-fold increase, respectively in percent vesicle (%VL) and basement membrane labeling (BML) by ferritin. This was reversible; at 60 mg/ml of circulating protein %VL and BML was indistinguishable from controls. Following a reduction of circulating protein to less than 1.1 mg/ml, the addition of 15 mg/ml LRSP reduced %VL but had no effect on BML. This suggests that in addition to shuttling vesicles there may be a second mechanism for the transport of ferritin, possibly involving transendothelial chains of vesicles.

2021 ◽  
pp. 247255522110138
Author(s):  
Önder Kartal ◽  
Fabio Andres ◽  
May Poh Lai ◽  
Rony Nehme ◽  
Kaspar Cottier

Surface-based biophysical methods for measuring binding kinetics of molecular interactions, such as surface plasmon resonance (SPR) or grating-coupled interferometry (GCI), are now well established and widely used in drug discovery. Increasing throughput is an often-cited need in the drug discovery process and this has been achieved with new instrument generations where multiple interactions are measured in parallel, shortening the total measurement times and enabling new application areas within the field. Here, we present the development of a novel technology called waveRAPID for a further—up to 10-fold—increase in throughput, consisting of an injection method using a single sample. Instead of sequentially injecting increasing analyte concentrations for constant durations, the analyte is injected at a single concentration in short pulses of increasing durations. A major advantage of the new method is its ability to determine kinetics from a single well of a microtiter plate, making it uniquely suitable for kinetic screening. We present the fundamentals of this approach using a small-molecule model system for experimental validation and comparing kinetic parameters to traditional methods. By varying experimental conditions, we furthermore assess the robustness of this new technique. Finally, we discuss its potential for improving hit quality and shortening cycle times in the areas of fragment screening, low-molecular-weight compound screening, and hit-to-lead optimization.


Plants ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 344
Author(s):  
Md Momtazur Rahman ◽  
Mikhail Vasiliev ◽  
Kamal Alameh

Manipulation of the LED illumination spectrum can enhance plant growth rate and development in grow tents. We report on the identification of the illumination spectrum required to significantly enhance the growth rate of sweet basil (Ocimum basilicum L.) plants in grow tent environments by controlling the LED wavebands illuminating the plants. Since the optimal illumination spectrum depends on the plant type, this work focuses on identifying the illumination spectrum that achieves significant basil biomass improvement compared to improvements reported in prior studies. To be able to optimize the illumination spectrum, several steps must be achieved, namely, understanding plant biology, conducting several trial-and-error experiments, iteratively refining experimental conditions, and undertaking accurate statistical analyses. In this study, basil plants are grown in three grow tents with three LED illumination treatments, namely, only white LED illumination (denoted W*), the combination of red (R) and blue (B) LED illumination (denoted BR*) (relative red (R) and blue (B) intensities are 84% and 16%, respectively) and a combination of red (R), blue (B) and far-red (F) LED illumination (denoted BRF*) (relative red (R), blue (B) and far-red (F) intensities are 79%, 11%, and 10%, respectively). The photosynthetic photon flux density (PPFD) was set at 155 µmol m−2 s−1 for all illumination treatments, and the photoperiod was 20 h per day. Experimental results show that a combination of blue (B), red (R), and far-red (F) LED illumination leads to a one-fold increase in the yield of a sweet basil plant in comparison with only white LED illumination (W*). On the other hand, the use of blue (B) and red (R) LED illumination results in a half-fold increase in plant yield. Understanding the effects of LED illumination spectrum on the growth of plant sweet basil plants through basic horticulture research enables farmers to significantly improve their production yield, thus food security and profitability.


Circulation ◽  
2000 ◽  
Vol 102 (16) ◽  
pp. 2011-2018 ◽  
Author(s):  
Stylianos E. Orfanos ◽  
Apostolos Armaganidis ◽  
Constantinos Glynos ◽  
Ekaterini Psevdi ◽  
Panagiotis Kaltsas ◽  
...  

Circulation ◽  
2007 ◽  
Vol 116 (suppl_16) ◽  
Author(s):  
Ramendra K Kundu ◽  
Ahmad Y Sheikh ◽  
Michael Y Ho ◽  
Hyung J Chun ◽  
Diem T Huynh ◽  
...  

Introduction: The APJ receptor and its ligand, apelin, comprise a homeostatic, cardio-regulatory pathway. Although cardiac apelin expression levels are altered in humans with cardiac failure, the cell type responsible for apelin expression and modulation in disease states remains unknown. Hypothesis: Apelin production is restricted to the endothelial compartment and is upregulated in states of cardiovascular stress. Methods: Transgenic apelin-LacZ reporter mice (SVJ background) were created by insertion of the nuclear localizing bacterial LacZ gene immediately downstream of the apelin promoter. Mice (n=12) were randomized to left anterior descending coronary artery (LAD) ligation, thoracic aortic constriction (TAC) or sham groups. Hearts were harvested 3 and 8 weeks post-TAC or LAD ligation, respectively. Localization of apelin expression was determined by Xgal staining. Endothelial phenotype of lacZ positive cells was confirmed by CD31 co-staining. Apelin expressing cells were quantified by histology. Apelin reporter results were confirmed by quantitating apelin expression in WT animals following LAD ligation (n=11) or sham (n=11) procedure by RT-PCR. Results: Extensive immunohistochemistry studies of heart tissue revealed lacZ reporter gene expression to be restricted to the coronary venous and capillary endothelium, with no expression by cardiomyocytes. Following both LAD ligation and TAC, the number of LacZ-apelin (+) endothelial cells significantly increased (p<0.002) in all chambers of the heart (Table ), with no evidence of apelin expression by other cell types. Evaluation of WT hearts by RT-PCR for the apelin gene confirmed the reporter gene findings with 1.3±0.3 fold increase (p<0.05) of apelin expression induced by LAD ligation compared to sham. Conclusions: Apelin is primary expressed by endothelial cells within the heart and is upregulated in response to myocardial stress. Apelin-LacZ Expressing Endothelial Cells are Increased Following Myocardial Injury


2003 ◽  
Vol 30 (12) ◽  
pp. 1233 ◽  
Author(s):  
Dev T. Britto ◽  
Herbert J. Kronzucker

The analysis of tracer efflux kinetics is fundamental to membrane transport studies, but requires the rigorous identification of subcellular tracer sources. We present a solution to this problem through the analysis of sharp increases in 13NH4+ efflux from roots of radiolabelled barley (Hordeum vulgare L.) seedlings, in response to a 100-fold increase in external [NH4+]. By comparing these trans-stimulation data with a mathematical model incorporating changes in subcellular NH4+ fluxes and pool sizes, we show that the cytosol of root cells is the origin of the tracer efflux. Our analysis provides new insight into the rapidly occurring events underlying compensatory flux regulation during transitions from one nutritional steady state to another, and confirms the validity of compartmental analysis by tracer efflux (CATE) in this important model system.


1995 ◽  
Vol 25 (1) ◽  
pp. 8-17 ◽  
Author(s):  
Stephen J. McCanny ◽  
Martin J. Lechowicz ◽  
Bill Shipley ◽  
William H. Hendershot

We used sand culture to examine the effects of aluminum (Al3+) on gas exchange and growth in 1+0 Picearubens Sarg. seedlings. Growth solutions were prepared to match the elemental concentrations of the soil solutions in a declining Abiesbalsamea (L.) Mill.–P. rubens stand. In our first experiment, we used a four-way factorial design to examine the interactions between Al, pH, nutrients, and CO2. All plants were nitrogen deficient. Photosynthetic rate and instantaneous water-use efficiency both declined at 250 μmol•L−1 Al, as did the root/shoot ratio of the seedlings. Plant weight, however, was not significantly affected by the 10-fold increase in Al solution concentration. These results were robust across a range of experimental conditions, suggesting that other aspects of soil acidification, such as NO3 fertilization in N deficient soils, could counteract the toxic effects of Al. In a second experiment, we examined the effects of subtoxic concentrations of Al at two levels of nutrient cation availability. Unlike in a previous study, cation fertilization affected neither gas exchange nor growth. While the decreases in root/shoot ratio and growth were expected at 250 μmol•L−1 Al, the peak in growth rates for intermediate concentrations of Al (25–100 μmol•L−1) was not. The latter result was interpreted in light of the positive effects that low concentrations of Al are known to have on other species.


1. The lungs of four species of bats, Phyllostomus hastatus (PH, mean body mass, 98 g), Pteropus lylei (PL,456 g), Pteropus alecto (PA, 667 g), and Pteropus poliocephalus (PP, 928 g) were analysed by morphometric methods. These data increase fivefold the range of body masses for which bat lung data are available, and allow more representative allometric equations to be formulated for bats. 2. Lung volume ranged from 4.9 cm 3 for PH to 39 cm 3 for PP. The volume density of the lung parenchyma (i.e. the volume proportion of the parenchyma in the lung) ranged from 94% in PP to 89% in PH. Of the components of the parenchyma, the alveoli composed 89% and the blood capillaries about 5% . 3. The surface area of the alveoli exceeded that of the blood—gas (tissue) barrier and that of the capillary endothelium whereas the surface area of the red blood cells as well as that of the capillary endothelium was greater than that of the tissue barrier. PH had the thinnest tissue barrier (0.1204 μm) and PP had the thickest (0.3033 μm). 4. The body mass specific volume of the lung, that of the volume of pulmonary capillary blood, the surface area of the blood-gas (tissue) barrier, the diffusing capacity of the tissue barrier, and the total morphometric pulmonary diffusing capacity in PH all substantially exceeded the corresponding values of the pteropid species (i.e. PL, PA and PP). This conforms with the smaller body mass and hence higher unit mass oxygen consumption of PH, a feature reflected in the functionally superior gas exchange performance of its lungs. 5. Morphometrically, the lungs of different species of bats exhibit remarkable differences which cannot always be correlated with body mass, mode of flight and phylogeny. Conclusive explanations of these pulmonary structural disparities in different species of bats must await additional physiological and flight biomechanical studies. 6. While the slope, the scaling factor (b), of the allometric equation fitted to bat lung volume data (b = 0.82) exceeds the value for flight Vo 2max , (b = 0.70), those for the surface area of the blood-gas (tissue) barrier (b = 0.74), the pulmonary capillary blood volume (b = 0.74), and the total morphometric lung diffusing capacity for oxygen (b = 0.69) all correspond closely to the Vo 2max , value. 7 Allometric comparisons of the morphometric pulmonary parameters of bats, birds and non-flying mammals reveal that superiority of the bat lung over that of the non-flying mammal. However, the bat parameters relative to those of non-flying mammals deteriorate towards the higher body size range, because of the generally steeper slopes of the equations for non-flying mammals. Allometric comparisons also reveal that small-size bats have, in general, better adapted lungs than birds of equivalent size but at the higher body mass scale, bats are generally inferior to birds.


2021 ◽  
Vol 118 (10) ◽  
pp. e2019845118
Author(s):  
Connor J. Thompson ◽  
Vinh H. Vu ◽  
Deborah E. Leckband ◽  
Daniel K. Schwartz

Cadherin transmembrane proteins are responsible for intercellular adhesion in all biological tissues and modulate tissue morphogenesis, cell motility, force transduction, and macromolecular transport. The protein-mediated adhesions consist of adhesive trans interactions and lateral cis interactions. Although theory suggests cooperativity between cis and trans bonds, direct experimental evidence of such cooperativity has not been demonstrated. Here, the use of superresolution microscopy, in conjunction with intermolecular single-molecule Förster resonance energy transfer, demonstrated the mutual cooperativity of cis and trans interactions. Results further demonstrate the consequent assembly of large intermembrane junctions, using a biomimetic lipid bilayer cell adhesion model. Notably, the presence of cis interactions resulted in a nearly 30-fold increase in trans-binding lifetimes between epithelial-cadherin extracellular domains. In turn, the presence of trans interactions increased the lifetime of cis bonds. Importantly, comparison of trans-binding lifetimes of small and large cadherin clusters suggests that this cooperativity is primarily due to allostery. The direct quantitative demonstration of strong mutual cooperativity between cis and trans interactions at intermembrane adhesions provides insights into the long-standing controversy of how weak cis and trans interactions act in concert to create strong macroscopic cell adhesions.


Pharmaceutics ◽  
2020 ◽  
Vol 12 (2) ◽  
pp. 162
Author(s):  
Anna Kwiecień ◽  
Jana Ruda-Kucerova ◽  
Kamil Kamiński ◽  
Zuzana Babinska ◽  
Iwona Popiołek ◽  
...  

The pharmacokinetic profile and tissue uptake of daidzein (DAI) was determined in rat serum and tissues (lungs, eyes, brain, heart, spleen, fat, liver, kidney, and testes) after intravenous and intraperitoneal administration of DAI in suspension or complexed with ethylenediamine-modified γ-cyclodextrin (GCD-EDA/DAI). The absolute and relative bioavailability of DAI suspended (20 mg/kg i.v. vs. 50 mg/kg i.p.) and complexed (0.54 mg/kg i.v. vs. 1.35 mg/kg i.p.) was determined. After i.p. administration, absorption of DAI complexed with GCD-EDA was more rapid (tmax = 15 min) than that of DAI in suspension (tmax = 45 min) with a ca. 3.6 times higher maximum concentration (Cmax = 615 vs. 173 ng/mL). The i.v. half-life of DAI was longer in GCD-EDA/DAI complex compared with DAI in suspension (t0.5 = 380 min vs. 230 min). The volume of distribution of DAI given i.v. in GCD-EDA/DAI complex was ca. 6 times larger than DAI in suspension (38.6 L/kg vs. 6.2 L/kg). Our data support the concept that the pharmacokinetics of DAI suspended in high doses are nonlinear. Increasing the intravenous dose 34 times resulted in a 5-fold increase in AUC. In turn, increasing the intraperitoneal dose 37 times resulted in a ca. 2-fold increase in AUC. The results of this study suggested that GCD-EDA complex may improve DAI bioavailability after i.p. administration. The absolute bioavailability of DAI in GCD-EDA inclusion complex was ca. 3 times greater (F = 82.4% vs. 28.2%), and the relative bioavailability was ca. 21 times higher than that of DAI in suspension, indicating the need to study DAI bioavailability after administration by routes other than intraperitoneal, e.g., orally, subcutaneously, or intramuscularly. The concentration of DAI released from GCD-EDA/DAI inclusion complex to all the rat tissues studied was higher than after administration of DAI in suspension. The concentration of DAI in brain and lungs was found to be almost 90 and 45 times higher, respectively, when administered in complex compared to the suspended DAI. Given the nonlinear relationship between DAI bioavailability and the dose released from the GCD-EDA complex, complexation of DAI may thus offer an effective approach to improve DAI delivery for treatment purposes, for example in mucopolysaccharidosis (MPS), allowing the reduction of ingested DAI doses.


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